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Int. J. Mol. Sci., Volume 18, Issue 5 (May 2017)

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Cover Story The figure shows the molecular basis and principles of TSPO-PET imaging. The 18 kDa Translocator [...] Read more.
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Open AccessArticle Interleukin 17A Promotes Lymphocytes Adhesion and Induces CCL2 and CXCL1 Release from Brain Endothelial Cells
Int. J. Mol. Sci. 2017, 18(5), 1000; doi:10.3390/ijms18051000
Received: 21 February 2017 / Revised: 31 March 2017 / Accepted: 1 May 2017 / Published: 8 May 2017
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Abstract
The nature of the interaction between Th17 cells and the blood–brain barrier (BBB) is critical for the development of autoimmune inflammation in the central nervous system (CNS). Tumor necrosis factor alpha (TNF-α) or interleukin 17 (IL-17) stimulation is known to enhance the adherence
[...] Read more.
The nature of the interaction between Th17 cells and the blood–brain barrier (BBB) is critical for the development of autoimmune inflammation in the central nervous system (CNS). Tumor necrosis factor alpha (TNF-α) or interleukin 17 (IL-17) stimulation is known to enhance the adherence of Th17 cells to the brain endothelium. The brain endothelial cells (bEnd.3) express Vascular cell adhesion molecule 1 (VCAM-1), the receptor responsible for inflammatory cell adhesion, which binds very late antigen 4 (VLA-4) on migrating effector lymphocytes at the early stage of brain inflammation. The present study examines the effect of the pro-inflammatory cytokines TNF-α and IL-17 on the adherence of Th17 cells to bEnd.3. The bEnd.3 cells were found to increase production of CCL2 and CXCL1 after stimulation by pro-inflammatory cytokines, while CCL2, CCL5, CCL20 and IL17 induced Th17 cell migration through a bEnd.3 monolayer. This observation may suggest potential therapeutic targets for the prevention of autoimmune neuroinflammation development in the CNS. Full article
(This article belongs to the Special Issue Cell-cell Interactions in Blood Vessels)
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Open AccessArticle A Long-Term Treatment with Arachidonyl-2′-Chloroethylamide Combined with Valproate Increases Neurogenesis in a Mouse Pilocarpine Model of Epilepsy
Int. J. Mol. Sci. 2017, 18(5), 900; doi:10.3390/ijms18050900
Received: 19 February 2017 / Revised: 12 April 2017 / Accepted: 19 April 2017 / Published: 25 April 2017
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Abstract
Rational polytherapy in the treatment of refractory epilepsy has been the main therapeutic modality for several years. In treatment with two or more antiepileptic drugs (AEDs), it is of particular importance that AEDs be selected based on their high anticonvulsant properties, minimal side
[...] Read more.
Rational polytherapy in the treatment of refractory epilepsy has been the main therapeutic modality for several years. In treatment with two or more antiepileptic drugs (AEDs), it is of particular importance that AEDs be selected based on their high anticonvulsant properties, minimal side effects, and impact on the formation of new neurons. The aim of the study was to conduct an in vivo evaluation of the relationship between treatments with synthetic cannabinoid arachidonyl-2′-chloroethylamide (ACEA) alone or in combination with valproic acid (VPA) and hippocampal neurogenesis in a mouse pilocarpine model of epilepsy. All studies were performed on adolescent male CB57/BL mice with using the following drugs: VPA (10 mg/kg), ACEA (10 mg/kg), phenylmethylsulfonyl fluoride (PMSF—a substance protecting ACEA against degradation by fatty acid hydrolase, 30 mg/kg), pilocarpine (PILO, a single dose of 290 mg/kg) and methylscopolamine (30 min before PILO to stop peripheral cholinergic effects of pilocarpine, 1 mg/kg). We evaluated the process of neurogenesis after a 10-day treatment with ACEA and VPA, alone and in combination. We observed a decrease of neurogenesis in the PILO control group as compared to the healthy control mice. Furthermore, ACEA + PMSF alone and in combination with VPA significantly increased neurogenesis compared to the PILO control group. In contrast, VPA 10-day treatment had no impact on the level of neurons in comparison to the PILO control group. The combination of ACEA, PMSF and VPA considerably stimulated the process of creating new cells, particularly neurons, while chronic administration of VPA itself had no influence on neurogenesis in the mouse pilocarpine model of epilepsy. The obtained results enabled an in vivo evaluation of neurogenesis after treatment with antiepileptic drugs in an experimental model of epilepsy. Full article
(This article belongs to the Special Issue Cannabinoid Signaling in Nervous System)
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Open AccessArticle Molecular Pathways Involved in the Amelioration of Myocardial Injury in Diabetic Rats by Kaempferol
Int. J. Mol. Sci. 2017, 18(5), 1001; doi:10.3390/ijms18051001
Received: 20 March 2017 / Revised: 24 April 2017 / Accepted: 2 May 2017 / Published: 15 May 2017
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Abstract
There is growing evidence that chronic hyperglycemia leads to the formation of advanced glycation end products (AGEs) which exerts its effect via interaction with the receptor for advanced glycation end products (RAGE). AGE-RAGE activation results in oxidative stress and inflammation. It is well
[...] Read more.
There is growing evidence that chronic hyperglycemia leads to the formation of advanced glycation end products (AGEs) which exerts its effect via interaction with the receptor for advanced glycation end products (RAGE). AGE-RAGE activation results in oxidative stress and inflammation. It is well known that this mechanism is involved in the pathogenesis of cardiovascular disease in diabetes. Kaempferol, a dietary flavonoid, is known to possess antioxidant, anti-apoptotic, and anti-inflammatory activities. However, little is known about the effect of kaempferol on myocardial ischemia-reperfusion (IR) injury in diabetic rats. Diabetes was induced in male albino Wistar rats using streptozotocin (70 mg/kg; i.p.), and rats with glucose level >250 mg/dL were considered as diabetic. Diabetic rats were treated with vehicle (2 mL/kg; i.p.) and kaempferol (20 mg/kg; i.p.) daily for a period of 28 days and on the 28th day, ischemia was produced by one-stage ligation of the left anterior descending coronary artery for 45 min followed by reperfusion for 60 min. After completion of surgery, rats were sacrificed and the heart tissue was processed for biochemical, morphological, and molecular studies. Kaempferol pretreatment significantly reduced hyperglycemia, maintained hemodynamic function, suppressed AGE-RAGE axis activation, normalized oxidative stress, and preserved morphological alterations. In addition, there was decreased level of inflammatory markers (tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and NF-κB), inhibition of active c-Jun N-terminal kinase (JNK) and p38 proteins, and activation of Extracellular signal regulated kinase 1/2 (ERK1/2) a prosurvival kinase. Furthermore, it also attenuated apoptosis by reducing the expression of pro-apoptotic proteins (Bax and Caspase-3), Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) positive cells, and increasing the level of anti-apoptotic protein (Bcl-2). In conclusion, kaempferol attenuated myocardial ischemia-reperfusion injury in diabetic rats by reducing AGE-RAGE/ mitogen activated protein kinase (MAPK) induced oxidative stress and inflammation. Full article
(This article belongs to the Special Issue Correlation between Nutrition, Oxidative Stress and Disease)
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Open AccessCommunication An Additive Effect of Promoting Thermogenic Gene Expression in Mice Adipose-Derived Stromal Vascular Cells by Combination of Rosiglitazone and CL316,243
Int. J. Mol. Sci. 2017, 18(5), 1002; doi:10.3390/ijms18051002
Received: 29 March 2017 / Revised: 2 May 2017 / Accepted: 3 May 2017 / Published: 8 May 2017
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Abstract
It is well-documented that CL316,243 (a β3 agonist) or rosiglitazone (a PPARγ agonist) can induce white adipocyte populations to brown-like adipocytes, thus increasing energy consumption and combating obesity. However, whether there is a combined effect remains unknown. In the present study, stromal vascular
[...] Read more.
It is well-documented that CL316,243 (a β3 agonist) or rosiglitazone (a PPARγ agonist) can induce white adipocyte populations to brown-like adipocytes, thus increasing energy consumption and combating obesity. However, whether there is a combined effect remains unknown. In the present study, stromal vascular cells of inguinal white adipose tissue (iWAT-SVCs for short) from mice were cultured and induced into browning by CL316,243, rosiglitazone, or both. Results showed that a combination of CL316,243 and rosiglitazone significantly upregulated the expression of the core thermogenic gene Ucp1 as well as genes related with mitochondrial function (Cidea, Cox5b, Cox7a1, Cox8b, and Cycs), compared with the treatment of CL316,243 or rosiglitazone alone. Moreover, co-treatment with rosiglitazone could reverse the downregulation of Adiponectin resulting from CL316,243 stimuli alone. Taken together, a combination of rosiglitazone and CL316,243 can produce an additive effect of promoting thermogenic gene expression and an improvement of insulin sensitivity in mouse iWAT-SVCs. Full article
(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)
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Open AccessArticle 188Re-Liposome Can Induce Mitochondrial Autophagy and Reverse Drug Resistance for Ovarian Cancer: From Bench Evidence to Preliminary Clinical Proof-of-Concept
Int. J. Mol. Sci. 2017, 18(5), 903; doi:10.3390/ijms18050903
Received: 6 March 2017 / Revised: 17 April 2017 / Accepted: 20 April 2017 / Published: 25 April 2017
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Abstract
Despite standard treatment, about 70% of ovarian cancer will recur. Cancer stem cells (CSCs) have been implicated in the drug-resistance mechanism. Several drug resistance mechanisms have been proposed, and among these, autophagy plays a crucial role for the maintenance and tumorigenicity of CSCs.
[...] Read more.
Despite standard treatment, about 70% of ovarian cancer will recur. Cancer stem cells (CSCs) have been implicated in the drug-resistance mechanism. Several drug resistance mechanisms have been proposed, and among these, autophagy plays a crucial role for the maintenance and tumorigenicity of CSCs. Compared to their differentiated counterparts, CSCs have been demonstrated to display a significantly higher level of autophagy flux. Moreover, mitophagy, a specific type of autophagy that selectively degrades excessive or damaged mitochondria, is shown to contribute to cancer progression and recurrence in several types of tumors. Nanomedicine has been shown to tackle the CSCs problem by overcoming drug resistance. In this work, we developed a nanomedicine, 188Re-liposome, which was demonstrated to target autophagy and mitophagy in the tumor microenvironment. Of note, the inhibition of autophagy and mitophagy could lead to significant tumor inhibition in two xenograft animal models. Lastly, we presented two cases of recurrent ovarian cancer, both in drug resistance status that received a level I dose from a phase I clinical trial. Both cases developing drug resistance showed drug sensitivity to 188Re-liposome. These results suggest that inhibition of autophagy and mitophagy by a nanomedicine may be a novel strategy to overcome drug resistance in ovarian cancer. Full article
(This article belongs to the Special Issue Gynecologic Oncology: From Molecular Mechanisms to Targeted Therapies)
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Open AccessArticle Cultivar-Specific Changes in Primary and Secondary Metabolites in Pak Choi (Brassica Rapa, Chinensis Group) by Methyl Jasmonate
Int. J. Mol. Sci. 2017, 18(5), 1004; doi:10.3390/ijms18051004
Received: 14 March 2017 / Revised: 1 May 2017 / Accepted: 2 May 2017 / Published: 7 May 2017
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Abstract
Glucosinolates, their hydrolysis products and primary metabolites were analyzed in five pak choi cultivars to determine the effect of methyl jasmonate (MeJA) on metabolite flux from primary metabolites to glucosinolates and their hydrolysis products. Among detected glucosinolates (total 14 glucosinolates; 9 aliphatic, 4
[...] Read more.
Glucosinolates, their hydrolysis products and primary metabolites were analyzed in five pak choi cultivars to determine the effect of methyl jasmonate (MeJA) on metabolite flux from primary metabolites to glucosinolates and their hydrolysis products. Among detected glucosinolates (total 14 glucosinolates; 9 aliphatic, 4 indole and 1 aromatic glucosinolates), indole glucosinolate concentrations (153–229%) and their hydrolysis products increased with MeJA treatment. Changes in the total isothiocyanates by MeJA were associated with epithiospecifier protein activity estimated as nitrile formation. Goitrin, a goitrogenic compound, significantly decreased by MeJA treatment in all cultivars. Changes in glucosinolates, especially aliphatic, significantly differed among cultivars. Primary metabolites including amino acids, organic acids and sugars also changed with MeJA treatment in a cultivar-specific manner. A decreased sugar level suggests that they might be a carbon source for secondary metabolite biosynthesis in MeJA-treated pak choi. The result of the present study suggests that MeJA can be an effective agent to elevate indole glucosinolates and their hydrolysis products and to reduce a goitrogenic compound in pak choi. The total glucosinolate concentration was the highest in “Chinese cabbage” in the control group (32.5 µmol/g DW), but indole glucosinolates increased the greatest in “Asian” when treated with MeJA. Full article
(This article belongs to the Special Issue Biological Activity of Natural Secondary Metabolite Products)
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Open AccessArticle Combining Low Temperature Fluorescence DNA-Hybridization, Immunostaining, and Super-Resolution Localization Microscopy for Nano-Structure Analysis of ALU Elements and Their Influence on Chromatin Structure
Int. J. Mol. Sci. 2017, 18(5), 1005; doi:10.3390/ijms18051005
Received: 16 February 2017 / Revised: 25 April 2017 / Accepted: 2 May 2017 / Published: 7 May 2017
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Abstract
Immunostaining and fluorescence in situ hybridization (FISH) are well established methods for specific labelling of chromatin in the cell nucleus. COMBO-FISH (combinatorial oligonucleotide fluorescence in situ hybridization) is a FISH method using computer designed oligonucleotide probes specifically co-localizing at given target sites. In
[...] Read more.
Immunostaining and fluorescence in situ hybridization (FISH) are well established methods for specific labelling of chromatin in the cell nucleus. COMBO-FISH (combinatorial oligonucleotide fluorescence in situ hybridization) is a FISH method using computer designed oligonucleotide probes specifically co-localizing at given target sites. In combination with super resolution microscopy which achieves spatial resolution far beyond the Abbe Limit, it allows new insights into the nano-scaled structure and organization of the chromatin of the nucleus. To avoid nano-structural changes of the chromatin, the COMBO-FISH labelling protocol was optimized omitting heat treatment for denaturation of the target. As an example, this protocol was applied to ALU elements—dispersed short stretches of DNA which appear in different kinds in large numbers in primate genomes. These ALU elements seem to be involved in gene regulation, genomic diversity, disease induction, DNA repair, etc. By computer search, we developed a unique COMBO-FISH probe which specifically binds to ALU consensus elements and combined this DNA–DNA labelling procedure with heterochromatin immunostainings in formaldehyde-fixed cell specimens. By localization microscopy, the chromatin network-like arrangements of ALU oligonucleotide repeats and heterochromatin antibody labelling sites were simultaneously visualized and quantified. This novel approach which simultaneously combines COMBO-FISH and immunostaining was applied to chromatin analysis on the nanoscale after low-linear-energy-transfer (LET) radiation exposure at different doses. Dose-correlated curves were obtained from the amount of ALU representing signals, and the chromatin re-arrangements during DNA repair after irradiation were quantitatively studied on the nano-scale. Beyond applications in radiation research, the labelling strategy of immunostaining and COMBO-FISH with localization microscopy will also offer new potentials for analyses of subcellular elements in combination with other specific chromatin targets. Full article
(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)
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Open AccessArticle Inflammation Downregulates UCP1 Expression in Brown Adipocytes Potentially via SIRT1 and DBC1 Interaction
Int. J. Mol. Sci. 2017, 18(5), 1006; doi:10.3390/ijms18051006
Received: 11 April 2017 / Revised: 26 April 2017 / Accepted: 2 May 2017 / Published: 8 May 2017
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Abstract
Brown adipose tissue thermogenesis at the cost of energy is not only important for the development of obesity, but also possesses great promise in anti-obesity treatment. Uncoupling protein 1 (UCP1) expression has been reported to be under control of the intracellular deacetylase SIRT1.
[...] Read more.
Brown adipose tissue thermogenesis at the cost of energy is not only important for the development of obesity, but also possesses great promise in anti-obesity treatment. Uncoupling protein 1 (UCP1) expression has been reported to be under control of the intracellular deacetylase SIRT1. Here, we investigated the effect and mechanism of inflammation and sirtuin-1 (SIRT1) activation on the induction of thermogenic genes in immortalized brown adipocytes incubated with LPS or IL1β and mice with elevated inflammatory tone. In vitro stimulation of brown adipocytes with dibutyryl cyclic adenosine monophosthate (dbcAMP) reduced the expression of deleted in breast cancer-1 (Dbc1) (SIRT1 inhibitor) and increased the Ucp1 expression. Silencing of SIRT1 attenuated dbcAMP induction of Ucp1. In contrast, IL1β increased the expression of Dbc1 and greatly reduced the induction of Ucp1. Similarly, in vivo studies revealed decreased expression of Ucp1 in brown adipose tissue (BAT) in mice chronically infused with LPS. Resveratrol, a known SIRT1 activator, partly rescued the Ucp1 downregulation by inflammation in both the cell cultures and mice. Here, we describe how the expression of Ucp1 in BAT is controlled via SIRT1 and is reduced under inflammation and can be rescued by SIRT1 activation by resveratrol. We suggest the reduced UCP1 expression under inflammation is mediated by the increased expression of DBC1, which inhibits SIRT1 activity. Full article
(This article belongs to the Section Molecular Pathology, Diagnostics, and Therapeutics)
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Open AccessArticle Real-World Experiences with the Combination Treatment of Ledipasvir plus Sofosbuvir for 12 Weeks in HCV Genotype 1-Infected Japanese Patients: Achievement of a Sustained Virological Response in Previous Users of Peginterferon plus Ribavirin with HCV NS3/4A Inhibitors
Int. J. Mol. Sci. 2017, 18(5), 906; doi:10.3390/ijms18050906
Received: 4 April 2017 / Revised: 19 April 2017 / Accepted: 21 April 2017 / Published: 25 April 2017
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Abstract
The aim of this study was to characterize the treatment response and serious adverse events of ledipasvir plus sofosbuvir therapies in Japanese patients infected with hepatitis C virus (HCV) genotype 1 (GT1). This retrospective study analyzed 240 Japanese HCV GT1 patients treated for
[...] Read more.
The aim of this study was to characterize the treatment response and serious adverse events of ledipasvir plus sofosbuvir therapies in Japanese patients infected with hepatitis C virus (HCV) genotype 1 (GT1). This retrospective study analyzed 240 Japanese HCV GT1 patients treated for 12 weeks with 90 mg of ledipasvir plus 400 mg of sofosbuvir daily. Sustained virological response at 12 weeks post-treatment (SVR12) was achieved in 236 of 240 (98.3%) patients. Among treatment-naïve patients, SVR12 was achieved in 136 of 138 (98.6%) patients, and among treatment-experienced patients, SVR12 was achieved in 100 of 102 (98.0%) patients. In patients previously treated with peginterferon plus ribavirin with various HCV NS3/4A inhibitors, 100% SVR rates (25/25) were achieved. Two relapsers had HCV NS5A resistance-associated variants (RAVs), but no HCV NS5B-S282 was observed after they relapsed. We experienced two patients with cardiac events during treatment. In conclusion, combination of ledipasvir plus sofosbuvir for 12 weeks is a potential therapy for HCV GT1 patients. Caution is needed for HCV NS5A RAVs, which were selected by HCV NS5A inhibitors and cardiac adverse events. Full article
(This article belongs to the Special Issue Hepatitis Virus Infection and Research)
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Open AccessArticle Chloramidine/Bisindolylmaleimide-I-Mediated Inhibition of Exosome and Microvesicle Release and Enhanced Efficacy of Cancer Chemotherapy
Int. J. Mol. Sci. 2017, 18(5), 1007; doi:10.3390/ijms18051007
Received: 22 February 2017 / Revised: 28 April 2017 / Accepted: 3 May 2017 / Published: 9 May 2017
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Abstract
Microvesicle (MV) release from tumour cells influences drug retention, contributing to cancer drug resistance. Strategically regulating MV release may increase drug retention within cancer cells and allow for lower doses of chemotherapeutic drugs. The contribution of exosomes to drug retention still remains unknown.
[...] Read more.
Microvesicle (MV) release from tumour cells influences drug retention, contributing to cancer drug resistance. Strategically regulating MV release may increase drug retention within cancer cells and allow for lower doses of chemotherapeutic drugs. The contribution of exosomes to drug retention still remains unknown. Potential exosome and MV (EMV) biogenesis inhibitors, tested on human prostate cancer (PC3) cells for their capacity to inhibit EMV release, were also tested on PC3 and MCF-7 (breast cancer) cells for improving chemotherapy. Agents inhibiting EMV release most significantly, whilst maintaining cell viability, were chloramidine (Cl-amidine; 50 µM) and bisindolylmaleimide-I (10 µM). Apoptosis mediated by the chemotherapy drug 5-fluorouracil (5-FU) was significantly enhanced in PC3 cells in the presence of both these EMV inhibitors, resulting in a 62% (Cl-amidine + 5-FU) and 59% (bisindolylmaleimide-I + 5-FU) decrease in numbers of viable PC3 cells compared to 5-FU alone after 24 h. For MCF-7 cells, there were similar increased reductions of viable cells compared to 5-FU treatment alone ranging from 67% (Cl-amidine + 5-FU) to 58% (bisindolylmaleimide-I + 5-FU). Using combinatory treatment, the two EMV inhibitors further reduced the number of viable cancer cells tested. Neither inhibitor affected cell viability. Combining selected EMV inhibitors may pose as a novel strategy to enhance the efficacy of chemotherapeutic drug-mediated apoptosis. Full article
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Open AccessArticle Carotid Artery Stenting and Blood–Brain Barrier Permeability in Subjects with Chronic Carotid Artery Stenosis
Int. J. Mol. Sci. 2017, 18(5), 1008; doi:10.3390/ijms18051008
Received: 13 March 2017 / Revised: 23 April 2017 / Accepted: 3 May 2017 / Published: 8 May 2017
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Abstract
Failure of the blood-brain barrier (BBB) is a critical event in the development and progression of diseases such as acute ischemic stroke, chronic ischemia or small vessels disease that affect the central nervous system. It is not known whether BBB breakdown in subjects
[...] Read more.
