Current Issues in Molecular Biology

15 pages, 1306 KiB

Open AccessBrief Report

High Caloric Diet Induces Memory Impairment and Disrupts Synaptic Plasticity in Aged Rats

by Sara L. Paulo, Catarina Miranda-Lourenço, Rita F. Belo, Rui S. Rodrigues, João Fonseca-Gomes, Sara R. Tanqueiro, Vera Geraldes, Isabel Rocha, Ana M. Sebastião, Sara Xapelli and Maria J. Diógenes

Curr. Issues Mol. Biol. 2021, 43(3), 2305-2319; https://doi.org/10.3390/cimb43030162 - 18 Dec 2021

Cited by 9 | Viewed by 3183

Abstract

The increasing consumption of sugar and fat seen over the last decades and the consequent overweight and obesity, were recently linked with a deleterious effect on cognition and synaptic function. A major question, which remains to be clarified, is whether obesity in the [...] Read more.

The increasing consumption of sugar and fat seen over the last decades and the consequent overweight and obesity, were recently linked with a deleterious effect on cognition and synaptic function. A major question, which remains to be clarified, is whether obesity in the elderly is an additional risk factor for cognitive impairment. We aimed at unravelling the impact of a chronic high caloric diet (HCD) on memory performance and synaptic plasticity in aged rats. Male rats were kept on an HCD or a standard diet (control) from 1 to 24 months of age. The results showed that under an HCD, aged rats were obese and displayed significant long-term recognition memory impairment when compared to age-matched controls. Ex vivo synaptic plasticity recorded from hippocampal slices from HCD-fed aged rats revealed a reduction in the magnitude of long-term potentiation, accompanied by a decrease in the levels of the brain-derived neurotrophic factor receptors TrkB full-length (TrkB-FL). No alterations in neurogenesis were observed, as quantified by the density of immature doublecortin-positive neurons in the hippocampal dentate gyrus. This study highlights that obesity induced by a chronic HCD exacerbates age-associated cognitive decline, likely due to impaired synaptic plasticity, which might be associated with deficits in TrkB-FL signaling. Full article

(This article belongs to the Topic Neuroprotection by Drugs, Nutraceuticals and Physical Activity)

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16 pages, 1891 KiB

Open AccessArticle

Improved Production of Streptomyces sp. FA1 Xylanase in a Dual-Plasmid Pichia pastoris System

by Wei Xia, Mengkai Hu, Yang Pan, Dan Wu and Jing Wu

Curr. Issues Mol. Biol. 2021, 43(3), 2289-2304; https://doi.org/10.3390/cimb43030161 - 18 Dec 2021

Cited by 8 | Viewed by 3108

Abstract

Methanol is considered as a potential hazard in the methanol-induced yeast expression of food-related enzymes. To increase the production efficiency of recombinant proteins in Pichia pastoris without methanol induction, a novel dual-plasmid system was constructed, for the first time, by a combining [...] Read more.

Methanol is considered as a potential hazard in the methanol-induced yeast expression of food-related enzymes. To increase the production efficiency of recombinant proteins in Pichia pastoris without methanol induction, a novel dual-plasmid system was constructed, for the first time, by a combining the strategies of genomic integration and episomal expression. To obtain a high copy number of the target gene, the autonomously replicating sequence derived from Kluyveromyces lactis (PARS) was used to construct episomal vectors carrying the constitutive promoters P_GAP and P_GCW14. In addition, an integrative vector carrying the P_GCW14 promoter was constructed by replacing the P_GAP promoter sequence with a partial P_GCW14 promoter. Next, using xylanase XynA from Streptomyces sp. FA1 as the model enzyme, recombination strains were transformed with different combinations of integrating and episomal vectors that were constructed to investigate the changes in the protein yield. Results in shake flasks indicated that the highest enzyme yield was achieved when integrated P_GAP and episomal P_GCW14 were simultaneously transformed into the host strain. Meanwhile, the copy number of xynA increased from 1.14 ± 0.46 to 3.06 ± 0.35. The yield of XynA was successfully increased to 3925 U·mL⁻¹ after 102 h of fermentation in a 3.6 L fermenter, which was 16.7-fold and 2.86-fold of the yields that were previously reported for the constitutive expression and methanol-induced expression of the identical protein, respectively. Furthermore, the high-cell-density fermentation period was shortened from 132 h to 102 h compared to that of methanol-induced system. Since the risk of methanol toxicity is removed, this novel expression system would be suitable for the production of proteins related to the food and pharmaceutical industries. Full article

(This article belongs to the Section Molecular Microbiology)

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13 pages, 961 KiB

Open AccessArticle

Construction of Three High-Density Genetic Linkage Maps and Dynamic QTL Mapping of Growth Traits in Yellow River Carp (Cyprinus carpio haematopterus)

by Lei Wang, Songpeng Jia, Yuxuan Zhang, Shuhong Jiang, Yuhan Chen, Junping Chen, Miao Yu, Lan Zhang, Zhigang Qiao and Xuejun Li

Curr. Issues Mol. Biol. 2021, 43(3), 2276-2288; https://doi.org/10.3390/cimb43030160 - 17 Dec 2021

Cited by 5 | Viewed by 2205

Abstract

To provide the theoretical basis for researching growth, development, and molecular marker-assisted breeding of the economically important Yellow River carp (Cyprinus carpio haematopterus) using dynamic quantitative trait locus (QTL) mapping, we constructed three genetic linkage maps from 207 progeny using a [...] Read more.

To provide the theoretical basis for researching growth, development, and molecular marker-assisted breeding of the economically important Yellow River carp (Cyprinus carpio haematopterus) using dynamic quantitative trait locus (QTL) mapping, we constructed three genetic linkage maps from 207 progeny using a new modified genotyping-by-sequencing method. The three maps contained 16,886, 16,548, and 7482 single nucleotide polymorphism markers, respectively, with an average interval of 0.36 cM, 0.45 cM, and 1.00 cM. We identified 148 QTLs related to four growth traits that were located on 25 chromosomes from three growth stages of Yellow River carp. A total of 32, 36, 43, and 37 QTLs were associated with body length, height, width, and weight, respectively. Among them, 47 QTLs were detected for only one growth trait in one stage, but all of the other QTLs were co-localized. Of the 14 main QTLs, 13 were located on chromosome 12, which suggests the presence of growth-related genes on this chromosome. We then detected 17 candidate genes within 50 K upstream and downstream of the 14 main QTLs. This is the first report of the dynamic QTL mapping of growth traits of Yellow River carp, and the results can be used in future studies of growth, development, and molecular-assisted breeding of this species. Full article

(This article belongs to the Section Bioinformatics and Systems Biology)

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10 pages, 5341 KiB

Open AccessCase Report

Case Report: Genetic Alterations Associated with the Progression of Carotid Paraganglioma

by Vladislav Pavlov, Anastasiya Snezhkina, Dmitry Kalinin, Alexander Golovyuk, Anastasiya Kobelyatskaya, Ildar Bakhtogarimov, Nadezhda Volchenko, George Krasnov and Anna Kudryavtseva

Curr. Issues Mol. Biol. 2021, 43(3), 2266-2275; https://doi.org/10.3390/cimb43030159 - 17 Dec 2021

Cited by 2 | Viewed by 2409

Abstract

Paragangliomas (PGLs) are rare neuroendocrine tumors that can develop from any paraganglion across the body. The carotid body is the most often location of PGLs in the head and neck region. Carotid PGLs (CPGLs) are characterized by predominantly non-aggressive behavior; however, all tumors [...] Read more.

