Biomolecules, Volume 13, Issue 11 (November 2023) – 116 articles

(1) Background: A large and diverse microbial population exists in the human intestinal tract, which supports gut homeostasis and the health of the host. Short-chain fatty acid (SCFA)-secreting microbes also generate several metabolites with favorable regulatory effects on various malignancies and immunological inflammations. The involvement of intestinal SCFAs in kidney diseases, such as various kidney malignancies and inflammations, has emerged as a fascinating area of study in recent years. However, the mechanisms of SCFAs and other metabolites produced by SCFA-producing bacteria against kidney cancer and inflammation have not yet been investigated. (2) Methods: We considered 177 different SCFA-producing microbial species and 114 metabolites from the gutMgene database. Further, we used different online-based database platforms to predict 1890 gene targets associated with metabolites. Moreover, DisGeNET, OMIM, and Genecard databases were used to consider 13,104 disease-related gene targets. We used a Venn diagram and various protein−protein interactions (PPIs), KEGG pathways, and GO analyses for the functional analysis of gene targets. Moreover, the subnetwork of protein−protein interactions (through string and cytoscape platforms) was used to select the top 20% of gene targets through degree centrality, betweenness centrality, and closeness centrality. To screen the possible candidate compounds, we performed an analysis of the ADMET (absorption, distribution, metabolism, excretion, and toxicity) properties of metabolites and then found the best binding affinity using molecular docking simulation. (3) Results: Finally, we found the key gene targets that interact with suitable compounds and function against kidney cancer and inflammation, such as MTOR (with glycocholic acid), PIK3CA (with 11-methoxycurvularin, glycocholic acid, and isoquercitrin), IL6 (with isoquercitrin), PTGS2 (with isoquercitrin), and IGF1R (with 2-amino-1-methyl-6-phenylimidazo[4,5-b] pyridine, isoquercitrin), showed a lower binding affinity. (4) Conclusions: This study provides evidence to support the positive effects of SCFA-producing microbial metabolites that function against kidney cancer and inflammation and makes integrative research proposals that may be used to guide future studies. Full article

mRNA-based therapeutics have been found to be a promising treatment strategy in immunotherapy, gene therapy, and cancer treatments. Effectiveness of mRNA therapeutics depends on the level and duration of a desired protein’s expression, which is determined by various cis- and trans-regulatory elements of the mRNA. Sequences of 5′ and 3′ untranslated regions (UTRs) are responsible for translational efficiency and stability of mRNA. An optimal combination of the regulatory sequences allows researchers to significantly increase the target protein’s expression. Using both literature data and previously obtained experimental data, we chose six sequences of 5′UTRs (adenoviral tripartite leader [TPL], HBB, rabbit β-globin [Rabb], H4C2, Moderna, and Neo2) and five sequences of 3′UTRs (mtRNR-EMCV, mtRNR-AES, mtRNR-mtRNR, BioNTech, and Moderna). By combining them, we constructed 30 in vitro transcribed RNAs encoding firefly luciferase with various combinations of 5′- and 3′UTRs, and the resultant bioluminescence was assessed in the DC2.4 cell line at 4, 8, 24, and 72 h after transfection. The cellular data enabled us to identify the best seven combinations of 5′- and 3′UTRs, whose translational efficiency was then assessed in BALB/c mice. Two combinations of 5′- and 3′UTRs (5′Rabb-3′mtRNR-EMCV and 5′TPL-3′Biontech) led to the most pronounced increase in the luciferase amount in the in vivo experiment in mice. Subsequent analysis of the stability of the mRNA indicated that the increase in luciferase expression is explained primarily by the efficiency of translation, not by the number of RNA molecules. Altogether, these findings suggest that 5′UTR-and-3′UTR combinations 5′Rabb-3′mtRNR- EMCV and 5′TPL-3′Biontech lead to high expression of target proteins and may be considered for use in preventive and therapeutic modalities based on mRNA. Full article

The use of template molecules as chemical scaffolds that significantly influence the course of the reaction has recently been intensively studied. Peptide nucleic acids (PNA) are molecules that mimic natural nucleic acids. They are a promising matrix in such reactions because they possess high affinity and specificity in their interactions. The manner of PNA interaction is predictable based on sequence complementarity. Recently, we report the visible light-induced metathesis reaction in peptides containing a diselenide bond. Herein, we present an efficient and straightforward method of the visible light-driven diselenide-based metathesis of peptide–nucleic acid conjugates. Compared to a similar photochemical transformation in peptides, a significant increase in the metathesis efficiency was obtained due to the template effect. Full article