Failure of the blood-brain barrier (BBB) is a critical event in the development and progression of diseases such as acute ischemic stroke, chronic ischemia or small vessels disease that affect the central nervous system. It is not known whether BBB breakdown in subjects with chronic carotid artery stenosis can be restrained with postoperative recovery of cerebral perfusion. The aim of the study was to assess the short-term effect of internal carotid artery stenting on basic perfusion parameters and permeability surface area-product (PS) in such a population. Forty subjects (23 males) with stenosis of >70% within a single internal carotid artery and neurological symptoms who underwent a carotid artery stenting procedure were investigated. Differences in the following computed tomography perfusion (CTP) parameters were compared before and after surgery: global cerebral blood flow (CBF), cerebral blood volume (CBV), mean transit time (MTT), time to peak (TTP) and PS. PS acquired by CTP is used to measure the permeability of the BBB to contrast material. In all baseline cases, the CBF and CBV values were low, while MTT and TTP were high on both the ipsi- and contralateral sides compared to reference values. PS was approximately twice the normal value. CBF was higher (+6.14%), while MTT was lower (−9.34%) on the contralateral than on the ipsilateral side. All perfusion parameters improved after stenting on both the ipsilateral (CBF +22.66%; CBV +18.98%; MTT −16.09%, TTP −7.62%) and contralateral (CBF +22.27%, CBV +19.72%, MTT −14.65%, TTP −7.46%) sides. PS decreased by almost half: ipsilateral −48.11%, contralateral −45.19%. The decline in BBB permeability was symmetrical on the ipsi- and contralateral sides to the stenosis. Augmented BBB permeability can be controlled by surgical intervention in humans. Full article
(This article belongs to the Special Issue Cerebral Blood Flow and Metabolism)
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Open AccessArticle Interaction of Biliverdin Chromophore with Near-Infrared Fluorescent Protein BphP1-FP Engineered from Bacterial Phytochrome
Int. J. Mol. Sci. 2017, 18(5), 1009; doi:10.3390/ijms18051009
Received: 12 April 2017 / Revised: 30 April 2017 / Accepted: 4 May 2017 / Published: 8 May 2017
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Abstract
Near-infrared (NIR) fluorescent proteins (FPs) designed from PAS (Per-ARNT-Sim repeats) and GAF (cGMP phosphodiesterase/adenylate cyclase/FhlA transcriptional activator) domains of bacterial phytochromes covalently bind biliverdin (BV) chromophore via one or two Cys residues. We studied BV interaction with a series of NIR FP variants
[...] Read more.
Near-infrared (NIR) fluorescent proteins (FPs) designed from PAS (Per-ARNT-Sim repeats) and GAF (cGMP phosphodiesterase/adenylate cyclase/FhlA transcriptional activator) domains of bacterial phytochromes covalently bind biliverdin (BV) chromophore via one or two Cys residues. We studied BV interaction with a series of NIR FP variants derived from the recently reported BphP1-FP protein. The latter was engineered from a bacterial phytochrome RpBphP1, and has two reactive Cys residues (Cys15 in the PAS domain and Cys256 in the GAF domain), whereas its mutants contain single Cys residues either in the PAS domain or in the GAF domain, or no Cys residues. We characterized BphP1-FP and its mutants biochemically and spectroscopically in the absence and in the presence of denaturant. We found that all BphP1-FP variants are monomers. We revealed that spectral properties of the BphP1-FP variants containing either Cys15 or Cys256, or both, are determined by the covalently bound BV chromophore only. Consequently, this suggests an involvement of the inter-monomeric allosteric effects in the BV interaction with monomers in dimeric NIR FPs, such as iRFPs. Likely, insertion of the Cys15 residue, in addition to the Cys256 residue, in dimeric NIR FPs influences BV binding by promoting the BV chromophore covalent cross-linking to both PAS and GAF domains. Full article
(This article belongs to the Special Issue Fluorescent Proteins)
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Open AccessArticle miR-103 Promotes Proliferation and Metastasis by Targeting KLF4 in Gastric Cancer
Int. J. Mol. Sci. 2017, 18(5), 910; doi:10.3390/ijms18050910
Received: 7 April 2017 / Revised: 19 April 2017 / Accepted: 23 April 2017 / Published: 26 April 2017
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Abstract
MicroRNAs (miRNAs) play important roles in the cancer development and progression; overexpression of miR-103 has been identified in various tumors. However, its biological function and regulatory mechanism involved in modulation of human gastric cancer (GC) remain largely unknown. This study aimed to confirm
[...] Read more.
MicroRNAs (miRNAs) play important roles in the cancer development and progression; overexpression of miR-103 has been identified in various tumors. However, its biological function and regulatory mechanism involved in modulation of human gastric cancer (GC) remain largely unknown. This study aimed to confirm clinical significance of miR-103 and investigate its biological role and underlying mechanism in GC. Real-time quantitative PCR (qRT-PCR) revealed miR-103 was highly expressed in GC tissues and cell lines. miR-103 expression was correlated closely with tumor size, Lauren’s classification, and lymph node metastasis. Importantly, Kaplan-Meier analysis revealed that high expression of miR-103 was significantly associated with poor overall survival and disease-free survival of GC patients. Downregulation of miR-103 by transfecting with miR-103 inhibitor significantly suppressed cell proliferation, induced apoptosis, inhibited migration and invasion in vitro and in vivo. Furthermore, miRNA target databases and luciferase reporter assay confirmed that Krüppel-like Factor-4 (KLF4) was a direct target of miR-103 in GC, and there was a significant inverse correlation between miR-103 and KLF4 expression in GC tissues. Moreover, KLF4 downregulation could rescue miR-103’s oncogenic effect on GC cell proliferation, apoptosis, migration, and invasion. Therefore, these results suggested that miR-103 overexpression could contribute to tumor progression by suppressing KLF4, and it might serve as a promising candidate for the prognosis of GC patients. Full article
(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)
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Open AccessArticle Inhibiting HDAC1 Enhances the Anti-Cancer Effects of Statins through Downregulation of GGTase-Iβ Expression
Int. J. Mol. Sci. 2017, 18(5), 1010; doi:10.3390/ijms18051010
Received: 26 March 2017 / Revised: 23 April 2017 / Accepted: 1 May 2017 / Published: 8 May 2017
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Abstract
Hydroxy-methyl-glutaryl-coenzyme A (HMG-CoA) reductase inhibitors, namely statins, are potential anti-tumor agents. Previously, we showed that a pan-histone deacetylase (HDAC) inhibitor enhances the anti-tumor effects of the HMG-CoA inhibitor. However, the underlying mechanisms were not fully understood. Cancer cell lines (CAL-27 and SACC-83) were
[...] Read more.
Hydroxy-methyl-glutaryl-coenzyme A (HMG-CoA) reductase inhibitors, namely statins, are potential anti-tumor agents. Previously, we showed that a pan-histone deacetylase (HDAC) inhibitor enhances the anti-tumor effects of the HMG-CoA inhibitor. However, the underlying mechanisms were not fully understood. Cancer cell lines (CAL-27 and SACC-83) were exposed to pan-HDAC inhibitor, or HDAC1 inhibitor, or geranylgeranyl transferase type I (GGTase-I) inhibitor alone or in combination with statin. Cell viability, apoptosis, migration, and invasion were assessed by Cell Count Kit-8, 4′,6-diamidino-2-phenylindole staining, and transwell assay, respectively. A xenograft model was used for assessing tumor growth in vivo. Western blot and real-time PCR were used to assess the expression of genes. We observed that inhibiting HDAC1 could enhance the anti-tumor effects of statins both in vitro and in vivo. Inhibiting HDAC1 blocked the statin-induced upregulation of geranylgeranyl transferase type Iβ subunit (GGTase-Iβ), resulting in an enhancement of the anti-cancer effects of statin. Overexpression of GGTase-Iβ or constitutively active RhoA abolished the enhancement by inhibiting HDAC1 on anti-tumor effects of statins. The HDAC1 inhibitor failed to enhance cytotoxicity in non-tumor primary cells treated with statin. Inhibiting HDAC1 enhanced the anti-cancer effects of statins through downregulation of GGTase-Iβ expression, and thus further inactivation of RhoA. A combination of statin with HDAC1 or GGTase-I inhibitor would be a new strategy for cancer chemotherapy. Full article
(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)
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Open AccessArticle Atractylenolide-I Protects Human SH-SY5Y Cells from 1-Methyl-4-Phenylpyridinium-Induced Apoptotic Cell Death
Int. J. Mol. Sci. 2017, 18(5), 1012; doi:10.3390/ijms18051012
Received: 24 January 2017 / Revised: 21 April 2017 / Accepted: 3 May 2017 / Published: 8 May 2017
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Abstract
Oxidative stress and apoptosis are the major mechanisms that induce dopaminergic cell death. Our study investigates the protective effects of atractylenolide-I (ATR-I) on 1-methyl-4-phenylpyridinium (MPP+)-induced cytotoxicity in human dopaminergic SH-SY5Y cells, as well as its underlying mechanism. Our experimental data indicates
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Oxidative stress and apoptosis are the major mechanisms that induce dopaminergic cell death. Our study investigates the protective effects of atractylenolide-I (ATR-I) on 1-methyl-4-phenylpyridinium (MPP+)-induced cytotoxicity in human dopaminergic SH-SY5Y cells, as well as its underlying mechanism. Our experimental data indicates that ATR-I significantly inhibits the loss of cell viability induced by MPP+ in SH-SY5Y cells. To further unravel the mechanism, we examined the effect of ATR-I on MPP+-induced apoptotic cell death characterized by an increase in the Bax/Bcl-2 mRNA ratio, the release of cytochrome-c, and the activation of caspase-3 leading to elevated levels of cleaved poly(ADP-ribose) polymerase (PARP) resulting in SH-SY5Y cell death. Our results demonstrated that ATR-I decreases the level of pro-apoptotic proteins induced by MPP+ and also restored Bax/Bcl-2 mRNA levels, which are critical for inducing apoptosis. In addition, ATR-I demonstrated a significant increase in the protein expression of heme-oxygenase in MPP+-treated SH-SY5Y cells. These results suggest that the pharmacological effect of ATR-I may be, at least in part, caused by the reduction in pro-apoptotic signals and also by induction of anti-oxidant protein. Full article
(This article belongs to the Section Bioactives and Nutraceuticals)
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Open AccessArticle Imipramine Protects against Bone Loss by Inhibition of Osteoblast-Derived Microvesicles
Int. J. Mol. Sci. 2017, 18(5), 1013; doi:10.3390/ijms18051013
Received: 8 March 2017 / Revised: 2 April 2017 / Accepted: 3 May 2017 / Published: 8 May 2017
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Abstract
The maintenance of bone homeostasis is largely dependent upon cellular communication between osteoclasts and osteoblasts. Microvesicles (MVs) represent a novel mechanism for osteoblasts and osteoclasts communication, as has been demonstrated in our previous study. Sphingomyelinases catalyze the hydrolysis of sphingomyelin, which leads to
[...] Read more.
The maintenance of bone homeostasis is largely dependent upon cellular communication between osteoclasts and osteoblasts. Microvesicles (MVs) represent a novel mechanism for osteoblasts and osteoclasts communication, as has been demonstrated in our previous study. Sphingomyelinases catalyze the hydrolysis of sphingomyelin, which leads to increased membrane fluidity and facilitates MV generation. This effect can be inhibited by imipramine, an inhibitor of acid sphingomyelinase (ASM), which is also known as a member of tricyclic antidepressants (TCAs). A recent study has reported that in vitro treatment of imipramine blocked MVs release from glial cells. However, whether imipramine has this effect on osteoblast-derived MVs and whether it is involved in MV generation in vivo is unclear. Here, our investigations found that imipramine slightly reduced the expression of osteoblast differentiation of related genes, but did not impact parathyroid hormone (PTH) regulation for these genes and also did not affect receptor activator of nuclear factor-κB ligand (RANKL)-mediated osteoclast formation; however, imipramine treatment blocked MVs released from osteoblasts and inhibited MV-induced osteoclast formation. In vivo, mice administrated with imipramine were protected from ovariectomy-induced bone loss as evaluated by various bone structural parameters and serum levels of biochemical markers. Our results suggest that inhibiting the production of MVs containing RANKL in vivo is very important for preventing bone loss. Full article
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Open AccessArticle Exploring the Behavioral and Metabolic Phenotype Generated by Re-Introduction of the Ghrelin Receptor in the Ventral Tegmental Area
Int. J. Mol. Sci. 2017, 18(5), 914; doi:10.3390/ijms18050914
Received: 22 February 2017 / Revised: 1 April 2017 / Accepted: 20 April 2017 / Published: 26 April 2017
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Abstract
Ghrelin receptor (Ghr-R) signaling in neurons of the ventral tegmental area (VTA) can modulate dopaminergic function and the reward-related effects of both palatable foods and drugs of abuse. In this study, we re-introduced the Ghr-R in VTA neurons in Ghr-R knockout mice (Ghr-R
[...] Read more.
Ghrelin receptor (Ghr-R) signaling in neurons of the ventral tegmental area (VTA) can modulate dopaminergic function and the reward-related effects of both palatable foods and drugs of abuse. In this study, we re-introduced the Ghr-R in VTA neurons in Ghr-R knockout mice (Ghr-RVTA mice) to specifically study the importance of the constitutively active Ghr-R for VTA neuronal signaling. Our results showed that re-introduction of the Ghr-R in the VTA had no impact on body weight or food intake under basal conditions. However, during novel environment stress Ghr-RVTA mice showed increased food intake and energy expenditure compared to Ghr-R knockout mice, demonstrating the significance of Ghr-R signaling in the response to stress. Ghr-RVTA mice also showed increased cocaine-induced locomotor activity compared to Ghr-R knockout mice, highlighting the importance of ghrelin signaling for the reward-related effects of activation of VTA neurons. Overall, our data suggest that re-introduction of the Ghr-R in the mesolimbic reward system of Ghr-R knockout mice increases the level of activation induced by both cocaine and novelty stress. Full article
(This article belongs to the Special Issue Neurobiological Perspectives on Ghrelin)
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Open AccessArticle The Dose–Response Association between Nitrogen Dioxide Exposure and Serum Interleukin-6 Concentrations
Int. J. Mol. Sci. 2017, 18(5), 1015; doi:10.3390/ijms18051015
Received: 31 March 2017 / Revised: 26 April 2017 / Accepted: 28 April 2017 / Published: 8 May 2017
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Abstract
Systemic inflammation is an integral part of chronic obstructive pulmonary disease (COPD), and air pollution is associated with cardiorespiratory mortality, yet the interrelationships are not fully defined. We examined associations between nitrogen dioxide (NO2) exposure (as a marker of traffic-related air
[...] Read more.
Systemic inflammation is an integral part of chronic obstructive pulmonary disease (COPD), and air pollution is associated with cardiorespiratory mortality, yet the interrelationships are not fully defined. We examined associations between nitrogen dioxide (NO2) exposure (as a marker of traffic-related air pollution) and pro-inflammatory cytokines, and investigated effect modification and mediation by post-bronchodilator airflow obstruction (post-BD-AO) and cardiovascular risk. Data from middle-aged participants in the Tasmanian Longitudinal Health Study (TAHS, n = 1389) were analyzed by multivariable logistic regression, using serum interleukin (IL)-6, IL-8 and tumor necrosis factor-α (TNF-α) as the outcome. Mean annual NO2 exposure was estimated at residential addresses using a validated satellite-based land-use regression model. Post-BD-AO was defined by post-BD forced expiratory ratio (FEV1/FVC) < lower limit of normal, and cardiovascular risk by a history of either cerebrovascular or ischaemic heart disease. We found a positive association with increasing serum IL-6 concentration (geometric mean 1.20 (95% CI: 1.1 to 1.3, p = 0.001) per quartile increase in NO2). This was predominantly a direct relationship, with little evidence for either effect modification or mediation via post-BD-AO, or for the small subgroup who reported cardiovascular events. However, there was some evidence consistent with serum IL-6 being on the causal pathway between NO2 and cardiovascular risk. These findings raise the possibility that the interplay between air pollution and systemic inflammation may differ between post-BD airflow obstruction and cardiovascular diseases. Full article
(This article belongs to the Special Issue Inhaled Pollutants Modulate Respiratory and Systemic Diseases)
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Open AccessArticle Antiplatelet Activity of a Newly Synthesized Novel Ruthenium (II): A Potential Role for Akt/JNK Signaling
Int. J. Mol. Sci. 2017, 18(5), 916; doi:10.3390/ijms18050916
Received: 13 March 2017 / Revised: 4 April 2017 / Accepted: 18 April 2017 / Published: 27 April 2017
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Abstract
In oncotherapy, ruthenium complexes are considered as potential alternatives for platinum compounds, and have been proved as promising anticancer drugs with high efficacy and lesser side effects. Platelet activation plays a major role in cancer metastasis and progression. Hence, this study explored the
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In oncotherapy, ruthenium complexes are considered as potential alternatives for platinum compounds, and have been proved as promising anticancer drugs with high efficacy and lesser side effects. Platelet activation plays a major role in cancer metastasis and progression. Hence, this study explored the effect of a newly synthesized ruthenium complex, [Ru(η6-cymene)(L)Cl]BF4(TQ5), where L = 4-phenyl-2-pyridin-2-yl-quinazoline), on human platelet activation. TQ5 (3–5 µM) inhibited concentration-dependent collagen-induced platelet aggregation in washed human platelets. However, this compound only inhibited platelet aggregation at a maximum concentration of 500 and 100 µM against thrombin and 9,11-dideoxy-11α, 9α-epoxymethanoprostaglandin (U46619)-induced stimulation, respectively. TQ5 inhibited collagen-induced ATP release and calcium mobilization ([Ca2+]i), without inducing cell cytotoxicity. In addition, neither SQ22536, an adenylate cyclase inhibitor, nor 1H-[1,2,4] oxadiazolo [4,3-a]quinoxalin-1-one (ODQ), a guanylate cyclase inhibitor, significantly reversed the TQ5-mediated inhibition of platelet aggregation. TQ5 inhibited the collagen-induced phosphorylation of protein kinase B (Akt) and c-Jun N-terminal kinase (JNK), but did not effectively inhibit extracellular signal-regulated kinase 1/2 (ERK1/2) and p38-mitogen-activated protein kinase (p38-MAPK) in human platelets. Additionally, TQ5 significantly prolonged the closure time in whole blood and increased the occlusion time of thrombotic platelet plug formation in mice. This study demonstrates, for the first time, that a newly synthesized ruthenium complex, TQ5, exhibits potent antiplatelet activity by hindering ATP release and [Ca2+]i, and by decreasing the activation of Akt/JNK signals. Together, these results suggest that TQ5 could be developed as a therapeutic agent that helps prevent or treat thromboembolic disorders, since it is found to be potently more effective than a well-established antithrombotic aspirin. Full article
(This article belongs to the Special Issue Recent Advances in Metal Based Drugs)
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Open AccessArticle Impaired Platelet Aggregation and Rebalanced Hemostasis in Patients with Chronic Hepatitis C Virus Infection
Int. J. Mol. Sci. 2017, 18(5), 1016; doi:10.3390/ijms18051016
Received: 3 March 2017 / Revised: 3 May 2017 / Accepted: 4 May 2017 / Published: 8 May 2017
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Abstract
Increased risk of both cardiovascular disease (CVD) and bleeding has been found in patients with chronic hepatitis C (CHC) infection, and a re-balanced hemostasis has been proposed. The aim of this study was to investigate functional whole blood coagulation and platelet function in
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Increased risk of both cardiovascular disease (CVD) and bleeding has been found in patients with chronic hepatitis C (CHC) infection, and a re-balanced hemostasis has been proposed. The aim of this study was to investigate functional whole blood coagulation and platelet function in CHC infection. The prospective study included 82 patients with CHC infection (39 with advanced liver fibrosis and 43 with no or mild liver fibrosis) and 39 healthy controls. A total of 33 patients were treated for CHC infection and achieved sustained virological response (SVR). Baseline and post-treatment blood samples were collected. Hemostasis was assessed by both standard coagulation tests and functional whole blood hemostatic assays (thromboelastograhy (TEG), and platelet aggregation (Multiplate). Patients with CHC and advanced fibrosis had impaired platelet aggregation both compared to patients with no or mild fibrosis and to healthy controls. Patients with CHC and advanced fibrosis also had lower antithrombin, platelet count, and coagulation factors II-VII-X compared to healthy controls. In contrast, TEG did not differ between groups. In treated patients achieving SVR, post-treatment platelet count was higher than pre-treatment counts (p = 0.033) and ADPtest, ASPItest, and RISTOhightest all increased post treatment (all p < 0.05). All Multiplate tests values, however, remained below those in the healthy controls. CHC-infected patients displayed evidence of rebalanced hemostasis with only partly hemostatic normalization in patients achieving SVR. The implications of rebalanced hemostasis and especially the impact on risk of CVD and bleeding warrants further studies. Full article
(This article belongs to the Section Molecular Pathology, Diagnostics, and Therapeutics)
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Open AccessArticle Sulfiredoxin May Promote Cervical Cancer Metastasis via Wnt/β-Catenin Signaling Pathway
Int. J. Mol. Sci. 2017, 18(5), 917; doi:10.3390/ijms18050917
Received: 19 March 2017 / Revised: 17 April 2017 / Accepted: 22 April 2017 / Published: 27 April 2017
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Abstract
The abnormal elevation of sulfiredoxin (Srx/SRXN1)—an antioxidant enzyme whose main function is to protect against oxidative stress—has been shown to be closely correlated with the progression of several types of cancer, including human cervical cancer. However, the molecular mechanism by which Srx promotes
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The abnormal elevation of sulfiredoxin (Srx/SRXN1)—an antioxidant enzyme whose main function is to protect against oxidative stress—has been shown to be closely correlated with the progression of several types of cancer, including human cervical cancer. However, the molecular mechanism by which Srx promotes tumor progression, especially cancer metastasis in cervical cancer, has not been elucidated. Here, we show that Srx expression gradually increases during the progression of human cervical cancer and its expression level is closely correlated with lymph node metastasis. Our study also reveals a significant positive correlation between the expression of Srx and β-catenin in cervical cancer tissues. Loss-of-function studies demonstrate that Srx knockdown using a lentiviral vector-mediated specific shRNA decreases the migration and invasion capacity in HeLa (human papilloma virus 18 type cervical cancer cell line) and SiHa SiHa (cervical squamous cancer cell line). Notably, the exact opposite effects were observed in gain-of-function experiments in C-33A cells. Mechanistically, downregulation or upregulation of Srx leads to an altered expression of proteins associated with the Wnt/β-catenin signaling pathway. Furthermore, blockage of the Wnt/β-catenin signaling pathway contributed to attenuated Srx expression and resulted in significant inhibition of cell migration and invasion in cervical cancer cell lines. Combined, Srx might be an oncoprotein in cervical cancer, playing critical roles in activating the Wnt/β-catenin signaling pathway; it may therefore be a therapeutic target for cervical cancer. Full article
(This article belongs to the Special Issue Chemical and Molecular Approach to Tumor Metastases)
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Open AccessArticle Thymoquinone Defeats Diabetes-Induced Testicular Damage in Rats Targeting Antioxidant, Inflammatory and Aromatase Expression
Int. J. Mol. Sci. 2017, 18(5), 919; doi:10.3390/ijms18050919
Received: 9 March 2017 / Revised: 14 April 2017 / Accepted: 21 April 2017 / Published: 27 April 2017
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Abstract
Antioxidants have valuable effects on the process of spermatogenesis, particularly with diabetes mellitus (DM). Therefore, the present study investigated the impact and the intracellular mechanisms by which thymoquinone (TQ) works against diabetes-induced testicular deteriorations in rats. Wistar male rats (n = 60)
[...] Read more.
