Elevated mRNA Levels Research Articles

High-fat diet induced obesity promotes inflammation, oxidative stress, and hepatotoxicity in female FVB/N mice.

Although obesity and subsequent liver injury are increasingly prevalent in women, female mouse models have generally shown resistance to high-fat diet (HFD)-induced obesity. We evaluated control and HFD-fed male and female FVB/N mice, a strain well-suited to transgenic analyses, for phenotypic, histological, and molecular markers related to control of glucose, lipids, and inflammation in serum, liver, and perigonadal white adipose tissues. Unlike many mouse models, HFD-fed FVB/N females gained more perigonadal and mesenteric fat mass and overall body weight than their male counterparts, with increased hepatic expression of lipogenic PPARγ target genes (Cd36, Fsp27, and Fsp27β), oxidative stress genes and protein (Nqo1 and CYP2E1), inflammatory gene (Mip-2), and the pro-fibrotic gene Pai-1, along with increases in malondialdehyde and serum ALT levels. Further, inherent to females (independently of HFD), hepatic antioxidant heme oxygenase-1 (HMOX1, HO-1) protein levels were reduced compared to their male counterparts. In contrast, males may have been relatively protected from HFD-induced oxidative stress and liver injury by elevated mRNA and protein levels of hepatic antioxidants BHMT and Gpx2, increased fatty acid oxidation genes in liver and adipocytes (Pparδ), despite disorganized and inflamed adipocytes. Thus, female FVB/N mice offer a valuable preclinical, genetically malleable model that recapitulates many of the features of diet-induced obesity and liver damage observed in human females.

Application of CD25 and CTLA4 gene transcription levels in early prediction of acute graft-versus-host disease.

Our study investigated the potential of peripheral blood T cell CD25, CD28, and CTLA-4 gene transcription levels as predictive biomarkers for acute graft-versus-host disease (aGVHD) following allogeneic hematopoietic stem cell transplantation (allo-HSCT). Real-time reverse transcription fluorescent quantitative PCR (RT-qPCR) analysis was conducted on day +7, +14, and +21 post-transplantation in patients undergoing allo-HSCT. Elevated levels of CD25 and CTLA-4 mRNA were found to be associated with the occurrence of aGVHD, as well as severe and gastrointestinal aGVHD. Receiver operating characteristic (ROC) curve analysis was utilized to assess the predictive value of each biomarker. Combined analysis of CD25 and CTLA-4 mRNA levels demonstrated promising predictive potential for aGVHD. Our results confirmed that the transcription levels of CD25 and CTLA-4 genes could be used as early predictive biomarkers for aGVHD post-allo-HSCT.

Canagliflozin Alleviates Atherosclerosis Progression through Inflammation, Oxidative Stress, and Autophagy in Western Diet-fed ApoE−/− Mice

Purpose: This study was aimed at investigating the effect of canagliflozin (Cana) on atherosclerosis and further exploring its potential mechanism. Methods: ApoE−/− mice were fed a Western diet (WD) and randomly divided into a WD group and WD+Cana group. After 15 weeks of canagliflozin treatment, serum levels of fasting insulin and inflammatory cytokines were determined with ELISA kits. HE, Oil Red O, and Masson staining were used to estimate the extent of atherosclerosis. Immunohistochemistry, immunofluorescence, ROS staining, and RT-PCR were used to further investigate Cana’s potential mechanism. Results: Histological analysis indicated that Cana restrained atherosclerotic plaque development. Furthermore, Cana decreased the percentage of F4/80 positive cells, and the areal density of ROS and relative fluorescence intensity of P62, but enhanced the relative fluorescence intensity of LC3 in the aortic root. Analysis of factors associated with the inflammatory response mediated by AP-1, oxidative stress mediated through the ROS/Nrf2 pathway, and autophagy in the aorta indicated elevated mRNA levels of F4/80, MCP-1, VCAM-1, AP-1, ROS, NOX4, P62, NLRP3, and IL-1β, but diminished mRNA levels of Nrf2, GST, eNOS, and LC3, in the WD+Cana group. Conclusion: Canagliflozin may attenuate atherosclerosis by decreasing the inflammatory response mediated by AP-1, alleviating oxidative stress through the ROS/Nrf2 pathway, and enhancing autophagy in WD-fed ApoE−/− mice.

Analysis of the Expression and Prognostic Value of SIRTs in Hepatocellular Carcinoma.