Paragangliomas (PGLs) are rare neuroendocrine tumors that can develop from any paraganglion across the body. The carotid body is the most often location of PGLs in the head and neck region. Carotid PGLs (CPGLs) are characterized by predominantly non-aggressive behavior; however, all tumors have the potential to metastasize. To date, molecular mechanisms of paraganglioma progression remain elusive. We report a case of a 38-year-old woman with metastatic CPGL manifesting as a recurrent tumor with lymph node metastasis. The tumor was fast-growing and had a high Ki-67 proliferation index. Immunohistochemical (IHC) examination and whole-exome sequencing were performed for both recurrent tumor and metastasis. A germline pathogenic splice acceptor variant in the SDHB gene was found in the patient. Immunoreactivity of the SDHB subunit was weak diffuse in both samples, indicating deficiency of the succinate dehydrogenase. Moreover, the recurrent tumor exhibited loss of heterozygosity (LOH) at the SDHB locus, that is according to Knudson’s "two-hit" hypothesis of cancer causation. We also identified a rare somatic promotor mutation in the TERT gene associated with the tumor progression. Obtained results confirmed the indicative role of the germline SDHB mutation for metastatic CPGLs, as well as the potential prognostic value of the TERT promoter mutation. Full article

(This article belongs to the Special Issue Linking Genomic Changes with Cancer in the NGS Era)

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13 pages, 1697 KiB

Open AccessArticle

Effect of Ethanol on Parthenogenetic Activation and α-Tocopherol Supplementation during In Vitro Maturation on Developmental Competence of Summer-Collected Bovine Oocytes

by Francisco Báez, Belén Gómez, Victoria de Brun, Nélida Rodríguez-Osorio and Carolina Viñoles

Curr. Issues Mol. Biol. 2021, 43(3), 2253-2265; https://doi.org/10.3390/cimb43030158 - 16 Dec 2021

Cited by 6 | Viewed by 2588

Abstract

The use of α-tocopherol during in vitro maturation (IVM) is an alternative to minimize the adverse effects of heat stress on oocyte competence. However, α-tocopherol is diluted in ethanol, which can induce oocyte parthenogenetic activation (PA). This study aimed to evaluate the role [...] Read more.

The use of α-tocopherol during in vitro maturation (IVM) is an alternative to minimize the adverse effects of heat stress on oocyte competence. However, α-tocopherol is diluted in ethanol, which can induce oocyte parthenogenetic activation (PA). This study aimed to evaluate the role of ethanol concentration on PA and the effect of α-tocopherol supplementation during IVM on the developmental competence and the expression of key genes in blastocysts derived from summer-collected oocytes. All in vitro embryo production was conducted at 5% O₂, 5% CO₂ at 38.5 °C. Experiment 1: oocytes were cultured with or without 0.05% ethanol. As positive PA control matured oocytes were subjected to 3% or 7% ethanol for 7 min. Oocytes from all groups were placed in fertilization medium (22 h) and culture medium (9 days). Ethanol at 0.05% during IVM did not induce oocyte PA, however, 3% and 7% ethanol were effective parthenogenetic inductors. Experiment 2: oocytes were cultured in maturation medium supplemented with 0, 50, 100 and 200 μM α-tocopherol, diluted in 0.05% ethanol. After in vitro fertilization and embryo culture, we assessed blastocyst apoptotic index and the transcription of a panel of genes. The results showed that supplementation with 100 μM α-tocopherol reduced apoptotic index and increased the expression of SOD2. In conclusion, 100 μM α-tocopherol, diluted in 0.05% ethanol, can be used during IVM to embryonic quality. Full article

(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)

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15 pages, 2490 KiB

Open AccessArticle

A Link between Mitochondrial Dysregulation and Idiopathic Autism Spectrum Disorder (ASD): Alterations in Mitochondrial Respiratory Capacity and Membrane Potential

by Hazirah Hassan, Fazaine Zakaria, Suzana Makpol and Norwahidah Abdul Karim

Curr. Issues Mol. Biol. 2021, 43(3), 2238-2252; https://doi.org/10.3390/cimb43030157 - 16 Dec 2021

Cited by 7 | Viewed by 3099

Abstract

Autism spectrum disorder (ASD) is a neurological disorder triggered by various factors through complex mechanisms. Research has been done to elucidate the potential etiologic mechanisms in ASD, but no single cause has been confirmed. The involvement of oxidative stress is correlated with ASD [...] Read more.

Autism spectrum disorder (ASD) is a neurological disorder triggered by various factors through complex mechanisms. Research has been done to elucidate the potential etiologic mechanisms in ASD, but no single cause has been confirmed. The involvement of oxidative stress is correlated with ASD and possibly affects mitochondrial function. This study aimed to elucidate the link between mitochondrial dysregulation and idiopathic ASD by focusing on mitochondrial respiratory capacity and membrane potential. Our findings showed that mitochondrial function in the energy metabolism pathway was significantly dysregulated in a lymphoblastoid cell line (LCL) derived from an autistic child (ALCL). Respiratory capacities of oxidative phosphorylation (OXPHOS), electron transfer of the Complex I and Complex II linked pathways, membrane potential, and Complex IV activity of the ALCL were analyzed and compared with control cell lines derived from a developmentally normal non-autistic sibling (NALCL). All experiments were performed using high-resolution respirometry. Respiratory capacities of OXPHOS, electron transfer of the Complex I- and Complex II-linked pathways, and Complex IV activity of the ALCL were significantly higher compared to healthy controls. Mitochondrial membrane potential was also significantly higher, measured in the Complex II-linked pathway during LEAK respiration and OXPHOS. These results indicate the abnormalities in mitochondrial respiratory control linking mitochondrial function with autism. Correlating mitochondrial dysfunction and autism is important for a better understanding of ASD pathogenesis in order to produce effective interventions. Full article

(This article belongs to the Topic Neuroprotection by Drugs, Nutraceuticals and Physical Activity)

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18 pages, 1417 KiB

Open AccessArticle

Genome Analysis of the Janthinobacterium sp. Strain SLB01 from the Diseased Sponge of the Lubomirskia baicalensis

by Sergei I. Belikov, Ivan S. Petrushin and Lubov I. Chernogor

Curr. Issues Mol. Biol. 2021, 43(3), 2220-2237; https://doi.org/10.3390/cimb43030156 - 11 Dec 2021

Cited by 4 | Viewed by 2902

Abstract

The strain Janthinobacterium sp. SLB01 was isolated from the diseased freshwater sponge Lubomirskia baicalensis (Pallas, 1776) and the draft genome was published previously. The aim of this work is to analyze the genome of the Janthinobacterium sp. SLB01 to search for pathogenicity factors [...] Read more.

The strain Janthinobacterium sp. SLB01 was isolated from the diseased freshwater sponge Lubomirskia baicalensis (Pallas, 1776) and the draft genome was published previously. The aim of this work is to analyze the genome of the Janthinobacterium sp. SLB01 to search for pathogenicity factors for Baikal sponges. We performed genomic analysis to determine virulence factors, comparing the genome of the strain SLB01 with genomes of other related J. lividum strains from the environment. The strain Janthinobacterium sp. SLB01 contained genes encoding violacein, alpha-amylases, phospholipases, chitinases, collagenases, hemolysin, and a type VI secretion system. In addition, the presence of conservative clusters of genes for the biosynthesis of secondary metabolites of tropodithietic acid and marinocine was found. We present genes for antibiotic resistance, including five genes encoding various lactamases and eight genes for penicillin-binding proteins, which are conserved in all analyzed strains. Major differences were found between the Janthinobacterium sp. SLB01 and J. lividum strains in the spectra of genes for glycosyltransferases and glycoside hydrolases, serine hydrolases, and trypsin-like peptidase, as well as some TonB-dependent siderophore receptors. Thus, the study of the analysis of the genome of the strain SLB01 allows us to conclude that the strain may be one of the pathogens of freshwater sponges. Full article

(This article belongs to the Section Molecular Microbiology)

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10 pages, 1421 KiB

Open AccessArticle

Simulated Microgravity Induces the Proliferative Inhibition and Morphological Changes in Porcine Granulosa Cells

by Truong Xuan Dai, Hoang Nghia Son, Ho Nguyen Quynh Chi, Hoang Nghia Quang Huy, Nguyen Thai Minh, Nguyen Thi Thuy Tram, Nguyen Thi Thuong Huyen, To Minh Quan, Doan Chinh Chung, Truong Hai Nhung, Tran Thi Minh, Tran Hong Diem, Nguyen Thi Phuong Mai and Le Thanh Long

Curr. Issues Mol. Biol. 2021, 43(3), 2210-2219; https://doi.org/10.3390/cimb43030155 - 10 Dec 2021

Cited by 3 | Viewed by 3065

Abstract

Astronauts are always faced with serious health problems during prolonged spaceflights. Previous studies have shown that weightlessness significantly affects the physiological function of female astronauts, including a change in reproductive hormones and ovarian cells, such as granulosa and theca cells. However, the effects [...] Read more.