Antioxidants have valuable effects on the process of spermatogenesis, particularly with diabetes mellitus (DM). Therefore, the present study investigated the impact and the intracellular mechanisms by which thymoquinone (TQ) works against diabetes-induced testicular deteriorations in rats. Wistar male rats (n = 60) were randomly allocated into four groups; Control, Diabetic (streptozotocin (STZ)-treated rats where diabetes was induced by intraperitoneal injection of STZ, 65 mg/kg), Diabetic + TQ (diabetic rats treated with TQ (50 mg/kg) orally once daily), and TQ (non-diabetic rats treated with TQ) for 12 weeks. Results revealed that TQ significantly improved the sperm parameters with a reduction in nitric oxide (NO) and malondialdehyde (MDA) levels in testicular tissue. Also, it increased testicular reduced glutathione (GSH) levels and superoxide dismutase (SOD) activity. Interestingly, TQ induced downregulation of testicular inducible nitric oxide synthase (iNOS) and nuclear factor kappa-B (NF-κB) and significantly upregulated the aromatase protein expression levels in testicles in comparison with the diabetic rats. In conclusion, TQ treatment exerted a protective effect against reproductive dysfunction induced by diabetes not only through its powerful antioxidant and hypoglycemic effects but also through its downregulation of testicular iNOS and NF-κB along with upregulation of aromatase expression levels in diabetic rats. Full article
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Open AccessArticle YKL-40-Induced Inhibition of miR-590-3p Promotes Interleukin-18 Expression and Angiogenesis of Endothelial Progenitor Cells
Int. J. Mol. Sci. 2017, 18(5), 920; doi:10.3390/ijms18050920
Received: 5 March 2017 / Revised: 19 April 2017 / Accepted: 21 April 2017 / Published: 27 April 2017
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Abstract
YKL-40, also known as human cartilage glycoprotein-39 or chitinase-3-like-1, is a pro-inflammatory protein that is highly expressed in rheumatoid arthritis (RA) patients. Angiogenesis is a critical step in the pathogenesis of RA, promoting the infiltration of inflammatory cells into joints and providing oxygen
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YKL-40, also known as human cartilage glycoprotein-39 or chitinase-3-like-1, is a pro-inflammatory protein that is highly expressed in rheumatoid arthritis (RA) patients. Angiogenesis is a critical step in the pathogenesis of RA, promoting the infiltration of inflammatory cells into joints and providing oxygen and nutrients to RA pannus. In this study, we examined the effects of YKL-40 in the production of the pro-inflammatory cytokine interleukin-18 (IL-18), and the stimulation of angiogenesis and accumulation of osteoblasts. We observed that YKL-40 induces IL-18 production in osteoblasts and thereby stimulates angiogenesis of endothelial progenitor cells (EPCs). We found that this process occurs through the suppression of miR-590-3p via the focal adhesion kinase (FAK)/PI3K/Akt signaling pathway. YKL-40 inhibition reduced angiogenesis in in vivo models of angiogenesis: the chick embryo chorioallantoic membrane (CAM) and Matrigel plug models. We report that YKL-40 stimulates IL-18 expression in osteoblasts and facilitates EPC angiogenesis. Full article
(This article belongs to the Section Molecular Pathology, Diagnostics, and Therapeutics)
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Open AccessArticle Characterization of Copy Number Variation’s Potential Role in Marek’s Disease
Int. J. Mol. Sci. 2017, 18(5), 1020; doi:10.3390/ijms18051020
Received: 27 March 2017 / Revised: 22 April 2017 / Accepted: 4 May 2017 / Published: 9 May 2017
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Abstract
Marek’s Disease (MD) is a highly contagious pathogenic and oncogenic disease primarily affecting chickens. Chicken Lines 63 and 72, as well as their recombinant congenic strains (RCS) with varied susceptibility to MD, are ideal models to study the complex mechanisms of genetic resistance
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Marek’s Disease (MD) is a highly contagious pathogenic and oncogenic disease primarily affecting chickens. Chicken Lines 63 and 72, as well as their recombinant congenic strains (RCS) with varied susceptibility to MD, are ideal models to study the complex mechanisms of genetic resistance to MD. In this study, we investigated copy number variation (CNV) in these inbred chicken lines using the Affymetrix Axiom HD 600 K SNP genotyping array. We detected 393 CNV segments across all ten chicken lines, of which 12 CNVs were specifically identified in Line 72. We then assessed genetic structure based on CNV and observed markedly different patterns. Finally, we validated two deletion events in Line 72 and correlated them with genes expression using qPCR and RNA-seq, respectively. Our combined results indicated that these two CNV deletions were likely to contribute to MD susceptibility. Full article
(This article belongs to the Special Issue Exploring the Genotype–Phenotype Map to Explain Complex Traits)
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Open AccessArticle Chemical Composition and Biological Activities of Mono- and Heterofloral Bee Pollen of Different Geographical Origins
Int. J. Mol. Sci. 2017, 18(5), 921; doi:10.3390/ijms18050921
Received: 27 February 2017 / Revised: 10 April 2017 / Accepted: 24 April 2017 / Published: 27 April 2017
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Abstract
Recent research shows variations in pollen chemical constituents and, consequently, in their therapeutic properties. Mono and multifloral bee pollen extracts were investigated for antioxidant and enzyme inhibitory activity properties, phenolic compounds and fatty acid composition. Generally, Eucalyptus spp. and multifloral extracts exhibited potent
[...] Read more.
Recent research shows variations in pollen chemical constituents and, consequently, in their therapeutic properties. Mono and multifloral bee pollen extracts were investigated for antioxidant and enzyme inhibitory activity properties, phenolic compounds and fatty acid composition. Generally, Eucalyptus spp. and multifloral extracts exhibited potent inhibitory activity against α-amylase, acetylcholinesterase, tyrosinase, lipoxygenase, lipase and hyaluronidase. On the other hand, Miconia spp. demonstrated higher antihemolytic activity. Cocos nucifera and Miconia spp. extracts exhibited important antioxidant properties in the different assays (ABTS, DPPH, β-carotene/linoleic acid and reducing power). Moreover, these extracts had greater amounts of total phenols and flavonoids in comparison to others. The increase in antioxidant activity (decrease in EC50 values) was accompanied by an increase in the amount of total phenols in the extracts. The pollen extracts contained linoleic acid and α-linolenic acid as major fatty acids, followed by palmitic acid, and oleic acid. In this study, differences were observed in both chemical constituents and biological activities of the samples related to the geographical and botanical origin of bee pollen. Full article
(This article belongs to the Special Issue Nutraceuticals in Human Health and Disease)
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Open AccessArticle Transcriptomic Analysis of Calcium Remodeling in Colorectal Cancer
Int. J. Mol. Sci. 2017, 18(5), 922; doi:10.3390/ijms18050922
Received: 8 March 2017 / Revised: 6 April 2017 / Accepted: 13 April 2017 / Published: 27 April 2017
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Abstract
Colorectal cancer (CRC) cells undergo the remodeling of intracellular Ca2+ homeostasis, which contributes to cancer hallmarks such as enhanced proliferation, invasion and survival. Ca2+ remodeling includes critical changes in store-operated Ca2+ entry (SOCE) and Ca2+ store content. Some changes
[...] Read more.
Colorectal cancer (CRC) cells undergo the remodeling of intracellular Ca2+ homeostasis, which contributes to cancer hallmarks such as enhanced proliferation, invasion and survival. Ca2+ remodeling includes critical changes in store-operated Ca2+ entry (SOCE) and Ca2+ store content. Some changes have been investigated at the molecular level. However, since nearly 100 genes are involved in intracellular Ca2+ transport, a comprehensive view of Ca2+ remodeling in CRC is lacking. We have used Next Generation Sequencing (NGS) to investigate differences in expression of 77 selected gene transcripts involved in intracellular Ca2+ transport in CRC. To this end, mRNA from normal human colonic NCM460 cells and human colon cancer HT29 cells was isolated and used as a template for transcriptomic sequencing and expression analysis using Ion Torrent technology. After data transformation and filtering, exploratory analysis revealed that both cell types were well segregated. In addition, differential gene expression using R and bioconductor packages show significant differences in expression of selected voltage-operated Ca2+ channels and store-operated Ca2+ entry players, transient receptor potential (TRP) channels, Ca2+ release channels, Ca2+ pumps, Na+/Ca2+ exchanger isoforms and genes involved in mitochondrial Ca2+ transport. These data provide the first comprehensive transcriptomic analysis of Ca2+ remodeling in CRC. Full article
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Open AccessArticle Genome-Wide Analyses of MicroRNA Profiling in Interleukin-27 Treated Monocyte-Derived Human Dendritic Cells Using Deep Sequencing: A Pilot Study
Int. J. Mol. Sci. 2017, 18(5), 925; doi:10.3390/ijms18050925
Received: 25 January 2017 / Revised: 20 April 2017 / Accepted: 25 April 2017 / Published: 28 April 2017
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Abstract
MicroRNAs (miRNAs) regulate gene expression and thereby influence cell fate and function. Recent studies suggest that an abundant class of miRNAs play important roles in immune cells, such as T cells, natural killer (NK) cells, B cells, and dendritic cells (DCs). Interleukin (IL)-27
[...] Read more.
MicroRNAs (miRNAs) regulate gene expression and thereby influence cell fate and function. Recent studies suggest that an abundant class of miRNAs play important roles in immune cells, such as T cells, natural killer (NK) cells, B cells, and dendritic cells (DCs). Interleukin (IL)-27 is a member of the IL-12 family of cytokines with broad anti-viral effects. It is a potent inhibitor of HIV-1 infection in CD4+ T cells and macrophages, as well as monocyte-derived immature dendritic cells (iDCs). This pilot study compared miRNA profiles between iDCs and IL-27-treated iDCs (27DCs) using deep sequencing methods and identified 46 known miRNAs that were significantly differentially expressed in 27DCs: 36 were upregulated and 10 downregulated by IL-27. Many of the potential target genes of these miRNAs are involved in IL-27 associated pathways, such as JAK/STAT, MAPKs, and PI3K and several were also previously reported to be involved in the regulation of human DC function. This study found that these miRNAs also potentially target several viral genomes and therefore may have antiviral effects. Four of these differential miRNAs (miR-99a-5p, miR-222-3p, miR-138-5p, and miR-125b-5p) were validated using quantitative reverse transcription PCR (RT-qPCR). Twenty-two novel miRNAs were discovered from deep sequencing and confirmed using RT-qPCR. This study furthers the understanding of the role of IL-27 in immunity and lays a foundation for future characterization of the role of specific miRNAs in DCs. Full article
(This article belongs to the Special Issue Transcriptome Profiling in Human Diseases)
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Open AccessArticle Estrogen Modulates Specific Life and Death Signals Induced by LH and hCG in Human Primary Granulosa Cells In Vitro
Int. J. Mol. Sci. 2017, 18(5), 926; doi:10.3390/ijms18050926
Received: 28 February 2017 / Revised: 31 March 2017 / Accepted: 25 April 2017 / Published: 28 April 2017
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Abstract
Luteinizing hormone (LH) and human chorionic gonadotropin (hCG) are glycoprotein hormones used for assisted reproduction acting on the same receptor (LHCGR) and mediating different intracellular signaling. We evaluated the pro- and anti-apoptotic effect of 100 pM LH or hCG, in the presence or
[...] Read more.
Luteinizing hormone (LH) and human chorionic gonadotropin (hCG) are glycoprotein hormones used for assisted reproduction acting on the same receptor (LHCGR) and mediating different intracellular signaling. We evaluated the pro- and anti-apoptotic effect of 100 pM LH or hCG, in the presence or in the absence of 200 pg/mL 17β-estradiol, in long-term, serum-starved human primary granulosa cells (hGLC) and a transfected granulosa cell line overexpressing LHCGR (hGL5/LHCGR). To this purpose, phospho-extracellular-regulated kinase 1/2 (pERK1/2), protein kinase B (pAKT), cAMP-responsive element binding protein (pCREB) activation and procaspase 3 cleavage were evaluated over three days by Western blotting, along with the expression of target genes by real-time PCR and cell viability by colorimetric assay. We found that LH induced predominant pERK1/2 and pAKT activation STARD1, CCND2 and anti-apoptotic XIAP gene expression, while hCG mediated more potent CREB phosphorylation, expression of CYP19A1 and procaspase 3 cleavage than LH. Cell treatment by LH is accompanied by increased (serum-starved) cell viability, while hCG decreased the number of viable cells. The hCG-specific, pro-apoptotic effect was blocked by a physiological dose of 17β-estradiol, resulting in pAKT activation, lack of procaspase 3 cleavage and increased cell viability. These results confirm that relatively high levels of steroidogenic pathway activation are linked to pro-apoptotic signals in vitro, which may be counteracted by other factors, i.e., estrogens. Full article
(This article belongs to the Special Issue hCG—An Endocrine, Regulator of Gestation and Cancer)
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Open AccessArticle Plasticizing Effects of Polyamines in Protein-Based Films
Int. J. Mol. Sci. 2017, 18(5), 1026; doi:10.3390/ijms18051026
Received: 7 April 2017 / Revised: 5 May 2017 / Accepted: 6 May 2017 / Published: 10 May 2017
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Abstract
Zeta potential and nanoparticle size were determined on film forming solutions of native and heat-denatured proteins of bitter vetch as a function of pH and of different concentrations of the polyamines spermidine and spermine, both in the absence and presence of the plasticizer
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Zeta potential and nanoparticle size were determined on film forming solutions of native and heat-denatured proteins of bitter vetch as a function of pH and of different concentrations of the polyamines spermidine and spermine, both in the absence and presence of the plasticizer glycerol. Our results showed that both polyamines decreased the negative zeta potential of all samples under pH 8.0 as a consequence of their ionic interaction with proteins. At the same time, they enhanced the dimension of nanoparticles under pH 8.0 as a result of macromolecular aggregations. By using native protein solutions, handleable films were obtained only from samples containing either a minimum of 33 mM glycerol or 4 mM spermidine, or both compounds together at lower glycerol concentrations. However, 2 mM spermidine was sufficient to obtain handleable film by using heat-treated samples without glycerol. Conversely, brittle materials were obtained by spermine alone, thus indicating that only spermidine was able to act as an ionic plasticizer. Lastly, both polyamines, mainly spermine, were found able to act as “glycerol-like” plasticizers at concentrations higher than 5 mM under experimental conditions at which their amino groups are undissociated. Our findings open new perspectives in obtaining protein-based films by using aliphatic polycations as components. Full article
(This article belongs to the Special Issue Biodegradable Materials 2017)
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Open AccessArticle The Effect of VPA on Increasing Radiosensitivity in Osteosarcoma Cells and Primary-Culture Cells from Chemical Carcinogen-Induced Breast Cancer in Rats
Int. J. Mol. Sci. 2017, 18(5), 1027; doi:10.3390/ijms18051027
Received: 30 March 2017 / Revised: 30 April 2017 / Accepted: 5 May 2017 / Published: 10 May 2017
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Abstract
This study explored whether valproic acid (VPA, a histone deacetylase inhibitor) could radiosensitize osteosarcoma and primary-culture tumor cells, and determined the mechanism of VPA-induced radiosensitization. The working system included osteosarcoma cells (U2OS) and primary-culture cells from chemical carcinogen (DMBA)-induced breast cancer in rats;
[...] Read more.