Petroselinic fatty acid (PeFA) is considered a rare fatty acid and one of the most important fatty acids in the Apiaceae family. Its content varies depending on plant species, geographical origin, extraction method, ripeness, etc. Indeed, reported levels of petroselinic fatty acid range from 10.4 to 75.6% (in anise seed oil), 1 to 81.9% (in coriander seed oil), 28.5 to 57.6% (in caraway seed oil), 49.4 to 75.6% (in celery seed oil), 41.3 to 61.8% (in caraway seed oil), 79.9 to 87.2% (in dill seed oil), 43.1 to 81.9% (in fennel seed oil), and 35 to 75.1% (parsley seed oil). In this review, we also show current knowledge about genes encoding biosynthesis, from the desaturation of 16:0-ACP to petroselinic acid stored in triacylglycerol in the seeds. Furthermore, petroselinic acid is not related to the synthesis of ABA. PeFA was successfully isolated from Apiaceae family plant seeds in order to study their reactivity and biological activities. Several investigations showed that this fatty acid has a wide range of biological potentials, including antidiabetic, antibacterial, and antifungal activities. In cosmetics, PeFA alone or in association with other active compounds has interesting applications as an anti-inflammatory agent for the treatment of skin, hair, and nail disorders. Full article

The gut is the body’s largest immune organ, and the intestinal barrier prevents harmful substances such as bacteria and toxins from passing through the gastrointestinal mucosa. Intestinal barrier dysfunction is closely associated with various diseases. However, there are currently no FDA-approved therapies targeting the intestinal epithelial barriers. Long noncoding RNAs (lncRNAs), a class of RNA transcripts with a length of more than 200 nucleotides and no coding capacity, are essential for the development and regulation of a variety of biological processes and diseases. lncRNAs are involved in the intestinal barrier function and homeostasis maintenance. This article reviews the emerging role of lncRNAs in the intestinal barrier and highlights the potential applications of lncRNAs in the treatment of various intestinal diseases by reviewing the literature on cells, animal models, and clinical patients. The aim is to explore potential lncRNAs involved in the intestinal barrier and provide new ideas for the diagnosis and treatment of intestinal barrier damage-associated diseases in the clinical setting. Full article

Many natural products have been acquired from plants for their helpful properties. Medicinal plants are used for treating a variety of pathologies or symptoms. The axes of many pathological processes are inflammation, oxidative stress, and senescence. This work is focused on identifying Mexican medicinal plants with potential anti-oxidant, anti-inflammatory, anti-aging, and anti-senescence effects through network analysis and chemoinformatic screening of their phytochemicals. We used computational methods to analyze drug-like phytochemicals in Mexican medicinal plants, multi-target compounds, and signaling pathways related to anti-oxidant, anti-inflammatory, anti-aging, and anti-senescence mechanisms. A total of 1373 phytochemicals are found in 1025 Mexican medicinal plants, and 148 compounds showed no harmful functionalities. These compounds displayed comparable structures with reference molecules. Based on their capacity to interact with pharmacological targets, three clusters of Mexican medicinal plants have been established. Curatella americana, Ximenia americana, Malvastrum coromandelianum, and Manilkara zapota all have anti-oxidant, anti-inflammatory, anti-aging, and anti-senescence effects. Plumeria rubra, Lonchocarpus yucatanensis, and Salvia polystachya contained phytochemicals with anti-oxidant, anti-inflammatory, anti-aging, and anti-senescence reported activity. Lonchocarpus guatemalensis, Vallesia glabra, Erythrina oaxacana, and Erythrina sousae have drug-like phytochemicals with potential anti-oxidant, anti-inflammatory, anti-aging, and anti-senescence effects. Between the drug-like phytochemicals, lonchocarpin, vallesine, and erysotrine exhibit potential anti-oxidant, anti-inflammatory, anti-aging, and anti-senescence effects. For the first time, we conducted an initial virtual screening of selected Mexican medicinal plants, which was subsequently confirmed in vivo, evaluating the anti-inflammatory activity of Lonchocarpus guatemalensis Benth in mice. Full article

The interaction of cold atmospheric plasma (CAP) with biotargets is accompanied by chemical reactions on their surfaces and insides, and it has great potential as an anticancer approach. This study discovers the molecular mechanisms that may explain the selective death of tumor cells under CAP exposure. To reach this goal, the transcriptional response to CAP treatment was analyzed in A549 lung adenocarcinoma cells and in lung-fibroblast Wi-38 cells. We found that the CAP treatment induced the common trend of response from A549 and Wi-38 cells—the p53 pathway, KRAS signaling, UV response, TNF-alpha signaling, and apoptosis-related processes were up-regulated in both cell lines. However, the amplitude of the response to CAP was more variable in the A549 cells. The CAP-dependent death of A549 cells was accompanied by DNA damage, cell-cycle arrest in G2/M, and the dysfunctional response of glutathione peroxidase 4 (GPx4). The activation of the genes of endoplasmic reticulum stress and ER lumens was detected only in the A549 cells. Transmission-electron microscopy confirmed the alteration of the morphology of the ER lumens in the A549 cells after the CAP exposure. It can be concluded that the responses to nuclear stress and ER stress constitute the main differences in the sensitivity of tumor and healthy cells to CAP exposure. Full article