Astronauts are always faced with serious health problems during prolonged spaceflights. Previous studies have shown that weightlessness significantly affects the physiological function of female astronauts, including a change in reproductive hormones and ovarian cells, such as granulosa and theca cells. However, the effects of microgravity on these cells have not been well characterized, especially in granulosa cells. This study aimed to investigate the effects of simulated microgravity (SMG) on the proliferation and morphology of porcine granulosa cells (pGCs). pGC proliferation from the SMG group was inhibited, demonstrated by the reduced O.D. value and cell density in the WST-1 assay and cell number counting. SMG-induced pGCs exhibited an increased ratio of cells in the G0/G1 phase and a decreased ratio of cells in the S and G2/M phase. Western blot analysis indicated a down-regulation of cyclin D1, cyclin-dependent kinase 4 (cdk4), and cyclin-dependent kinase 6 (cdk6), leading to the prevention of the G1-S transition and inducing the arrest phase. pGCs under the SMG condition showed an increase in nuclear area. This caused a reduction in nuclear shape value in pGCs under the SMG condition. SMG-induced pGCs exhibited different morphologies, including fibroblast-like shape, rhomboid shape, and pebble-like shape. These results revealed that SMG inhibited proliferation and induced morphological changes in pGCs. Full article

(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)

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11 pages, 2907 KiB

Open AccessArticle

Effect of Regorafenib on P2X7 Receptor Expression and Different Oncogenic Signaling Pathways in a Human Breast Cancer Cell Line: A Potential of New Insight of the Antitumor Effects of Regorafenib

by Muhammed M. Salahuddin, Gamal A. Omran, Maged W. Helmy and Maha E. Houssen

Curr. Issues Mol. Biol. 2021, 43(3), 2199-2209; https://doi.org/10.3390/cimb43030154 - 9 Dec 2021

Cited by 10 | Viewed by 3447

Abstract

Background: Breast cancer is the most common malignancy in women worldwide. P2X7 is a transmembrane receptor expressed in breast cancer and activated by the ATP tumor microenvironment, driving cell proliferation, angiogenesis, and metastasis via different signaling pathways. The role of the P2X7 receptor, [...] Read more.

Background: Breast cancer is the most common malignancy in women worldwide. P2X7 is a transmembrane receptor expressed in breast cancer and activated by the ATP tumor microenvironment, driving cell proliferation, angiogenesis, and metastasis via different signaling pathways. The role of the P2X7 receptor, hypoxia, and autophagy in regulating tumor progression is controversial. The multikinase inhibitor regorafenib prevents the activation of numerous kinases involved in angiogenesis, proliferation, and metastasis. The present study aimed to evaluate the modulatory effect of regorafenib on the hypoxia/angiogenesis/P2X7R/autophagy axis on the MCF7 breast cancer cell line and its impact on different signaling pathways involved in breast cancer pathogenesis. Methods: The levels of VEGF, VEGFR, PI3K, NF-κB, HIF-1α, and LC3-II were analyzed using ELISA, and caspase-3 activity was also assessed colorimetrically. Phosphorylated (p)-p38 MAPK and purinergic ligand-gated ion channel 7 (P2X7) receptor protein expression levels were analyzed via Western blotting. Reverse transcription-quantitative PCR was used to determine the mRNA expression levels of Beclin 1 (BECN1), LC3-II, and sequestosome 1 (p62). Results: Regorafenib reduced MCF7 cell viability in a dose-dependent manner. Furthermore, regorafenib significantly reduced levels of PI3K, NF-κB, VEGF, VEGFR, P2X7 receptor, and p-p38 MAPK protein expression, and markedly reduced p62 mRNA expression levels. However, regorafenib significantly increased caspase-3 activity, as well as BECN1 and LC3-II mRNA expression levels. Conclusions: Regorafenib was demonstrated to possibly exhibit antitumor activity on the breast cancer cell line via modulation of the P2X7/HIF-1α/VEGF, P2X7/P38, P2X7/ERK/NF-κB, and P2X7/beclin 1 pathways. Full article

(This article belongs to the Section Molecular Pharmacology)

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10 pages, 1488 KiB

Open AccessArticle

Permeability of Gemcitabine and PBPK Modeling to Assess Oral Administration

by Abigail Ferreira, Rui Lapa and Nuno Vale

Curr. Issues Mol. Biol. 2021, 43(3), 2189-2198; https://doi.org/10.3390/cimb43030153 - 7 Dec 2021

Cited by 3 | Viewed by 3463

Abstract

Gemcitabine is a nucleoside analog effective against several solid tumors. Standard treatment consists of an intravenous infusion over 30 min. This is an invasive, uncomfortable and often painful method, involving recurring visits to the hospital and costs associated with medical staff and equipment. [...] Read more.

Gemcitabine is a nucleoside analog effective against several solid tumors. Standard treatment consists of an intravenous infusion over 30 min. This is an invasive, uncomfortable and often painful method, involving recurring visits to the hospital and costs associated with medical staff and equipment. Gemcitabine’s activity is significantly limited by numerous factors, including metabolic inactivation, rapid systemic clearance of gemcitabine and transporter deficiency-associated resistance. As such, there have been research efforts to improve gemcitabine-based therapy efficacy, as well as strategies to enhance its oral bioavailability. In this work, gemcitabine in vitro and clinical data were analyzed and in silico tools were used to study the pharmacokinetics of gemcitabine after oral administration following different regimens. Several physiologically based pharmacokinetic (PBPK) models were developed using simulation software GastroPlus™, predicting the PK parameters and plasma concentration–time profiles. The integrative biomedical data analyses presented here are promising, with some regimens of oral administration reaching higher AUC in comparison to the traditional IV infusion, supporting this route of administration as a viable alternative to IV infusions. This study further contributes to personalized health care based on potential new formulations for oral administration of gemcitabine, as well nanotechnology-based drug delivery systems. Full article

(This article belongs to the Collection Application of Natural and Pseudo Natural Products in Drug Discovery and Development)

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12 pages, 1421 KiB

Open AccessArticle

De Novo Assembly and Species-Specific Marker Development as a Useful Tool for the Identification of Scutellaria L. Species

by Hakjoon Choi, Wan Seok Kang, Jin Seok Kim, Chang-Su Na and Sunoh Kim

Curr. Issues Mol. Biol. 2021, 43(3), 2177-2188; https://doi.org/10.3390/cimb43030152 - 1 Dec 2021

Cited by 1 | Viewed by 3147

Abstract

Scutellaria L. (family Lamiaceae) includes approximately 470 species found in most parts of the world and is commonly known as skullcaps. Scutellaria L. is a medicinal herb used as a folk remedy in Korea and East Asia, but it is difficult to [...] Read more.

Scutellaria L. (family Lamiaceae) includes approximately 470 species found in most parts of the world and is commonly known as skullcaps. Scutellaria L. is a medicinal herb used as a folk remedy in Korea and East Asia, but it is difficult to identify and classify various subspecies by morphological methods. Since Scutellaria L. has not been studied genetically, to expand the knowledge of species in the genus Scutellaria L., de novo whole-genome assembly was performed in Scutellaria indica var. tsusimensis (H. Hara) Ohwi using the Illumina sequencing platform. We aimed to develop a molecular method that could be used to classify S.indica var. tsusimensis (H. Hara) Ohwi, S. indica L. and three other Scutellaria L. species. The assembly results for S.indica var. tsusimensis (H. Hara) Ohwi revealed a genome size of 318,741,328 bp and a scaffold N50 of 78,430. The assembly contained 92.08% of the conserved BUSCO core gene set and was estimated to cover 94.65% of the genome. The obtained genes were compared with previously registered Scutellaria nucleotide sequences and similar regions using the NCBI BLAST service, and a total of 279 similar nucleotide sequences were detected. By selecting the 279 similar nucleotide sequences and nine chloroplast DNA barcode genes, primers were prepared so that the size of the PCR product was 100 to 1000 bp. As a result, a species-specific primer set capable of distinguishing five species of Scutellaria L. was developed. Full article

(This article belongs to the Section Bioinformatics and Systems Biology)

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10 pages, 1080 KiB

Open AccessArticle

Gene Expression Studies in Formalin-Fixed Paraffin-Embedded Samples of Cutaneous Cancer: The Need for Reference Genes

by Omar García-Pérez, Leticia Melgar-Vilaplana, Elizabeth Córdoba-Lanús and Ricardo Fernández-de-Misa

Curr. Issues Mol. Biol. 2021, 43(3), 2167-2176; https://doi.org/10.3390/cimb43030151 - 30 Nov 2021

Cited by 3 | Viewed by 2709

Abstract

Formalin-fixed paraffin-embedded (FFPE) tumour samples may provide crucial data regarding biomarkers for neoplasm progression. Analysis of gene expression is frequently used for this purpose. Therefore, mRNA expression needs to be normalized through comparison to reference genes. In this study, we establish which of [...] Read more.