This study explored whether valproic acid (VPA, a histone deacetylase inhibitor) could radiosensitize osteosarcoma and primary-culture tumor cells, and determined the mechanism of VPA-induced radiosensitization. The working system included osteosarcoma cells (U2OS) and primary-culture cells from chemical carcinogen (DMBA)-induced breast cancer in rats; and clonogenic survival, immunofluorescence, fluorescent in situ hybridization (FISH) for chromosome aberrations, and comet assays were used in this study. It was found that VPA at the safe or critical safe concentration of 0.5 or 1.0 mM VPA could result in the accumulation of more ionizing radiation (IR)-induced DNA double strand breaks, and increase the cell radiosensitivity. VPA-induced radiosensitivity was associated with the inhibition of DNA repair activity in the working systems. In addition, the chromosome aberrations including chromosome breaks, chromatid breaks, and radial structures significantly increased after the combination treatment of VPA and IR. Importantly, the results obtained by primary-culture cells from the tissue of chemical carcinogen-induced breast cancer in rats further confirmed our findings. The data in this study demonstrated that VPA at a safe dose was a radiosensitizer for osteosarcoma and primary-culture tumor cells through suppressing DNA-double strand breaks repair function. Full article
(This article belongs to the Special Issue Chemically-Induced DNA Damage, Mutagenesis, and Cancer)
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Open AccessArticle Identification and Expression Profiling of the Auxin Response Factors in Dendrobium officinale under Abiotic Stresses
Int. J. Mol. Sci. 2017, 18(5), 927; doi:10.3390/ijms18050927
Received: 14 March 2017 / Revised: 24 April 2017 / Accepted: 25 April 2017 / Published: 4 May 2017
Cited by 1 | PDF Full-text (12922 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Auxin response factor (ARF) proteins play roles in plant responses to diverse environmental stresses by binding specifically to the auxin response element in the promoters of target genes. Using our latest public Dendrobium transcriptomes, a comprehensive characterization and analysis of 14 DnARF genes
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Auxin response factor (ARF) proteins play roles in plant responses to diverse environmental stresses by binding specifically to the auxin response element in the promoters of target genes. Using our latest public Dendrobium transcriptomes, a comprehensive characterization and analysis of 14 DnARF genes were performed. Three selected DnARFs, including DnARF1, DnARF4, and DnARF6, were confirmed to be nuclear proteins according to their transient expression in epidermal cells of Nicotiana benthamiana leaves. Furthermore, the transcription activation abilities of DnARF1, DnARF4, and DnARF6 were tested in a yeast system. Our data showed that DnARF6 is a transcriptional activator in Dendrobium officinale. To uncover the basic information of DnARF gene responses to abiotic stresses, we analyzed their expression patterns under various hormones and abiotic treatments. Based on our data, several hormones and significant stress responsive DnARF genes have been identified. Since auxin and ARF genes have been identified in many plant species, our data is imperative to reveal the function of ARF mediated auxin signaling in the adaptation to the challenging Dendrobium environment. Full article
(This article belongs to the Section Molecular Botany)
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Open AccessArticle Molecular Ghrelin System in the Pancreatic Acinar Cells: The Role of the Polypeptide, Caerulein and Sensory Nerves
Int. J. Mol. Sci. 2017, 18(5), 929; doi:10.3390/ijms18050929
Received: 28 February 2017 / Revised: 9 April 2017 / Accepted: 19 April 2017 / Published: 2 May 2017
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Abstract
Ghrelin (GHRL) is an endogenous ligand for the growth hormone secretagogue receptor (GHS-R). Experimental studies showed that GHRL protects the stomach and pancreas against acute damage, but the effect of GHRL on pancreatic acinar cells was still undetermined. Aim: To investigate the effect
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Ghrelin (GHRL) is an endogenous ligand for the growth hormone secretagogue receptor (GHS-R). Experimental studies showed that GHRL protects the stomach and pancreas against acute damage, but the effect of GHRL on pancreatic acinar cells was still undetermined. Aim: To investigate the effect of GHRL and caerulein on the functional ghrelin system in pancreatic acinar cells taking into account the role of sensory nerves (SN). Methods: Experiments were carried out on isolated pancreatic acinar cells and AR42J cells. Before acinar cells isolation, GHRL was administered intraperitoneally at a dose of 50 µg/kg to rats with intact SN or with capsaicin deactivation of SN (CDSN). After isolation, pancreatic acinar cells were incubated in caerulein-free or caerulein containing solution. AR42J cells were incubated under basal conditions and stimulated with caerulein, GHRL or a combination of the above. Results: Incubation of isolated acinar cells with caerulein inhibited GHS-R and GHRL expression at the level of mRNA and protein in those cells. Either in rats with intact SN or with CDSN, administration of GHRL before isolation of acinar cells increased expression of GHRL and GHS-R in those cells and reversed the caerulein-induced reduction in expression of those parameters. Similar upregulation of GHS-R and GHRL was observed after administration of GHRL in AR42J cells. Conclusions: GHRL stimulates its own expression and expression of its receptor in isolated pancreatic acinar cells and AR42J cells on the positive feedback pathway. This mechanism seems to participate in the pancreatoprotective effect of GHRL in the course of acute pancreatitis. Full article
(This article belongs to the Special Issue Pancreatic Disorders)
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Open AccessArticle PCVMZM: Using the Probabilistic Classification Vector Machines Model Combined with a Zernike Moments Descriptor to Predict Protein–Protein Interactions from Protein Sequences
Int. J. Mol. Sci. 2017, 18(5), 1029; doi:10.3390/ijms18051029
Received: 24 March 2017 / Revised: 24 April 2017 / Accepted: 29 April 2017 / Published: 11 May 2017
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Abstract
Protein–protein interactions (PPIs) are essential for most living organisms’ process. Thus, detecting PPIs is extremely important to understand the molecular mechanisms of biological systems. Although many PPIs data have been generated by high-throughput technologies for a variety of organisms, the whole interatom is
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Protein–protein interactions (PPIs) are essential for most living organisms’ process. Thus, detecting PPIs is extremely important to understand the molecular mechanisms of biological systems. Although many PPIs data have been generated by high-throughput technologies for a variety of organisms, the whole interatom is still far from complete. In addition, the high-throughput technologies for detecting PPIs has some unavoidable defects, including time consumption, high cost, and high error rate. In recent years, with the development of machine learning, computational methods have been broadly used to predict PPIs, and can achieve good prediction rate. In this paper, we present here PCVMZM, a computational method based on a Probabilistic Classification Vector Machines (PCVM) model and Zernike moments (ZM) descriptor for predicting the PPIs from protein amino acids sequences. Specifically, a Zernike moments (ZM) descriptor is used to extract protein evolutionary information from Position-Specific Scoring Matrix (PSSM) generated by Position-Specific Iterated Basic Local Alignment Search Tool (PSI-BLAST). Then, PCVM classifier is used to infer the interactions among protein. When performed on PPIs datasets of Yeast and H. Pylori, the proposed method can achieve the average prediction accuracy of 94.48% and 91.25%, respectively. In order to further evaluate the performance of the proposed method, the state-of-the-art support vector machines (SVM) classifier is used and compares with the PCVM model. Experimental results on the Yeast dataset show that the performance of PCVM classifier is better than that of SVM classifier. The experimental results indicate that our proposed method is robust, powerful and feasible, which can be used as a helpful tool for proteomics research. Full article
(This article belongs to the Special Issue Special Protein Molecules Computational Identification)
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Open AccessArticle The Potential of Triterpenoids from Loquat Leaves (Eriobotrya japonica) for Prevention and Treatment of Skin Disorder
Int. J. Mol. Sci. 2017, 18(5), 1030; doi:10.3390/ijms18051030
Received: 25 February 2017 / Revised: 5 May 2017 / Accepted: 6 May 2017 / Published: 11 May 2017
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Abstract
The leaves of loquat (Eriobotrya japonica) possess high medicinal value and have been used as traditional medicines. However, there are no evidence-based studies on the skin-care effects of E. japonica leaves. To explore new biological activities of E. japonica leaves against
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The leaves of loquat (Eriobotrya japonica) possess high medicinal value and have been used as traditional medicines. However, there are no evidence-based studies on the skin-care effects of E. japonica leaves. To explore new biological activities of E. japonica leaves against skin disorder and to gain a better understanding of the chemical components associated with bioactivities, we evaluated 18 triterpenoids from E. japonica leaves on anti-melanogenesis, anti-acne, anti-allergy and anti-aging activities. Our results revealed that eight compounds showed anti-melanogenesis activity, of which ursolic acid (1) and maslinic acid (7) were the most potent with the similar selective index to that of arbutin. Structure–activity relationship and possible mechanism of active compounds were proposed. Twelve compounds exhibited anti-acne effect; ursolic acid (1), maslinic acid (7), corosolic acid (8) and euscaphic acid (12) showed highest activities against P. acnes. Four compounds displayed anti-allergy and anti-inflammatory activity; 3-epicorosolic acid (9) and euscaphic acid (12) showed marked activity against β-hexosaminidase release. Finally, ursolic acid (1), pomolic acid (10), colosolic acid (8) and its methylated derivative (6) exhibited the highest anti-aging activity by stimulating collagen and hyaluronic acid (HA) production. Our findings provide valuable evidence that E. japonica leaves have potential applications as ingredients of function foods or cosmetics for health benefits and a number of triterpenoids may play an important role in these bioactivities. Full article
(This article belongs to the Section Bioactives and Nutraceuticals)
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Open AccessArticle Redistribution of Cerebral Blood Flow during Severe Hypovolemia and Reperfusion in a Sheep Model: Critical Role of α1-Adrenergic Signaling
Int. J. Mol. Sci. 2017, 18(5), 1031; doi:10.3390/ijms18051031
Received: 22 March 2017 / Revised: 5 May 2017 / Accepted: 8 May 2017 / Published: 11 May 2017
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Abstract
Background: Maintenance of brain circulation during shock is sufficient to prevent subcortical injury but the cerebral cortex is not spared. This suggests area-specific regulation of cerebral blood flow (CBF) during hemorrhage. Methods: Cortical and subcortical CBF were continuously measured during blood loss (≤50%)
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Background: Maintenance of brain circulation during shock is sufficient to prevent subcortical injury but the cerebral cortex is not spared. This suggests area-specific regulation of cerebral blood flow (CBF) during hemorrhage. Methods: Cortical and subcortical CBF were continuously measured during blood loss (≤50%) and subsequent reperfusion using laser Doppler flowmetry. Blood gases, mean arterial blood pressure (MABP), heart rate and renal blood flow were also monitored. Urapidil was used for α1A-adrenergic receptor blockade in dosages, which did not modify the MABP-response to blood loss. Western blot and quantitative reverse transcription polymerase chain reactions were used to determine adrenergic receptor expression in brain arterioles. Results: During hypovolemia subcortical CBF was maintained at 81 ± 6% of baseline, whereas cortical CBF decreased to 40 ± 4% (p < 0.001). Reperfusion led to peak CBFs of about 70% above baseline in both brain regions. α1A-Adrenergic blockade massively reduced subcortical CBF during hemorrhage and reperfusion, and prevented hyperperfusion during reperfusion in the cortex. α1A-mRNA expression was significantly higher in the cortex, whereas α1D-mRNA expression was higher in the subcortex (p < 0.001). Conclusions: α1-Adrenergic receptors are critical for perfusion redistribution: activity of the α1A-receptor subtype is a prerequisite for redistribution of CBF, whereas the α1D-receptor subtype may determine the magnitude of redistribution responses. Full article
(This article belongs to the Special Issue Cerebral Blood Flow and Metabolism)
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Open AccessArticle Fasting Enhances the Contrast of Bone Metastatic Lesions in 18F-Fluciclovine-PET: Preclinical Study Using a Rat Model of Mixed Osteolytic/Osteoblastic Bone Metastases
Int. J. Mol. Sci. 2017, 18(5), 934; doi:10.3390/ijms18050934
Received: 27 March 2017 / Revised: 21 April 2017 / Accepted: 26 April 2017 / Published: 29 April 2017
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Abstract
18F-fluciclovine (trans-1-amino-3-18F-fluorocyclobutanecarboxylic acid) is an amino acid positron emission tomography (PET) tracer used for cancer staging (e.g., prostate and breast). Patients scheduled to undergo amino acid-PET are usually required to fast before PET tracer administration. However, there have
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18F-fluciclovine (trans-1-amino-3-18F-fluorocyclobutanecarboxylic acid) is an amino acid positron emission tomography (PET) tracer used for cancer staging (e.g., prostate and breast). Patients scheduled to undergo amino acid-PET are usually required to fast before PET tracer administration. However, there have been no reports addressing whether fasting improves fluciclovine-PET imaging. In this study, the authors investigated the influence of fasting on fluciclovine-PET using triple-tracer autoradiography with 14C-fluciclovine, [5,6-3H]-2-fluoro-2-deoxy-d-glucose (3H-FDG), and 99mTc-hydroxymethylene diphosphonate (99mTc-HMDP) in a rat breast cancer model of mixed osteolytic/osteoblastic bone metastases in which the animals fasted overnight. Lesion accumulation of each tracer was evaluated using the target-to-background (muscle) ratio. The mean ratios of 14C-fluciclovine in osteolytic lesions were 4.6 ± 0.8 and 2.8 ± 0.6, respectively, with and without fasting, while those for 3H-FDG were 6.9 ± 2.5 and 5.1 ± 2.0, respectively. In the peri-tumor bone formation regions (osteoblastic), where 99mTc-HMDP accumulated, the ratios of 14C-fluciclovine were 4.3 ± 1.4 and 2.4 ± 0.7, respectively, and those of 3H-FDG were 6.2 ± 3.8 and 3.3 ± 2.2, respectively, with and without fasting. These results suggest that fasting before 18F-fluciclovine-PET improves the contrast between osteolytic and osteoblastic bone metastatic lesions and background, as well as 18F-FDG-PET. Full article
(This article belongs to the Special Issue Diagnostic, Prognostic and Predictive Biomarkers in Prostate Cancer)
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Open AccessArticle An AAAG-Rich Oligodeoxynucleotide Rescues Mice from Bacterial Septic Peritonitis by Interfering Interferon Regulatory Factor 5
Int. J. Mol. Sci. 2017, 18(5), 1034; doi:10.3390/ijms18051034
Received: 9 March 2017 / Revised: 4 May 2017 / Accepted: 6 May 2017 / Published: 11 May 2017
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Abstract
A previous study found that an AAAG-rich Oligodeoxynucleotide (ODN), designated as MS19, could lessen the acute lung inflammatory injury (ALII) in mice infected by influenza viruses. Bioinformatics analysis found that MS19 is consensus with the binding site of interferon regulatory factor 5 (IRF5)
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A previous study found that an AAAG-rich Oligodeoxynucleotide (ODN), designated as MS19, could lessen the acute lung inflammatory injury (ALII) in mice infected by influenza viruses. Bioinformatics analysis found that MS19 is consensus with the binding site of interferon regulatory factor 5 (IRF5) in the regulatory elements of pro-inflammatory genes. This study established a septic peritonitis model in Institute of Cancer Research (ICR) mice infected with Escherichia coli (E. coli), and found that MS19 prolonged the survival of the mice and down-regulated the expression of inducible nitric oxide synthase (iNOS), interleukin-6 (IL-6), and tumor necrosis factor α (TNF-α). In cultured RAW264.7 cells, MS19 significantly reduced the expression of iNOS, IRF5, IL-6, and TNF-α and inhibited the nuclear translocation of IRF5. This data may provide a new insight for understanding how MS19 reduces the excessive inflammatory responses in sepsis. Full article
(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)
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Open AccessArticle Nobiletin Inhibits Angiogenesis by Regulating Src/FAK/STAT3-Mediated Signaling through PXN in ER+ Breast Cancer Cells
Int. J. Mol. Sci. 2017, 18(5), 935; doi:10.3390/ijms18050935
Received: 9 March 2017 / Revised: 25 April 2017 / Accepted: 26 April 2017 / Published: 30 April 2017
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Abstract
Tumor angiogenesis is one of the major hallmarks of tumor progression. Nobiletin is a natural flavonoid isolated from citrus peel that has anti-angiogenic activity. Steroid receptor coactivator (Src) is an intracellular tyrosine kinase so that focal adhesion kinase (FAK) binds to Src to
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Tumor angiogenesis is one of the major hallmarks of tumor progression. Nobiletin is a natural flavonoid isolated from citrus peel that has anti-angiogenic activity. Steroid receptor coactivator (Src) is an intracellular tyrosine kinase so that focal adhesion kinase (FAK) binds to Src to play a role in tumor angiogenesis. Signal transducer and activator of transcription 3 (STAT3) is a marker for tumor angiogenesis which interacts with Src. Paxillin (PXN) acts as a downstream target for both FAK and STAT3. The main goal of this study was to assess inhibition of tumor angiogenesis by nobiletin in estrogen receptor positive (ER+) breast cancer cells via Src, FAK, and STAT3-mediated signaling through PXN. Treatment with nobiletin in MCF-7 and T47D breast cancer cells inhibited angiogenesis markers, based on western blotting and RT-PCR. Validation of in vitro angiogenesis in the human umbilical vein endothelial cells (HUVEC) endothelial cell line proved the anti-angiogenic activity of nobiletin. Electrophoretic mobility shift assay and the ChIP assay showed that nobiletin inhibits STAT3/DNA binding activity and STAT3 binding to a novel binding site of the PXN gene promoter. We also investigated the migration and invasive ability of nobiletin in ER+ cells. Nobiletin inhibited tumor angiogenesis by regulating Src, FAK, and STAT3 signaling through PXN in ER+ breast cancer cells. Full article
(This article belongs to the Special Issue Tumor Targeting Therapy and Selective Killing)
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Open AccessArticle Circulating Cell-Free DNA and Circulating Tumor Cells as Prognostic and Predictive Biomarkers in Advanced Non-Small Cell Lung Cancer Patients Treated with First-Line Chemotherapy
Int. J. Mol. Sci. 2017, 18(5), 1035; doi:10.3390/ijms18051035
Received: 7 April 2017 / Revised: 4 May 2017 / Accepted: 5 May 2017 / Published: 11 May 2017
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Abstract
Cell-free DNA (cfDNA) and circulating tumor cells (CTCs) are promising prognostic and predictive biomarkers in non-small cell lung cancer (NSCLC). In this study, we examined the prognostic role of cfDNA and CTCs, in separate and joint analyses, in NSCLC patients receiving first line
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Cell-free DNA (cfDNA) and circulating tumor cells (CTCs) are promising prognostic and predictive biomarkers in non-small cell lung cancer (NSCLC). In this study, we examined the prognostic role of cfDNA and CTCs, in separate and joint analyses, in NSCLC patients receiving first line chemotherapy. Seventy-three patients with advanced NSCLC were enrolled in this study. CfDNA and CTC were analyzed at baseline and after two cycles of chemotherapy. Plasma cfDNA quantification was performed by quantitative PCR (qPCR) whereas CTCs were isolated by the ScreenCell Cyto (ScreenCell, Paris, France) device and enumerated according to malignant features. Patients with baseline cfDNA higher than the median value (96.3 hTERT copy number) had a significantly worse overall survival (OS) and double the risk of death (hazard ratio (HR): 2.14; 95% confidence limits (CL) = 1.24–3.68; p-value = 0.006). Conversely, an inverse relationship between CTC median baseline number (6 CTC/3 mL of blood) and OS was observed. In addition, we found that in patients reporting stable disease (SD), the baseline cfDNA and CTCs were able to discriminate patients at high risk of poor survival. cfDNA demonstrated a more reliable biomarker than CTCs in the overall population. In the subgroup of SD patients, both biomarkers identified patients at high risk of poor prognosis who might deserve additional/alternative therapeutic interventions. Full article
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Open AccessArticle Pan-Cancer Mutational and Transcriptional Analysis of the Integrator Complex
Int. J. Mol. Sci. 2017, 18(5), 936; doi:10.3390/ijms18050936
Received: 10 April 2017 / Revised: 20 April 2017 / Accepted: 23 April 2017 / Published: 29 April 2017
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Abstract
The integrator complex has been recently identified as a key regulator of RNA Polymerase II-mediated transcription, with many functions including the processing of small nuclear RNAs, the pause-release and elongation of polymerase during the transcription of protein coding genes, and the biogenesis of
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The integrator complex has been recently identified as a key regulator of RNA Polymerase II-mediated transcription, with many functions including the processing of small nuclear RNAs, the pause-release and elongation of polymerase during the transcription of protein coding genes, and the biogenesis of enhancer derived transcripts. Moreover, some of its components also play a role in genome maintenance. Thus, it is reasonable to hypothesize that their functional impairment or altered expression can contribute to malignancies. Indeed, several studies have described the mutations or transcriptional alteration of some Integrator genes in different cancers. Here, to draw a comprehensive pan-cancer picture of the genomic and transcriptomic alterations for the members of the complex, we reanalyzed public data from The Cancer Genome Atlas. Somatic mutations affecting Integrator subunit genes and their transcriptional profiles have been investigated in about 11,000 patients and 31 tumor types. A general heterogeneity in the mutation frequencies was observed, mostly depending on tumor type. Despite the fact that we could not establish them as cancer drivers, INTS7 and INTS8 genes were highly mutated in specific cancers. A transcriptome analysis of paired (normal and tumor) samples revealed that the transcription of INTS7, INTS8, and INTS13 is significantly altered in several cancers. Experimental validation performed on primary tumors confirmed these findings. Full article
(This article belongs to the Special Issue Transcriptome Profiling in Human Diseases)
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Open AccessArticle Selective Expression of Flt3 within the Mouse Hematopoietic Stem Cell Compartment
Int. J. Mol. Sci. 2017, 18(5), 1037; doi:10.3390/ijms18051037
Received: 20 April 2017 / Revised: 5 May 2017 / Accepted: 5 May 2017 / Published: 12 May 2017
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Abstract
The fms-like tyrosine kinase 3 (Flt3) is a cell surface receptor that is expressed by various hematopoietic progenitor cells (HPC) and Flt3-activating mutations are commonly present in acute myeloid and lymphoid leukemias. These findings underscore the importance of Flt3 to steady-state and malignant
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The fms-like tyrosine kinase 3 (Flt3) is a cell surface receptor that is expressed by various hematopoietic progenitor cells (HPC) and Flt3-activating mutations are commonly present in acute myeloid and lymphoid leukemias. These findings underscore the importance of Flt3 to steady-state and malignant hematopoiesis. In this study, the expression of Flt3 protein and Flt3 mRNA by single cells within the hematopoietic stem cell (HSC) and HPC bone marrow compartments of C57/BL6 mice was investigated using flow cytometry and the quantitative reverse transcription polymerase chain reaction. Flt3 was heterogeneously expressed by almost all of the populations studied, including long-term reconstituting HSC and short-term reconstituting HSC. The erythropoietin receptor (EpoR) and macrophage colony-stimulating factor receptor (M-CSFR) were also found to be heterogeneously expressed within the multipotent cell compartments. Co-expression of the mRNAs encoding Flt3 and EpoR rarely occurred within these compartments. Expression of both Flt3 and M-CSFR protein at the surface of single cells was more commonly observed. These results emphasize the heterogeneous nature of HSC and HPC and the new sub-populations identified are important to understanding the origin and heterogeneity of the acute myeloid leukemias. Full article
(This article belongs to the Special Issue The Biology and Treatment of Myeloid Leukaemias)
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Open AccessArticle Applying Unconventional Secretion in Ustilago maydis for the Export of Functional Nanobodies
Int. J. Mol. Sci. 2017, 18(5), 937; doi:10.3390/ijms18050937
Received: 28 February 2017 / Revised: 21 April 2017 / Accepted: 24 April 2017 / Published: 29 April 2017
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Abstract
Exploiting secretory pathways for production of heterologous proteins is highly advantageous with respect to efficient downstream processing. In eukaryotic systems the vast majority of heterologous proteins for biotechnological application is exported via the canonical endoplasmic reticulum–Golgi pathway. In the endomembrane system target proteins
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Exploiting secretory pathways for production of heterologous proteins is highly advantageous with respect to efficient downstream processing. In eukaryotic systems the vast majority of heterologous proteins for biotechnological application is exported via the canonical endoplasmic reticulum–Golgi pathway. In the endomembrane system target proteins are often glycosylated and may thus be modified with foreign glycan patterns. This can be destructive for their activity or cause immune reactions against therapeutic proteins. Hence, using unconventional secretion for protein expression is an attractive alternative. In the fungal model Ustilago maydis, chitinase Cts1 is secreted via an unconventional pathway connected to cell separation which can be used to co-export heterologous proteins. Here, we apply this mechanism for the production of nanobodies. First, we achieved expression and unconventional secretion of a functional nanobody directed against green fluorescent protein (Gfp). Second, we found that Cts1 binds to chitin and that this feature can be applied to generate a Gfp-trap. Thus, we demonstrated the dual use of Cts1 serving both as export vehicle and as purification tag. Finally, we established and optimized the production of a nanobody against botulinum toxin A and hence describe the first pharmaceutically relevant target exported by Cts1-mediated unconventional secretion. Full article
(This article belongs to the Special Issue Unconventional Proteins and Membranes Traffic)
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Open AccessArticle The Selective Centrifugation Ensures a Better In Vitro Isolation of ASCs and Restores a Soft Tissue Regeneration In Vivo
Int. J. Mol. Sci. 2017, 18(5), 1038; doi:10.3390/ijms18051038
Received: 31 January 2017 / Revised: 26 March 2017 / Accepted: 28 April 2017 / Published: 12 May 2017
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Abstract
Autologous fat grafting procedures in plastic surgery have been extensively used to reinforce soft tissue in congenital or acquired tissue impairments. With this background, the aim of this study is firstly to examine the impact of a selective centrifugation on existing adipose stem
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Autologous fat grafting procedures in plastic surgery have been extensively used to reinforce soft tissue in congenital or acquired tissue impairments. With this background, the aim of this study is firstly to examine the impact of a selective centrifugation on existing adipose stem cells (ASCs) in terms of stemness profile maintenance and, secondly, to investigate the effect of restoring volume in reconstruction on patients affected by soft tissue damage. After centrifugation, the fat graft products were separated into two layers and subsequently examined in vitro for the expression of CD34, CD90, CD117, CD105, CD29, CD31, CD44, CD73, CD133, CD14 and CD45 markers by flow cytometry and gene expression analyses were performed for Sox2, WNT3A, END, CD44, FUT4, COLL1, CTNNB1, hbEGF, KRTLG, MMP2 and VIM genes. The results showed that in the middle-high density (MHD) layer there was a peak concentration of ASCs, compared to another layer obtained after centrifugation. Research carried out on patients under treatment for soft tissue regeneration using cells obtained from MHD layer selection will be fundamental in comparative analysis. These studies will lead to an adequate standardization of outcomes, provided that treatment is performed through cell selection. Therefore, a unique procedure in tissue reconstruction and regeneration through fat grafting is presented here. Full article
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Open AccessArticle Clock Genes and Altered Sleep–Wake Rhythms: Their Role in the Development of Psychiatric Disorders
Int. J. Mol. Sci. 2017, 18(5), 938; doi:10.3390/ijms18050938
Received: 19 October 2016 / Revised: 4 March 2017 / Accepted: 9 March 2017 / Published: 29 April 2017
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Abstract
In mammals, the circadian clocks network (central and peripheral oscillators) controls circadian rhythms and orchestrates the expression of a range of downstream genes, allowing the organism to anticipate and adapt to environmental changes. Beyond their role in circadian rhythms, several studies have highlighted
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In mammals, the circadian clocks network (central and peripheral oscillators) controls circadian rhythms and orchestrates the expression of a range of downstream genes, allowing the organism to anticipate and adapt to environmental changes. Beyond their role in circadian rhythms, several studies have highlighted that circadian clock genes may have a more widespread physiological effect on cognition, mood, and reward-related behaviors. Furthermore, single nucleotide polymorphisms in core circadian clock genes have been associated with psychiatric disorders (such as autism spectrum disorder, schizophrenia, anxiety disorders, major depressive disorder, bipolar disorder, and attention deficit hyperactivity disorder). However, the underlying mechanisms of these associations remain to be ascertained and the cause–effect relationships are not clearly established. The objective of this article is to clarify the role of clock genes and altered sleep–wake rhythms in the development of psychiatric disorders (sleep problems are often observed at early onset of psychiatric disorders). First, the molecular mechanisms of circadian rhythms are described. Then, the relationships between disrupted circadian rhythms, including sleep–wake rhythms, and psychiatric disorders are discussed. Further research may open interesting perspectives with promising avenues for early detection and therapeutic intervention in psychiatric disorders. Full article
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Open AccessArticle Magnetic Hyperthermia and Oxidative Damage to DNA of Human Hepatocarcinoma Cells
Int. J. Mol. Sci. 2017, 18(5), 939; doi:10.3390/ijms18050939
Received: 1 March 2017 / Revised: 13 April 2017 / Accepted: 23 April 2017 / Published: 29 April 2017
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Abstract
Nanotechnology is addressing major urgent needs for cancer treatment. We conducted a study to compare the frequency of 3-(2-deoxy-β-d-erythro-pentafuranosyl)pyrimido[1,2-α]purin-10(3H)-one deoxyguanosine (M1dG) and 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG) adducts, biomarkers of oxidative stress and/or lipid peroxidation, on human hepatocarcinoma HepG2 cells exposed to
[...] Read more.