Atypical hemolytic uremic syndrome (HUS) develops as a result of damage to the endothelium of microvasculature vessels by Shiga toxin produced by enterohemorrhagic Escherichia coli (STEC-HUS). STEC-HUS remains the leading cause of acute kidney injury (AKI) in children aged 6 months to 5 years. The pathomorphological essence of the disease is the development of thrombotic microangiopathy (TMA). One of the key causes of TMA is an imbalance in the ADAMTS13–von Willebrand factor (vWF)–platelet system. The goal of the work was to clarify the role of a moderate decrease in ADAMTS13 activity in the pathogenesis of STEC-HUS. The activity of ADAMTS13 was determined in 138 children (4 months–14.7 years) in the acute period of STEC-HUS and the features of the course of the disease in these patients were analyzed. The study revealed a decrease in the activity and concentration of ADAMTS13 in 79.8% and 90.6% of patients, respectively. Measurements of von Willebrand factor antigen content and the activity of von Willebrand factor in the blood plasma of part of these patients were carried out. In 48.6% and 34.4% of cases, there was an increase in the antigen concentration and the activity of the Willebrand factor, respectively. Thrombocytopenia was diagnosed in 97.8% of children. We have demonstrated that moderately reduced ADAMTS13 activity correlates with the risk of severe manifestations of STEC-HUS in children; the rate of developing multiple organ failure, cerebral disorders, pulmonary edema, and acute kidney injury with the need for dialysis increases. It is assumed that reduction in ADAMTS13 activity may serve as a predictor of disease severity. Full article

Fatty acid transport protein 1 (FATP1) is an integral transmembrane protein that is involved in facilitating the translocation of long-chain fatty acids (LCFA) across the plasma membrane, thereby orchestrating the importation of LCFA into the cell. FATP1 also functions as an acyl-CoA ligase, catalyzing the ATP-dependent formation of fatty acyl-CoA using LCFA and VLCFA (very-long-chain fatty acids) as substrates. It is expressed in various types of tissues and is involved in the regulation of crucial signalling pathways, thus playing a vital role in numerous physiological and pathological conditions. Structural insight about FATP1 is, thus, extremely important for understanding the mechanism of action of this protein and developing efficient treatments against its anomalous expression and dysregulation, which are often associated with pathological conditions such as breast cancer. As of now, there has been no prior prediction or evaluation of the 3D configuration of the human FATP1 protein, hindering a comprehensive understanding of the distinct functional roles of its individual domains. In our pursuit to unravel the structure of the most commonly expressed isoforms of FATP1, we employed the cutting-edge ALPHAFOLD 2 model for an initial prediction of the entire protein’s structure. This prediction was complemented by molecular dynamics simulations, focusing on the most promising model. We predicted the structure of FATP1 in silico and thoroughly refined and validated it using coarse and molecular dynamics in the absence of the complete crystal structure. Their relative dynamics revealed the different properties of the characteristic FATP1. Full article

Azacarbazoles have attracted significant interest due to their valuable properties, such as anti-pathogenic and antitumor activity. In this study, a series of structurally related tricyclic benzo[4,5]- and tertacyclic naphtho[2′,1′:4,5]imidazo[1,2-c]pyrimidinone derivatives with one or two positively charged tethers were synthesized and evaluated for anti-proliferative activity. Lead tetracyclic derivative 5b with two amino-bearing arms inhibited the metabolic activity of A549 lung adenocarcinoma cells with a CC₅₀ value of 3.6 μM, with remarkable selectivity (SI = 17.3) over VA13 immortalized fibroblasts. Cell-cycle assays revealed that 5b triggers G2/M arrest without signs of apoptosis. A study of its interaction with various DNA G4s and duplexes followed by dual luciferase and intercalator displacement assays suggests that intercalation, rather than the modulation of G4-regulated oncogene expression, might contribute to the observed activity. Finally, a water-soluble salt of 5b was shown to cause no acute toxic effects, changes in mice behavior, or any decrease in body weight after a 72 h treatment at concentrations up to 20 mg/kg. Thus, 5b is a promising candidate for studies in vivo; however, further investigations are needed to elucidate its molecular target(s). Full article