Formalin-fixed paraffin-embedded (FFPE) tumour samples may provide crucial data regarding biomarkers for neoplasm progression. Analysis of gene expression is frequently used for this purpose. Therefore, mRNA expression needs to be normalized through comparison to reference genes. In this study, we establish which of the usually reported reference genes is the most reliable one in cutaneous malignant melanoma (MM) and cutaneous squamous cell carcinoma (CSCC). ACTB, TFRC, HPRT1 and TBP expression was quantified in 123 FFPE samples (74 MM and 49 CSCC biopsies) using qPCR. Expression stability was analysed by NormFinder and Bestkeeper softwares, and the direct comparison method between means and SD. The in-silico analysis with BestKeeper indicated that HPRT1 was more stable than ACTB and TFRC in MM (1.85 vs. 2.15) and CSCC tissues (2.09 vs. 2.33). The best option to NormFinder was ACTB gene (0.56) in MM and TFRC (0.26) in CSCC. The direct comparison method showed lower SD means of ACTB expression in MM (1.17) and TFRC expression in CSCC samples (1.00). When analysing the combination of two reference genes for improving stability, NormFinder indicated HPRT1 and ACTB to be the best for MM samples, and HPRT1 and TFRC genes for CSCC. In conclusion, HPRT1 and ACTB genes in combination are the most appropriate choice for normalization in gene expression studies in MM FFPE tissue, while the combination of HPRT1 and TFRC genes are the best option in analysing CSCC FFPE samples. These may be used consistently in forthcoming studies on gene expression in both tumours. Full article

(This article belongs to the Topic Cancer Biology and Therapy)

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10 pages, 1039 KiB

Open AccessArticle

The Identification of Marker Genes for Predicting the Osteogenic Differentiation Potential of Mesenchymal Stromal Cells

by Masami Kanawa, Akira Igarashi, Katsumi Fujimoto, Tania Saskianti, Ayumu Nakashima, Yukihito Higashi, Hidemi Kurihara, Yukio Kato and Takeshi Kawamoto

Curr. Issues Mol. Biol. 2021, 43(3), 2157-2166; https://doi.org/10.3390/cimb43030150 - 30 Nov 2021

Cited by 7 | Viewed by 2673

Abstract

Mesenchymal stromal cells (MSCs) have the potential to differentiate into a variety of mature cell types and are a promising source of regenerative medicine. The success of regenerative medicine using MSCs strongly depends on their differentiation potential. In this study, we sought to [...] Read more.

Mesenchymal stromal cells (MSCs) have the potential to differentiate into a variety of mature cell types and are a promising source of regenerative medicine. The success of regenerative medicine using MSCs strongly depends on their differentiation potential. In this study, we sought to identify marker genes for predicting the osteogenic differentiation potential by comparing ilium MSC and fibroblast samples. We measured the mRNA levels of 95 candidate genes in nine ilium MSC and four fibroblast samples before osteogenic induction, and compared them with alkaline phosphatase (ALP) activity as a marker of osteogenic differentiation after induction. We identified 17 genes whose mRNA expression levels positively correlated with ALP activity. The chondrogenic and adipogenic differentiation potentials of jaw MSCs are much lower than those of ilium MSCs, although the osteogenic differentiation potential of jaw MSCs is comparable with that of ilium MSCs. To select markers suitable for predicting the osteogenic differentiation potential, we compared the mRNA levels of the 17 genes in ilium MSCs with those in jaw MSCs. The levels of 7 out of the 17 genes were not substantially different between the jaw and ilium MSCs, while the remaining 10 genes were expressed at significantly lower levels in jaw MSCs than in ilium MSCs. The mRNA levels of the seven similarly expressed genes were also compared with those in fibroblasts, which have little or no osteogenic differentiation potential. Among the seven genes, the mRNA levels of IGF1 and SRGN in all MSCs examined were higher than those in any of the fibroblasts. These results suggest that measuring the mRNA levels of IGF1 and SRGN before osteogenic induction will provide useful information for selecting competent MSCs for regenerative medicine, although the effectiveness of the markers is needed to be confirmed using a large number of MSCs, which have various levels of osteogenic differentiation potential. Full article

(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)

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10 pages, 896 KiB

Open AccessArticle

Molecular Genetics of Pre-B Acute Lymphoblastic Leukemia Sister Cell Lines during Disease Progression

by Hilmar Quentmeier, Claudia Pommerenke and Hans G. Drexler

Curr. Issues Mol. Biol. 2021, 43(3), 2147-2156; https://doi.org/10.3390/cimb43030149 - 30 Nov 2021

Viewed by 2601

Abstract

For many years, immortalized tumor cell lines have been used as reliable tools to understand the function of oncogenes and tumor suppressor genes. Today, we know that tumors can comprise subclones with common and with subclone-specific genetic alterations. We sequenced DNA and RNA [...] Read more.

For many years, immortalized tumor cell lines have been used as reliable tools to understand the function of oncogenes and tumor suppressor genes. Today, we know that tumors can comprise subclones with common and with subclone-specific genetic alterations. We sequenced DNA and RNA of sequential sister cell lines obtained from patients with pre-B acute lymphoblastic leukemia at different phases of the disease. All five pairs of cell lines carry alterations that are typical for this disease: loss of tumor suppressors (CDKN2A, CDKN2B), expression of fusion genes (ETV6-RUNX1, BCR-ABL1, MEF2D-BCL9) or of genes targeted by point mutations (KRAS A146T, NRAS G12C, PAX5 R38H). MEF2D-BCL9 and PAX R38H mutations in cell lines have hitherto been undescribed, suggesting that YCUB-4 (MEF2D-BCL9), PC-53 (PAX R38H) and their sister cell lines will be useful models to elucidate the function of these genes. All aberrations mentioned above occur in both sister cell lines, demonstrating that the sisters derive from a common ancestor. However, we also found mutations that are specific for one sister cell line only, pointing to individual subclones of the primary tumor as originating cells. Our data show that sequential sister cell lines can be used to study the clonal development of tumors and to elucidate the function of common and clone-specific mutations. Full article

(This article belongs to the Section Molecular Medicine)

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12 pages, 1043 KiB

Open AccessArticle

Evaluation of Microbiome Alterations Following Consumption of BIOHM, a Novel Probiotic

by Mahmoud A. Ghannoum, Thomas S. McCormick, Mauricio Retuerto, Gurkan Bebek, Susan Cousineau, Lynn Hartman, Charles Barth and Kory Schrom

Curr. Issues Mol. Biol. 2021, 43(3), 2135-2146; https://doi.org/10.3390/cimb43030148 - 29 Nov 2021

Cited by 7 | Viewed by 7454

Abstract

Gastrointestinal microbiome dysbiosis may result in harmful effects on the host, including those caused by inflammatory bowel diseases (IBD). The novel probiotic BIOHM, consisting of Bifidobacterium breve, Saccharomyces boulardii, Lactobacillus acidophilus, L. rhamnosus, and amylase, was developed to rebalance the [...] Read more.