This study contributes to the evolving understanding of the pivotal involvement of Sirtuins (SIRTs) in various human cancers, with a particular focus on elucidating their expression patterns and clinical relevance within the context of hepatocellular carcinoma (HCC). The investigation involves a comprehensive analysis of mRNA expression and prognostic implications associated with distinct SIRTs in HCC. Initial data pertaining to SIRT expression in HCC patients were collated from publicly accessible databases. Subsequently, the expression levels of select members of the SIRT family were validated using clinicopathological specimens from HCC patients. Additionally, HCC tissue microarray was employed to scrutinize the correlation between SIRT7 expression and HCC prognosis. The findings indicated a substantial upregulation of SIRT2, SIRT3, SIRT4, SIRT6, and SIRT7 in HCC tissues. Survival analysis underscored a pronounced association between elevated mRNA levels of SIRT3, SIRT6, and SIRT7 and an adverse prognosis for HCC patients. Particularly, SIRT7 emerged as a potential independent risk factor for poor prognosis in HCC patients. Examination of the HCC tissue microarray revealed heightened expression of SIRT7 in 68 cases (54.8%) of HCC tissues. Multivariate analysis established high SIRT7 expression as an independent risk factor for diminished Disease-Free Survival (DFS) and Overall Survival (OS) in HCC patients. The aberrant expression of SIRT7 presents itself may be as a novel biomarker for predicting the prognosis of HCC patients.

Lysine tRNA fragments and miR-194-5p co-regulate hepatic steatosis via β-Klotho and perilipin 2

ObjectiveNon-alcoholic fatty liver disease (NAFLD) involves hepatic accumulation of intracellular lipid droplets via incompletely understood processes. Here, we report distinct and cooperative NAFLD roles of LysTTT-5′tRF transfer RNA fragments and microRNA miR-194-5p. MethodsCombined use of diet induced obese mice with human-derived oleic acid-exposed Hep G2 cells revealed new NAFLD roles of LysTTT-5′tRF and miR-194-5p. ResultsUnlike lean animals, dietary-induced NAFLD mice showed concurrent hepatic decrease of both LysTTT-5′tRF and miR-194-5p levels, which were restored following miR-132 antisense oligonucleotide treatment which suppresses hepatic steatosis. Moreover, exposing human-derived Hep G2 cells to oleic acid for 7 days co-suppressed miR-194-5p and LysTTT-5′tRF levels while increasing lipid accumulation. Inversely, transfecting fattened cells with a synthetic LysTTT-5′tRF mimic elevated mRNA levels of the metabolic regulator β-Klotho while decreasing triglyceride amounts by 30% within 24 h. In contradistinction, antisense suppression of miR-194-5p induced accumulation of its novel target, the NAFLD-implicated lipid droplet-coating PLIN2 protein. Further, two out of 15 steatosis-alleviating screened drug-repurposing compounds, Danazol and Latanoprost, elevated miR-194-5p or LysTTT-5′tRF levels. ConclusionOur findings highlight the different yet complementary roles of miR-194-5p and LysTTT-5′tRF and offer new insights into the complex roles of small non-coding RNAs and the multiple pathways involved in NAFLD pathogenesis.

Evaluation of the osteogenic potential of demineralized and decellularized bovine bone granules following implantation in rat calvaria critical-size defect model.

The aim of this study was to compare the ability of demineralized (DMB) and decellularized (DCC) bovine bone granules to support bone regeneration in rat calvaria critical-size defects. DMB and DCC were prepared using a previously published method. The granule size used ranged between 500 and 750 μm. A total of forty-eight Sprague-Dawley rats were divided into two groups (n = 24). A pair of 5 mm diameter defects were created on the calvaria of the rats in the right and left parietal bone in both groups. Group A animals received DMB granules and Group B received DCC granules in the right parietal defect side while the left parietal untreated defect acted as sham surgery for both groups. Four animals per group were euthanized in a CO2 chamber at day 7, 14 and 21 post-surgery and the calvaria implantation site biopsy harvested was subjected to osteogenic gene expression analysis. Another four animals per group were euthanized at days 15, 30 and 60 post surgery and the calvaria implantation site biopsy harvested was subjected to histological, immunohistochemistry, RAMAN spectroscopy and Micro-CT analysis at the mentioned time points. Statistical analysis was conducted using t-tests and ANOVA. Histomorphometry showed significantly higher new bone formation in the DCC sites (p<0.05) compared to DMB. Both DMB and DCC implantation sites showed distinct staining for osteocalcin and osteopontin proteins compared to their respective sham sites. By day 21 after implantation, DCC sites demonstrated significantly elevated mRNA levels of osteonectin (p<0.001), osteopontin (p<0.001), osteocalcin (p<0.0001), ALP (p<0.01), and BMP-2 (p<0.001) compared to DMB. However, VEGF expression showed no significant differences at this time point between the two groups. Micro-CT analysis also showed enhanced defect closure and higher bone density in DCC implanted sites while RAMAN spectra demonstrated increased abundance of collagen and bone minerals, especially, PO43- ions than DMB. In conclusion, both DMB and DCC granules demonstrated favorable osteogenic potential in critical-sized defects, with DCC exhibited superior osteoconductive, osteoinductive and osteogenesis properties.

