Apelin Signaling Research Articles

Single-Cell Transcriptomics of Endothelial Cells in Upper and Lower Human Esophageal Squamous Cell Carcinoma.

Esophageal squamous cell carcinoma (ESCC) is a type of progressive and distant metastatic tumor. Targeting anti-angiogenic genes could effectively hinder ESCC development and metastasis, whereas ESCC locating on the upper or the lower esophagus showed different response to the same clinical treatment, suggesting ESCC location should be taken into account when exploring new therapeutic targets. In the current study, to find novel anti-angiogenic therapeutic targets, we identified endothelial cell subsets in upper and lower human ESCC using single-cell RNA sequencing (scRNA-seq), screened differentially expressed genes (DEGs), and performed gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. The results showed that common DEGs shared in the upper and the lower endothelial cells mainly are involved in vessel development, angiogenesis, and cell motility of endothelial cells by regulating PI3K-AKT, Rap1, Ras, TGF-beta, and Apelin signaling pathways. The critical regulatory genes were identified as ITGB1, Col4A1, Col4A2, ITGA6, LAMA4, LAMB1, LAMC1, VWF, ITGA5, THBS1, PDGFB, PGF, RHOC, and CTNNB1. Cell metabolism-relevant genes, e.g., MGST3, PNP, UPP1, and HYAL2 might be the prospective therapeutic targets. Furthermore, we found that DEGs only in the upper endothelial cells, such as MAPK3, STAT3, RHOA, MAPK11, HIF1A, FGFR1, GNG5, GNB1, and ARHGEF12, mainly regulated cell adhesion, structure morphogenesis, and motility through Phospholipase D, Apelin, and VEGF signaling pathways. Moreover, DEGs only in the lower endothelial cells, for instance PLCG2, EFNA1, CALM1, and RALA, mainly regulated cell apoptosis and survival by targeting calcium ion transport through Rap1, Ras, cAMP, Phospholipase D, and Phosphatidylinositol signaling pathways. In addition, the upper endothelial cells showed significant functional diversity such as cytokine-responsive, migratory, and proliferative capacity, presenting a better angiogenic capacity and making it more sensitive to anti-angiogenic therapy compared with the lower endothelial cells. Our study has identified the potential targeted genes for anti-angiogenic therapy for both upper and lower ESCC, and further indicated that anti-angiogenic therapy might be more effective for upper ESCC, which still need to be further examined in the future.

Maternal exercise improves epithelial development of fetal intestine by enhancing apelin signaling and oxidative metabolism.

Obesity in pregnancy is currently the leading cause of gestational complications for the mother and fetus worldwide. Maternal obesity (MO), common in western societies, impedes development of intestinal epithelium in the fetuses, which causes disorders in the nutrient absorption and intestine-related immune responses in offspring. Here, using a mouse model of maternal exercise (ME), we found that exercise during pregnancy protects the impairment of fetal intestinal morphometrical formation and epithelial development due to MO. MO decreased villus length and epithelial proliferation markers in E18.5 fetal small intestine, which was increased due to ME. The expression of the epithelial differentiation markers, Lyz1, Muc2, and Tff3, in fetal small intestine was decreased due to MO, but protected by ME. Consistently, the biomarkers related to mitochondrial biogenesis and oxidative metabolism were downregulated in MO fetal small intestine but recovered by ME. Apelin injection to dams partially mirrored the beneficial effects of ME. ME and apelin injection activated AMPK, the downstream target of apelin receptor signaling, which might mediate the improvement of fetal epithelial development and oxidative metabolism. These findings suggest that ME, a highly accessible intervention, is effective in improving fetal intestinal epithelium of obese dams. Apelin-AMPK-mitochondrial biogenesis axis provides amenable therapeutic targets to facilitate fetal intestinal development of obese mothers.

Network pharmacology approach to screen hypoglycemic extracts from Coptidis rhizoma and stu dy on their targets and pathways

Purpose: To screen the hypoglycemic active ingredients from Coptidis rhizoma, and study their targets as well as signal pathways via network pharmacology.
 Methods: The fifty-nine ingredients of Coptidis rhizoma were screened for. Their targets were confirmed by comparing with the hypoglycemic targets in DrugBank databases. The relationship between ingredients and targets was revealed through String database. The ingredient-target-passageway network was constructed. Coptidis rhizoma was soaked in boiling water and concentrated. Rat models were rendered diabetic by the administration of streptozotocin (STZ) intraperitoneal injection, and administered Coptidis rhizoma (0.40 g/kg, once a day by gavage), and tested for antidiabetic activity.
 Results: After four weeks of treatment, their blood glucose levels (BG) of all treated hyperglycemic rats decreased (p < 0.05). Twenty-four hypoglycemic compounds screened from Coptidis rhizome via network pharmacology could activate 13 targets, such as D (2) dopamine receptor (DRD2), insulin-like growth factor 1 receptor (IGF1R), 5-hydroxytryptamine receptor 2C (HTR2C), 5-hydroxytryptamine receptor 3A (HTR3A) and sodium-dependent noradrenaline transporter (SLC6A2). These targets were involved in 141 pathways, e.g., cAMP signaling pathway, chemokine signaling pathway, Rap1 signaling pathway, estrogen signaling pathway, and Apelin signaling pathway.
 Conclusion: This study suggests that Coptidis rhizoma contains several active compounds that show significant hypoglycemic effects. Furthermore, this study has established the basis for future investigations on the hypoglycemic effects of Coptidis rhizoma.