High-density planting can increase crop productivity per unit area of cultivated land. However, the application of this technology is limited by the inhibition of plant growth in the presence of neighbors, which is not only due to their competition for resources but is also caused by growth regulators. Specifically, the abscisic acid (ABA) accumulated in plants under increased density of planting has been shown to inhibit their growth. The goal of the present study was to test the hypothesis that bacteria capable of degrading ABA can reduce the growth inhibitory effect of competition among plants by reducing concentration of this hormone in plants and their environment. Lettuce plants were grown both individually and three per pot; the rhizosphere was inoculated with a strain of Pseudomonas plecoglossicida 2.4-D capable of degrading ABA. Plant growth was recorded in parallel with immunoassaying ABA concentration in the pots and plants. The presence of neighbors indeed inhibited the growth of non-inoculated lettuce plants. Bacterial inoculation positively affected the growth of grouped plants, reducing the negative effects of competition. The bacteria-induced increase in the mass of competing plants was greater than that in the single ones. ABA concentration was increased by the presence of neighbors both in soil and plant shoots associated with the inhibition of plant growth, but accumulation of this hormone as well as inhibition of the growth of grouped plants was prevented by bacteria. The results confirm the role of ABA in the response of plants to the presence of competitors as well as the possibility of reducing the negative effect of competition on plant productivity with the help of bacteria capable of degrading this hormone. Full article

Tardive dyskinesia (TD) is a hyperkinetic movement disorder that displays unusual involuntary movement along with orofacial dysfunction. It is predominantly associated with the long-term use of antipsychotic medications, particularly typical or first-generation antipsychotic drugs such as haloperidol. Oxidative stress, mitochondrial dysfunction, neuroinflammation, and apoptosis are major pathophysiological mechanisms of TD. The BCG vaccine has been reported to suppress inflammation, oxidative stress, and apoptosis and exert neuroprotection via several mechanisms. Our study aimed to confirm the neuroprotective effect of the BCG vaccine against haloperidol-induced TD-like symptoms in rats. The rats were given haloperidol (1 mg/kg, i.p.) for 21 days after 1 h single administration of the BCG vaccine (2 × 10⁷ cfu). Various behavioral parameters for orofacial dyskinesia and locomotor activity were assessed on the 14th and 21st days after haloperidol injection. On the 22nd day, all rats were euthanized, and the striatum was isolated to estimate the biochemical, apoptotic, inflammatory, and neurotransmitter levels. The administration of the BCG vaccine reversed orofacial dyskinesia and improved motor function in regard to haloperidol-induced TD-like symptoms in rats. The BCG vaccine also enhanced the levels of antioxidant enzymes (SOD, GSH) and reduced prooxidants (MDA, nitrite) and pro-apoptotic markers (Cas-3, Cas-6, Cas-9) in rat brains. Besides this, BCG treatment also restored the neurotransmitter (DA, NE, 5-HT) levels and decreased the levels of HVA in the striatum. The study findings suggest that the BCG vaccine has antioxidant, antiapoptotic, and neuromodulatory properties that could be relevant in the management of TD. Full article

Vitamin D has been known to exert a wide range of physiological effects, including calcemic, osteogenic, anticancer, and immune responses. We previously generated genetically modified (GM) rats and performed a comparative analysis of their physiological properties to elucidate the roles of vitamin D and vitamin D receptor (VDR). In this study, our primary goal was to investigate the manifestations of type II rickets in rats with the VDR(H301Q) mutation, analogous to the human VDR(H305Q). Additionally, we created a double-mutant rat with the VDR(R270L/H301Q) mutation, resulting in almost no affinity for 1,25-dihydroxy-vitamin D3 (1,25D3) or 25-hydroxy-vitamin D3 (25D3). Notably, the plasma calcium concentration in Vdr(R270L/H301Q) rats was significantly lower than in wild-type (WT) rats. Meanwhile, Vdr(H301Q) rats had calcium concentrations falling between those of Vdr(R270L/H301Q) and WT rats. GM rats exhibited markedly elevated plasma parathyroid hormone and 1,25D3 levels compared to those of WT rats. An analysis of bone mineral density in the cortical bone of the femur in both GM rats revealed significantly lower values than in WT rats. Conversely, the bone mineral density in the trabecular bone was notably higher, indicating abnormal bone formation. This abnormal bone formation was more pronounced in Vdr(R270L/H301Q) rats than in Vdr(H301Q) rats, highlighting the critical role of the VDR-dependent function of 1,25D3 in bone formation. In contrast, neither Vdr(H301Q) nor Vdr(R270L/H301Q) rats exhibited symptoms of alopecia or cyst formation in the skin, which were observed in the Vdr-KO rats. These findings strongly suggest that unliganded VDR is crucial for maintaining the hair cycle and normal skin. Our GM rats hold significant promise for comprehensive analyses of vitamin D and VDR functions in future research. Full article