Biotransformed bear bile powder alleviates diet-induced nonalcoholic fatty liver disease in mice by regulating the gut microbiota and reversing lipid metabolism

BackgroundThe pathogenesis of metabolic syndrome was strongly associated with compromised metabolism homeostasis and gut microbiota imbalance. NAFLD is a progressive metabolic liver disease for which effective interventions are lacking. Bile acids exhibited appreciable metabolic regulatory effects and selective antimicrobial activity. Aim of the studyThis study was designed to investigate the effect of BBBP, which mainly contained bile acids, on NAFLD from the perspectives of gut microbiota and metabolomics. Materials and methodsThe present study was initiated on the anti-NAFLD effect of BBBP in HFD-fed mice. The efficacy of BBBP was evaluated by mice phenotypes, liver histopathological analysis and serum lipid and glucose levels. The activation of bile acid receptors such as Nr1h4, Nr1i2 and S1pr2 were detected by qRT-PCR analysis. Subsequently, untargeted metabolomics coupled with microbiomics was used to explore the mechanism of BBBP against NAFLD. Human L02 hepatocytes induced by OA and PA were used to investigate the effect of GABA on reducing lipid accumulation in vitro. ResultsBBBP significantly and dose-dependently alleviated the obese phenotype, lipid accumulation and liver injury in mice subjected to 18 weeks HFD diet. Untargeted metabolomics and microbiomics analysis revealed that BBBP could alleviate the disturbance of lipid and amino acid metabolism and the imbalance of gut microbiota. Furthermore, BBBP oral gavage activated liver bile acid receptors, as indicated by elevated mRNA levels of Nr1h4, Nr1i2 and S1pr2. Surprisingly, we determined that BBBP, which mainly contained bile acids that possessed antimicrobial activity, could promote the growth of Lactobacillus. Correlation analysis showed a remarkable correlation between Lactobacillus and endogenous metabolites such as valine, serine, glutamine, et al. Among them, GABA which could be produced by Lactobacillus significantly reduced the lipid accumulation in L02 cells. ConclusionsThe role of BBBP in regulating lipid metabolism might be achieved by activating bile acid receptors, or partially by promoting the levels of Lactobacillus and its metabolites such as GABA. Our study provided evidence that BBBP could be a novel therapeutic candidate for the treatment of NAFLD.

MRNA Levels of Aromatase, 5α-Reductase Isozymes, and Prostate Cancer-Related Genes in Plucked Hair from Young Men with Androgenic Alopecia.

Androgenic alopecia (AGA) is the most prevalent type of progressive hair loss and has psychological repercussions. Nevertheless, the effectiveness of current pharmacological treatments remains limited, in part because the molecular basis of the disease has not been fully elucidated. Our group previously highlighted the important roles of aromatase and 5α-reductase (5α-R) in alopecia in young women with female pattern hair loss. Additionally, an association has been proposed between AGA and prostate cancer (PCa), suggesting that genes implicated in PCa would also be involved in AGA. A low-invasive, sensitive, and precise method was used to determine mRNA levels of aromatase, 5α-R isozymes, and 84 PCa-related genes in samples of plucked hair from young men with AGA and controls. Samples were obtained with a trichogram from the vertex scalp, and mRNA levels were quantified using real-time RT-PCR. The men with AGA had significantly higher 5α-R2 mRNA levels in comparison to controls; interestingly, some of them also showed markedly elevated mRNA levels of 5α-R1 or 5α-R3 or of both, which may explain the varied response to 5α-R inhibitor treatments. The men with AGA also showed significant changes versus controls in 6 out of the 84 genes implicated in PCa. This study contributes greater knowledge of the molecular bases of AGA, facilitating early selection of the most appropriate pharmacological therapy and opening the way to novel treatments.

AGER-1 Long Non-Coding RNA Levels Correlate with the Expression of the Advanced Glycosylation End-Product Receptor, a Regulator of the Inflammatory Response in Visceral Adipose Tissue of Women with Obesity and Type 2 Diabetes Mellitus.