Gastrointestinal microbiome dysbiosis may result in harmful effects on the host, including those caused by inflammatory bowel diseases (IBD). The novel probiotic BIOHM, consisting of Bifidobacterium breve, Saccharomyces boulardii, Lactobacillus acidophilus, L. rhamnosus, and amylase, was developed to rebalance the bacterial–fungal gut microbiome, with the goal of reducing inflammation and maintaining a healthy gut population. To test the effect of BIOHM on human subjects, we enrolled a cohort of 49 volunteers in collaboration with the Fermentation Festival group (Santa Barbara, CA, USA). The profiles of gut bacterial and fungal communities were assessed via stool samples collected at baseline and following 4 weeks of once-a-day BIOHM consumption. Mycobiome analysis following probiotic consumption revealed an increase in Ascomycota levels in enrolled individuals and a reduction in Zygomycota levels (p value < 0.01). No statistically significant difference in Basidiomycota was detected between pre- and post-BIOHM samples and control abundance profiles (p > 0.05). BIOHM consumption led to a significant reduction in the abundance of Candida genus in tested subjects (p value < 0.013), while the abundance of C. albicans also trended lower than before BIOHM use, albeit not reaching statistical significance. A reduction in the abundance of Firmicutes at the phylum level was observed following BIOHM use, which approached levels reported for control individuals reported in the Human Microbiome Project data. The preliminary results from this clinical study suggest that BIOHM is capable of significantly rebalancing the bacteriome and mycobiome in the gut of healthy individuals, suggesting that further trials examining the utility of the BIOHM probiotic in individuals with gastrointestinal symptoms, where dysbiosis is considered a source driving pathogenesis, are warranted. Full article

(This article belongs to the Section Molecular Microbiology)

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11 pages, 2942 KiB

Open AccessArticle

Bone Morphogenetic Protein 4 (BMP4) Enhances the Differentiation of Human Induced Pluripotent Stem Cells into Limbal Progenitor Cells

by Hyun Soo Lee, Jeewon Mok and Choun-Ki Joo

Curr. Issues Mol. Biol. 2021, 43(3), 2124-2134; https://doi.org/10.3390/cimb43030147 - 29 Nov 2021

Cited by 4 | Viewed by 3032

Abstract

Corneal epithelium maintains visual acuity and is regenerated by the proliferation and differentiation of limbal progenitor cells. Transplantation of human limbal progenitor cells could restore the integrity and functionality of the corneal surface in patients with limbal stem cell deficiency. However, multiple protocols [...] Read more.

Corneal epithelium maintains visual acuity and is regenerated by the proliferation and differentiation of limbal progenitor cells. Transplantation of human limbal progenitor cells could restore the integrity and functionality of the corneal surface in patients with limbal stem cell deficiency. However, multiple protocols are employed to differentiate human induced pluripotent stem (iPS) cells into corneal epithelium or limbal progenitor cells. The aim of this study was to optimize a protocol that uses bone morphogenetic protein 4 (BMP4) and limbal cell-specific medium. Human dermal fibroblast-derived iPS cells were differentiated into limbal progenitor cells using limbal cell-specific (PI) medium and varying doses (1, 10, and 50 ng/mL) and durations (1, 3, and 10 days) of BMP4 treatment. Differentiated human iPS cells were analyzed by real-time polymerase chain reaction (RT-PCR), Western blotting, and immunocytochemical studies at 2 or 4 weeks after BMP4 treatment. Culturing human dermal fibroblast-derived iPS cells in limbal cell-specific medium and BMP4 gave rise to limbal progenitor and corneal epithelial-like cells. The optimal protocol of 10 ng/mL and three days of BMP4 treatment elicited significantly higher limbal progenitor marker (ABCG2, ∆Np63α) expression and less corneal epithelial cell marker (CK3, CK12) expression than the other combinations of BMP4 dose and duration. In conclusion, this study identified a successful reprogramming strategy to induce limbal progenitor cells from human iPS cells using limbal cell-specific medium and BMP4. Additionally, our experiments indicate that the optimal BMP4 dose and duration favor limbal progenitor cell differentiation over corneal epithelial cells and maintain the phenotype of limbal stem cells. These findings contribute to the development of therapies for limbal stem cell deficiency disorders. Full article

(This article belongs to the Special Issue Multipotent and Multisystem Biomolecules—Associated Interlinking Networks and Crossing Organ Differentiation)

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13 pages, 2599 KiB

Open AccessArticle

Matrix Metalloproteinases in Human Decidualized Endometrial Stromal Cells

by Yoji Hisamatsu, Hiromi Murata, Hiroaki Tsubokura, Yoshiko Hashimoto, Masaaki Kitada, Susumu Tanaka and Hidetaka Okada

Curr. Issues Mol. Biol. 2021, 43(3), 2111-2123; https://doi.org/10.3390/cimb43030146 - 26 Nov 2021

Cited by 10 | Viewed by 3207

Abstract

Cyclic changes, such as growth, decidualization, shedding, and regeneration, in the human endometrium are regulated by the reciprocal action of female hormones, such as estradiol (E₂), and progesterone (P₄). Matrix metalloproteases (MMPs) and tissue inhibitors of MMPs (TIMPs) control [...] Read more.

Cyclic changes, such as growth, decidualization, shedding, and regeneration, in the human endometrium are regulated by the reciprocal action of female hormones, such as estradiol (E₂), and progesterone (P₄). Matrix metalloproteases (MMPs) and tissue inhibitors of MMPs (TIMPs) control the invasion of extravillous trophoblast cells after implantation. Several MMPs and TIMPs function in the decidua and endometrial stromal cells (ESCs). Here, we aimed to systematically investigate the changes in MMPs and TIMPs associated with ESC decidualization. We evaluated the expression of 23 MMPs, four TIMPs, and four anti-sense non-coding RNAs from MMP loci. Primary ESC cultures treated with E₂ + medroxyprogesterone acetate (MPA), a potent P₄ receptor agonist, showed significant down-regulation of MMP3, MMP10, MMP11, MMP12, MMP20, and MMP27 in decidualized ESCs, as assessed by quantitative reverse transcription PCR. Further, MMP15 and MMP19 were significantly upregulated in decidualized ESCs. siRNA-mediated silencing of Heart and Neural Crest Derivatives Expressed 2 (HAND2), a master transcriptional regulator in ESC decidualization, significantly increased MMP15 expression in untreated human ESCs. These results collectively indicate the importance of MMP15 and MMP19 in ESC decidualization and highlight the role of HAND2 in repressing MMP15 transcription, thereby regulating decidualization. Full article

(This article belongs to the Collection Feature Papers in Current Issues in Molecular Biology)

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13 pages, 3272 KiB

Open AccessArticle

Linking Pedigree Information to the Gene Expression Phenotype to Understand Differential Family Survival Mechanisms in Highly Fecund Fish: A Case Study in the Larviculture of Pacific Bluefin Tuna

by Motoshige Yasuike, Kazunori Kumon, Yosuke Tanaka, Kenji Saitoh and Takuma Sugaya

Curr. Issues Mol. Biol. 2021, 43(3), 2098-2110; https://doi.org/10.3390/cimb43030145 - 26 Nov 2021

Cited by 1 | Viewed by 2807

Abstract

Mass spawning in fish culture often brings about a marked variance in family size, which can cause a reduction in effective population sizes in seed production for stock enhancement. This study reports an example of combined pedigree information and gene expression phenotypes to [...] Read more.

Mass spawning in fish culture often brings about a marked variance in family size, which can cause a reduction in effective population sizes in seed production for stock enhancement. This study reports an example of combined pedigree information and gene expression phenotypes to understand differential family survival mechanisms in early stages of Pacific bluefin tuna, Thunnus orientalis, in a mass culture tank. Initially, parentage was determined using the partial mitochondrial DNA control region sequence and 11 microsatellite loci at 1, 10, 15, and 40 days post-hatch (DPH). A dramatic proportional change in the families was observed at around 15 DPH; therefore, transcriptome analysis was conducted for the 15 DPH larvae using a previously developed oligonucleotide microarray. This analysis successfully addressed the family-specific gene expression phenotypes with 5739 differentially expressed genes and highlighted the importance of expression levels of gastric-function-related genes at the developmental stage for subsequent survival. This strategy demonstrated herein can be broadly applicable to species of interest in aquaculture to comprehend the molecular mechanism of parental effects on offspring survival, which will contribute to the optimization of breeding technologies. Full article

(This article belongs to the Section Bioinformatics and Systems Biology)

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16 pages, 4835 KiB

Open AccessArticle

Effect of Metformin and Simvastatin in Inhibiting Proadipogenic Transcription Factors

by Jelena Jakab, Milorad Zjalić, Štefica Mikšić, Ivan Tušek, Vesna Ćosić, Nikola Volarić, Dario Nakić, Aleksandar Včev and Blaženka Miškić

Curr. Issues Mol. Biol. 2021, 43(3), 2082-2097; https://doi.org/10.3390/cimb43030144 - 25 Nov 2021

Cited by 4 | Viewed by 3099

Abstract

Obesity is a multifactorial chronic disease characterized by the excessive accumulation of fat in adipose tissue driven by hypertrophy and hyperplasia of adipocytes through adipogenesis. Adipogenesis plays a key role in the development of obesity and related metabolic disorders, which makes it potential [...] Read more.