Local cell therapy has recently gained attention for the treatment of joint diseases and fractures. Mesenchymal stem cells (MSCs) are not only involved in osteogenesis and angiogenesis, but they also have immunomodulatory functions, such as inducing macrophage migration during bone regeneration via macrophage crosstalk. C-C motif chemokine ligand 2 (CCL2), a known inflammatory mediator, is associated with the migration of macrophages during inflammation. This study examined the utility of CCL2 as a therapeutic target for local cell therapy. Using lentiviral vectors for rabbit MSCs, genetically modified CCL2 overexpressing MSCs were generated. Osteogenic differentiation assays were performed using MSCs with or without macrophages in co-culture, and cell migration assays were also performed. Additionally, co-cultures were performed with endothelial cells (ECs), and angiogenesis was evaluated using a tube formation assay. Overexpression of CCL2 did not affect bone formation under monoculture conditions but promoted chemotaxis and osteogenesis when co-cultured with macrophages. Furthermore, CCL2-overexpression promoted tube formation in co-culture with ECs. These results suggest that CCL2 induces macrophage chemotaxis and osteogenesis by promoting crosstalk between MSCs and macrophages; CCL2 also stimulates ECs to induce angiogenesis. These findings indicate that CCL2 may be a useful therapeutic target for local cell therapy in areas of bone loss. Full article

Pyroptosis is a form of programmed cell death mediated by gasdermins, particularly gasdermin D (GSDMD), which is widely expressed in tissues throughout the body. GSDMD belongs to the gasdermin family, which is expressed in a variety of cell types including epithelial cells and immune cells. It is involved in the regulation of anti-inflammatory responses, leading to its differential expression in a wide range of diseases. In this review, we provide an overview of the current understanding of the major activation mechanisms and effector pathways of GSDMD. Subsequently, we examine the importance and role of GSDMD in different diseases, highlighting its potential as a pan-biomarker. We specifically focus on the biological characteristics of GSDMD in several diseases and its promising role in diagnosis, early detection, and differential diagnosis. Furthermore, we discuss the application of GSDMD in predicting prognosis and monitoring treatment efficacy in cancer. This review proposes a new strategy to guide therapeutic decision-making and suggests potential directions for further research into GSDMD. Full article

Background: Despite patients undergoing chronic hemodialysis (HD) being notoriously prone to adverse cardiovascular (CV) events, risk prediction in this population remains challenging. miRNA 122-5p, a short, non-coding RNA predominantly involved in lipid and carbohydrate metabolism, has recently been related to the onset and progression of CV disease. Methods: We run a pilot, multicenter, longitudinal, observational study to evaluate the clinical significance and prognostic usefulness of circulating miRNA 122-5p in a multicentric cohort of 74 individuals on maintenance HD. Results: Patients displayed lower circulating miRNA 122-5p as compared to healthy controls (p = 0.004). At correlation analyses, ALT (β = 0.333; p = 0.02), E/e’ (β = 0.265; p = 0.02) and CRP (β = −0.219; p = 0.041) were independent predictors of miRNA 122-5p levels. During a median follow-up of 22 months (range of 1–24), 30 subjects (40.5%) experienced a composite endpoint of all-cause mortality and fatal/non-fatal CV events. Baseline circulating miRNA 122-5p was higher in these subjects (p = 0.01) and it predicted a significantly higher risk of endpoint occurrence (Kaplan–Meier crude HR 3.192; 95% CI 1.529–6.663; p = 0.002; Cox regression adjusted HR 1.115; 95% CI 1.009–1.232; p = 0.03). Conclusions: Altered miRNA 122-5p levels in HD patients may reflect hepatic and CV damage and may impart important prognostic information for improving CV risk prediction in this particular setting. Full article

(1) Background: The LEO1 (Left open reading frame 1) protein is a conserved subunit of the PAF1C complex (RNA polymerase II-associated factor 1 complex). PAF1C has well-established mechanistic functions in elongation of transcription and RNA processing. We previously showed, in fission yeast, that LEO1 controls histone H3K9 methylation levels by affecting the turnover of histone H3 in chromatin, and that it is essential for the proper regulation of gene expression during cellular quiescence. Human fibroblasts enter a reversible quiescence state upon serum deprivation in the growth media. Here we investigate the function of LEO1 in human fibroblasts. (2) Methods: We knocked out the LEO1 gene using CRISPR/Cas9 methodology in human fibroblasts and verified that the LEO1 protein was undetectable by Western blot. We characterized the phenotype of the ΔLEO1 knockout cells with FACS analysis and cell growth assays. We used RNA-sequencing using spike-in controls to measure gene expression and spike-in controlled ChIP-sequencing experiments to measure the histone modification H3K9me2 genome-wide. (3) Results: Gene expression levels are altered in quiescent cells, however factors controlling chromatin and gene expression changes in quiescent human cells are largely unknown. The ΔLEO1 knockout fibroblasts are viable but have reduced metabolic activity compared to wild-type cells. ΔLEO1 cells showed a slower entry into quiescence and a different morphology compared to wild-type cells. Gene expression was generally reduced in quiescent wild-type cells. The downregulated genes included genes involved in cell proliferation. A small number of genes were upregulated in quiescent wild-type cells including several genes involved in ERK1/ERK2 and Wnt signaling. In quiescent ΔLEO1 cells, many genes were mis-regulated compared to wild-type cells. This included genes involved in Calcium ion transport and cell morphogenesis. Finally, spike-in controlled ChIP-sequencing experiments demonstrated that the histone modification H3K9me2 levels are globally increased in quiescent ΔLEO1 cells. (4) Conclusions: Thus, LEO1 is important for proper entry into cellular quiescence, control of H3K9me2 levels, and gene expression in human fibroblasts. Full article