The advanced glycosylation end-product receptor (AGER) is involved in the development of metabolic inflammation and related complications in type 2 diabetes mellitus (T2DM). Tissue expression of the AGER gene (AGER) is regulated by epigenetic mediators, including a long non-coding RNA AGER-1 (lncAGER-1). This study aimed to investigate whether human obesity and T2DM are associated with an altered expression of AGER and lncAGER-1 in adipose tissue and, if so, whether these changes affect the local inflammatory milieu. The expression of genes encoding AGER, selected adipokines, and lncAGER-1 was assessed using real-time PCR in visceral (VAT) and subcutaneous (SAT) adipose tissue. VAT and SAT samples were obtained from 62 obese (BMI > 40 kg/m2; N = 24 diabetic) and 20 normal weight (BMI = 20-24.9 kg/m2) women, while a further 15 SAT samples were obtained from patients who were 18 to 24 months post-bariatric surgery. Tissue concentrations of adipokines were measured at the protein level using an ELISA-based method. Obesity was associated with increased AGER mRNA levels in SAT compared to normal weight status (p = 0.04) and surgical weight loss led to their significant decrease compared to pre-surgery levels (p = 0.01). Stratification by diabetic status revealed that AGER mRNA levels in VAT were higher in diabetic compared to non-diabetic women (p = 0.018). Elevated AGER mRNA levels in VAT of obese diabetic patients correlated with lncAGER-1 (p = 0.04, rs = 0.487) and with interleukin 1β (p = 0.008, rs = 0.525) and resistin (p = 0.004, rs = 0.6) mRNA concentrations. In conclusion, obesity in women is associated with increased expression of AGER in SAT, while T2DM is associated with increased AGER mRNA levels and pro-inflammatory adipokines in VAT.

The mRNA of solute carrier family 22, member 17 (SLC22A17) plays the potential diagnostic and prognostic role in advantage non-small cell lung cancer patients.

This study aims to evaluate the mRNA levels of solute carrier family 22, member 17 (SLC22A17) and its potential clinical value as a diagnostic and prognostic biomarker in non-small cell lung cancer. This prospective study measured SLC22A17 mRNA levels in lung cancer and paracancer tissues using quantitative reverse transcription-polymerase chain reaction (PCR). The levels of SLC22A17 mRNA in plasma samples from healthy control subjects and patients with lung cancer were also measured. The association between SLC22A17 mRNA levels in plasma and clinicopathological characteristics was determined. Receiver operating characteristic (ROC) curve analysis was performed to determine the diagnostic value of SLC22A17 in plasma. Survival curve analysis was performed using the Kaplan-Meier method. SLC22A17 mRNA levels were significantly higher in lung cancer samples than in the paired paracancerous tissues. Plasma SLC22A17 mRNA levels were also significantly higher in patients with lung cancer than in healthy controls. The COX analysis indicated that there was a significant correlation between elevated plasma SLC22A17 mRNA levels and lymph node metastasis, distant metastasis, and TNM stage. Furthermore, the ROC curve analysis demonstrated that plasma SLC22A17 had high diagnostic value. High plasma SLC22A17 mRNA levels are associated with a significantly shorter survival time. SLC22A17 is upregulated in lung cancer and may serve as a novel diagnostic and prognostic biomarker.

Molecular Mechanisms of the Regulation of Liver Cytochrome P450 by Brain NMDA Receptors and via the Neuroendocrine Pathway—A Significance for New Psychotropic Therapies

Recent investigations have highlighted the potential utility of the selective antagonist of the NMDA receptor GluN2B subunit for addressing major depressive disorders. Our previous study showed that the systemic administration of the antagonist of the GluN2B subunit of the NMDA receptor, the compound CP-101,606, affected liver cytochrome P450 expression and activity. To discern between the central and peripheral mechanisms of enzyme regulation, our current study aimed to explore whether the intracerebral administration of CP-101,606 could impact cytochrome P450. The injection of CP-101,606 to brain lateral ventricles (6, 15, or 30 µg/brain) exerted dose-dependent effects on liver cytochrome P450 enzymes and hypothalamic or pituitary hormones. The lowest dose led to an increase in the activity, protein, and mRNA level of CYP2C11 compared to the control. The activities of CYP2A, CYP2B, CYP2C11, CYP2C6, CYP2D, and protein levels of CYP2B, CYP2C11 were enhanced compared to the highest dose. Moreover, CP-101,606 increased the CYP1A protein level coupled with elevated CYP1A1 and CYP1A2 mRNA levels, but not activity. The antagonist decreased the pituitary somatostatin level and increased the serum growth hormone concentration after the lowest dose, while independently decreasing the serum corticosterone concentration of the dose. The findings presented here unveil a novel physiological regulatory mechanism whereby the brain glutamatergic system, via the NMDA receptor, influences liver cytochrome P450. This regulatory process appears to involve the endocrine system. These results may have practical applications in predicting alterations in cytochrome P450 activity and endogenous metabolism, and potential metabolic drug–drug interactions elicited by drugs that cross the blood–brain barrier and affect NMDA receptors.