Upregulation of circ_0008812 and circ_0001583 predicts poor prognosis and promotes breast cancer proliferation.

Background: Accumulating evidence suggests that circular RNAs (circRNAs) are highly correlated with tumor progression and pathogenesis in breast cancer. Whereas, their regulatory roles and corresponding mechanisms in breast cancer are still not exhaustive. Thus, we intended to establish circRNA-mediated competive endogenous RNA (ceRNA) network to uncover the possible roles and clinical implications of circRNAs in breast cancer. Methods: Microarray and RNA-sequencing (RNA-seq) data were download from GEO and TCGA database to screen for differentially expressed RNAs (DEcircRNAs, DEmiRNAs, DEmRNAs) in breast cancer. By implementing online databases, we established ceRNA networks, performed gene set enrichment analysis, constructed protein-protein interaction (PPI) networks, and assessed the expression levels and prognostic significance of hub genes. Subsequently, we explored the functions of prognosis-related genes and constructed gene-drug interaction networks. Finally, the functional roles of DEcircRNAs in breast cancer were revealed via MTT and colony formation assay. Results: Based on the identified 8 DEcircRNAs, 25 miRNAs and 216 mRNAs, a ceRNA regulatory network was established. Further analysis revealed that prominent enrichments were transcription factor binding, transforming growth factor-beta (TGF-β) and Apelin signaling pathway etc. PPI network and survival curves analysis showed that elevated levels of hub genes (RACGAP1 and KPNA2) were associated with poorer prognosis. They were found to be positively relevant to cell cycle and proliferation. Then a prognostic sub-network of ceRNA was constructed, consisting of 2 circRNAs, 4 miRNAs and 2 mRNAs. The gene-drug interaction network showed that numerous drugs could regulate the expression of these two prognosis-related genes. Functional experiments showed that depletion of circ_0008812 and circ_0001583 could significantly inhibit the proliferation of MCF-7 cells. Conclusion: Our study constructed 4 prognostic regulatory axes that are significantly correlated with tumor prognosis in breast cancer patients, and uncover the roles of circ_0008812 and circ_0001583 in breast cancer, providing a new perspective into the molecular mechanisms of breast cancer pathogenesis.

Upregulated Apelin Signaling in Pancreatic Cancer Activates Oncogenic Signaling Pathways to Promote Tumor Development.

Despite decades of effort in understanding pancreatic ductal adenocarcinoma (PDAC), there is still a lack of innovative targeted therapies for this devastating disease. Herein, we report the expression of apelin and its receptor, APJ, in human pancreatic adenocarcinoma and its protumoral function. Apelin and APJ protein expression in tumor tissues from patients with PDAC and their spatiotemporal pattern of expression in engineered mouse models of PDAC were investigated by immunohistochemistry. Apelin signaling function in tumor cells was characterized in pancreatic tumor cell lines by Western blot as well as proliferation, migration assays and in murine orthotopic xenograft experiments. In premalignant lesions, apelin was expressed in epithelial lesions whereas APJ was found in isolated cells tightly attached to premalignant lesions. However, in the invasive stage, apelin and APJ were co-expressed by tumor cells. In human tumor cells, apelin induced a long-lasting activation of PI3K/Akt, upregulated β-catenin and the oncogenes c-myc and cyclin D1 and promoted proliferation, migration and glucose uptake. Apelin receptor blockades reduced cancer cell proliferation along with a reduction in pancreatic tumor burden. These findings identify the apelin signaling pathway as a new actor for PDAC development and a novel therapeutic target for this incurable disease.

Postnatal developmental expression of apelin receptor proteins and its role in juvenile mice testis