Currently, the symptomatic status and left ventricular ejection fraction (LVEF) play a crucial role in aortic stenosis (AS) assessment. However, the symptoms are often subjective, and LVEF is not a sensitive marker of left ventricle (LV) decompensation. Over the past years, the cardiac structure and function research on AS has increased due to advanced imaging modalities and potential therapies. New imaging parameters emerged as predictors of disease progression in AS. LV global longitudinal strain has proved useful for risk stratification in asymptomatic severe AS patients with preserved LVEF. The assessment of myocardial fibrosis by cardiac magnetic resonance is the most studied application and offers prognostic information on AS. Moreover, the usage of biomarkers in AS as objective measures of LV decompensation has recently gained more interest. The present review focuses on the transition from compensatory LV hypertrophy (H) to LV dysfunction and the biomarkers associated with myocardial wall stress, fibrosis, and myocyte death. Moreover, we discuss the potential impact of non-invasive imaging parameters for optimizing the timing of aortic valve replacement and provide insight into novel biomarkers for possible prognostic use in AS. However, data from randomized clinical trials are necessary to define their utility in daily practice. Full article

Ribosomal frameshifting (RFS) at the slippery site of SARS-CoV-2 RNA is essential for the biosynthesis of the viral replication machinery. It requires the formation of a pseudoknot (PK) structure near the slippery site and can be inhibited by PK-disrupting oligonucleotide-based antivirals. We obtained and compared three types of such antiviral candidates, namely locked nucleic acids (LNA), LNA–DNA gapmers, and G-clamp-containing phosphorothioates (CPSs) complementary to PK stems. Using optical and electrophoretic methods, we showed that stem 2-targeting oligonucleotide analogs induced PK unfolding at nanomolar concentrations, and this effect was particularly pronounced in the case of LNA. For the leading PK-unfolding LNA and CPS oligonucleotide analogs, we also demonstrated dose-dependent RSF inhibition in dual luciferase assays (DLAs). Finally, we showed that the leading oligonucleotide analogs reduced SARS-CoV-2 replication at subtoxic concentrations in the nanomolar range in two human cell lines. Our findings highlight the promise of PK targeting, illustrate the advantages and limitations of various types of DNA modifications and may promote the future development of oligonucleotide-based antivirals. Full article

Immature starfish oocytes isolated from the ovary are susceptible to polyspermy due to the structural organization of the vitelline layer covering the oocyte plasma membrane, as well as the distribution and biochemical properties of the actin cytoskeleton of the oocyte cortex. After the resumption of the meiotic cycle of the oocyte triggered by the hormone 1-methyladenine, the maturing oocyte reaches fertilizable conditions to be stimulated by only one sperm with a normal Ca²⁺ response and cortical reaction. This cytoplasmic ripening of the oocyte, resulting in normal fertilization and development, is due to the remodeling of the cortical actin cytoskeleton and germinal vesicle breakdown (GVBD). Since disulfide-reducing agents such as dithiothreitol (DTT) are known to induce the maturation and GVBD of oocytes in many species of starfish, we analyzed the pattern of the fertilization response displayed by Astropecten aranciacus oocytes pre-exposed to DTT with or without 1-MA stimulation. Short treatment of A. aranciacus immature oocytes with DTT reduced the rate of polyspermic fertilization and altered the sperm-induced Ca²⁺ response by changing the morphology of microvilli, cortical granules, and biochemical properties of the cortical F-actin. At variance with 1-MA, the DTT treatment of immature starfish oocytes for 70 min did not induce GVBD. On the other hand, the DTT treatment caused an alteration in microvilli morphology and a drastic depolymerization of the cortical F-actin, which impaired the sperm-induced Ca²⁺ response at fertilization and the subsequent embryonic development. Full article

L-DOPA is the mainstay of treatment for Parkinson’s disease (PD). However, over time this drug can produce dyskinesia. A useful acute PD model for screening novel compounds for anti-parkinsonian and L-DOPA-induced dyskinesia (LID) are dopamine-depleted dopamine-transporter KO (DDD) mice. Treatment with α-methyl-para-tyrosine rapidly depletes their brain stores of DA and renders them akinetic. During sensitization in the open field (OF), their locomotion declines as vertical activities increase and upon encountering a wall they stand on one leg or tail and engage in climbing behavior termed “three-paw dyskinesia”. We have hypothesized that L-DOPA induces a stereotypic activation of locomotion in DDD mice, where they are unable to alter the course of their locomotion, and upon encountering walls engage in “three-paw dyskinesia” as reflected in vertical counts or beam-breaks. The purpose of our studies was to identify a valid index of LID in DDD mice that met three criteria: (a) sensitization with repeated L-DOPA administration, (b) insensitivity to a change in the test context, and (c) stimulatory or inhibitory responses to dopamine D1 receptor agonists (5 mg/kg SKF81297; 5 and 10 mg/kg MLM55-38, a novel compound) and amantadine (45 mg/kg), respectively. Responses were compared between the OF and a circular maze (CM) that did not hinder locomotion. We found vertical counts and climbing were specific for testing in the OF, while oral stereotypies were sensitized to L-DOPA in both the OF and CM and responded to D1R agonists and amantadine. Hence, in DDD mice oral stereotypies should be used as an index of LID in screening compounds for PD. Full article