(−)-Epigallocatechin-3-Gallate Reduces Perfluorodecanoic Acid-Exacerbated Adiposity and Hepatic Lipid Accumulation in High-Fat Diet-Fed Male C57BL/6J Mice

Perfluorodecanoic acid (PFDA), an enduring and harmful organic pollutant, is widely employed in diverse food-related sectors. Our previous studies have provided evidence that PFDA has the potential to facilitate obesity and hepatic fat accumulation induced by high-fat diet (HFD) intake. Epigallocatechin-3-gallate (EGCG), a polyphenol found in green tea, has been suggested to possess potential preventive effects against metabolic abnormalities and fatty liver. The purpose of this research was to investigate the effects of EGCG on PFDA-exacerbated adiposity and hepatic lipid accumulation in HFD-fed mice. The results showed that EGCG reduced body weight gain; tissue and organ weights; blood glucose, serum insulin, HOMA-IR, leptin, and lipid parameters; serum inflammatory cytokines (IL−1β, IL−18, IL−6, and TNF−α); and hepatic lipid accumulation in PFDA-exposed mice fed an HFD. Further work showed that EGCG improved liver function and glucose homeostasis in mice fed an HFD and co-exposed to PFDA. The elevated hepatic mRNA levels of SREBP-1 and associated lipogenic genes, NLRP3, and caspase−1 in PFDA-exposed mice fed an HFD were significantly decreased by EGCG. Our work provides evidence for the potential anti-obesity effect of EGCG on co-exposure to HFD and PFDA and may call for further research on the bioactivity of EGCG to attenuate the endocrine disruption effects of long-term exposure to pollutants.

Protein Phosphatase 6 Complex: Novel Regulator of Fetal Hemoglobin and Potential Therapeutic Target in Sickle Cell Disease

Although increasing levels of fetal hemoglobin (HbF) in sickle cell disease (SCD) significantly reduces mortality, effective pharmacologic induction of HbF has remained elusive. To identify potentially druggable molecules involved in HbF control, we carried out a domain-focused CRISPR-Cas9-based genetic screen in HUDEP2 cells targeting all 218 serine/threonine protein phosphatases with 6 independent sgRNAs each. A single phosphatase, PPP6C, emerged as a potential HbF repressor. PPP6C is the catalytic subunit of protein phosphatase 6 (PP6), a serine/threonine phosphatase complex that has been implicated in cell cycle regulation, autophagy, and innate immunity mechanisms in other cell types, but its role in erythropoiesis and HbF regulation has not previously been described. CRISPR-Cas9-based depletion of PPP6C in primary human erythroid cells elevated mRNA levels of HBG (the fetal form of β-globin) in a dose-dependent manner up to 3-4 fold, increased HbF levels 3-fold as measured by HPLC (up to 15-20% of total hemoglobin), and doubled the number of HbF-expressing cells. PPP6C depletion caused relatively few changes in the erythroid transcriptome by RNA-seq analysis, and did not measurably impair erythroid maturation. In addition, PPP6C loss in primary SCD patient-derived cells was well-tolerated, led to robust levels of HbF induction, and reduced cell sickling in vitro by up to 60%. Mechanistically, loss of PPP6C reduced the levels of the HbF repressor BCL11A by nearly 50% but left unchanged the levels of other HbF regulators, such as HRI, LRF, EKLF, NFIA/X, ZNF410, or HIC2, suggesting that PPP6C-mediated HbF regulation proceeds at least in part via loss of BCL11A. Importantly, xenotransplantation data (NBSGW) showed ~4-fold induction of HBG at 16 weeks post-transplant, suggesting that PPP6C deficiency leads to effective, sustained HbF induction in vivo. We next set out to determine whether any PP6 subunits have erythroid specificity that could be exploited therapeutically. In contrast to PPP6C, which is highly expressed in all hematopoietic lineages, the PP6 facultative subunit PPP6R1 appears to be enriched in erythroid precursors, suggesting that depletion of PPP6R1 may function as an erythroid-selective target to increase HbF levels. CRISPR-Cas9-based depletion of PPP6R1 in primary human erythroid cells showed a 2-3-fold increase in HBG and near-doubling of HbF-expressing cells, recapitulating the majority of PPP6C-mediated HbF induction. Functional studies are currently in progress to specifically determine whether additional PP6 subunits play a key role in HbF regulation. Furthermore, ongoing work utilizing dTAG-based acute depletion of PPP6C and PPP6R1 paired with phospho-proteomic studies will narrow down direct targets of the PP6 complex and provide insights into erythropoietic and HbF regulatory pathways impacted by PP6. Taken together, our data indicate that PP6, including its catalytic subunit PPP6C and erythroid-enriched regulatory subunit PPP6R1, inhibit HbF production in part via modulating BCL11A levels, and may serve as a red-cell specific therapeutic target in the treatment of SCD.