The expression of apelin system has been shown in the adult testis of rat and mice. It has also been emphasized that regulation of testicular activity in early stages is important to sustain normal testicular activity in adulthood. Since the expression of apelin receptor (APJ) has been shown in the adult testis, moreover, developmental expression of APJ and its role has not been explored yet. Thus, we have examined the testicular expression of APJ during postnatal stages with special reference to proliferation, apoptosis and hormone secretion in early postnatal stage. Postnatal analysis showed that circulating apelin was lowest at PND1 and maximum at PND42. Among testosterone, estrogen and androstenedione, only circulating testosterone showed a gradual increase from PND1 to PND42. Testicular expression of APJ was also developmenatly regulated from PND1 to PND42, revealing a positive correlation with circulating apelin, testosterone, and androstenedione. Immunohistochemical study showed that APJ was mainly confined to Leydig cells of early postnatal stages, whereas, seminiferous tubules at PND42 showed immunostaining in the round spermatids. APJ inhibition from PND14-PND20 by ML221 suppressed the testicular proliferation, increased apoptosis and increased estrogen secretion. However, expression of AR was down-regulated by ML221 treatment. Furthermore, ML221 decreased the abundance of p-Akt. In vitro study also showed that APJ antagonist, ML221 decreased AR expression. These results suggests that apelin signaling during early developmental stages might be required to stimulate the germ cell proliferation, and inhibition of apoptosis. Both in vivo and in vitro study have shown that expression of AR was regulated by apelin signaling. Since the first wave spermatogenesis involves proliferation and apoptosis, therefore, further study would be required to unravel the exact mechanism of apelin mediated regulation of testicular activity during early postnatal stages. In conclusion, the present results are an indicative of apelin mediated signaling during early postnatal stage for regulation of germ cell proliferation, apoptosis and AR expression.

Proteomic Analysis of Effects of Spironolactone in Heart Failure With Preserved Ejection Fraction.

The TOPCAT trial (Treatment of Preserved Cardiac Function Heart Failure With an Aldosterone Antagonist Trial) suggested clinical benefits of spironolactone treatment among patients with heart failure with preserved ejection fraction enrolled in the Americas. However, a comprehensive assessment of biologic pathways impacted by spironolactone therapy in heart failure with preserved ejection fraction has not been performed. We conducted aptamer-based proteomic analysis utilizing 5284 modified aptamers to 4928 unique proteins on plasma samples from TOPCAT participants from the Americas (n=164 subjects with paired samples at baseline and 1 year) to identify proteins and pathways impacted by spironolactone therapy in heart failure with preserved ejection fraction. Mean percentage change from baseline was calculated for each protein. Additionally, we conducted pathway analysis of proteins altered by spironolactone. Spironolactone therapy was associated with proteome-wide significant changes in 7 proteins. Among these, CARD18 (caspase recruitment domain-containing protein 18), PKD2 (polycystin 2), and PSG2 (pregnancy-specific glycoprotein 2) were upregulated, whereas HGF (hepatic growth factor), PLTP (phospholipid transfer protein), IGF2R (insulin growth factor 2 receptor), and SWP70 (switch-associated protein 70) were downregulated. CARD18, a caspase-1 inhibitor, was the most upregulated protein by spironolactone (-0.5% with placebo versus +66.5% with spironolactone, P<0.0001). The top canonical pathways that were significantly associated with spironolactone were apelin signaling, stellate cell activation, glycoprotein 6 signaling, atherosclerosis signaling, liver X receptor activation, and farnesoid X receptor activation. Among the top pathways, collagens were a consistent theme that increased in patients receiving placebo but decreased in patients randomized to spironolactone. Proteomic analysis in the TOPCAT trial revealed proteins and pathways altered by spironolactone, including the caspase inhibitor CARD18 and multiple pathways that involved collagens. In addition to effects on fibrosis, our studies suggest potential antiapoptotic effects of spironolactone in heart failure with preserved ejection fraction, a hypothesis that merits further exploration.

Integrated Analysis of Transcriptome, microRNAs, and Chromatin Accessibility Revealed Potential Early B-Cell Factor1-Regulated Transcriptional Networks during the Early Development of Fetal Brown Adipose Tissues in Rabbits.

In domestic mammals, cold stress decreases the survival rate of newborns and increases the cost of management. Brown adipose tissue (BAT) is the main thermogenic organ for cubs, and well-developed fetal BAT (FBAT) is beneficial for newborns to maintain core temperatures during the first several days of life. However, our knowledge of the epigenetic mechanisms during the early development of FBAT remains largely unknown. Rabbits (Oryctolagus cuniculus) are economically important domestic animals. In this study, a histological analysis shows that the tissue content, thermogenic capacity, and lipid content of FBAT dramatically increases from gestational day 21 (G21) to gestational day 24 (G24) in rabbits. RNA-seq, microRNA-seq (miRNA-seq), and the assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) show that many genes, miRNAs, and chromatin-accessible regions (referred to as peaks) were identified as significantly changed from G21 to G24, respectively. The upregulated genes from G21 to G24 were significantly enriched in the mitochondrial metabolism and thermogenesis-related signal pathways. The integrated analysis of transcriptome and chromatin accessibility reveals that the peaks in the promoters have a more regulatory effect than peaks in other genomic elements on the expression of their nearby genes in FBATs. The upregulated genes that are associated with increased chromatin accessibility in the promoter regions are involved in the energy metabolism-related signaling pathways. The genes that have a greater tendency to be regulated by miRNAs than the chromatin accessibility in gene promoters are involved in the apelin, insulin, and endocytosis signaling pathways. Furthermore, genome-wide transcription factor (TF) footprinting analysis identifies early B-cell factor1 (EBF1) as playing a key role during early FBAT development. The carbon metabolism, citrate cycle, and PPAR signaling pathways are significantly enriched by the predicted EBF1-regulated cascade TF-network. In conclusion, our work provides a framework for understanding epigenetics regulatory mechanisms underlying the early development of FBAT and identifies potential TF involved in the early development of FBAT in rabbits.