Honey bees are typical model organisms for the study of caste differentiation, and the juvenile hormone (JH) is a crucial link in the regulatory network of caste differentiation in honey bees. To investigate the mechanism of JH-mediated caste differentiation, we analyzed the effect of the JH response gene AmKr-h1 on this process. We observed that AmKr-h1 expression levels were significantly higher in queen larvae than in worker larvae at the 48 h, 84 h, and 120 h larval stages, and were regulated by JH. Inhibiting AmKr-h1 expression in honey bee larvae using RNAi could lead to the development of larvae toward workers. We also analyzed the transcriptome changes in honey bee larvae after AmKr-h1 RNAi and identified 191 differentially expressed genes (DEGs) and 682 differentially expressed alternative splicing events (DEASEs); of these, many were related to honey bee caste differentiation. Our results indicate that AmKr-h1 regulates caste differentiation in honey bees by acting as a JH-responsive gene. Full article

Despite advances in treatment options, such as corticosteroid administration and less invasive respiratory support, bronchopulmonary dysplasia (BPD) remains an important prognostic factor in preterm infants. We previously reported that furin regulates changes in lung smooth muscle cell phenotypes, suggesting that it plays a critical role in BPD pathogenesis. Therefore, in this study, we aimed to evaluate whether it regulates the alveolarization of immature lungs through activating alveolarization-driving proteins. We first examined furin expression levels, and its functions, using an established hyperoxia-induced BPD mouse model. Thereafter, we treated mice pups, as well as primary myofibroblast cell cultures, with furin inhibitors. Finally, we administered the hyperoxia-exposed mice pups with recombinant furin. Immunofluorescence revealed the co-expression of furin with alpha-smooth muscle actin. Hyperoxia exposure for 10 d decreased alveolar formation, as well as the expression of furin and its target, IGF-1R. Hexa-D-arginine administration also significantly inhibited alveolar formation. Another furin inhibitor, decanoyl-RVKR-chloromethylketone, accumulated pro-IGF-1R, and decreased IGF-1R phosphorylation in myofibroblast primary cultures. Finally, recombinant furin treatment significantly improved alveolarization in hyperoxia-exposed mice pups. Furin regulates alveolarization in immature lungs. Therefore, this study provides novel insights regarding the involvement of furin in BPD pathogenesis, and highlights a potential treatment target for ameliorating the impact of BPD. Full article

In eukaryotes, the Dph1•Dph2 dimer is a non-canonical radical SAM enzyme. Using iron-sulfur (FeS) clusters, it cleaves the cosubstrate S-adenosyl-methionine (SAM) to form a 3-amino-3-carboxy-propyl (ACP) radical for the synthesis of diphthamide. The latter decorates a histidine residue on elongation factor 2 (EF2) conserved from archaea to yeast and humans and is important for accurate mRNA translation and protein synthesis. Guided by evidence from archaeal orthologues, we searched for a putative SAM-binding pocket in Dph1•Dph2 from Saccharomyces cerevisiae. We predict an SAM-binding pocket near the FeS cluster domain that is conserved across eukaryotes in Dph1 but not Dph2. Site-directed DPH1 mutagenesis and functional characterization through assay diagnostics for the loss of diphthamide reveal that the SAM pocket is essential for synthesis of the décor on EF2 in vivo. Further evidence from structural modeling suggests particularly critical residues close to the methionine moiety of SAM. Presumably, they facilitate a geometry specific for SAM cleavage and ACP radical formation that distinguishes Dph1•Dph2 from classical radical SAM enzymes, which generate canonical 5′-deoxyadenosyl (dAdo) radicals. Full article

Published evidence over the past few decades suggests that general anesthetics could be neurotoxins especially when administered at the extremes of age. The reported pathology is not only at the morphological level when examined in very young and aged brains, given that, importantly, newly developing evidence suggests a variety of behavioral impairments. Since anesthesia is unavoidable in certain clinical settings, we should consider the development of new anesthetics. A promising and safe solution could be a new family of anesthetics referred to as neuroactive steroids. In this review, we summarize the currently available evidence regarding their anesthetic and analgesic properties. Full article