Obesity is a multifactorial chronic disease characterized by the excessive accumulation of fat in adipose tissue driven by hypertrophy and hyperplasia of adipocytes through adipogenesis. Adipogenesis plays a key role in the development of obesity and related metabolic disorders, which makes it potential target for the therapeutic approach to obesity. An increasing number of studies confirm the pleiotropic action of the combined treatment with metformin and statins, suggesting their anti-hypertensive, anti-inflammatory, and anti-adipogenic effect. The aim of this study was to analyze the effect of different doses of metformin (MET) and simvastatin (SIM) on the expression of key transcription factors of adipogenesis. Mouse 3T3-L1 preadipocytes were induced to differentiation in adipogenic medium with sustained MET and SIM treatment to assess the effect on adipogenesis. Nine days after initiating adipogenesis, the cells were prepared for further experiments, including Oil Red O staining, RT-PCR, Western blotting, and immunocytochemistry. Treating the cells with the combination of MET and SIM slightly reduced the intensity of Oil Red O staining compared with the control group, and down-regulated mRNA and protein expression of PPARγ, C/EBPα, and SREBP-1C. In conclusion, the inhibitory effect of MET and SIM on adipocyte differentiation, as indicated by decreased lipid accumulation, appears to be mediated through the down-regulation of adipogenic transcription factors, peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer binding pro-tein α (C/EBPα), and sterol regulatory element-binding protein 1 (SREBP-1C). Full article

(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)

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14 pages, 1999 KiB

Open AccessArticle

Recombinant Destabilase from Hirudo medicinalis Is Able to Dissolve Human Blood Clots In Vitro

by Pavel Bobrovsky, Valentin Manuvera, Izolda Baskova, Svetlana Nemirova, Alexandr Medvedev and Vassili Lazarev

Curr. Issues Mol. Biol. 2021, 43(3), 2068-2081; https://doi.org/10.3390/cimb43030143 - 20 Nov 2021

Cited by 4 | Viewed by 13060

Abstract

Leeches are amazing animals that can be classified as conditionally poisonous animals since the salivary cocktail they produce is injected directly into the victim, and its components have strictly defined biological purposes, such as preventing blood clot formation. Thrombolytic drugs are mainly aimed [...] Read more.

Leeches are amazing animals that can be classified as conditionally poisonous animals since the salivary cocktail they produce is injected directly into the victim, and its components have strictly defined biological purposes, such as preventing blood clot formation. Thrombolytic drugs are mainly aimed at treating newly formed blood clots. Aged clots are stabilized by a large number of isopeptide bonds that prevent the action of thrombolytics. These bonds are destroyed by destabilase, an enzyme of the leech’s salivary glands. Here, we conducted a pilot study to evaluate the feasibility and effectiveness of the use of destabilase in relation to blood clots formed during real pathological processes. We evaluated the isopeptidase activity of destabilase during the formation of a stabilized fibrin clot. We showed that destabilase does not affect the internal and external coagulation cascades. We calculated the dose–response curve and tested the ability of destabilase to destroy isopeptide bonds in natural blood clots. The effect of aged and fresh clots dissolving ability after treatment with destabilase coincided with the morphological characteristics of clots during surgery. Thus, recombinant destabilase can be considered as a potential drug for the treatment of aged clots, which are difficult to treat with known thrombolytics. Full article

(This article belongs to the Topic Blood Physiology: Molecular Mechanisms of Vascular Wall Functioning)

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9 pages, 1839 KiB

Open AccessArticle

Advanced Glycation End Products Increase Salivary Gland Hypofunction in d-Galactose-Induced Aging Rats and Its Prevention by Physical Exercise

by Woo Kwon Jung, Su-Bin Park, Hyung Rae Kim, Hwa Young Ryu, Yong Hwan Kim and Junghyun Kim

Curr. Issues Mol. Biol. 2021, 43(3), 2059-2067; https://doi.org/10.3390/cimb43030142 - 19 Nov 2021

Cited by 8 | Viewed by 3062

Abstract

A declined salivary gland function is commonly observed in elderly people. Advanced glycation end products (AGEs) are believed to contribute to the pathogenesis of aging. Although physical exercise is shown to increase various organ functions in human and experimental models, it is not [...] Read more.

A declined salivary gland function is commonly observed in elderly people. Advanced glycation end products (AGEs) are believed to contribute to the pathogenesis of aging. Although physical exercise is shown to increase various organ functions in human and experimental models, it is not known whether it has a similar effect in the salivary glands. In the present study, we evaluated the AGEs burden in the salivary gland in the aging process and the protective effect of physical exercise on age-related salivary hypofunction. To accelerate the aging process, rats were peritoneally injected with D-galactose for 6 weeks. Young control rats and d-galactose-induced aging rats in the old group were not exercised. The rats in the physical exercise group ran on a treadmill (12 m/min, 60 min/day, 3 days/week for 6 weeks). The results showed that the salivary flow rate and total protein levels in the saliva of the d-galactose-induced aging rats were reduced compared to those of the young control rats. Circulating AGEs in serum and secreted AGEs in saliva increased with d-galactose-induced aging. AGEs also accumulated in the salivary glands of these aging rats. The salivary gland of aging rats showed increased reactive oxygen species (ROS) generation, loss of acinar cells, and apoptosis compared to young control mice. However, physical exercise suppressed all of these age-related salivary changes. Overall, physical exercise could provide a beneficial option for age-related salivary hypofunction. Full article

(This article belongs to the Special Issue Advances of Molecular Sciences in Dental Diseases Detection and Treatment)

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11 pages, 3447 KiB

Open AccessArticle

Comprehensive Draft Genome Analyses of Three Rockfishes (Scorpaeniformes, Sebastiscus) via Genome Survey Sequencing

by Chenghao Jia, Tianyan Yang, Takashi Yanagimoto and Tianxiang Gao

Curr. Issues Mol. Biol. 2021, 43(3), 2048-2058; https://doi.org/10.3390/cimb43030141 - 18 Nov 2021

Cited by 4 | Viewed by 2592

Abstract

Sebastiscus species, marine rockfishes, are of essential economic value. However, the genomic data of this genus is lacking and incomplete. Here, whole genome sequencing of all species of Sebastiscus was conducted to provide fundamental genomic information. The genome sizes were estimated to be [...] Read more.

Sebastiscus species, marine rockfishes, are of essential economic value. However, the genomic data of this genus is lacking and incomplete. Here, whole genome sequencing of all species of Sebastiscus was conducted to provide fundamental genomic information. The genome sizes were estimated to be 802.49 Mb (S. albofasciatus), 786.79 Mb (S. tertius), and 776.00 Mb (S. marmoratus) by using k-mer analyses. The draft genome sequences were initially assembled, and genome-wide microsatellite motifs were identified. The heterozygosity, repeat ratios, and numbers of microsatellite motifs all suggested possibly that S. tertius is more closely related to S. albofasciatus than S. marmoratus at the genetic level. Moreover, the complete mitochondrial genome sequences were assembled from the whole genome data and the phylogenetic analyses genetically supported the validation of Sebastiscus species. This study provides an important genome resource for further studies of Sebastiscus species. Full article

(This article belongs to the Section Bioinformatics and Systems Biology)

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12 pages, 3534 KiB

Open AccessArticle

A Comparative Study between Spanish and British SARS-CoV-2 Variants

by Jose A. Jimenez Ruiz, Cecilia Lopez Ramirez and Jose Luis Lopez-Campos

Curr. Issues Mol. Biol. 2021, 43(3), 2036-2047; https://doi.org/10.3390/cimb43030140 - 16 Nov 2021

Cited by 2 | Viewed by 2976

Abstract

The study of the interaction between the SARS-CoV-2 spike protein and the angiotensin-converting enzyme 2 (ACE2) receptor is key to understanding binding affinity and stability. In the present report, we sought to investigate the differences between two already sequenced genome variants (Spanish and [...] Read more.