Title: Repeated implantation failure is associated with increased Th17/Treg cell ratio, during the secretory phase of the human endometrium

Repeated implantation failure (RIF) is a significant limiting factor in assisted reproduction. Chronic endometrial inflammation has been noted in RIF women, therefore we sought to investigate the potential association of endometrial Th17/Treg ratio and endometrial inflammation in these cases. Endometrial pipelle biopsies were obtained from volunteers, 29 women with RIF (failure to achieve pregnancy following at least 3 transfers of high-grade embryos in IVF-cycles) and 27 fertile women (at least one child) in total, at the secretory phase of the menstrual cycle. Using tissues from 17 fertile and 18 RIF endometrial samples, stromal and immune cells were isolated and flow cytometry analysis was performed to determine Th17 and CD4+CD25high FOXP3+ cell populations in endometrial stromal cell suspensions. Another group of tissues from 10 fertile and 11 RIF samples were used for mRNA expression levels of Treg and Th17-cell transcription factors, FOXP3 and RORγt respectively. Endometrial inflammatory mediators’ mRNA expression was also analyzed. A statistically significant increase in protein flow cytometry analysis of Th17/Treg ratio (p≤0.05) as well as a reduction in absolute Treg cells in the endometrium (p≤0.05) was noted in women with RIF. Additionally, RNA analysis on the same set of women indicated RORγt/FOXP3 significantly increased in women with RIF compared to fertile ones (p≤0.05). Finally, women with RIF exhibited significantly (p≤0.05) elevated mRNA levels of pro-inflammatory mediators (ΤΝF-a, ΙL-6, IL-8 and CCl2). Women with RIF exhibit elevated Th17/Treg ratio, mostly due to endometrial Treg depletion, as well as a pro-inflammatory state in the endometrium.

Analysis of the mRNA export protein ZC3H11A in HCMV infection and pan-cancer.

We have previously reported that human cytomegalovirus (HCMV) infection could promote the progression of glioma. Here we discovered a stress-induced nuclear protein ZC3H11A (ZC3) through high-throughput sequencing after HCMV infection, which has been reported recently by our research group in regulating mRNA export under stress conditions. And also, a thorough analysis of ZC3 in pan-cancer and the omics data of ZC3 are yet to be conducted. The transcriptomes of glioma cells after HCMV infection were assessed by RNA sequencing. ZC3 mRNA and protein level following HCMV infection were validated and measured by qRT-PCR and Western-blot. The RNA sequencing and protein expression information of ZC3 across pan-cancer were analyzed and visualized by R packages. The localization of ZC3 protein was assessed by IHC images from HPA. The ZC3 proteomics and transcriptomics data in different cancers were extracted through the CPTAC data portal, and comparisons were conducted with a Python script. The genetic alteration, survival prognosis, immune infiltration analysis of ZC3 in pan-cancer were analyzed by cBioPortal, TCGA, and TIMER2 databases. The protein interaction networks were revealed by STRING, GEPIA2 and TCGA. Genes in mRNA processing pathways were upregulated after HCMV infection and ZC3 expression in mRNA and protein level was validated. We also discovered that the status of ZC3 were generally at high levels in cancers, although varied among different cancer types. ZC3 protein in tumor cells localized to the nuclear whereas in normal cells it was mainly found in cytoplasmic/membranous. However, from ZC3 proteomics and transcriptomics data in some cancer types, the increase in ZC3 protein was not accompanied by a significant elevation in mRNA level. Additionally, our analysis indicated that elevated ZC3 expression was primarily linked to a negative prognosis in majority cancers but still varied depending on the cancer types. Our annotation analysis suggested that ZC3-related proteins are mainly involved in mRNA processing clusters. We demonstrated that ZC3 significantly impacted by HCMV infection in gliomas. Furthermore, we identified a set of genes exhibiting analogous expression patterns to ZC3H11A in TCGA pan-cancer cohorts, implying a potential functional role for ZC3H11A in mRNA processing. Our study provided valuable insights into the role of a new mRNA export protein ZC3 in HCMV infection and pan-cancer progression. These results lay the foundation for our next research on the regulatory mechanism of ZC3 in virus-infected tumors.