Cancer is a complex and multifaceted disease with a high global incidence and mortality rate. Although cancer therapy has evolved significantly over the years, numerous challenges persist on the path to effectively combating this multifaceted disease. Natural compounds derived from plants, fungi, or marine organisms have garnered considerable attention as potential therapeutic agents in the field of cancer research. Ellagic acid (EA), a natural polyphenolic compound found in various fruits and nuts, has emerged as a potential cancer prevention and treatment agent. This review summarizes the experimental evidence supporting the role of EA in targeting key hallmarks of cancer, including proliferation, angiogenesis, apoptosis evasion, immune evasion, inflammation, genomic instability, and more. We discuss the molecular mechanisms by which EA modulates signaling pathways and molecular targets involved in these cancer hallmarks, based on in vitro and in vivo studies. The multifaceted actions of EA make it a promising candidate for cancer prevention and therapy. Understanding its impact on cancer biology can pave the way for developing novel strategies to combat this complex disease. Full article

Many vascular diseases are linked to lipid metabolism disorders, which cause lipid accumulation and peroxidation in the vascular wall. These processes lead to degenerative changes in the vessel, such as phenotypic transformation of smooth muscle cells and dysfunction and apoptosis of endothelial cells. In intracranial aneurysms, the coexistence of lipid plaques is often observed, indicating localized lipid metabolism disorders. These disorders may impair the function of the vascular wall or result from it. We summarize the literature on the relationship between lipid metabolism disorders and intracranial aneurysms below. Full article

Histamine is a neuromodulator that affects gut motility and visceral sensitivity through intrinsic and extrinsic neural pathways, yet the mechanisms regulating histamine availability in these pathways remain poorly understood. Here, we show that enteric glia contribute to histamine clearance in the enteric nervous system (ENS) through their expression of the enzyme histamine N-methyltransferase (HNMT). Glial HNMT expression was initially assessed using immunolabeling and gene expression, and functionally tested using CRISPR-Cas9 to create a Cre-dependent conditional Hnmt ablation model targeting glia. Immunolabeling, calcium imaging, and visceromotor reflex recordings were used to assess the effects on ENS structure and visceral hypersensitivity. Immunolabeling and gene expression data show that enteric neurons and glia express HNMT. Deleting Hnmt in Sox10+ enteric glia increased glial histamine levels and altered visceromotor responses to colorectal distension in male mice, with no effect in females. Interestingly, deleting glial Hnmt protected males from histamine-driven visceral hypersensitivity. These data uncover a significant role for glial HNMT in histamine degradation in the gut, which impacts histamine-driven visceral hypersensitivity in a sex-dependent manner. Changes in the capacity of glia to clear histamines could play a role in the susceptibility to developing visceral pain in disorders of the gut–brain interaction. Full article

Recalcitrant rice blast disease is caused by Magnaporthe oryzae, which has a significant negative economic reverberation on crop productivity. In order to induce the disease onto the host, M. oryzae positively generates many types of small secreted proteins, here named as effectors, to manipulate the host cell for the purpose of stimulating pathogenic infection. In M. oryzae, by engaging with specific receptors on the cell surface, effectors activate signaling channels which control an array of cellular activities, such as proliferation, differentiation and apoptosis. The most recent research on effector identification, classification, function, secretion, and control mechanism has been compiled in this review. In addition, the article also discusses directions and challenges for future research into an effector in M. oryzae. Full article

Orchid seeds lack endosperms and depend on mycorrhizal fungi for germination and nutrition acquisition under natural conditions. Piriformospora indica is a mycorrhizal fungus that promotes seed germination and seedling development in epiphytic orchids, such as Dendrobium nobile. To understand the impact of P. indica on D. nobile seed germination, we examined endogenous hormone levels by using liquid chromatography–mass spectrometry. We performed transcriptomic analysis of D. nobile protocorm at two developmental stages under asymbiotic germination (AG) and symbiotic germination (SG) conditions. The result showed that the level of endogenous IAA in the SG protocorm treatments was significantly higher than that in the AG protocorm treatments. Meanwhile, GA₃ was only detected in the SG protocorm stages. IAA and GA synthesis and signaling genes were upregulated in the SG protocorm stages. Exogenous GA₃ application inhibited fungal colonization inside the protocorm, and a GA biosynthesis inhibitor (PAC) promoted fungal colonization. Furthermore, we found that PAC prevented fungal hyphae collapse and degeneration in the protocorm, and differentially expressed genes related to cell wall metabolism were identified between the SG and AG protocorm stages. Exogenous GA₃ upregulated SRC2 and LRX4 expression, leading to decreased fungal colonization. Meanwhile, GA inhibitors upregulated EXP6, EXB16, and EXP10-2 expression, leading to increased fungal colonization. Our findings suggest that GA regulates the expression of cell wall metabolism genes in D. nobile, thereby inhibiting the establishment of mycorrhizal symbiosis. Full article