Nanotechnology is addressing major urgent needs for cancer treatment. We conducted a study to compare the frequency of 3-(2-deoxy-β-d-erythro-pentafuranosyl)pyrimido[1,2-α]purin-10(3H)-one deoxyguanosine (M1dG) and 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG) adducts, biomarkers of oxidative stress and/or lipid peroxidation, on human hepatocarcinoma HepG2 cells exposed to increasing levels of Fe3O4-nanoparticles (NPs) versus untreated cells at different lengths of incubations, and in the presence of increasing exposures to an alternating magnetic field (AMF) of 186 kHz using 32P-postlabeling. The levels of oxidative damage tended to increase significantly after ≥24 h of incubations compared to controls. The oxidative DNA damage tended to reach a steady-state after treatment with 60 μg/mL of Fe3O4-NPs. Significant dose–response relationships were observed. A greater adduct production was observed after magnetic hyperthermia, with the highest amounts of oxidative lesions after 40 min exposure to AMF. The effects of magnetic hyperthermia were significantly increased with exposure and incubation times. Most important, the levels of oxidative lesions in AMF exposed NP treated cells were up to 20-fold greater relative to those observed in nonexposed NP treated cells. Generation of oxidative lesions may be a mechanism by which magnetic hyperthermia induces cancer cell death. Full article
(This article belongs to the Special Issue Chemically-Induced DNA Damage, Mutagenesis, and Cancer)
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Open AccessArticle Structure, Expression, and Functional Analysis of the Hexokinase Gene Family in Cassava
Int. J. Mol. Sci. 2017, 18(5), 1041; doi:10.3390/ijms18051041
Received: 9 April 2017 / Revised: 8 May 2017 / Accepted: 8 May 2017 / Published: 12 May 2017
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Abstract
Hexokinase (HXK) proteins play important roles in catalyzing hexose phosphorylation and sugar sensing and signaling. To investigate the roles of HXKs in cassava tuber root development, seven HXK genes (MeHXK1–7) were isolated and analyzed. A phylogenetic analysis revealed that the MeHXK
[...] Read more.
Hexokinase (HXK) proteins play important roles in catalyzing hexose phosphorylation and sugar sensing and signaling. To investigate the roles of HXKs in cassava tuber root development, seven HXK genes (MeHXK1–7) were isolated and analyzed. A phylogenetic analysis revealed that the MeHXK family can be divided into five subfamilies of plant HXKs. MeHXKs were clearly divided into type A (MeHXK1) and type B (MeHXK2–7) based on their N-terminal sequences. MeHXK1–5 all had typical conserved regions and similar protein structures to the HXKs of other plants; while MeHXK6–7 lacked some of the conserved regions. An expression analysis of the MeHXK genes in cassava organs or tissues demonstrated that MeHXK2 is the dominant HXK in all the examined tissues (leaves, stems, fruits, tuber phloems, and tuber xylems). Notably, the expression of MeHXK2 and the enzymatic activity of HXK were higher at the initial and expanding tuber stages, and lower at the mature tuber stage. Furthermore, the HXK activity of MeHXK2 was identified by functional complementation of the HXK-deficient yeast strain YSH7.4-3C (hxk1, hxk2, glk1). The gene expression and enzymatic activity of MeHXK2 suggest that it might be the main enzyme for hexose phosphorylation during cassava tuber root development, which is involved in sucrose metabolism to regulate the accumulation of starch. Full article
(This article belongs to the Section Molecular Botany)
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Open AccessArticle BDNF Binds Its Pro-Peptide with High Affinity and the Common Val66Met Polymorphism Attenuates the Interaction
Int. J. Mol. Sci. 2017, 18(5), 1042; doi:10.3390/ijms18051042
Received: 7 April 2017 / Revised: 6 May 2017 / Accepted: 8 May 2017 / Published: 12 May 2017
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Abstract
Most growth factors are initially synthesized as precursors then cleaved into bioactive mature domains and pro-domains, but the biological roles of pro-domains are poorly understood. In the present study, we investigated the pro-domain (or pro-peptide) of brain-derived neurotrophic factor (BDNF), which promotes neuronal
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Most growth factors are initially synthesized as precursors then cleaved into bioactive mature domains and pro-domains, but the biological roles of pro-domains are poorly understood. In the present study, we investigated the pro-domain (or pro-peptide) of brain-derived neurotrophic factor (BDNF), which promotes neuronal survival, differentiation and synaptic plasticity. The BDNF pro-peptide is a post-processing product of the precursor BDNF. Using surface plasmon resonance and biochemical experiments, we first demonstrated that the BDNF pro-peptide binds to mature BDNF with high affinity, but not other neurotrophins. This interaction was more enhanced at acidic pH than at neutral pH, suggesting that the binding is significant in intracellular compartments such as trafficking vesicles rather than the extracellular space. The common Val66Met BDNF polymorphism results in a valine instead of a methionine in the pro-domain, which affects human brain functions and the activity-dependent secretion of BDNF. We investigated the influence of this variation on the interaction between BDNF and the pro-peptide. Interestingly, the Val66Met polymorphism stabilized the heterodimeric complex of BDNF and its pro-peptide. Furthermore, compared with the Val-containing pro-peptide, the complex with the Met-type pro-peptide was more stable at both acidic and neutral pH, suggesting that the Val66Met BDNF polymorphism forms a more stable complex. A computational modeling provided an interpretation to the role of the Val66Met mutation in the interaction of BDNF and its pro-peptide. Lastly, we performed electrophysiological experiments, which indicated that the BDNF pro-peptide, when pre-incubated with BDNF, attenuated the ability of BDNF to inhibit hippocampal long-term depression (LTD), suggesting a possibility that the BDNF pro-peptide may interact directly with BDNF and thereby inhibit its availability. It was previously reported that the BDNF pro-domain exerts a chaperone-like function and assists the folding of the BDNF protein. However, our results suggest a new role for the BDNF pro-domain (or pro-peptide) following proteolytic cleave of precursor BDNF, and provide insight into the Val66Met polymorphism. Full article
(This article belongs to the Special Issue Neurotrophic Factors—Historical Perspective and New Directions)
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Open AccessArticle Rutin-Enriched Extract from Coriandrum sativum L. Ameliorates Ionizing Radiation-Induced Hematopoietic Injury
Int. J. Mol. Sci. 2017, 18(5), 942; doi:10.3390/ijms18050942
Received: 16 December 2016 / Revised: 22 April 2017 / Accepted: 24 April 2017 / Published: 29 April 2017
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Abstract
Hematopoietic injury is a major cause of mortality in radiation accidents and a primary side effect in patients undergoing radiotherapy. Ionizing radiation (IR)-induced myelosuppression is largely attributed to the injury of hematopoietic stem and progenitor cells (HSPCs). Coriander is a culinary herb with
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Hematopoietic injury is a major cause of mortality in radiation accidents and a primary side effect in patients undergoing radiotherapy. Ionizing radiation (IR)-induced myelosuppression is largely attributed to the injury of hematopoietic stem and progenitor cells (HSPCs). Coriander is a culinary herb with multiple pharmacological effects and has been widely used in traditional medicine. In this study, flavonoids were identified as the main component of coriander extract with rutin being the leading compound (rutin-enriched coriander extract; RE-CE). We evaluated the radioprotective effect of RE-CE against IR-induced HSPCs injury. Results showed that RE-CE treatment markedly improved survival, ameliorated organ injuries and myelosuppression, elevated HSPCs frequency, and promoted differentiation and proliferation of HSPCs in irradiated mice. The protective role of RE-CE in hematopoietic injury is probably attributed to its anti-apoptotic and anti-DNA damage effect in irradiated HSPCs. Moreover, these changes were associated with reduced reactive oxygen species (ROS) and enhanced antioxidant enzymatic activities in irradiated HSPCs. Collectively, these findings demonstrate that RE-CE is able to ameliorate IR-induced hematopoietic injury partly by reducing IR-induced oxidative stress. Full article
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Open AccessArticle Hypoxia Mediates Differential Response to Anti-EGFR Therapy in HNSCC Cells
Int. J. Mol. Sci. 2017, 18(5), 943; doi:10.3390/ijms18050943
Received: 22 February 2017 / Revised: 19 April 2017 / Accepted: 24 April 2017 / Published: 29 April 2017
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Abstract
Despite advances in the head and neck squamous cell carcinoma (HNSCC) treatment modalities, drug resistance and cancer recurrence are often reported. Hypoxia signaling through hypoxia-inducible factor 1 (HIF-1) promotes angiogenesis and metastasis by inducing epithelial-mesenchymal-transition (EMT). The aim of this study was to
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Despite advances in the head and neck squamous cell carcinoma (HNSCC) treatment modalities, drug resistance and cancer recurrence are often reported. Hypoxia signaling through hypoxia-inducible factor 1 (HIF-1) promotes angiogenesis and metastasis by inducing epithelial-mesenchymal-transition (EMT). The aim of this study was to evaluate the impact of hypoxia on response to therapy as well as EMT and expression of stem cell markers in HNSCC cells. Five HNSCC cell lines (UT-SCC-2, UT-SCC-14, LK0412, LK0827, and LK0923) were selected for this study. The treatment sensitivity for radiation, cisplatin, cetuximab, and dasatinib was assessed by crystal violet assay. Gene expression of EMT and cancer stem cell (CSC) markers as well as protein level of EGFR signaling molecules were analyzed by qPCR and western blotting, respectively. Unlike UT-SCC-14 and LK0827, the LK0412 cell line became significantly more sensitive to cetuximab in hypoxic conditions. This cetuximab sensitivity was efficiently reversed after suppression of HIF-1α with siRNA. Additionally, hypoxia-induced EMT and expression of stem cell markers in HNSCC cells was partially revoked by treatment with cetuximab or knockdown of HIF-1α. In summary, our study shows that hypoxia might have a positive influence on the anti-EGFR therapy effectiveness in HNSCC. However, due to heterogeneity of HNSCC lesions, targeting HIF-1α may not be sufficient to mediate such a response. Further studies identifying a trait of hypoxia-specific response to cetuximab in HNSCC are advisable. Full article
(This article belongs to the Special Issue Cancer Stem Cells)
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Open AccessArticle HepPar1-Positive Circulating Microparticles Are Increased in Subjects with Hepatocellular Carcinoma and Predict Early Recurrence after Liver Resection
Int. J. Mol. Sci. 2017, 18(5), 1043; doi:10.3390/ijms18051043
Received: 10 February 2017 / Revised: 14 April 2017 / Accepted: 1 May 2017 / Published: 12 May 2017
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Abstract
Circulating microparticles (MPs) are novel potential biomarkers in cancer patients. Their role in hepatocellular carcinoma (HCC) is under intensive investigation. In this study, we tested the hypothesis that MPs expressing the antigen HepPar1 are increased in the blood of subjects with HCC and
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Circulating microparticles (MPs) are novel potential biomarkers in cancer patients. Their role in hepatocellular carcinoma (HCC) is under intensive investigation. In this study, we tested the hypothesis that MPs expressing the antigen HepPar1 are increased in the blood of subjects with HCC and may serve as markers of early recurrence after liver resection (LR). We studied 15 patients affected by HCC undergoing LR, and used flow cytometry to assess the number of circulating HepPar1+ MPs. Ten subjects without HCC (five with liver cirrhosis and five with healthy livers) were used as controls. After LR, HCC patients underwent a follow-up to check for early recurrence, which occurred in seven cases. The number of circulating HepPar1+ MPs was significantly higher in subjects affected by HCC, compared to individuals without cancer (p < 0.01). We also found that, among HCC patients, the number of circulating HepPar1+ MPs, measured before LR, was significantly higher in those who displayed early recurrence compared to those without recurrence (p = 0.02). Of note, other types of circulating MPs, such as those derived from endothelial cells (CD144+) or those produced by the activated endothelium (CD144+/CD62+), were not associated with HCC, nor could they predict HCC recurrence. HepPar1+ MPs deserve further investigation as novel biomarkers of disease and prognosis in HCC patients. Full article
(This article belongs to the collection Molecular Mechanisms of Human Liver Diseases)
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Open AccessArticle Adiponectin Is Involved in Connective Tissue Growth Factor-Induced Proliferation, Migration and Overproduction of the Extracellular Matrix in Keloid Fibroblasts
Int. J. Mol. Sci. 2017, 18(5), 1044; doi:10.3390/ijms18051044
Received: 17 March 2017 / Revised: 5 May 2017 / Accepted: 8 May 2017 / Published: 12 May 2017
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Abstract
Adiponectin, an adipocyte-derived hormone, exerts pleiotropic biological effects on metabolism, inflammation, vascular homeostasis, apoptosis and immunity. Recently, adiponectin has been suggested to attenuate the progression of human dermal fibrosis. Connective tissue growth factor (CTGF) is induced in keloids and is thought to be
[...] Read more.
Adiponectin, an adipocyte-derived hormone, exerts pleiotropic biological effects on metabolism, inflammation, vascular homeostasis, apoptosis and immunity. Recently, adiponectin has been suggested to attenuate the progression of human dermal fibrosis. Connective tissue growth factor (CTGF) is induced in keloids and is thought to be participated in the formation of keloid fibrosis. However, the roles played by adiponectin in keloids remain unclear. In this study, we explored the effects of adiponectin on CTGF-induced cell proliferation, migration and the deposition of extracellular matrix (ECM) and their associated intracellular signalling pathways in keloid fibroblasts (KFs). We also explored possible mechanisms of keloid pathogenesis. Primary fibroblast cultures were established from foreskin biopsies and skin biopsies from patients with keloids. The expression of adiponectin and adiponectin receptors (adipoRs) was evaluated by reverse transcription-PCR (RT-PCR), quantitative real-time RT-PCR, immunofluorescence staining, and immunohistochemical analysis. Next, KFs and normal dermal fibroblasts (NFs) were treated with CTGF in the presence or absence of adiponectin. A cell counting kit-8 (CCK-8) and the Transwell assay were used to examine cell proliferation and migration. The level of the collagen I, fibronectin (FN) and α-smooth muscle actin (α-SMA) mRNAs and proteins were determined by quantitative real-time RT-PCR and western blotting. The effects of RNA interference (RNAi) targeting the adipoR genes were detected. Phosphorylation of adenosine 5′-monophosphate (AMP)-activated protein kinase (AMPK), mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3 kinase-protein kinase (PI3K-Akt) were examined by western blotting to further investigate the signalling pathways. Furthermore, inhibitors of signal transduction pathways were investigated. The expression levels of adiponectin and adipoRs were significantly decreased in keloids compared with those in normal skin tissue. Adiponectin suppressed the CTGF-induced KFs, but not NFs, proliferation, migration and ECM production. Moreover, adiponectin inhibited the phosphorylation of AMPK, p38 and extracellular-regulated kinase (ERK), but not that of Jun N-terminal kinase (JNK) or Akt, in CTGF-treated KFs. The activity of adiponectin-mediated signalling pathways was attenuated by small interfering RNAs (siRNAs) targeting adipoR1 (but not siRNAs targeting adipoR2, T-cadherin or calreticulin), AMPK (Compound C), p38 (SB203580) inhibitors, and mitogen-activated protein kinase kinase (MEK) inhibitor (PD98059). Based on our results, adiponectin suppresses CTGF-induced KFs proliferation, migration and ECM overproduction. One of the underlying mechanisms is the activation of the adipoR1, AMPK, p38, and ERK signalling pathways. Therefore, adiponectin may play an important role in the progression of keloids, suggesting a potential novel target for keloid treatment. Full article
(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)
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Open AccessArticle Determination of Genes Related to Uveitis by Utilization of the Random Walk with Restart Algorithm on a Protein–Protein Interaction Network
Int. J. Mol. Sci. 2017, 18(5), 1045; doi:10.3390/ijms18051045
Received: 9 April 2017 / Revised: 8 May 2017 / Accepted: 9 May 2017 / Published: 13 May 2017
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Abstract
Uveitis, defined as inflammation of the uveal tract, may cause blindness in both young and middle-aged people. Approximately 10–15% of blindness in the West is caused by uveitis. Therefore, a comprehensive investigation to determine the disease pathogenesis is urgent, as it will thus
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Uveitis, defined as inflammation of the uveal tract, may cause blindness in both young and middle-aged people. Approximately 10–15% of blindness in the West is caused by uveitis. Therefore, a comprehensive investigation to determine the disease pathogenesis is urgent, as it will thus be possible to design effective treatments. Identification of the disease genes that cause uveitis is an important requirement to achieve this goal. To begin to answer this question, in this study, a computational method was proposed to identify novel uveitis-related genes. This method was executed on a large protein–protein interaction network and employed a popular ranking algorithm, the Random Walk with Restart (RWR) algorithm. To improve the utility of the method, a permutation test and a procedure for selecting core genes were added, which helped to exclude false discoveries and select the most important candidate genes. The five-fold cross-validation was adopted to evaluate the method, yielding the average F1-measure of 0.189. In addition, we compared our method with a classic GBA-based method to further indicate its utility. Based on our method, 56 putative genes were chosen for further assessment. We have determined that several of these genes (e.g., CCL4, Jun, and MMP9) are likely to be important for the pathogenesis of uveitis. Full article
(This article belongs to the Special Issue Special Protein Molecules Computational Identification)
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Open AccessArticle CIK Cells and HDAC Inhibitors in Multiple Myeloma
Int. J. Mol. Sci. 2017, 18(5), 945; doi:10.3390/ijms18050945
Received: 8 February 2017 / Revised: 7 April 2017 / Accepted: 25 April 2017 / Published: 29 April 2017
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Abstract
Multiple myeloma is the second most common hematological malignancy. Despite all the progress made in treating multiple myeloma, it still remains an incurable disease. Patients are left with a median survival of 4–5 years. The combined treatment of multiple myeloma with histone deacetylase
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Multiple myeloma is the second most common hematological malignancy. Despite all the progress made in treating multiple myeloma, it still remains an incurable disease. Patients are left with a median survival of 4–5 years. The combined treatment of multiple myeloma with histone deacetylase inhibitors and cytokine-induced killer cells provides a promising targeted treatment option for patients. This study investigated the impact of a combined treatment compared to treatment with histone deacetylase inhibitors. The experiments revealed that a treatment with histone deacetylase (HDAC) inhibitors could reduce cell viability to 59% for KMS 18 cell line and 46% for the U-266 cell line. The combined treatment led to a decrease of cell viability to 33% for KMS 18 and 27% for the U-266 cell line, thus showing a significantly better efficacy than the single treatment. Full article
(This article belongs to the Special Issue Natural Killer T (NKT) Cells)
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Open AccessArticle Identifying the Genes Regulated by AtWRKY6 Using Comparative Transcript and Proteomic Analysis under Phosphorus Deficiency
Int. J. Mol. Sci. 2017, 18(5), 1046; doi:10.3390/ijms18051046
Received: 8 February 2017 / Revised: 26 April 2017 / Accepted: 4 May 2017 / Published: 12 May 2017
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Abstract
Phosphorus (P) is an important mineral nutrient for plant growth and development. Overexpressing AtWRKY6 (35S:WRKY6-9) was more sensitive and wrky6 (wrky6-1) was more resistant under low Pi conditions. To better understand the function of AtWRKY6 under low phosphate stress conditions, we
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Phosphorus (P) is an important mineral nutrient for plant growth and development. Overexpressing AtWRKY6 (35S:WRKY6-9) was more sensitive and wrky6 (wrky6-1) was more resistant under low Pi conditions. To better understand the function of AtWRKY6 under low phosphate stress conditions, we applied two-dimensional gel electrophoresis (2-DE) to analyse differentially expressed proteins in the shoots and roots between wild type, 35S:WRKY6-9 and wrky6-1 after phosphorus deficiency treatment for three days. The results showed 88 differentially abundant protein spots, which were identified between the shoots and roots of 35S:WRKY6-9 and wrky6-1 plants. In addition, 59 differentially expressed proteins were identified in the leaves and roots of 35S:WRKY6-9 plants. After analysis, 9 genes with W-box elements in their promoter sequences were identified in the leaves, while 6 genes with W-box elements in their promoter sequences were identified in the roots. A total of 8 genes were identified as potential target genes according to the quantitative PCR (QPCR) and two dimension difference gel electrophoresis, (2D-DIGE) results, including ATP synthase, gln synthetase, nitrilase, 14-3-3 protein, carbonic anhydrases 2, and tryptophan synthase. These results provide important information concerning the AtWRKY6 regulation network and reveal potential vital target genes of AtWRKY6 under low phosphorus stress. two dimension difference gel electrophoresis, 2D-DIGE Full article
(This article belongs to the Special Issue Selected Papers from the 6th National Plant Protein Research Congress)
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Open AccessArticle CTC-mRNA (AR-V7) Analysis from Blood Samples—Impact of Blood Collection Tube and Storage Time
Int. J. Mol. Sci. 2017, 18(5), 1047; doi:10.3390/ijms18051047
Received: 31 March 2017 / Revised: 2 May 2017 / Accepted: 8 May 2017 / Published: 12 May 2017
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Abstract
Circulating tumour cells (CTCs) are an emerging resource for monitoring cancer biomarkers. New technologies for CTC isolation and biomarker detection are increasingly sensitive, however, the ideal blood storage conditions to preserve CTC-specific mRNA biomarkers remains undetermined. Here we tested the preservation of tumour
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Circulating tumour cells (CTCs) are an emerging resource for monitoring cancer biomarkers. New technologies for CTC isolation and biomarker detection are increasingly sensitive, however, the ideal blood storage conditions to preserve CTC-specific mRNA biomarkers remains undetermined. Here we tested the preservation of tumour cells and CTC-mRNA over time in common anticoagulant ethylene-diamine-tetra-acetic acid (EDTA) and acid citrate dextrose solution B (Citrate) blood tubes compared to preservative-containing blood tubes. Blood samples spiked with prostate cancer cells were processed after 0, 24, 30, and 48 h storage at room temperature. The tumour cell isolation efficiency and the mRNA levels of the prostate cancer biomarkers androgen receptor variant 7 (AR-V7) and total AR, as well as epithelial cell adhesion molecule (EpCAM) were measured. Spiked cells were recovered across all storage tube types and times. Surprisingly, tumour mRNA biomarkers were readily detectable after 48 h storage in EDTA and Citrate tubes, but not in preservative-containing tubes. Notably, AR-V7 expression was detected in prostate cancer patient blood samples after 48 h storage in EDTA tubes at room temperature. This important finding presents opportunities for measuring AR-V7 expression from clinical trial patient samples processed within 48 h—a much more feasible timeframe compared to previous recommendations. Full article
(This article belongs to the Special Issue Diagnostic, Prognostic and Predictive Biomarkers in Prostate Cancer)
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Open AccessArticle Clinical Correlates and Prognostic Value of Plasma Galectin-3 Levels in Degenerative Aortic Stenosis: A Single-Center Prospective Study of Patients Referred for Invasive Treatment
Int. J. Mol. Sci. 2017, 18(5), 947; doi:10.3390/ijms18050947
Received: 31 January 2017 / Revised: 24 March 2017 / Accepted: 25 April 2017 / Published: 29 April 2017
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Abstract
Galectin-3 (Gal-3), a β-galactoside-binding lectin, has been implicated in myocardial fibrosis, development of left ventricular (LV) dysfunction and transition from compensated LV hypertrophy to overt heart failure (HF), being a novel prognostic marker in HF. Risk stratification is crucial for the choice of
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Galectin-3 (Gal-3), a β-galactoside-binding lectin, has been implicated in myocardial fibrosis, development of left ventricular (LV) dysfunction and transition from compensated LV hypertrophy to overt heart failure (HF), being a novel prognostic marker in HF. Risk stratification is crucial for the choice of the optimal therapy in degenerative aortic stenosis (AS), affecting elderly subjects with coexistent diseases. Our aim was to assess correlates and prognostic value of circulating Gal-3 in real-world patients with degenerative AS referred for invasive treatment. Gal-3 levels were measured at admission in 80 consecutive patients with symptomatic degenerative AS (mean age: 79 ± 8 years; aortic valve area (AVA) index: 0.4 ± 0.1 cm2/m2). The therapeutic strategy was chosen following a dedicated multidisciplinary team-oriented approach, including surgical valve replacement (n = 11), transcatheter valve implantation (n = 19), balloon aortic valvuloplasty (BAV) (n = 25) and optimal medical therapy (n = 25). Besides routine echocardiographic indices, valvulo-arterial impedance (Zva), an index of global LV afterload, was computed. There were 22 deaths over a median follow-up of 523 days. Baseline Gal-3 correlated negatively with estimated glomerular filtration rate (eGFR) (r = −0.61, p < 0.001) and was unrelated to age, symptomatic status, AVA index, LV ejection fraction, LV mass index or Zva. For the study group as a whole, Gal-3 tended to predict mortality (Gal-3 >17.8 vs. Gal-3 <17.8 ng/mL; hazard ratio (HR): 2.03 (95% confidence interval, 0.88–4.69), p = 0.09), which was abolished upon adjustment for eGFR (HR: 1.70 (0.61–4.73), p = 0.3). However, in post-BAV patients multivariate-adjusted pre-procedural Gal-3 was associated with worse survival (HR: 7.41 (1.52–36.1), p = 0.01) regardless of eGFR. In conclusion, the inverse eGFR–Gal-3 relationship underlies a weak association between Gal-3 and adverse outcome in patients with degenerative AS referred for invasive therapy irrespective of type of treatment employed. In contrast, pre-procedural Gal-3 appears an independent mortality predictor in high-risk AS patients undergoing BAV. Full article
(This article belongs to the Special Issue Improvement of Cardiac Function in Heart Failure 2017)
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Open AccessArticle Valproic Acid Induces Endocytosis-Mediated Doxorubicin Internalization and Shows Synergistic Cytotoxic Effects in Hepatocellular Carcinoma Cells
Int. J. Mol. Sci. 2017, 18(5), 1048; doi:10.3390/ijms18051048
Received: 31 March 2017 / Revised: 3 May 2017 / Accepted: 8 May 2017 / Published: 12 May 2017
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Abstract
Valproic acid (VPA), a well-known histone deacetylase (HDAC) inhibitor, is used as an anti-cancer drug for various cancers, but the synergistic anti-cancer effect of VPA and doxorubicin (DOX) combination treatment and its potential underlying mechanism in hepatocellular carcinoma (HCC) remain to be elucidated.