DNA methylation profiling of circulating CD4+ T cells and network analysis reveal that JUNB, SETD7, and MEF2D genes predict patients with heart failure preserved vs. reduced ejection fraction

Abstract Background Heart failure (HF) with preserved ejection fraction (HFpEF) represents about 50% of new HF diagnosis but still lacks specific biomarkers and effective therapy. Epigenetic-sensitive changes accumulated over time, mainly guided by DNA methylation and demethylation, can lead to chromatin remodelling and affect transcriptional molecular networks underlying microvascular inflammation, oxidative stress, and maladaptive cardiac remodelling which predispose synergistically to HFpEF. Purpose We hypothesize that circulating genome-wide DNA methylation profiles may identify novel biomarkers for discriminating HFpEF vs. HF with reduced EF (HFrEF) and provide insight into the underlying pathophysiology of disease. Methods A total of n=47 subjects were enrolled including patients with HFpEF (n=22) and HFrEF (n=13) as well as healthy subjects (n=12) serving as control group (CON). Starting from pheripheral blood biospecimens, we isolated CD4+ T cells and extracted genomic DNA for genome-wide methylome analysis and total RNA for experimental validation set. Results The reduced representation bisulfite sequencing (RRBS) revealed that patients with HFpEF had unique variations in DNA methylation profiles as compared to HFrEF and CON groups and reflected distinct pathogenic mechanisms. Using the list of differentially methylated CpG sites in HFpEF, we performed a complex network analysis to indentify the "HFpEF interactome" (Fig.1 A-B). Then, using the DisGeNET database we extracted n=12 candidate genes which were already associated with diastolic dysfunction, cardiac hypertrophy, cardiac fibrosis, inflammation, obesity, and hypertension, as key pathophenotypes of HFpEF (Fig.1 C). The hub genes included the ACTB, VAV2, JUNB, HOOK2, SETD7, PNKP, ASAP1, PAICS, DNM1L, HIF1AN, MEF2D, and NRG1. Validation experiment set by qRT-PCR revealed that elevated mRNA levels of the JUNB, SETD7, and MEF2D hypermethylated genes significantly discriminated HFpEF patients vs. CON (p&amp;lt;0.01, p&amp;lt;0.01, p&amp;lt;0.001, respectively) and vs. HFrEF patients (p&amp;lt;0.05, p&amp;lt;0.01, p&amp;lt;0.001, respectively) (Fig. 2A). Receiver operating characteristic (ROC) curve showed that network-oriented JUNB (AUC: 1, p&amp;lt;0.001), SETD7 (AUC: 0.97, p&amp;lt;0.01), and MEF2D (AUC: 1, p&amp;lt;0.001) genes have a high diagnostic accuracy in discriminating HFpEF vs. CON as well as HFpEF vs. HFrEF (AUC: 1, p&amp;lt;0.01, AUC: 0.97, p&amp;lt;0.01, AUC: 1, p&amp;lt;0.001, respectively) (Fig. 2B). Conclusions Circulating CD4+ T cell-derived hypermethylation of JUNB, SETD7, and MEF2D may reveal novel molecular drivers of HFpEF and suggest possible biomarkers to optimize patient phenotyping.

PDE3A Is a Highly Expressed Therapy Target in Myxoid Liposarcoma.

Liposarcomas (LPSs) are a heterogeneous group of malignancies that arise from adipose tissue. Although LPSs are among the most common soft-tissue sarcoma subtypes, precision medicine treatments are not currently available. To discover LPS-subtype-specific therapy targets, we investigated RNA sequenced transcriptomes of 131 clinical LPS tissue samples and compared the data with a transcriptome database that contained 20,218 samples from 95 healthy tissues and 106 cancerous tissue types. The identified genes were referred to the NCATS BioPlanet library with Enrichr to analyze upregulated signaling pathways. PDE3A protein expression was investigated with immunohistochemistry in 181 LPS samples, and PDE3A and SLFN12 mRNA expression with RT-qPCR were investigated in 63 LPS samples. Immunoblotting and cell viability assays were used to study LPS cell lines and their sensitivity to PDE3A modulators. We identified 97, 247, and 37 subtype-specific, highly expressed genes in dedifferentiated, myxoid, and pleomorphic LPS subtypes, respectively. Signaling pathway analysis revealed a highly activated hedgehog signaling pathway in dedifferentiated LPS, phospholipase c mediated cascade and insulin signaling in myxoid LPS, and pathways associated with cell proliferation in pleomorphic LPS. We discovered a strong association between high PDE3A expression and myxoid LPS, particularly in high-grade tumors. Moreover, myxoid LPS samples showed elevated expression levels of SLFN12 mRNA. In addition, PDE3A- and SLFN12-coexpressing LPS cell lines SA4 and GOT3 were sensitive to PDE3A modulators. Our results indicate that PDE3A modulators are promising drugs to treat myxoid LPS. Further studies are required to develop these drugs for clinical use.