Network pharmacology approach to screen hypoglycemic extracts from Coptidis rhizoma and study their targets and pathways

Purpose: To screen the hypoglycemic active ingredients of Coptidis rhizoma, and study their targets as well as signal pathways via network pharmacology.&#x0D; Methods: Fifty-nine ingredients of Coptidis rhizoma were screened. Their targets were confirmed by comparing with the hypoglycemic targets in DrugBank databases. The relationship between ingredients and targets was revealed through String database. The ingredient-target-passageway network was constructed. Coptidis rhizoma was soaked in boiling water and concentrated. Rat models were rendered diabetic by the administration of streptozotocin (STZ) intraperitoneal injection. The hyperglycemic rats received Coptidis rhizoma extract (0.40 g/kg, once a day by gavage).&#x0D; Results: After four weeks of treatment, the blood glucose levels (BG) of all treated hyperglycemic rats decreased (p &lt; 0.05). Twenty-four hypoglycemic active compounds were obtained after screening the extract of Coptidis rhizome via network pharmacology. These active compounds activated 13 targets, including D (2) dopamine receptor (DRD2), Insulin-like growth factor 1 receptor (IGF1R), 5- hydroxytryptamine receptor 2C (HTR2C), 5-hydroxytryptamine receptor 3A (HTR3A) and sodiumdependent noradrenaline transporter (SLC6A2). These targets were involved in 141 pathways, e.g., cAMP signaling pathway, chemokine signaling pathway, Rap1 signaling pathway, estrogen signaling pathway, and apelin signaling pathway.&#x0D; Conclusion: Coptidis rhizoma contains several active compounds that exhibit good hypoglycemic effects. Thus, there is a need for human studies on the hypoglycemic effects of Coptidis rhizome extracts.&#x0D; Keywords: Coptidis rhizome; Hypoglycemic effects; Network pharmacology; D (2) dopamine receptor (DRD2); Insulin-like growth factor 1 receptor

Differentially CTCF-Binding Sites in Cattle Rumen Tissue during Weaning.

The weaning transition in calves is characterized by major structural changes such as an increase in the rumen capacity and surface area due to diet changes. Studies evaluating rumen development in calves are vital to identify genetic mechanisms affected by weaning. This study aimed to provide a genome-wide characterization of CTCF-binding sites and differentially CTCF-binding sites (DCBS) in rumen tissue during the weaning transition of four Holstein calves to uncover regulatory elements in rumen epithelial tissue using ChIP-seq. Our study generated 67,280 CTCF peaks for the before weaning (BW) and 39,891 for after weaning (AW). Then, 7401 DCBS were identified for the AW vs. BW comparison representing 0.15% of the cattle genome, comprising ~54% of induced DCBS and ~46% of repressed DCBS. Most of the induced and repressed DCBS were in distal intergenic regions, showing a potential role as insulators. Gene ontology enrichment revealed many shared GO terms for the induced and the repressed DCBS, mainly related to cellular migration, proliferation, growth, differentiation, cellular adhesion, digestive tract morphogenesis, and response to TGFβ. In addition, shared KEGG pathways were obtained for adherens junction and focal adhesion. Interestingly, other relevant KEGG pathways were observed for the induced DCBS like gastric acid secretion, salivary secretion, bacterial invasion of epithelial cells, apelin signaling, and mucin-type O-glycan biosynthesis. IPA analysis further revealed pathways with potential roles in rumen development during weaning, including TGFβ, Integrin-linked kinase, and Integrin signaling. When DCBS were further integrated with RNA-seq data, 36 putative target genes were identified for the repressed DCBS, including KRT84, COL9A2, MATN3, TSPAN1, and AJM1. This study successfully identified DCBS in cattle rumen tissue after weaning on a genome-wide scale and revealed several candidate target genes that may have a role in rumen development, such as TGFβ, integrins, keratins, and SMADs. The information generated in this preliminary study provides new insights into bovine genome regulation and chromatin landscape.

Use of transcriptome sequencing to explore the effect of CSRP3 on chicken myoblasts