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Valproic acid (VPA), a well-known histone deacetylase (HDAC) inhibitor, is used as an anti-cancer drug for various cancers, but the synergistic anti-cancer effect of VPA and doxorubicin (DOX) combination treatment and its potential underlying mechanism in hepatocellular carcinoma (HCC) remain to be elucidated. Here, we evaluate the mono- and combination-therapy effects of VPA and DOX in HCC and identify a specific and efficient, synergistic anti-proliferative effect of the VPA and DOX combination in HCC cells, especially HepG2 cells; this effect was not apparent in MIHA cells, a normal hepatocyte cell line. The calculation of the coefficient of drug interaction confirmed the significant synergistic effect of the combination treatment. Concurrently, the synergistic apoptotic cell death caused by the VPA and DOX combination treatment was confirmed by Hoechst nuclear staining and Western blot analysis of caspase-3 and poly (ADP-ribose) polymerase (PARP) activation. Co-treatment with VPA and DOX enhanced reactive oxygen species (ROS) generation and autophagy, which were clearly attenuated by ROS and autophagy inhibitors, respectively. Furthermore, as an indication of the mechanism underlying the synergistic effect, we observed that DOX internalization, which was induced in the VPA and DOX combination-treated group, occurred via by the caveolae-mediated endocytosis pathway. Taken together, our study uncovered the potential effect of the VPA and DOX combination treatment with regard to cell death, including induction of cellular ROS, autophagy, and the caveolae-mediated endocytosis pathway. Therefore, these results present novel implications in drug delivery research for the treatment of HCC. Full article
(This article belongs to the Section Molecular Toxicology)
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Open AccessArticle Clinical Utility of the Adrenocorticotropin Stimulation Test with/without Dexamethasone Suppression for Definitive and Subtype Diagnosis of Primary Aldosteronism
Int. J. Mol. Sci. 2017, 18(5), 948; doi:10.3390/ijms18050948
Received: 23 January 2017 / Revised: 11 March 2017 / Accepted: 27 April 2017 / Published: 30 April 2017
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Abstract
The adrenocorticotropin (ACTH) stimulation test (AST) has been reported to be useful for diagnosing primary aldosteronism (PA), particularly for differentiating PA subtypes under 1-mg dexamethasone suppression (DS). The aim of our study was to clarify the effect of 1-mg DS on AST results.
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The adrenocorticotropin (ACTH) stimulation test (AST) has been reported to be useful for diagnosing primary aldosteronism (PA), particularly for differentiating PA subtypes under 1-mg dexamethasone suppression (DS). The aim of our study was to clarify the effect of 1-mg DS on AST results. A retrospective cohort study was conducted using data for 48 patients (PA: 30/48). We estimated the difference in plasma aldosterone concentration (PAC) responsiveness to ACTH stimulation with single (AST alone) and combined (AST under 1-mg DS) tests within the same patient. We compared the diagnostic accuracy of these two tests for PA and the laterality of hyperaldosteronism. We found no differences in PAC responsiveness to ACTH stimulation between single and combined tests, and observed a significant positive linear relationship (30 min, R2 = 0.75, p-value < 0.01). Both tests showed the highest diagnostic accuracy for PA following 30 min of ACTH stimulation. The ability to detect the laterality of hyperaldosteronism was inconsistent and differed according to the two definitions: lateralization ratio and the absolute aldosterone levels in adrenal venous sampling. PAC responsiveness to ACTH stimulation was similar for AST with and without 1-mg DS. AST can be performed under both conditions with similar accuracy to detect PA. Full article
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Open AccessArticle A Histopathological Scheme for the Quantitative Scoring of Intervertebral Disc Degeneration and the Therapeutic Utility of Adult Mesenchymal Stem Cells for Intervertebral Disc Regeneration
Int. J. Mol. Sci. 2017, 18(5), 1049; doi:10.3390/ijms18051049
Received: 8 March 2017 / Revised: 28 April 2017 / Accepted: 8 May 2017 / Published: 12 May 2017
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Abstract
The purpose of this study was to develop a quantitative histopathological scoring scheme to evaluate disc degeneration and regeneration using an ovine annular lesion model of experimental disc degeneration. Toluidine blue and Haematoxylin and Eosin (H&E) staining were used to evaluate cellular morphology:
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The purpose of this study was to develop a quantitative histopathological scoring scheme to evaluate disc degeneration and regeneration using an ovine annular lesion model of experimental disc degeneration. Toluidine blue and Haematoxylin and Eosin (H&E) staining were used to evaluate cellular morphology: (i) disc structure/lesion morphology; (ii) proteoglycan depletion; (iii) cellular morphology; (iv) blood vessel in-growth; (v) cell influx into lesion; and (vi) cystic degeneration/chondroid metaplasia. Three study groups were examined: 5 × 5 mm lesion; 6 × 20 mm lesion; and 6 × 20 mm lesion plus mesenchymal stem cell (MSC) treatment. Lumbar intervertebral discs (IVDs) were scored under categories (i–vi) to provide a cumulative score, which underwent statistical analysis using STATA software. Focal proteoglycan depletion was associated with 5 × 5 mm annular rim lesions, bifurcations, annular delamellation, concentric and radial annular tears and an early influx of blood vessels and cells around remodeling lesions but the inner lesion did not heal. Similar features in 6 × 20 mm lesions occurred over a 3–6-month post operative period. MSCs induced a strong recovery in discal pathology with a reduction in cumulative histopathology degeneracy score from 15.2 to 2.7 (p = 0.001) over a three-month recovery period but no recovery in carrier injected discs. Full article
(This article belongs to the Special Issue Advances in Bone and Cartilage Research)
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Open AccessArticle Specific MicroRNA Pattern in Colon Tissue of Young Children with Eosinophilic Colitis
Int. J. Mol. Sci. 2017, 18(5), 1050; doi:10.3390/ijms18051050
Received: 14 March 2017 / Revised: 15 April 2017 / Accepted: 5 May 2017 / Published: 12 May 2017
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Abstract
Eosinophilic colitis (EC) is a common cause of haematochezia in infants and young children. The exact pathomechanism is not understood, and the diagnosis is challenging. The role of microRNAs as key class of regulators of mRNA expression and translation in patients with EC
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Eosinophilic colitis (EC) is a common cause of haematochezia in infants and young children. The exact pathomechanism is not understood, and the diagnosis is challenging. The role of microRNAs as key class of regulators of mRNA expression and translation in patients with EC has not been explored. Therefore, the aim of the present study was to explore the miRNA profile in EC with respect to eosinophilic inflammation. Patients enrolled in the study (n = 10) had persistent rectal bleeding, and did not respond to elimination dietary treatment. High-throughput microRNA sequencing was carried out on colonic biopsy specimens of children with EC (EC: n = 4) and controls (C: n = 4) as a preliminary screening of the miRNA profile. Based on the next-generation sequencing (NGS) results and literature data, a potentially relevant panel of miRNAs were selected for further measurements by real-time reverse transcription (RT)-PCR (EC: n = 14, C: n = 10). Validation by RT-PCR resulted in significantly altered expression of miR-21, -31, -99b, -125a, -146a, -184, -221, -223, and -559 compared to controls (p ≤ 0.05). Elevation in miR-21, -99b, -146a, -221, and -223 showed statistically significant correlation to the extent of tissue eosinophilia. Based on our results, we conclude that the dysregulated miRNAs have a potential role in the regulation of apoptosis by targeting Protein kinase B/Mechanistic target of rapamycin (AKT/mTOR)-related pathways in inflammation by modulating Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)-related signalling and eosinophil cell recruitment and activation, mainly by regulating the expression of the chemoattractant eotaxin and the adhesion molecule CD44. Our results could serve as a basis for further extended research exploring the pathomechanism of EC. Full article
(This article belongs to the collection Regulation by Non-Coding RNAs)
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Open AccessArticle Expression of Root Genes in Arabidopsis Seedlings Grown by Standard and Improved Growing Methods
Int. J. Mol. Sci. 2017, 18(5), 951; doi:10.3390/ijms18050951
Received: 31 March 2017 / Revised: 25 April 2017 / Accepted: 27 April 2017 / Published: 3 May 2017
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Abstract
Roots of Arabidopsis thaliana seedlings grown in the laboratory using the traditional plant-growing culture system (TPG) were covered to maintain them in darkness. This new method is based on a dark chamber and is named the improved plant-growing method (IPG). We measured the
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Roots of Arabidopsis thaliana seedlings grown in the laboratory using the traditional plant-growing culture system (TPG) were covered to maintain them in darkness. This new method is based on a dark chamber and is named the improved plant-growing method (IPG). We measured the light conditions in dark chambers, and found that the highest light intensity was dramatically reduced deeper in the dark chamber. In the bottom and side parts of dark chambers, roots were almost completely shaded. Using the high-throughput RNA sequencing method on the whole RNA extraction from roots, we compared the global gene expression levels in roots of seedlings from these two conditions and identified 141 differently expressed genes (DEGs) between them. According to the KEGG (Kyoto Encyclopedia of Genes and Genomes) enrichment, the flavone and flavonol biosynthesis and flavonoid biosynthesis pathways were most affected among all annotated pathways. Surprisingly, no genes of known plant photoreceptors were identified as DEGs by this method. Considering that the light intensity was decreased in the IPG system, we collected four sections (1.5 cm for each) of Arabidopsis roots grown in TPG and IPG conditions, and the spatial-related differential gene expression levels of plant photoreceptors and polar auxin transporters, including CRY1, CRY2, PHYA, PHYB, PHOT1, PHOT2, and UVR8 were analyzed by qRT-PCR. Using these results, we generated a map of the spatial-related expression patterns of these genes under IPG and TPG conditions. The expression levels of light-related genes in roots is highly sensitive to illumination and it provides a background reference for selecting an improved culture method for laboratory-maintained Arabidopsis seedlings. Full article
(This article belongs to the Special Issue Selected Papers from the 6th National Plant Protein Research Congress)
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Open AccessArticle Isolation of Rice Bran Lectins and Characterization of Their Unique Behavior in Caco-2 Cells
Int. J. Mol. Sci. 2017, 18(5), 1052; doi:10.3390/ijms18051052
Received: 29 March 2017 / Revised: 5 May 2017 / Accepted: 10 May 2017 / Published: 13 May 2017
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Abstract
Rice bran lectins, named as RBA1 and RBA2, were isolated from Oryza sativa in two chromatography steps: affinity chromatography and cation-exchange chromatography. RBA1 was found to be composed of a covalently linked heterodimer of 20- and 12-kDa subunits, and RBA2 was a noncovalently
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Rice bran lectins, named as RBA1 and RBA2, were isolated from Oryza sativa in two chromatography steps: affinity chromatography and cation-exchange chromatography. RBA1 was found to be composed of a covalently linked heterodimer of 20- and 12-kDa subunits, and RBA2 was a noncovalently linked dimer of 12-kDa subunits. Both RBA1 and RBA2 bound to desialylated complex glycoproteins such as fetuin, α1-acid glycoprotein, and transferrin, and agalactosylated complex glycoproteins such as agalacto fetuin, agalacto-α1-acid glycoprotein, and agalacto-transferrin, in addition to chitooligosacchrides. RBAs were heat stable up to 80 °C and stable at pH 4–10. RBA1 increased the transport of the fluorescent marker, rhodamine 123, which is known to be transported via the P-glycoprotein-mediated efflux pathway across human intestinal Caco-2 cell monolayers. Furthermore, RBA1 itself was transported to the basolateral side of the monolayers via an endocytotic pathway. Full article
(This article belongs to the Special Issue Plant Lectins: From Model Species to Crop Plants)
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Open AccessArticle Inhibitory Effects of Dimethyllirioresinol, Epimagnolin A, Eudesmin, Fargesin, and Magnolin on Cytochrome P450 Enzyme Activities in Human Liver Microsomes
Int. J. Mol. Sci. 2017, 18(5), 952; doi:10.3390/ijms18050952
Received: 21 March 2017 / Revised: 25 April 2017 / Accepted: 27 April 2017 / Published: 1 May 2017
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Abstract
Magnolin, epimagnolin A, dimethyllirioresinol, eudesmin, and fargesin are pharmacologically active tetrahydrofurofuranoid lignans found in Flos Magnoliae. The inhibitory potentials of dimethyllirioresinol, epimagnolin A, eudesmin, fargesin, and magnolin on eight major human cytochrome P450 (CYP) enzyme activities in human liver microsomes were evaluated using
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Magnolin, epimagnolin A, dimethyllirioresinol, eudesmin, and fargesin are pharmacologically active tetrahydrofurofuranoid lignans found in Flos Magnoliae. The inhibitory potentials of dimethyllirioresinol, epimagnolin A, eudesmin, fargesin, and magnolin on eight major human cytochrome P450 (CYP) enzyme activities in human liver microsomes were evaluated using liquid chromatography–tandem mass spectrometry to determine the inhibition mechanisms and inhibition potency. Fargesin inhibited CYP2C9-catalyzed diclofenac 4’-hydroxylation with a Ki value of 16.3 μM, and it exhibited mechanism-based inhibition of CYP2C19-catalyzed [S]-mephenytoin 4’-hydroxylation (Ki, 3.7 μM; kinact, 0.102 min−1), CYP2C8-catalyzed amodiaquine N-deethylation (Ki, 10.7 μM; kinact, 0.082 min−1), and CYP3A4-catalyzed midazolam 1’-hydroxylation (Ki, 23.0 μM; kinact, 0.050 min−1) in human liver microsomes. Fargesin negligibly inhibited CYP1A2-catalyzed phenacetin O-deethylation, CYP2A6-catalyzed coumarin 7-hydroxylation, CYP2B6-catalyzed bupropion hydroxylation, and CYP2D6-catalyzed bufuralol 1’-hydroxylation at 100 μM in human liver microsomes. Dimethyllirioresinol weakly inhibited CYP2C19 and CYP2C8 with IC50 values of 55.1 and 85.0 μM, respectively, without inhibition of CYP1A2, CYP2A6, CYP2B6, CYP2C9, CYP2D6, and CYP3A4 activities at 100 μM. Epimagnolin A, eudesmin, and magnolin showed no the reversible and time-dependent inhibition of eight major CYP activities at 100 μM in human liver microsomes. These in vitro results suggest that it is necessary to investigate the potentials of in vivo fargesin-drug interaction with CYP2C8, CYP2C9, CYP2C19, and CYP3A4 substrates. Full article
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Open AccessArticle Melatonin Secretion Is Increased in Children with Severe Traumatic Brain Injury
Int. J. Mol. Sci. 2017, 18(5), 1053; doi:10.3390/ijms18051053
Received: 7 April 2017 / Revised: 9 May 2017 / Accepted: 11 May 2017 / Published: 13 May 2017
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Abstract
Background: Traumatic brain injury (TBI) is a leading cause of death and disability in children. Oxidative stress plays a significant role in brain damage and melatonin exhibits both direct and indirect antioxidant effects. The primary aim of the present study was to evaluate
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Background: Traumatic brain injury (TBI) is a leading cause of death and disability in children. Oxidative stress plays a significant role in brain damage and melatonin exhibits both direct and indirect antioxidant effects. The primary aim of the present study was to evaluate serum melatonin levels in children with severe TBI in comparison to critically ill children admitted to the Pediatric Intensive Care Unit for conditions other than TBI. Methods: Twenty-four children were evaluated, equally divided into severe TBI and no-TBI. Blood samples for serum melatonin analysis were collected at 22:00, 01:00, 03:00, 05:00, 08:00, and 12:00. Results: Mean serum melatonin peaks in children of the TBI group were higher compared to the values of no-TBI critically ill children (495 ± 102 vs. 294 ± 119 pg/mL, p = 0.0002). Furthermore, the difference was even more significant in comparison to values reported in literature for healthy age-matched children (495 ± 102 vs. 197 ± 71 pg/mL, p < 0.0001). Conclusion: This study has shown that endogenous serum melatonin levels dramatically increase in children after severe TBI. This elevation is likely to represent a response to oxidative stress and/or inflammation due to severe head injury. Full article
(This article belongs to the Special Issue Melatonin and Its Analogues: Experimental and Clinical Aspects)
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Open AccessArticle Chemical Profile and Antioxidant, Anti-Inflammatory, Antimutagenic and Antimicrobial Activities of Geopropolis from the Stingless Bee Melipona orbignyi
Int. J. Mol. Sci. 2017, 18(5), 953; doi:10.3390/ijms18050953
Received: 4 April 2017 / Accepted: 27 April 2017 / Published: 3 May 2017
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Abstract
Geopropolis is a resin mixed with mud, produced only by stingless bees. Despite being popularly known for its medicinal properties, few scientific studies have proven its biological activities. In this context, the objective of this study was to determine the chemical composition and
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Geopropolis is a resin mixed with mud, produced only by stingless bees. Despite being popularly known for its medicinal properties, few scientific studies have proven its biological activities. In this context, the objective of this study was to determine the chemical composition and antioxidant, anti-inflammatory, antimutagenic and antimicrobial activities of the Melipona orbignyi geopropolis. The hydroalcoholic extract of geopropolis (HEGP) was prepared and its chemical composition determined by high performance liquid chromatography coupled to diode array detector and mass spectrometry (HPLC-DAD-MS). The antioxidant activity was determined by the capture of free radicals and inhibition of lipid peroxidation in human erythrocytes. The anti-inflammatory activity was evaluated by the inhibition of the hyaluronidase enzyme and the antimutagenic action was investigated in Saccharomyces cerevisiae colonies. The antimicrobial activities were determined against bacteria and yeasts, isolated from reference strains and hospital origin. The chemical composition of HEGP included flavonoids, derivatives of glycosylated phenolic acids and terpenoids. HEGP showed high antioxidant activity, it inhibited the activity of the inflammatory enzyme hyaluronidase and reduced the mutagenic effects in S. cerevisiae. In relation to the antimicrobial activity, it promoted the death of all microorganisms evaluated. In conclusion, this study reveals for the first time the chemical composition of the HEGP of M. orbignyi and demonstrates its pharmacological properties. Full article
(This article belongs to the Special Issue Nutraceuticals in Human Health and Disease)
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Open AccessCommunication First Insights into Human Fingertip Regeneration by Echo-Doppler Imaging and Wound Microenvironment Assessment
Int. J. Mol. Sci. 2017, 18(5), 1054; doi:10.3390/ijms18051054
Received: 9 April 2017 / Revised: 3 May 2017 / Accepted: 6 May 2017 / Published: 13 May 2017
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Abstract
Fingertip response to trauma represents a fascinating example of tissue regeneration. Regeneration derives from proliferative mesenchymal cells (blastema) that subsequently differentiate into soft and skeletal tissues. Clinically, conservative treatment of the amputated fingertip under occlusive dressing can shift the response to tissue loss
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Fingertip response to trauma represents a fascinating example of tissue regeneration. Regeneration derives from proliferative mesenchymal cells (blastema) that subsequently differentiate into soft and skeletal tissues. Clinically, conservative treatment of the amputated fingertip under occlusive dressing can shift the response to tissue loss from a wound repair process towards regeneration. When analyzing by Immunoassay the wound exudate from occlusive dressings, the concentrations of brain-derived neurotrophic factor (BDNF) and leukemia inhibitory factor (LIF) were higher in fingertip exudates than in burn wounds (used as controls for wound repair versus regeneration). Vascular endothelial growth factor A (VEGF-A) and platelet-derived growth factor (PDGF) were highly expressed in both samples in comparable levels. In our study, pro-inflammatory cytokines were relatively higher expressed in regenerative fingertips than in the burn wound exudates while chemokines were present in lower levels. Functional, vascular and mechanical properties of the regenerated fingertips were analyzed three months after trauma and the data were compared to the corresponding fingertip on the collateral uninjured side. While sensory recovery and morphology (pulp thickness and texture) were similar to uninjured sides, mechanical parameters (elasticity, vascularization) were increased in the regenerated fingertips. Further studies should be done to clarify the importance of inflammatory cells, immunity and growth factors in determining the outcome of the regenerative process and its influence on the clinical outcome. Full article
(This article belongs to the Special Issue Wound Repair and Regeneration)
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Open AccessArticle Tubacin, an HDAC6 Selective Inhibitor, Reduces the Replication of the Japanese Encephalitis Virus via the Decrease of Viral RNA Synthesis
Int. J. Mol. Sci. 2017, 18(5), 954; doi:10.3390/ijms18050954
Received: 31 March 2017 / Revised: 26 April 2017 / Accepted: 28 April 2017 / Published: 1 May 2017
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Abstract
Japanese encephalitis virus (JEV), a neurotropic flavivirus, annually causes over 30,000 Japanese Encephalitis (JE) cases in East and Southeast Asia. Histone deacetylases (HDACs) modulate lysine acetylation of histones and non-histone proteins, regulating many processes including inflammation and antiviral immune response. This study investigated