The study of the interaction between the SARS-CoV-2 spike protein and the angiotensin-converting enzyme 2 (ACE2) receptor is key to understanding binding affinity and stability. In the present report, we sought to investigate the differences between two already sequenced genome variants (Spanish and British) of SARS-CoV-2. Methods: In silico model evaluating the homology, identity and similarity in the genome sequence and the structure and alignment of the predictive spike by computational docking methods. Results: The identity results between the Spanish and British variants of the Spike protein were 28.67%. This close correspondence in the results between the Spanish and British SARS-CoV-2 variants shows that they are very similar (99.99%). The alignment obtained results in four deletions. There were 23 nucleotide substitutions also predicted which could affect the functionality of the proteins produced from this sequence. The interaction between the binding receptor domain from the spike protein and the ACE2 receptor produces some of the mutations found and, therefore, the energy of this ligand varies. However, the estimated antigenicity of the British variant is higher than its Spanish counterpart. Conclusions: Our results indicate that minimal mutations could interfere in the infectivity of the virus due to changes in the fitness between host cell recognition and interaction proteins. In particular, the N501Y substitution, situated in the RBD of the spike of the British variant, might be the reason for its extraordinary infective potential. Full article

(This article belongs to the Section Bioinformatics and Systems Biology)

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14 pages, 2133 KiB

Open AccessArticle

Inflammation and Liver Cell Death in Patients with Hepatitis C Viral Infection

by Manuela G. Neuman and Lawrence B. Cohen

Curr. Issues Mol. Biol. 2021, 43(3), 2022-2035; https://doi.org/10.3390/cimb43030139 - 16 Nov 2021

Cited by 2 | Viewed by 3549

Abstract

Hepatitis C virus (HCV)-induced liver disease contributes to chronic hepatitis. The immune factors identified in HCV include changes in the innate and adaptive immune system. The inflammatory mediators, known as “inflammasome”, are a consequence of the metabolic products of cells and commensal or [...] Read more.

Hepatitis C virus (HCV)-induced liver disease contributes to chronic hepatitis. The immune factors identified in HCV include changes in the innate and adaptive immune system. The inflammatory mediators, known as “inflammasome”, are a consequence of the metabolic products of cells and commensal or pathogenic bacteria and viruses. The only effective strategy to prevent disease progression is eradication of the viral infection. Immune cells play a pivotal role during liver inflammation, triggering fibrogenesis. The present paper discusses the potential role of markers in cell death and the inflammatory cascade leading to the severity of liver damage. We aim to present the clinical parameters and laboratory data in a cohort of 88 HCV-infected non-cirrhotic and 25 HCV cirrhotic patients, to determine the characteristic light microscopic (LM) and transmission electron microscopic (TEM) changes in their liver biopsies and to present the link between the severity of liver damage and the serum levels of cytokines and caspases. A matched HCV non-infected cohort was used for the comparison of serum inflammatory markers. We compared the inflammation in HCV individuals with a control group of 280 healthy individuals. We correlated the changes in inflammatory markers in different stages of the disease and the histology. We concluded that the serum levels of cytokine, chemokine, and cleaved caspase markers reveal the inflammatory status in HCV. Based upon the information provided by the changes in biomarkers the clinician can monitor the severity of HCV-induced liver damage. New oral well-tolerated treatment regimens for chronic hepatitis C patients can achieve cure rates of over 90%. Therefore, using the noninvasive biomarkers to monitor the evolution of the liver damage is an effective personalized medicine procedure to establish the severity of liver injury and its repair. Full article

(This article belongs to the Topic Clinical, Translational and Basic Research on Liver Diseases)

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11 pages, 2194 KiB

Open AccessArticle

Farnesol Ameliorates Demyelinating Phenotype in a Cellular and Animal Model of Charcot-Marie-Tooth Disease Type 1A

by Na-Young Park, Geon Kwak, Hyun-Myung Doo, Hye-Jin Kim, So-Young Jang, Yun-Il Lee, Byung-Ok Choi and Young-Bin Hong

Curr. Issues Mol. Biol. 2021, 43(3), 2011-2021; https://doi.org/10.3390/cimb43030138 - 13 Nov 2021

Cited by 3 | Viewed by 3795

Abstract

Charcot-Marie-Tooth disease (CMT) is a genetically heterogeneous disease affecting the peripheral nervous system that is caused by either the demyelination of Schwann cells or degeneration of the peripheral axon. Currently, there are no treatment options to improve the degeneration of peripheral nerves in [...] Read more.

Charcot-Marie-Tooth disease (CMT) is a genetically heterogeneous disease affecting the peripheral nervous system that is caused by either the demyelination of Schwann cells or degeneration of the peripheral axon. Currently, there are no treatment options to improve the degeneration of peripheral nerves in CMT patients. In this research, we assessed the potency of farnesol for improving the demyelinating phenotype using an animal model of CMT type 1A. In vitro treatment with farnesol facilitated myelin gene expression and ameliorated the myelination defect caused by PMP22 overexpression, the major causative gene in CMT. In vivo administration of farnesol enhanced the peripheral neuropathic phenotype, as shown by rotarod performance in a mouse model of CMT1A. Electrophysiologically, farnesol-administered CMT1A mice exhibited increased motor nerve conduction velocity and compound muscle action potential compared with control mice. The number and diameter of myelinated axons were also increased by farnesol treatment. The expression level of myelin protein zero (MPZ) was increased, while that of the demyelination marker, neural cell adhesion molecule (NCAM), was reduced by farnesol administration. These data imply that farnesol is efficacious in ameliorating the demyelinating phenotype of CMT, and further elucidation of the underlying mechanisms of farnesol’s effect on myelination might provide a potent therapeutic strategy for the demyelinating type of CMT. Full article

(This article belongs to the Topic Neuroprotection by Drugs, Nutraceuticals and Physical Activity)

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14 pages, 1895 KiB

Open AccessArticle

Repetitive Treatment with Volatile Anesthetics Does Not Affect the In Vivo Plasma Concentration and Composition of Extracellular Vesicles in Rats

by Christian Bleilevens, Christian Beckers, Alexander Theissen, Tamara Fechter, Eva Miriam Buhl, Johannes Greven, Sandra Kraemer and Sebastian Wendt

Curr. Issues Mol. Biol. 2021, 43(3), 1997-2010; https://doi.org/10.3390/cimb43030137 - 13 Nov 2021

Cited by 5 | Viewed by 2904

Abstract

Background: Anesthetic-induced preconditioning (AIP) with volatile anesthetics is a well-known experimental technique to protect tissues from ischemic injury or oxidative stress. Additionally, plasmatic extracellular vesicle (EV) populations and their cargo are known to be affected by AIP in vitro, and to provide organ [...] Read more.

Background: Anesthetic-induced preconditioning (AIP) with volatile anesthetics is a well-known experimental technique to protect tissues from ischemic injury or oxidative stress. Additionally, plasmatic extracellular vesicle (EV) populations and their cargo are known to be affected by AIP in vitro, and to provide organ protective properties via their cargo. We investigated whether AIP would affect the generation of EVs in an in vivo rat model. Methods: Twenty male Sprague Dawley rats received a repetitive treatment with either isoflurane or with sevoflurane for a duration of 4 or 8 weeks. EVs from blood plasma were characterized by nanoparticle tracking analysis, transmission electron microscopy (TEM) and Western blot. A scratch assay (H9C2 cardiomyoblast cell line) was performed to investigate the protective capabilities of the isolated EVs. Results: TEM images as well as Western blot analysis indicated that EVs were successfully isolated. The AIP changed the flotillin and CD63 expression on the EV surface, but not the EV concentration. The scratch assay did not show increased cell migration and/or proliferation after EV treatment. Conclusion: AIP in rats changed the cargo of EVs but had no effect on EV concentration or cell migration/proliferation. Future studies are needed to investigate the cargo on a miRNA level and to investigate the properties of these EVs in additional functional experiments. Full article

(This article belongs to the Section Molecular Medicine)

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20 pages, 1700 KiB

Open AccessArticle

Quantitative Trait Locus Mapping of Seed Vigor in Soybean under −20 °C Storage and Accelerated Aging Conditions via RAD Sequencing

by Rongfan Wang, Fengqi Wu, Xianrong Xie and Cunyi Yang

Curr. Issues Mol. Biol. 2021, 43(3), 1977-1996; https://doi.org/10.3390/cimb43030136 - 12 Nov 2021

Cited by 6 | Viewed by 3048

Abstract

Due to its fast deterioration, soybean (Glycine max L.) has an inherently poor seed vigor. Vigor loss occurring during storage is one of the main obstacles to soybean production in the tropics. To analyze the genetic background of seed vigor, soybean seeds [...] Read more.