The mechanisms that regulate the specificity and maintenance of chicken muscle fiber types remain largely unknown. In mammals, CSRP3 has been shown to play a vital role in the maintenance of typical muscle structure and function. This study investigated the role that CSRP3 plays in chicken skeletal muscle. First, the antibody against chicken CSRP3 protein was prepared, and the expression levels of the mRNA and protein of the CSRP3 gene in four chicken skeletal muscles with different myofiber compositions were compared. Then the effects of CSRP3 silencing on the expression profile of chicken myoblast transcriptomes were analyzed. The results showed that the expression levels of the mRNA and protein of the CSRP3 gene were both associated with the composition of fiber types in chicken skeletal muscles. A total of 650 genes with at least 1.5-fold differences (Q<0.05) were identified, of which 255 genes were upregulated and 395 genes were downregulated by CSRP3 silencing. Functional enrichment showed that several pathways, including adrenergic signaling in cardiomyocytes, adipocytokine signaling pathway and apelin signaling pathway, were significantly (P<0.05) enriched both in differentially expressed genes and all expressed genes. The co-expressed gene network suggested that CSRP3 silencing caused a compensatory upregulation (Q<0.05) of genes related to the assembly of myofibrils, muscle differentiation, and contraction. Meanwhile, two fast myosin heavy chain genes (MyH1B and MyH1E) were upregulated (Q<0.05) upon CSRP3 silencing. These results suggested that CSRP3 plays a crucial role in chicken myofiber composition, and affects the distribution of chicken myofiber types, probably by regulating the expression of MyH1B and MyH1E.

Identification of key genes for hypertrophic cardiomyopathy using integrated network analysis of differential lncRNA and gene expression.

ObjectiveHypertrophic cardiomyopathy (HCM) is a complex heterogeneous heart disease. Recent reports found that long non-coding RNAs (lncRNAs) play an important role in the progression of cardiovascular diseases. The present study aimed to identify the novel lncRNAs and messenger RNAs (mRNAs) and determine the key pathways involved in HCM.MethodsThe lncRNA and mRNA sequencing datasets of GSE68316 and GSE130036 were downloaded from the Gene Expression Omnibus (GEO) database. An integrated co-expression network analysis was conducted to identify differentially expressed lncRNAs and differentially expressed mRNAs in patients with HCM. Then, gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were explored to identify the biological functions and signaling pathways of the co-expression network. Protein–protein interaction (PPI) and hub gene networks were constructed by using Cytoscape software. Plasma samples of patients with HCM and the GSE89714 dataset were used to validate the bioinformatics results.ResultsA total of 1,426 differentially expressed long non-coding RNAs (lncRNAs) and 1,715 differentially expressed mRNAs were obtained from GSE68316, of which 965 lncRNAs and 896 mRNAs were upregulated and 461 lncRNAs and 819 mRNAs were downregulated. A total of 469 differentially expressed lncRNAs and 2,407 differentially expressed mRNAs were screened from GSE130036, of which 183 lncRNAs and 1,283 mRNAs were upregulated and 286 lncRNAs and 1,124 mRNAs were downregulated. A co-expression network was constructed and contained 30 differentially expressed lncRNAs and 63 differentially expressed mRNAs, which were primarily involved in ‘G-protein beta/gamma-subunit complex binding,' ‘polyubiquitin modification-dependent protein binding,' ‘Apelin signaling pathway,' and ‘Wnt signaling pathway.' The 10 hub genes in the upregulated network [G Protein Subunit Alpha I2 (GNAI2), G Protein Subunit Alpha I1 (GNAI1), G Protein Subunit Alpha I3 (GNAI3), G Protein Subunit Gamma 2 (GNG2), G Protein Subunit Beta 1 (GNB1), G Protein Subunit Gamma 13 (GNG13), G Protein Subunit Gamma Transducin 1 (GNGT1), G Protein Subunit Gamma 12 (GNG12), AKT Serine/Threonine Kinase 1 (AKT1) and GNAS Complex Locus (GNAS)] and the 10 hub genes in the downregulated network [Nucleotide-Binding Oligomerization Domain Containing Protein 2 (NOD2), Receptor-Interacting Serine/Threonine Kinase 2 (RIPK2), Nucleotide-Binding Oligomerization Domain Containing Protein 1 (NOD1), Mitochondrial Antiviral Signaling Protein (MAVS), Autophagy Related 16-Like 1 (ATG16L1), Interferon Induced With Helicase C Domain 1 (IFIH1), Autophagy Related 5 (ATG5), TANK-Binding Kinase 1 (TBK1), Caspase Recruitment Domain Family Member 9 (CARD9), and von Willebrand factor (VWF)] were screened using cytoHubba. The expression of LA16c-312E8.2 and RP5-1160K1.3 in the plasma of patients with HCM was elevated, and the expression of the MIR22 host gene (MIR22HG) was decreased, which was consistent with our analysis, while the expression of LINC00324 and Small Nucleolar RNA Host Gene 12 (SNHG12) was not significantly different between the two groups. Verification analyses performed on GSE89714 showed the upregulated mRNAs of Chloride Voltage-Gated Channel 7 (CLCN7), N-Acetylglucosamine-1-Phosphate Transferase Subunit Gamma (GNPTG), Unk Like Zinc Finger (UNKL), Adenosine Monophosphate Deaminase 2 (AMPD2), GNAI3, WD Repeat Domain 81 (WDR81), and Serpin Family F Member 1 (SERPINF1) and downregulated mRNAs of TATA-Box Binding Protein Associated Factor 12 (TAF12) co-expressed with five crucial lncRNAs. Moreover, GNAI2, GNAI3, GNG12, and vWF were upregulated and GNAS was downregulated in the top 10 hub genes of upregulated and downregulated PPI networks.ConclusionThese findings from integrative biological analysis of lncRNA-mRNA co-expression networks explored the key genes and pathways and provide new insights into the understanding of the mechanism and discovering new therapeutic targets for HCM. Three differentially expressed pivotal lncRNAs (LA16c-312E8.2, RP5-1160K1.3, and MIR22HG) in the co-expression network may serve as biomarkers and intervention targets for the diagnosis and treatment of HCM.