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Japanese encephalitis virus (JEV), a neurotropic flavivirus, annually causes over 30,000 Japanese Encephalitis (JE) cases in East and Southeast Asia. Histone deacetylases (HDACs) modulate lysine acetylation of histones and non-histone proteins, regulating many processes including inflammation and antiviral immune response. This study investigated antiviral activity of pan- and selective-HDAC inhibitors as host-targeting agents against JEV. Among HDAC inhibitors, selective HDAC6 inhibitors (tubastatin-A (TBSA) and tubacin) concentration-dependently inhibited JEV-induced cytopathic effect and apoptosis, as well as reduced virus yield in human cerebellar medulloblastoma cells. The 50% inhibitory concentration (IC50) values of virus yield was 0.26 μM for tubacin and 1.75 μM for TBSA, respectively. Tubacin (IC50 of 1.52 μM), but not TBSA, meaningfully blocked the production of intracellular infectious virus particles. In time-of-addition assays, the greatest potency of antiviral activity was observed in the mode of pre-treatment with tubacin (IC50 of 1.89 μM) compared to simultaneous (IC50 of 4.88 μM) and post-treatment (IC50 of 2.05 μM) modes. Interestingly, tubacin induced the hyperacetylation of a HDAC6 substrate Hsp90 and reduced the interaction of Hsp90 with JEV NS5 protein. Novobiocin, an Hsp90 inhibitor, diminished the NS5 protein amount and virus replication in JEV-infected cells. Meantime, tubacin suppressed the NS5 expression and antisense RNA genome synthesis in infected cells. Tubacin-induced Hsp90 hyperacetylation was suggested to influence the NS5 activity in JEV replication. Therefore, tubacin had a high potential of a host-targeting agent against JEV, exhibiting preventive and therapeutic activities against JEV infection. Full article
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Open AccessArticle Supervised Clustering of Adipokines and Hormonal Receptors Predict Prognosis in a Population of Obese Women with Type 1 Endometrial Cancer
Int. J. Mol. Sci. 2017, 18(5), 1055; doi:10.3390/ijms18051055
Received: 24 March 2017 / Revised: 6 May 2017 / Accepted: 10 May 2017 / Published: 13 May 2017
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Abstract
Obesity is a major risk factor for endometrial cancer (EC). Yet, its impact on prognosis is controversial. Obesity is associated with metabolic and hormonal dysregulation as well as adipokines increase. The aim of this study was to characterize the expression of biological factors
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Obesity is a major risk factor for endometrial cancer (EC). Yet, its impact on prognosis is controversial. Obesity is associated with metabolic and hormonal dysregulation as well as adipokines increase. The aim of this study was to characterize the expression of biological factors related to obesity within the tumor and evaluate their impact on prognosis. One hundred and thirty-six patients, including 55 obese patients, with endometrioid type I EC operated by total hysterectomy were included in this retrospective study conducted in a Tertiary teaching hospital between 2000 and 2013. Immunohistochemistry (IHC) study was performed on type I EC tumor samples using five adipokines (SPARC, RBP4 (Retinol Binding Protein 4), adiponectin, TNF α, IL-6) and hormonal receptors (estrogen receptor and progesterone receptor). Supervised clustering of immunohistochemical markers was performed to identify clusters that could be associated with prognostic groups. The prognosis of the obese population was not different from the prognosis of the general population. Adipokine expression within tumors was not different in these two populations. In obese population, we found three clusters where co-expression was associated with a recurrence group in comparison with a non-recurrence group and four clusters where co-expression was associated with the high risk FIGO (International Federation of Gynecology and Obstetrics) stage I group in comparison of low risk FIGO stage I group. While obesity does not appear as a prognostic factor in endometrioid type I EC, the co-expression of biological factors in IHC on hysterectomy specimens allowed to distinguish two prognostic groups in obese population. Full article
(This article belongs to the Special Issue Gynecologic Oncology: From Molecular Mechanisms to Targeted Therapies)
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Open AccessArticle Induced Pluripotent Stem Cells Derived from a CLN5 Patient Manifest Phenotypic Characteristics of Neuronal Ceroid Lipofuscinoses
Int. J. Mol. Sci. 2017, 18(5), 955; doi:10.3390/ijms18050955
Received: 17 February 2017 / Revised: 12 April 2017 / Accepted: 26 April 2017 / Published: 1 May 2017
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Abstract
Neuronal ceroid lipofuscinoses (NCLs) are autosomal recessive progressive encephalopathies caused by mutations in at least 14 different genes. Despite extensive studies performed in different NCL animal models, the molecular mechanisms underlying neurodegeneration in NCLs remain poorly understood. To model NCL in human cells,
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Neuronal ceroid lipofuscinoses (NCLs) are autosomal recessive progressive encephalopathies caused by mutations in at least 14 different genes. Despite extensive studies performed in different NCL animal models, the molecular mechanisms underlying neurodegeneration in NCLs remain poorly understood. To model NCL in human cells, we generated induced pluripotent stem cells (iPSCs) by reprogramming skin fibroblasts from a patient with CLN5 (ceroid lipofuscinosis, neuronal, 5) disease, the late infantile variant form of NCL. These CLN5 patient-derived iPSCs (CLN5Y392X iPSCs) harbouring the most common CLN5 mutation, c.1175_1176delAT (p.Tyr392X), were further differentiated into neural lineage cells, the most affected cell type in NCLs. The CLN5Y392X iPSC-derived neural lineage cells showed accumulation of autofluorescent storage material and subunit C of the mitochondrial ATP synthase, both representing the hallmarks of many forms of NCLs, including CLN5 disease. In addition, we detected abnormalities in the intracellular organelles and aberrations in neuronal sphingolipid transportation, verifying the previous findings obtained from Cln5-deficient mouse macrophages. Therefore, patient-derived iPSCs provide a suitable model to study the mechanisms of NCL diseases. Full article
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Open AccessArticle Renalase Protects against Renal Fibrosis by Inhibiting the Activation of the ERK Signaling Pathways
Int. J. Mol. Sci. 2017, 18(5), 855; doi:10.3390/ijms18050855
Received: 5 February 2017 / Revised: 30 March 2017 / Accepted: 4 April 2017 / Published: 27 April 2017
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Abstract
Renal interstitial fibrosis is a common pathway for the progression of chronic kidney disease (CKD) to end-stage renal disease. Renalase, acting as a signaling molecule, has been reported to have cardiovascular and renal protective effects. However, its role in renal fibrosis remains unknown.
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Renal interstitial fibrosis is a common pathway for the progression of chronic kidney disease (CKD) to end-stage renal disease. Renalase, acting as a signaling molecule, has been reported to have cardiovascular and renal protective effects. However, its role in renal fibrosis remains unknown. In this study, we evaluated the therapeutic efficacy of renalase in rats with complete unilateral ureteral obstruction (UUO) and examined the inhibitory effects of renalase on transforming growth factor-β1 (TGF-β1)-induced epithelial–mesenchymal transition (EMT) in human proximal renal tubular epithelial (HK-2) cells. We found that in the UUO model, the expression of renalase was markedly downregulated and adenoviral-mediated expression of renalase significantly attenuated renal interstitial fibrosis, as evidenced by the maintenance of E-cadherin expression and suppressed expression of α-smooth muscle actin (α-SMA), fibronectin and collagen-I. In vitro, renalase inhibited TGF-β1-mediated upregulation of α-SMA and downregulation of E-cadherin. Increased levels of Phospho-extracellular regulated protein kinases (p-ERK1/2) in TGF-β1-stimulated cells were reversed by renalase cotreatment. When ERK1 was overexpressed, the inhibition of TGF-β1-induced EMT and fibrosis mediated by renalase was attenuated. Our study provides the first evidence that renalase can ameliorate renal interstitial fibrosis by suppression of tubular EMT through inhibition of the ERK pathway. These results suggest that renalase has potential renoprotective effects in renal interstitial fibrosis and may be an effective agent for slowing CKD progression. Full article
(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)
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Open AccessArticle Natural Killer Cells Response to IL-2 Stimulation Is Distinct between Ascites with the Presence or Absence of Malignant Cells in Ovarian Cancer Patients
Int. J. Mol. Sci. 2017, 18(5), 856; doi:10.3390/ijms18050856
Received: 23 February 2017 / Revised: 10 April 2017 / Accepted: 13 April 2017 / Published: 17 May 2017
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Abstract
Peritoneal ascites are a distinguishable feature of patients with advanced epithelial ovarian cancer (EOC). The presence of different lymphocyte subsets has been reported in EOC-associated ascites, which also can or not contain malignant cells. The goal of this study was to analyze the
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Peritoneal ascites are a distinguishable feature of patients with advanced epithelial ovarian cancer (EOC). The presence of different lymphocyte subsets has been reported in EOC-associated ascites, which also can or not contain malignant cells. The goal of this study was to analyze the functional characteristics of natural killer (NK) cells from EOC-associated ascites in terms of their expression of activating receptors and ascites’ contents of lymphocyte subtypes, cytokine profile and presence of EOC cells. NK cell function was evaluated by the expression of the degranulation marker CD107a in resting and interleukin (IL)-2 stimulated NK cells from ascites and blood. Degranulation of NK cells from EOC cell-free ascites was significantly (p < 0.05) higher than all the other groups, either in their resting state or after IL-2 stimulation, suggesting a previous local stimulation. In contrast, treatment with IL-2 had no effect on NK cells from ascites with EOC cells. The amount of regulatory T cells was significantly higher in ascites with EOC cells compared to EOC cell-free ascites. Ascites with EOC cells also had higher levels of tumor necrosis factor (TNF)-α, suggesting inflammation related to the malignancy. In conclusion, the functional performance of NK cells was distinct between EOC cell-free ascites and ascites with EOC cells. The impairment of NK cell response to IL-2 in ascites with EOC cells was consistent with an immunosuppressive tumor microenvironment. Full article
(This article belongs to the Special Issue Natural Killer (NK) Cells)
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Open AccessArticle Molecular Characterization and Expression Profiling of Tomato GRF Transcription Factor Family Genes in Response to Abiotic Stresses and Phytohormones
Int. J. Mol. Sci. 2017, 18(5), 1056; doi:10.3390/ijms18051056
Received: 24 March 2017 / Revised: 7 May 2017 / Accepted: 9 May 2017 / Published: 13 May 2017
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Abstract
Growth regulating factors (GRFs) are plant-specific transcription factors that are involved in diverse biological and physiological processes, such as growth, development and stress and hormone responses. However, the roles of GRFs in vegetative and reproductive growth, development and stress responses in tomato (
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Growth regulating factors (GRFs) are plant-specific transcription factors that are involved in diverse biological and physiological processes, such as growth, development and stress and hormone responses. However, the roles of GRFs in vegetative and reproductive growth, development and stress responses in tomato (Solanum lycopersicum) have not been extensively explored. In this study, we characterized the 13 SlGRF genes. In silico analysis of protein motif organization, intron–exon distribution, and phylogenetic classification confirmed the presence of GRF proteins in tomato. The tissue-specific expression analysis revealed that most of the SlGRF genes were preferentially expressed in young and growing tissues such as flower buds and meristems, suggesting that SlGRFs are important during growth and development of these tissues. Some of the SlGRF genes were preferentially expressed in fruits at distinct developmental stages suggesting their involvement in fruit development and the ripening process. The strong and differential expression of different SlGRFs under NaCl, drought, heat, cold, abscisic acid (ABA), and jasmonic acid (JA) treatment, predict possible functions for these genes in stress responses in addition to their growth regulatory functions. Further, differential expression of SlGRF genes upon gibberellic acid (GA3) treatment indicates their probable function in flower development and stress responses through a gibberellic acid (GA)-mediated pathway. The results of this study provide a basis for further functional analysis and characterization of this important gene family in tomato. Full article
(This article belongs to the Special Issue Abiotic Stress and Gene Networks in Plants 2017)
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Open AccessArticle Protective Effect of Fragaria ananassa Crude Extract on Cadmium-Induced Lipid Peroxidation, Antioxidant Enzymes Suppression, and Apoptosis in Rat Testes
Int. J. Mol. Sci. 2017, 18(5), 957; doi:10.3390/ijms18050957
Received: 15 March 2017 / Revised: 12 April 2017 / Accepted: 20 April 2017 / Published: 5 May 2017
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Abstract
Cadmium is a deleterious environmental pollutant that threats both animals and human health. Oxidative stress and elevated levels of reactive oxygen species (ROS) have recently been reported to be the main cause of cellular damage as a result of cadmium exposure. We investigate,
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Cadmium is a deleterious environmental pollutant that threats both animals and human health. Oxidative stress and elevated levels of reactive oxygen species (ROS) have recently been reported to be the main cause of cellular damage as a result of cadmium exposure. We investigate, here, the protective effect of strawberry crude extracts on cadmium-induced oxidative damage of testes in rats. Four groups (n = 8) of 32 adult male Wistar rats weighing 160–180 g were used. The control group received 0.9% saline solution all over the experimental period (5 days). Group 2 was intraperitoneally injected with 6.5 mg/kg CdCl2. Group 3 was provided only with an oral administration of strawberry methanolic extract (SME) at a dose of 250 mg/kg. Group 4 was treated with SME before cadmium injection with the same mentioned doses. It was shown that cadmium exposure results in a significant decrease in both relative testicular weight and serum testosterone level. Analyzing the oxidative damaging effect of cadmium on the testicular tissue revealed the induction of oxidative stress markers represented in the elevated level of lipid peroxidation (LPO), nitric oxide (NO), and a decrease in the reduced glutathione (GSH) content. Considering cadmium toxicity, the level of the antioxidant enzyme activities including catalase (CAT), superoxide dismutase (SOD2), glutathione peroxidase (GPx1), and glutathione reductase (GR) were markedly decreased. Moreover, gene expression analysis indicated significant upregulation of the pro-apoptotic proteins, bcl-2-associated-X-protein (BAX), and tumor necrosis factor-α (TNFA) in response to cadmium intoxication, while significant downregulation of the anti-apoptotic, B-cell lymphoma 2 (BCL2) gene was detected. Immunohistochemistry of the testicular tissue possessed positive immunostaining for the increased level of TNF-α, but decreased number of proliferating cell nuclear antigen (PCNA) stained cells. Administration of SME debilitated the deleterious effect of cadmium via reduction of both LPO and NO levels followed by a significant enhancement in the gene expression level of CAT, SOD2, GPX1, GR, nuclear factor-erythroid 2-related factor 2 (NFE2L2), heme oxygenase-1 (HMOX1), Bcl-2, and PCNA. In addition, the SME treated group revealed a significant increase in the level of testosterone and GSH accompanied by a marked decrease in the gene expression level of Bax and TNF-α. In terms of the summarized results, the SME of Fragaria ananassa has a protective effect against cadmium-induced oxidative damage of testes. Full article
(This article belongs to the Special Issue Correlation between Nutrition, Oxidative Stress and Disease)
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Open AccessArticle Osteopontin Deficiency Suppresses Intestinal Tumor Development in Apc-Deficient Min Mice
Int. J. Mol. Sci. 2017, 18(5), 1058; doi:10.3390/ijms18051058
Received: 29 March 2017 / Revised: 5 May 2017 / Accepted: 9 May 2017 / Published: 14 May 2017
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Abstract
Osteopontin (OPN) is a secreted phosphoglycoprotein, and is a transcriptional target of aberrant Wnt signaling. OPN is upregulated in human colon cancers, and is suggested to enhance cancer progression. In this study, the effect of deficiency of OPN on intestinal tumor development in
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Osteopontin (OPN) is a secreted phosphoglycoprotein, and is a transcriptional target of aberrant Wnt signaling. OPN is upregulated in human colon cancers, and is suggested to enhance cancer progression. In this study, the effect of deficiency of OPN on intestinal tumor development in Apc-deficient Min mice was investigated. At 16 weeks of age, the number of small intestinal polyps in Min/OPN(+/−) and Min/OPN(−/−) mice was lower than that of Min/OPN(+/+) mice. Colorectal tumor incidences and multiplicities in Min/OPN(+/−) and Min/OPN(−/−) mice were significantly lower than those in Min/OPN(+/+) mice, being 48% and 0.6 ± 0.8, 50% and 0.8 ± 0.9 vs. 80% and 1.6 ± 1.7, respectively. OPN expression in colorectal tumors was strongly upregulated in Min/OPN(+/+) compared to adjacent non-tumor parts, but was decreased in Min/OPN(+/−) and not detected in Min/OPN(−/−). Targets of OPN, matrix metalloproteinases (MMPs)-3, -9, and -13 were lowered by OPN deficiency. Macrophage marker F4/80 in colorectal tumors was also lowered by OPN deficiency. MMP-9 expression was observed in tumor cells and tumor-infiltrating neutrophils. These results indicate that induction of OPN by aberrant Wnt signaling could enhance colorectal tumor development in part by upregulation of MMP-3, -9, and -13 and infiltration of macrophage and neutrophils. Suppression of OPN expression could contribute to tumor prevention, but complete deficiency of OPN may cause some adverse effects. Full article
(This article belongs to the Special Issue Inflammation and Cancer)
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Open AccessArticle Comparative Proteomics of Rubber Latex Revealed Multiple Protein Species of REF/SRPP Family Respond Diversely to Ethylene Stimulation among Different Rubber Tree Clones
Int. J. Mol. Sci. 2017, 18(5), 958; doi:10.3390/ijms18050958
Received: 23 January 2017 / Revised: 3 April 2017 / Accepted: 21 April 2017 / Published: 2 May 2017
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Abstract
Rubber elongation factor (REF) and small rubber particle protein (SRPP) are two key factors for natural rubber biosynthesis. To further understand the roles of these proteins in rubber formation, six different genes for latex abundant REF or SRPP proteins, including REF138,175,258 and
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Rubber elongation factor (REF) and small rubber particle protein (SRPP) are two key factors for natural rubber biosynthesis. To further understand the roles of these proteins in rubber formation, six different genes for latex abundant REF or SRPP proteins, including REF138,175,258 and SRPP117,204,243, were characterized from Hevea brasiliensis Reyan (RY) 7-33-97. Sequence analysis showed that REFs have a variable and long N-terminal, whereas SRPPs have a variable and long C-terminal beyond the REF domain, and REF258 has a β subunit of ATPase in its N-terminal. Through two-dimensional electrophoresis (2-DE), each REF/SRPP protein was separated into multiple protein spots on 2-DE gels, indicating they have multiple protein species. The abundance of REF/SRPP proteins was compared between ethylene and control treatments or among rubber tree clones with different levels of latex productivity by analyzing 2-DE gels. The total abundance of each REF/SRPP protein decreased or changed a little upon ethylene stimulation, whereas the abundance of multiple protein species of the same REF/SRPP changed diversely. Among the three rubber tree clones, the abundance of the protein species also differed significantly. Especially, two protein species of REF175 or REF258 were ethylene-responsive only in the high latex productivity clone RY 8-79 instead of in RY 7-33-97 and PR 107. Some individual protein species were positively related to ethylene stimulation and latex productivity. These results suggested that the specific protein species could be more important than others for rubber production and post-translational modifications might play important roles in rubber biosynthesis. Full article
(This article belongs to the Special Issue Selected Papers from the 6th National Plant Protein Research Congress)
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Open AccessArticle Subcutaneous and Visceral Adipose Tissue Secretions from Extremely Obese Men and Women both Acutely Suppress Muscle Insulin Signaling
Int. J. Mol. Sci. 2017, 18(5), 959; doi:10.3390/ijms18050959
Received: 17 February 2017 / Revised: 14 April 2017 / Accepted: 21 April 2017 / Published: 2 May 2017
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Abstract
Adipose tissue plays a key role in the development of type-2 diabetes via the secretion of adipokines. The current study investigated if secretion media derived from intact visceral (VAT) and subcutaneous (SAT) adipose tissues from extremely obese men and women differently suppressed insulin
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Adipose tissue plays a key role in the development of type-2 diabetes via the secretion of adipokines. The current study investigated if secretion media derived from intact visceral (VAT) and subcutaneous (SAT) adipose tissues from extremely obese men and women differently suppressed insulin signaling in human skeletal myotubes derived from a healthy, non-diabetic male and female donor, respectively. Adipose tissue samples were collected from men and women during laparoscopic bariatric surgery. In general, secretion media collected from both SAT and VAT depots caused impaired insulin signaling in myotubes, independent of sex. In females, this was true regardless of the protein kinase B (Akt) phosphorylation site (Akt Thr308 and Akt Ser473) assessed (p < 0.01). In males, both SAT and VAT secretion media reduced Akt Thr308 activation in insulin-stimulated myotubes compared to controls (p < 0.001); however, only the VAT secretion media impaired Akt Ser473 phosphorylation. Independent of sex, 13 out of 18 detected cytokines, chemokines, and growth factors were more abundant in VAT versus SAT secretion media (p < 0.01). Both SAT and VAT secretion media from obese men and women acutely suppress insulin signaling in myotubes, despite different secretion profiles. We propose that this crosstalk model will help to extend our understanding of the interplay between adipose and muscle, as well as the pathogenesis of type-2 diabetes. Full article
(This article belongs to the Special Issue Adipokines)
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Open AccessArticle Analysis of Different Approaches for the Selection of Reference Genes in RT-qPCR Experiments: A Case Study in Skeletal Muscle of Growing Mice