Preventive Effects of Collagen-Derived Dipeptide Prolyl-Hydroxyproline against Dexamethasone-Induced Muscle Atrophy in Mouse C2C12 Skeletal Myotubes.

Glucocorticoids, commonly used to manage inflammatory diseases, can induce muscle atrophy by accelerating the breakdown of muscle proteins. This research delves into the influence of Prolyl-hydroxyproline (Pro-Hyp), a collagen-derived peptide, on muscle atrophy induced with dexamethasone (DEX), a synthetic glucocorticoid, in mouse C2C12 skeletal myotubes. Exposure to DEX (10 μM) for 6 days resulted in a decrease in myotube diameter, along with elevated mRNA and protein levels of two muscle-atrophy-related ubiquitin ligases, muscle atrophy F-box (MAFbx, also known as atrogin-1) and muscle ring finger 1 (MuRF-1). Remarkably, treatment with 0.1 mM of Pro-Hyp mitigated the reduction in myotube thickness caused by DEX, while promoting the phosphorylation of Akt, mammalian target of rapamycin (mTOR), and forkhead box O3a (Foxo3a). This led to the inhibition of the upregulation of the ubiquitin ligases atrogin-1 and MuRF-1. These findings indicate the potential significance of Pro-Hyp as a promising therapeutic target for countering DEX-induced muscle atrophy.

Synaptotagmin-11 regulates immune functions of microglia invivo.

Membrane trafficking pathways mediate key microglial activities such as cell migration, cytokine secretion, and phagocytosis. However, the underlying molecular mechanism remains poorly understood. Previously, we found that synaptotagmin-11 (Syt11), a non-Ca2+ -binding Syt associated with Parkinson's disease (PD) and schizophrenia, inhibits cytokine release and phagocytosis in primary microglia. Here we reported the invivo function of Syt11 in microglial immune responses using an inducible microglia-specific Syt11-conditional-knockout (cKO) mouse strain. Syt11-cKO resulted in activation of microglia and elevated mRNA levels of IL-6, TNF-α, IL-1β, and iNOS in various brain regions under both resting state and LPS-induced acute inflammation state in adult mice. In a PD mouse model generated by microinjection of preformed α-synuclein fibrils into the striatum, a reduced number of microglia migrated toward the injection sites and an enhanced phagocytosis of α-synuclein fibrils by microglia were found in Syt11-cKO mice. To understand the molecular mechanism of Syt11 function, we identified its direct binding proteins vps10p-tail-interactor-1a (vti1a) and vti1b. The linker domain of Syt11 interacted with both proteins and a peptide derived from it competitively inhibited the interaction of Syt11 with vti1a/vti1b invitro and in cells. Importantly, application of this peptide induced more cytokine secretion in wild-type microglia upon LPS treatment, phenocopying defects in Syt11 knockdown cells. Altogether, we propose that Syt11 inhibits microglial activation invivo and regulates cytokine secretion through interactions with vti1a and vti1b.

Energy and macronutrient restriction regulate bile acid homeostasis

As we reported previously, caloric restriction (CR) results in an increased concentration of bile acids (BA) in the intestinal mucosa. We now investigated the background of this phenotype, trying to identify nutrition-related factors modulating BA levels. Male mice were submitted to various types of restrictive diets and BA levels and expression of associated factors were measured. We found that BA concentration is increased in the liver of CR mice, which corresponds to reduced expression of the Shp gene and elevated mRNA levels of Cyp27a1, Bal, and Ntcp, as well as CYP7A1 protein and gene expression. Correlation between decreased concentration of BAs in the feces, increased BAs levels in plasma, and elevated gene expression of BAs transporters in the ileum mucosa suggests enhanced BA uptake in the intestine of CR mice. Corresponding to CR upregulation of liver and ileum mucosa, BA concentration was found in animals submitted to other types of prolonged energy-restricting dietary protocols, including intermittent fasting and fasting-mimicking diet. While over-night fasting had negligible impact on BAs levels. Manipulation of macronutrient levels partly affected BA balance. Low-carbohydrate and ketogenic diet increased BAs in the liver but not in the intestine. Carbohydrate restriction stimulates BA synthesis in the liver, but energy restriction is required for the increase in BA levels in the intestine and its uptake.