Effect of Rhizoma Drynariae on differential gene expression in ovariectomized rats with osteoporosis based on transcriptome sequencing.

Osteoporosis is increasingly becoming a serious problem affecting the quality of life of the older population. Several experimental studies have shown that Chinese medicine has a definite effect on improving osteoporosis. Based on transcriptome sequencing, we analyzed the differential gene expression and mechanism of the related signaling pathways. Fifteen rats were randomly divided into an experimental group, a model group, and a sham surgery group. The rat model for menopausal osteoporosis was established using an ovariectomy method. One week after modeling, the experimental group was administered(intragastric administration)8.1 g/kg of Rhizoma drynariae, whereas the model and sham groups received 0.9% saline solution twice daily for 12 weeks. Subsequently, the rats were sacrificed, and the left femur of each group was removed for computerized tomography testing, while right femurs were used for hematoxylin and eosin staining. High-throughput RNA sequencing and functional and pathway enrichment analyses were performed. Comparing the gene expression between the experimental and model groups, 149 differential genes were identified, of which 44 were downregulated and 105 were upregulated. The criteria for statistical significance were |log2 Fold Change| > 1 and P < 0.05. Gene ontology analysis showed that the differentially expressed genes were enriched in cell component terms such as cell part and outer cell membrane part, and the genes were associated with cell process, biological regulation, metabolic processes, DNA transcription, and catalytic activity. Enrichment analysis of Kyoto Encyclopedia of Genes and Genomes pathways showed significantly enriched pathways associated with systemic lupus erythematosus, herpes simplex infection, circadian rhythm, vascular smooth muscle contraction, the AGE-RAGE signaling pathway in diabetic complications, and the TNF, Apelin, and Ras signaling pathways. Our results revealed that the Npas2, Dbp, Rt1, Arntl, Grem2, H2bc9, LOC501233, Pla2g2c, Hpgd, Pde6c, and Dner genes, and the circadian rhythm, lipid metabolism, inflammatory signaling pathway, and immune pathways may be the key targets and pathways for traditional Chinese medicine therapy of Rhizoma Drynariae in osteoporosis.

Fecal microbiome transplantation attenuates manganese-induced neurotoxicity through regulation of the apelin signaling pathway by inhibition of autophagy in mouse brain

Manganese (Mn) is a common environmental pollutant. Mn exposure can lead to neurodegenerative diseases resembling Parkinson's disease, and has become a major public health concern. However, the mechanism of Mn-induced neurotoxicity in the brain is not clear. Fecal microbiome transplantation (FMT) may alleviate the neurotoxicity of Mn exposure by remodeling the gut microbiota. In this study, MnCl2 (manganese chloride) was administered to mice as in drinking water (Mn: 200 mg/L), and fecal matter from donor mice was administered by oral gavage every other day to the recipient mice. The Mn exposure model (Mn group) and FMT model (Mn+FMT group) were established and analyzed 5 weeks post-exposure. The Wipi1 gene exhibited the most significant increase associated with Mn exposure and Mn+FMT treatment groups based on transcriptome analysis. Combined analysis of transcriptomics and proteomics demonstrated that the apelin signaling pathway is the main pathway affected by FMT during Mn exposure. Immunofluorescence and Western blot showed that the expression of key proteins (Beclin-1, LC-3B, and PINK1) associated with autophagy in the hippocampus was robustly activated in the Mn exposure group, but attenuation was observed in Mn+FMT mice, suggesting a critical role of autophagy in neurotoxicity induced by Mn exposure. Our research provides evidence for the neurotoxic effects of Mn exposure through autophagy activation and provides an underlying mechanism of FMT protection against Mn-induced neurotoxicity through regulation of the apelin signaling pathway.

Liuwei Dihuang Decoction Alleviates Cognitive Dysfunction in Mice With D-Galactose-Induced Aging by Regulating Lipid Metabolism and Oxidative Stress via the Microbiota-Gut-Brain Axis.