Int. J. Mol. Sci. 2017, 18(5), 1060; doi:10.3390/ijms18051060
Received: 2 April 2017 / Revised: 9 May 2017 / Accepted: 11 May 2017 / Published: 16 May 2017
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Abstract
The reliability of reverse transcription-quantitative PCR (RT-qPCR) results in gene expression studies depends on the approaches used to account for non-biological variations. In order to find a proper normalization strategy for the study of genes related to growth hormone signaling in skeletal muscle
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The reliability of reverse transcription-quantitative PCR (RT-qPCR) results in gene expression studies depends on the approaches used to account for non-biological variations. In order to find a proper normalization strategy for the study of genes related to growth hormone signaling in skeletal muscle of growing mice, nine unrelated genes were evaluated as internal controls. According to the most used algorithms–geNorm, the Comparative ΔCq method, NormFinder and BestKeeper–GSK3B, YWHAZ, RPL13A and RN18S were found as the most stable. However, the relative expression levels of eight of the potential reference genes assessed decreased with age in cDNA samples obtained from the same amount of total RNA. In a different approach to analyze this apparent discrepancy, experiments were performed with cDNA obtained from equal amounts of purified mRNA. Since the decline was still observed, the hypothesis of an age-related change in mRNA to total RNA ratio that could account for the systematic decrease was rejected. Differences among experimental groups could be due to a substantial increase with age in highly expressed mRNAs, which would bias the quantitation of the remaining genes. Consequently, those reference genes reflecting this dilution effect, which would have been discarded considering their variable relative expression levels, arose as suitable internal controls. Full article
(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)
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Open AccessArticle Upregulated Autophagy in Sertoli Cells of Ethanol-Treated Rats Is Associated with Induction of Inducible Nitric Oxide Synthase (iNOS), Androgen Receptor Suppression and Germ Cell Apoptosis
Int. J. Mol. Sci. 2017, 18(5), 1061; doi:10.3390/ijms18051061
Received: 23 February 2017 / Revised: 22 April 2017 / Accepted: 10 May 2017 / Published: 15 May 2017
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Abstract
This study was conducted to investigate the autophagic response of Sertoli cells (SCs) to acute ethanol toxicity using in vivo and in vitro models. Adult Wistar rats were intraperitoneally injected with either 5 g/kg ethanol or phosphate-buffered saline (for the control group) and
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This study was conducted to investigate the autophagic response of Sertoli cells (SCs) to acute ethanol toxicity using in vivo and in vitro models. Adult Wistar rats were intraperitoneally injected with either 5 g/kg ethanol or phosphate-buffered saline (for the control group) and sacrificed 0, 3, 6 and 24 h after injection. Compared to the control group, enhanced germ cell apoptosis was observed in the ethanol-treated rats (ETRs) in association with upregulation of iNOS and reduced expression of androgen receptor protein levels in SCs, which were resistant to apoptosis. Meanwhile, autophagy was upregulated in ETR SCs (peaking at 24 h) compared to the control group, as evidenced by transcription factor EB (TFEB) nuclear translocation, enhanced expression of microtubule-associated protein 1 light chain3-II (LC3-II), lysosome-associated membrane protein-2 (LAMP-2), pan cathepsin protein levels and reduced expression of p62. This upregulation of SC autophagy was confirmed ultrastructurally by enhanced formation of autophagic vacuoles and by immunofluorescent double labelling of autophagosomal and lysosomal markers. Study of cultured SCs confirmed enhanced autophagic response to ethanol toxicity, which was cytoprotective based on decreased viability of SCs upon blocking autophagy with 3-methyladenine (3-MA). The results highlighted the molecular mechanisms of prosurvival autophagy in ETR SCs for the first time, and may have significant implications for male fertility. Full article
(This article belongs to the collection Programmed Cell Death and Apoptosis)
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Open AccessArticle 3D-QSAR, Molecular Docking and Molecular Dynamics Simulation of Pseudomonas aeruginosa LpxC Inhibitors
Int. J. Mol. Sci. 2017, 18(5), 761; doi:10.3390/ijms18050761
Received: 11 February 2017 / Revised: 17 March 2017 / Accepted: 26 March 2017 / Published: 6 May 2017
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Abstract
As an important target for the development of novel antibiotics, UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase (LpxC) has been widely studied. Pyridone methylsulfone hydroxamate (PMH) compounds can effectively inhibit the catalytic activity of LpxC. In this work, the three-dimensional quantitative structure-activity
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As an important target for the development of novel antibiotics, UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase (LpxC) has been widely studied. Pyridone methylsulfone hydroxamate (PMH) compounds can effectively inhibit the catalytic activity of LpxC. In this work, the three-dimensional quantitative structure-activity relationships of PMH inhibitors were explored and models with good predictive ability were established using comparative molecular field analysis and comparative molecular similarity index analysis methods. The effect of PMH inhibitors’ electrostatic potential on the inhibitory ability of Pseudomonas aeruginosa LpxC (PaLpxC) is revealed at the molecular level via molecular electrostatic potential analyses. Then, two molecular dynamics simulations for the PaLpxC and PaLpxC-inhibitor systems were also performed respectively to investigate the key residues of PaLpxC hydrolase binding to water molecules. The results indicate that orderly alternative water molecules can form stable hydrogen bonds with M62, E77, T190, and H264 in the catalytic center, and tetracoordinate to zinc ion along with H78, H237, and D241. It was found that the conformational transition space of PaLpxC changes after association with PMH inhibitors through free energy landscape and cluster analyses. Finally, a possible inhibitory mechanism of PMH inhibitors was proposed, based on our molecular simulation. This paper will provide a theoretical basis for the molecular design of LpxC-targeting antibiotics. Full article
(This article belongs to the Section Molecular Recognition)
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Open AccessArticle Long-Term Follow-Up of Resistance-Associated Substitutions in Hepatitis C Virus in Patients in Which Direct Acting Antiviral-Based Therapy Failed
Int. J. Mol. Sci. 2017, 18(5), 962; doi:10.3390/ijms18050962
Received: 22 March 2017 / Revised: 26 April 2017 / Accepted: 26 April 2017 / Published: 3 May 2017
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Abstract
We evaluated the transition of dominant resistance-associated substitutions (RASs) in hepatitis C virus during long-term follow-up after the failure of DAAs (direct acting antivirals)-based therapy. RASs in non-structure (NS)3/4A, NS5A, NS5B, and deletions in NS5A from 20 patients who failed simeprevir/pegylated-interferon/ribavirin (SMV/PEG-IFN/RBV) and
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We evaluated the transition of dominant resistance-associated substitutions (RASs) in hepatitis C virus during long-term follow-up after the failure of DAAs (direct acting antivirals)-based therapy. RASs in non-structure (NS)3/4A, NS5A, NS5B, and deletions in NS5A from 20 patients who failed simeprevir/pegylated-interferon/ribavirin (SMV/PEG-IFN/RBV) and 25 patients who failed daclatasvir/asunaprevir (DCV/ASV) treatment were examined by direct sequencing. With respect to SMV/PEG-IFN/RBV treatment, RAS was detected at D168 in NS3/4A but not detected in NS5A and NS5B at treatment failure in 16 of 20 patients. During the median follow-up period of 64 weeks, the RAS at D168 became less dominant in 9 of 16 patients. Among 25 DCV/ASV failures, RASs at D168, L31, and Y93 were found in 57.1%, 72.2%, and 76.9%, respectively. NS5A deletions were detected in 3 of 10 patients treated previously with SMV/PEG-IFN/RBV. The number of RASs in the breakthrough patients exceeded that in relapsers (mean 3.9 vs. 2.7, p < 0.05). RAS at D168 in NS3/4A became less dominant in 6 of 15 patients within 80 weeks. Y93H emerged at the time of relapse, then decreased gradually by 99% at 130 weeks post-treatment. Emerged RASs were associated with the clinical course of treatment and could not be detected during longer follow-up. Full article
(This article belongs to the Special Issue Hepatitis Virus Infection and Research)
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Open AccessArticle Ginsenoside Rb2 Alleviates Hepatic Lipid Accumulation by Restoring Autophagy via Induction of Sirt1 and Activation of AMPK
Int. J. Mol. Sci. 2017, 18(5), 1063; doi:10.3390/ijms18051063
Received: 15 February 2017 / Revised: 26 April 2017 / Accepted: 9 May 2017 / Published: 19 May 2017
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Abstract
Although Panax ginseng is a famous traditional Chinese medicine and has been widely used to treat a variety of metabolic diseases including hyperglycemia, hyperlipidemia, and hepatosteatosis, the effective mediators and molecular mechanisms remain largely unknown. In this study we found that ginsenoside Rb2,
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Although Panax ginseng is a famous traditional Chinese medicine and has been widely used to treat a variety of metabolic diseases including hyperglycemia, hyperlipidemia, and hepatosteatosis, the effective mediators and molecular mechanisms remain largely unknown. In this study we found that ginsenoside Rb2, one of the major ginsenosides in Panax ginseng, was able to prevent hepatic lipid accumulation through autophagy induction both in vivo and in vitro. Treatment of male db/db mice with Rb2 significantly improved glucose tolerance, decreased hepatic lipid accumulation, and restored hepatic autophagy. In vitro, Rb2 (50 µmol/L) obviously increased autophagic flux in HepG2 cells and primary mouse hepatocytes, and consequently reduced the lipid accumulation induced by oleic acid in combination with high glucose. Western blotting analysis showed that Rb2 partly reversed the high fatty acid in combination with high glucose (OA)-induced repression of autophagic pathways including AMP-activated protein kinase (AMPK) and silent information regulator 1 (sirt1). Furthermore, pharmacological inhibition of the sirt1 or AMPK pathways attenuated these beneficial effects of Rb2 on hepatic autophagy and lipid accumulation. Taken together, these results suggested that Rb2 alleviated hepatic lipid accumulation by restoring autophagy via the induction of sirt1 and activation of AMPK, and resulted in improved nonalcoholic fatty liver disease (NAFLD) and glucose tolerance. Full article
(This article belongs to the Section Bioactives and Nutraceuticals)
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Open AccessCommunication Mycoviruses in the Plant Pathogen Ustilaginoidea virens Are Not Correlated with the Genetic Backgrounds of Its Hosts
Int. J. Mol. Sci. 2017, 18(5), 963; doi:10.3390/ijms18050963
Received: 23 February 2017 / Revised: 21 April 2017 / Accepted: 26 April 2017 / Published: 3 May 2017
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Abstract
Ustilaginoidea virens, the causal agent of rice false smut, is one of the most devastating grain diseases that causes loss of yield in most rice-growing areas worldwide. In this study, we performed a dsRNA screen to isolate mycoviruses from 35 U. virens
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Ustilaginoidea virens, the causal agent of rice false smut, is one of the most devastating grain diseases that causes loss of yield in most rice-growing areas worldwide. In this study, we performed a dsRNA screen to isolate mycoviruses from 35 U. virens strains. The results revealed that 34 of the tested isolates were infected by various dsRNA elements, displaying highly viral diversity and mixed infections. We characterized a 5.3 kbp dsRNA from a typical isolate containing dsRNA segments with sizes ranging from 0.5 to 5.3 kbp. Sequence analysis of its genomic properties indicated that it is a novel victorivirus, named Ustilaginoidea virens RNA virus 5 (UvRV5), that belongs to the family Totiviridae. RT-PCR detection was performed and indicated that not all the dsRNA bands that were 5.3 kbp in size contained UvRV5. Moreover, the genetic relatedness of all the U. virens strains was estimated according to phylogenetic analysis of the partial intergenic spacer region (IGS) sequences. However, concordance was not found between the dsRNA profiles and the IGS-based genetic relatedness of their host fungi. Full article
(This article belongs to the Section Molecular Botany)
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Open AccessArticle Nimodipine but Not Nifedipine Promotes Expression of Fatty Acid 2-Hydroxylase in a Surgical Stress Model Based on Neuro2a Cells
Int. J. Mol. Sci. 2017, 18(5), 964; doi:10.3390/ijms18050964
Received: 28 March 2017 / Revised: 20 April 2017 / Accepted: 26 April 2017 / Published: 3 May 2017
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Abstract
Nimodipine is well characterized for the management of aneurysmal subarachnoid hemorrhage and has been shown to promote a better outcome and less delayed ischemic neurological deficits. Animal and clinical trials show neuroprotective efficacy following nerve injuries. We showed a neuroprotective effect on Neuro2a
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Nimodipine is well characterized for the management of aneurysmal subarachnoid hemorrhage and has been shown to promote a better outcome and less delayed ischemic neurological deficits. Animal and clinical trials show neuroprotective efficacy following nerve injuries. We showed a neuroprotective effect on Neuro2a cells. Subsequent microarray analysis revealed—among others—fatty acid 2-hydroxylase (FA2H) upregulated by nimodipine in vitro, which is a component of myelin synthesis. Differentiated Neuro2a cells were analyzed for nimodipine-mediated survival considering stress treatment in comparison to nifedipine-treatment. Cell survival was determined by measurement of LDH activity in the culture medium. Nimodipine decreased surgery-like stress-induced cell death of differentiated Neuro2a cells. Neuro2a cell culture was analyzed for changes in FA2H expression induced by nimodipine or nifedipine in surgery-like stress conditions. We analyzed expression levels of FA2H mRNA and protein by qPCR using fa2h specific primers or a FA2H-specific antibody in nimodipine or nifedipine non- and pre-treated Neuro2a cell culture, respectively. Nimodipine but not nifedipine increases FA2H protein levels and also significantly increases mRNA levels of FA2H in both undifferentiated and differentiated Neuro2a cells. Our findings indicate that higher expression of FA2H induced by nimodipine may cause higher survival of Neuro2a cells stressed with surgery-like stressors. Full article
(This article belongs to the Special Issue Neuroprotective Strategies 2017)
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Open AccessArticle First Report of a Fatal Case Associated with EV-D68 Infection in Hong Kong and Emergence of an Interclade Recombinant in China Revealed by Genome Analysis
Int. J. Mol. Sci. 2017, 18(5), 1065; doi:10.3390/ijms18051065
Received: 19 April 2017 / Revised: 4 May 2017 / Accepted: 9 May 2017 / Published: 16 May 2017
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Abstract
A fatal case associated with enterovirus D68 (EV-D68) infection affecting a 10-year-old boy was reported in Hong Kong in 2014. To examine if a new strain has emerged in Hong Kong, we sequenced the partial genome of the EV-D68 strain identified from the
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A fatal case associated with enterovirus D68 (EV-D68) infection affecting a 10-year-old boy was reported in Hong Kong in 2014. To examine if a new strain has emerged in Hong Kong, we sequenced the partial genome of the EV-D68 strain identified from the fatal case and the complete VP1, and partial 5′UTR and 2C sequences of nine additional EV-D68 strains isolated from patients in Hong Kong. Sequence analysis indicated that a cluster of strains including the previously recognized A2 strains should belong to a separate clade, clade D, which is further divided into subclades D1 and D2. Among the 10 EV-D68 strains, 7 (including the fatal case) belonged to the previously described, newly emerged subclade B3, 2 belonged to subclade B1, and 1 belonged to subclade D1. Three EV-D68 strains, each from subclades B1, B3, and D1, were selected for complete genome sequencing and recombination analysis. While no evidence of recombination was noted among local strains, interclade recombination was identified in subclade D2 strains detected in mainland China in 2008 with VP2 acquired from clade A. This study supports the reclassification of subclade A2 into clade D1, and demonstrates interclade recombination between clades A and D2 in EV-D68 strains from China. Full article
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Open AccessArticle Nanosecond-Pulsed DBD Plasma-Generated Reactive Oxygen Species Trigger Immunogenic Cell Death in A549 Lung Carcinoma Cells through Intracellular Oxidative Stress
Int. J. Mol. Sci. 2017, 18(5), 966; doi:10.3390/ijms18050966
Received: 1 March 2017 / Revised: 27 April 2017 / Accepted: 28 April 2017 / Published: 3 May 2017
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Abstract
A novel application for non-thermal plasma is the induction of immunogenic cancer cell death for cancer immunotherapy. Cells undergoing immunogenic death emit danger signals which facilitate anti-tumor immune responses. Although pathways leading to immunogenic cell death are not fully understood; oxidative stress is
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A novel application for non-thermal plasma is the induction of immunogenic cancer cell death for cancer immunotherapy. Cells undergoing immunogenic death emit danger signals which facilitate anti-tumor immune responses. Although pathways leading to immunogenic cell death are not fully understood; oxidative stress is considered to be part of the underlying mechanism. Here; we studied the interaction between dielectric barrier discharge plasma and cancer cells for oxidative stress-mediated immunogenic cell death. We assessed changes to the intracellular oxidative environment after plasma treatment and correlated it to emission of two danger signals: surface-exposed calreticulin and secreted adenosine triphosphate. Plasma-generated reactive oxygen and charged species were recognized as the major effectors of immunogenic cell death. Chemical attenuators of intracellular reactive oxygen species successfully abrogated oxidative stress following plasma treatment and modulated the emission of surface-exposed calreticulin. Secreted danger signals from cells undergoing immunogenic death enhanced the anti-tumor activity of macrophages. This study demonstrated that plasma triggers immunogenic cell death through oxidative stress pathways and highlights its potential development for cancer immunotherapy. Full article
(This article belongs to the Special Issue Tumor Targeting Therapy and Selective Killing)
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Open AccessArticle Studies on Trans-Resveratrol/Carboxymethylated (1,3/1,6)-β-d-Glucan Association for Aerosol Pharmaceutical Applications
Int. J. Mol. Sci. 2017, 18(5), 967; doi:10.3390/ijms18050967
Received: 21 March 2017 / Revised: 20 April 2017 / Accepted: 30 April 2017 / Published: 3 May 2017
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Abstract
A resveratrol/carboxymethylated glucan (CM-glucan) combination is known to inhibit rhinovirus replication and expression of inflammatory mediators in nasal epithelia. The aim of this study was to develop an aerosol formulation containing an association of the two molecules which could reach the lower respiratory
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A resveratrol/carboxymethylated glucan (CM-glucan) combination is known to inhibit rhinovirus replication and expression of inflammatory mediators in nasal epithelia. The aim of this study was to develop an aerosol formulation containing an association of the two molecules which could reach the lower respiratory tract. Mass median aerodynamic diameter (MMAD) of a resveratrol/CM-glucan combination was lower than that shown by resveratrol or CM-glucan alone (2.83 versus 3.28 and 2.96 µm, respectively). The resveratrol/CM-glucan association results in the finest and most monodispersed particles in comparison to the two single components. The association also evidenced lower values for all particle size distribution parameters, suggesting that the pharmacological synergy observed in previous studies may be accompanied by a pharmaceutical one. Moreover, we showed that the CM-glucan matrix did not exert an inhibitory effect on resveratrol nebulization, demonstrating the good suitability of these two molecules in association for simultaneous aerosol volatilization. Full article
(This article belongs to the Special Issue Glucan: New Perspectives on Biochemistry and Application)
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Open AccessArticle Atg7 Regulates Brain Angiogenesis via NF-κB-Dependent IL-6 Production
Int. J. Mol. Sci. 2017, 18(5), 968; doi:10.3390/ijms18050968
Received: 24 March 2017 / Revised: 25 April 2017 / Accepted: 28 April 2017 / Published: 3 May 2017
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Abstract
The formation of brain vasculature is an essential step during central nervous system development. The molecular mechanism underlying brain angiogenesis remains incompletely understood. The role of Atg7, an autophagy-related protein, in brain angiogenesis was investigated in this study. We found that the microvessel
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The formation of brain vasculature is an essential step during central nervous system development. The molecular mechanism underlying brain angiogenesis remains incompletely understood. The role of Atg7, an autophagy-related protein, in brain angiogenesis was investigated in this study. We found that the microvessel density in mice brains with endothelial-specific knockout of Atg7 (Atg7 EKO) was significantly decreased compared to wild-type control. Consistently, in vitro angiogenesis assays showed that Atg7 knockdown impaired angiogenesis in brain microvascular endothelial cells. Further results indicated that knockdown of Atg7 reduced interleukin-6 (IL-6) expression in brain microvascular endothelial cells, which is mediated by NF-κB-dependent transcriptional control. Interestingly, exogenous IL-6 restored the impaired angiogenesis and reduced cell motility caused by Atg7 knockdown. These results demonstrated that Atg7 has proangiogenic activity in brain angiogenesis which is mediated by IL-6 production in a NF-κB-dependent manner. Full article
(This article belongs to the Special Issue Vascular Biology and Therapeutics)
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Open AccessArticle The Regulatory Capacity of Bivalent Genes—A Theoretical Approach
Int. J. Mol. Sci. 2017, 18(5), 1069; doi:10.3390/ijms18051069
Received: 3 April 2017 / Revised: 8 May 2017 / Accepted: 10 May 2017 / Published: 17 May 2017
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