Due to its fast deterioration, soybean (Glycine max L.) has an inherently poor seed vigor. Vigor loss occurring during storage is one of the main obstacles to soybean production in the tropics. To analyze the genetic background of seed vigor, soybean seeds of a recombinant inbred line (RIL) population derived from the cross between Zhonghuang24 (ZH24, low vigor cultivar) and Huaxia3hao (HX3, vigorous cultivar) were utilized to identify the quantitative trait loci (QTLs) underlying the seed vigor under −20 °C conservation and accelerated aging conditions. According to the linkage analysis, multiple seed vigor-related QTLs were identified under both −20 °C and accelerated aging storage. Two major QTLs and eight QTL hotspots localized on chromosomes 3, 6, 9, 11, 15, 16, 17, and 19 were detected that were associated with seed vigor across two storage conditions. The indicators of seed vigor did not correlate well between the two aging treatments, and no common QTLs were detected in RIL populations stored in two conditions. These results indicated that deterioration under accelerated aging conditions was not reflective of natural aging at −20 °C. Additionally, we suggest 15 promising candidate genes that could possibly determine the seed vigor in soybeans, which would help explore the mechanisms responsible for maintaining high seed vigor. Full article

(This article belongs to the Section Biochemistry, Molecular and Cellular Biology)

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27 pages, 1591 KiB

Open AccessReview

A Critical Review: Recent Advancements in the Use of CRISPR/Cas9 Technology to Enhance Crops and Alleviate Global Food Crises

by Adnan Rasheed, Rafaqat Ali Gill, Muhammad Umair Hassan, Athar Mahmood, Sameer Qari, Qamar U. Zaman, Muhammad Ilyas, Muhammad Aamer, Maria Batool, Huijie Li and Ziming Wu

Curr. Issues Mol. Biol. 2021, 43(3), 1950-1976; https://doi.org/10.3390/cimb43030135 - 11 Nov 2021

Cited by 51 | Viewed by 10595

Abstract

Genome editing (GE) has revolutionized the biological sciences by creating a novel approach for manipulating the genomes of living organisms. Many tools have been developed in recent years to enable the editing of complex genomes. Therefore, a reliable and rapid approach for increasing [...] Read more.

Genome editing (GE) has revolutionized the biological sciences by creating a novel approach for manipulating the genomes of living organisms. Many tools have been developed in recent years to enable the editing of complex genomes. Therefore, a reliable and rapid approach for increasing yield and tolerance to various environmental stresses is necessary to sustain agricultural crop production for global food security. This critical review elaborates the GE tools used for crop improvement. These tools include mega-nucleases (MNs), such as zinc-finger nucleases (ZFNs), and transcriptional activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeats (CRISPR). Specifically, this review addresses the latest advancements in the role of CRISPR/Cas9 for genome manipulation for major crop improvement, including yield and quality development of biotic stress- and abiotic stress-tolerant crops. Implementation of this technique will lead to the production of non-transgene crops with preferred characteristics that can result in enhanced yield capacity under various environmental stresses. The CRISPR/Cas9 technique can be combined with current and potential breeding methods (e.g., speed breeding and omics-assisted breeding) to enhance agricultural productivity to ensure food security. We have also discussed the challenges and limitations of CRISPR/Cas9. This information will be useful to plant breeders and researchers in the thorough investigation of the use of CRISPR/Cas9 to boost crops by targeting the gene of interest. Full article

(This article belongs to the Section Molecular Plant Sciences)

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13 pages, 312 KiB

Open AccessArticle

Strategy to Develop and Evaluate a Multiplex RT-ddPCR in Response to SARS-CoV-2 Genomic Evolution

by Laura A. E. Van Poelvoorde, Mathieu Gand, Marie-Alice Fraiture, Sigrid C. J. De Keersmaecker, Bavo Verhaegen, Koenraad Van Hoorde, Ann Brigitte Cay, Nadège Balmelle, Philippe Herman and Nancy Roosens

Curr. Issues Mol. Biol. 2021, 43(3), 1937-1949; https://doi.org/10.3390/cimb43030134 - 6 Nov 2021

Cited by 8 | Viewed by 3608

Abstract

The worldwide emergence and spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) since 2019 has highlighted the importance of rapid and reliable diagnostic testing to prevent and control the viral transmission. However, inaccurate results may occur due to false negatives (FN) caused [...] Read more.

The worldwide emergence and spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) since 2019 has highlighted the importance of rapid and reliable diagnostic testing to prevent and control the viral transmission. However, inaccurate results may occur due to false negatives (FN) caused by polymorphisms or point mutations related to the virus evolution and compromise the accuracy of the diagnostic tests. Therefore, PCR-based SARS-CoV-2 diagnostics should be evaluated and evolve together with the rapidly increasing number of new variants appearing around the world. However, even by using a large collection of samples, laboratories are not able to test a representative collection of samples that deals with the same level of diversity that is continuously evolving worldwide. In the present study, we proposed a methodology based on an in silico and in vitro analysis. First, we used all information offered by available whole-genome sequencing data for SARS-CoV-2 for the selection of the two PCR assays targeting two different regions in the genome, and to monitor the possible impact of virus evolution on the specificity of the primers and probes of the PCR assays during and after the development of the assays. Besides this first essential in silico evaluation, a minimal set of testing was proposed to generate experimental evidence on the method performance, such as specificity, sensitivity and applicability. Therefore, a duplex reverse-transcription droplet digital PCR (RT-ddPCR) method was evaluated in silico by using 154 489 whole-genome sequences of SARS-CoV-2 strains that were representative for the circulating strains around the world. The RT-ddPCR platform was selected as it presented several advantages to detect and quantify SARS-CoV-2 RNA in clinical samples and wastewater. Next, the assays were successfully experimentally evaluated for their sensitivity and specificity. A preliminary evaluation of the applicability of the developed method was performed using both clinical and wastewater samples. Full article

(This article belongs to the Section Bioinformatics and Systems Biology)

31 pages, 14880 KiB

Open AccessArticle

Elucidating Drug-Like Compounds and Potential Mechanisms of Corn Silk (Stigma Maydis) against Obesity: A Network Pharmacology Study

by Ki-Kwang Oh, Md. Adnan and Dong-Ha Cho

Curr. Issues Mol. Biol. 2021, 43(3), 1906-1936; https://doi.org/10.3390/cimb43030133 - 6 Nov 2021

Cited by 2 | Viewed by 3476

Abstract

Corn silk (Stigma Maydis) has been utilized as an important herb against obesity by Chinese, Korean, and Native Americans, but its phytochemicals and mechanisms(s) against obesity have not been deciphered completely. This study aimed to identify promising bioactive constituents and mechanism [...] Read more.

Corn silk (Stigma Maydis) has been utilized as an important herb against obesity by Chinese, Korean, and Native Americans, but its phytochemicals and mechanisms(s) against obesity have not been deciphered completely. This study aimed to identify promising bioactive constituents and mechanism of action(s) of corn silk (CS) against obesity via network pharmacology. The compounds from CS were identified using Gas Chromatography Mass Spectrometry (GC-MS) and were confirmed ultimately by Lipinski’s rule via SwissADME. The relationships of the compound-targets or obesity-related targets were confirmed by public bioinformatics. The signaling pathways related to obesity, protein-protein interaction (PPI), and signaling pathways-targets-bioactives (STB) were constructed, visualized, and analyzed by RPackage. Lastly, Molecular Docking Test (MDT) was performed to validate affinity between ligand(s) and protein(s) on key signaling pathway(s). We identified a total of 36 compounds from CS via GC-MS, all accepted by Lipinski’s rule. The number of 36 compounds linked to 154 targets, 85 among 154 targets related directly to obesity-targets (3028 targets). Of the final 85 targets, we showed that the PPI network (79 edges, 357 edges), 12 signaling pathways on a bubble chart, and STB network (67 edges, 239 edges) are considered as therapeutic components. The MDT confirmed that two key activators (β-Amyrone, β-Stigmasterol) bound most stably to PPARA, PPARD, PPARG, FABP3, FABP4, and NR1H3 on the PPAR signaling pathway, also, three key inhibitors (Neotocopherol, Xanthosine, and β-Amyrone) bound most tightly to AKT1, IL6, FGF2, and PHLPP1 on the PI3K-Akt signaling pathway. Overall, we provided promising key signaling pathways, targets, and bioactives of CS against obesity, suggesting crucial pharmacological evidence for further clinical testing. Full article

(This article belongs to the Section Molecular Pharmacology)

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Curr. Issues Mol. Biol., Volume 43, Issue 3 (December 2021) – 78 articles

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