BackgroundAging is an important cause of cognitive dysfunction. Liuwei Dihuang decoction (LW), a commonly applied Chinese medicine formula, is widely used for the treatment of aging-related diseases in China. Previously, LW was confirmed to be effective in prolonging life span and reducing oxidative stress in aged mice. Unfortunately, the underlying mechanism of LW remains unclear. The aim of this study was to interpret the mechanism by which LW alleviates cognitive dysfunction related to aging from the perspective of the microbiota-gut-brain axis.MethodAll C57BL/6 mice (n = 60) were randomly divided into five groups: the control, model, vitamin E (positive control group), low-dose LW and high-dose LW groups (n = 12 in each group). Except for those in the control group, D-galactose was subcutaneously injected into mice in the other groups to induce the aging model. The antiaging effect of LW was evaluated by the water maze test, electron microscopy, 16S rRNA sequencing, combined LC–MS and GC–MS metabolomics, and ELISA.ResultsLiuwei Dihuang decoction ameliorated cognitive dysfunction and hippocampal synaptic ultrastructure damage in aging mice. Moreover, LW decreased Proteobacteria abundance and increased gut microbiota diversity in aging mice. Metabolomic analysis showed that LW treatment was associated with the significantly differential abundance of 14 metabolites, which were mainly enriched in apelin signaling, sphingolipid metabolism, glycerophospholipid and other metabolic pathways. Additionally, LW affected lipid metabolism and oxidative stress in aging mice. Finally, we also found that LW-regulated microbial species such as Proteobacteria and Fibrobacterota had potential relationships with lipid metabolism, oxidative stress and hippocampal metabolites.ConclusionIn brief, LW improved cognitive function in aging mice by regulating lipid metabolism and oxidative stress through restoration of the homeostasis of the microbiota-gut-brain axis.

Genomic landscape and clonal evolution between adenocarcinoma and squamous components in adenosquamous carcinoma of the pancreas.

e16265 Background: Adenosquamous carcinoma of the pancreas (ASCP) is a rare histological subtype of pancreatic cancer with high metastasis rate and poor prognosis, which is characterized by a variable proportion of adenocarcinoma and squamous carcinoma components with at least 30% of the tumor showing squamous differentiation. To get further insight into the tumorigenesis and progression, we profiled the genetic characterization and explored the evolutionary model of this subtype. Methods: A total of 10 patients with ASCP were enrolled. For each sample, glandular and squamous carcinoma tissues were separated by microdissection. Whole exome sequencing for 20 samples with matched peri-carcinomatous tissues were performed by next-generation sequencing. Results: Genetic profiling revealed that top 5 mutated genes of adenocarcinoma and squamous components were highly consistent including TP53 (100% vs 100%), KRAS (100% vs 90%), CDKN2A (40% vs 40%), KDM6A (20% vs 20%) and LRP1B (20% vs 20%). KEGG pathway analysis showed that PI3K-Akt, MAPK and FoxO signaling pathways were the most frequently mutated pathways in adenocarcinoma. In contrast, Apelin and Rap1 signaling pathways were highly mutated in squamous components, advocating that these pathways may play important roles in the transformation of squamous carcinomas. A higher proportion of indels were identified in adenocarcinoma than squamous components (12.1% vs 3%). The chromosome regions of 3p21.3, 9p21.3, 11p15.5, 16p13.11 and 21q22.3 were the most commonly deleted regions in the adenocarcinoma components, while losses of 1p36.21, 3p25.3, 3q13.13, 6p22.3 and 11p15.5 had been observed in the squamous components. Some of these regions had important effects on development and progression of cancer. For example, 9p21.3 had well-established impact on cell proliferation and apoptosis containing two crucial tumor suppressor genes, CDKN2A and CDKN2B. Additionally, the C &gt; T mutation was predominant among the 6 types of single nucleotide variations. Most common signatures were related to aging (Single base substitutions, SBS1), BRCA1/2 mutations and responders to platinum therapy (SBS3), and defective DNA mismatch repair (SBS6). Tumor evolution analysis showed that the glandular and squamous components shared a common origin. There were two evolutionary scenarios in the early stages of tumor evolution, with a third of nine patients ware parallel evolution and the remaining patients were linear evolution. KRAS (100%), TP53 (66.7%) and TTN (33.3%) were frequently identified as clonal mutations. Conclusions: This is the first study to elucidate the genomic alteration as well as the clonal evolution using whole-exome sequencing in ASCP patients. It provides evidence for more comprehensive understanding of pathophysiology of ASCP at genomic level, which could contribute to the amelioration for ASCP patients’ management.

Kynurenine-PARP-1 Link Mediated by MicroRNA 210 May Be Dysregulated in Pulmonary Hypertension.

BackgroundDysregulation of microRNAs is associated with pulmonary hypertension. The present study aimed to determine the alterations in microRNA and microRNA expressions and their role in signaling pathways and investigate the relationship with serum levels of apelin, kynurenine, and endocan in pulmonary hypertension.MethodsThe study design was prospective and single-centered. The study included 32 consecutive treatment-naive patients with precapillary pulmonary hypertension and 55 age and sex-matched healthy controls. All subjects underwent right heart catheterization. mRNA expressions of hypoxia-inducible factor-1 alpha, hypoxia-inducible factor-2 alpha, signal transducer and activator of transcription-3, fibroblast growth factor-2, fibroblast growth factor receptor-1, and poly-ADP-ribose polymerase-1 and microRNA expressions of miRNA-210, miRNA-130a, miRNA-424, miRNA-204, and miRNA-223 were determined by RT-PCR. Concentrations of kynurenine, apelin, and endocan were analyzed by ELISA.ResultsmRNA expressions of hypoxia-inducible factor-2 alpha, signal transducer and activator of transcription-33, and FGF-2 were increased; miRNA-210 and miRNA-130a were increased; miRNA-223 and miRNA-204 were decreased in pulmonary hypertension. Apelin and kynurenine concentrations were decreased in pulmonary hypertension. There were positive correlations: hypoxia-inducible factor-2 alpha-miRNA-424, Apelin-miRNA-424, kynurenine-miRNA-210, signal transducer and activator of transcription-3-PVR, miRNA-210-right atrial pressure, and kynurenine-right atrial pressure. There were negative correlations: poly-ADP-ribose polymerase-1-miRNA-210 and poly-ADP-ribose polymerase-1-right atrial pressure. On multiple logistic regression analyses, miRNA-130a and Apelin were independent risk factors for PH.ConclusionsWe report a novel relationship between the kynurenine and poly-ADP-ribose polymerase-1 signaling pathways that could be mediated by miRNA-210. We also report a connection between the Apelin and hypoxia-inducible factor-2 alpha signaling pathways that could be mediated by miRNA-424. Reduced levels of Apelin and elevated levels of miRNA-130a are associated with pulmonary hypertension. We also find that elevated levels of signal transducer and activator of transcription-3, miRNA-210, and kynurenine and reduced levels of poly-ADP-ribose polymerase-1 correlate with more severe hemodynamics.

HOXC6/8/10/13 predict poor prognosis and associate with immune infiltrations in glioblastoma

BackgroundGlioblastoma (GBM), characterized by deregulated cell proliferation and immune cells infiltration, is a common and lethal tumor of the central nervous system. Recently, the infiltration of immune cells has attracted attention as a potential novel GBM immunotherapy option. Homeobox C cluster (HOXC) is an evolutionarily conserved family of transcriptional factors that are involved in embryogenesis and tumorigenesis. Nevertheless, the correlations of HOXCs with the prognosis and immune infiltration of GBM remain blurred. MethodsThe RNA-seq data with corresponding clinical characteristics were downloaded from TCGA and GTEx databases. The correlations between HOXCs and clinical characteristics were calculated using univariable and multivariate Cox regression. R language with ggplot2, survminer, survival, GSVA, and pROC packages were employed to analyze the data and present the plots. MethSurv, UALCAN and cBioPortal were employed to evaluate the DNA methylation and mutation status of HOXCs in GBM. We also verified the expression and prognosis of HOXCs by qPCR and immunohistochemistry in a cohort of 36 patients. ResultsWe identified that HOXC6/8/10/13 were crucial biomarkers for diagnosis and prognostic judgement in GBM. Gene set variation analysis revealed that levels of expression of HOXCs were associated with the infiltration of various immune cells. The qPCR and immunohistochemistry data validated the prognostic values of HOXC6/8/10/13 in GBM. Finally, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that HOXCs might be involved in DNA-binding transcription activator activity and the apelin signaling pathway. ConclusionThis research highlights that HOXC6/8/10/13 are involved in the immune infiltrates, also provide potential clinical utility as therapeutic targets in GBM.

Fractionated Seminal Plasma of Boar Ejaculates Analyzed by LC-MS/MS: Its Effects on Post-Thaw Semen Quality.

This study aimed to characterize the protein composition of fractionated seminal plasma (SP) by liquid chromatography mass spectrometry (LC–MS/MS) analysis and investigate its effects on survival of frozen-thaw (FT) boar spermatozoa following storage. Seminal plasma (SP) was fractionated by gel filtration chromatography to give two fractions, SP1 with more than 40 kDa (>40 kDa) and SP2 with less than 40 kDa (<40 kDa). SP1 and SP2 were subjected to LC–MS/MS and bioinformatics analysis. Following cryopreservation, FT boar semen (n = 7) was thawed in Beltsville Thawing Solution (BTS), BTS + SP1 or BTS + SP2, stored at different periods and subjected to post-thaw (PT) quality assessment. A total of 52 and 22 abundant proteins were detected in SP1 and SP2, respectively. FN1, ANGPTL1, and KIF15 proteins were more abundance in SP1, whereas a high abundance of spermadhesins (PSP-I and PSP-II) was detected in SP2. Proteins of the fractionated SP were involved in various biological processes, such as cell motility and signal transduction. The dominant pathway of SP1 proteins was the apelin signaling pathway (GNA13, MEF2D, SPHK2, and MEF2C), whereas a pathway related to lysosome (CTSH, CTSB, and NPC2) was mainly represented by SP2 proteins. In most of the boars, significantly higher motility characteristics, membrane integrity, and viability were observed in FT spermatozoa exposed to SP1 or SP2 compared with BTS. The results of our study confirm that a combination of several proteins from the fractionated SP exerted beneficial effects on the sperm membrane, resulting in improved quality characteristics following PT storage.

Metabolic endophenotype associated with right ventricular glucose uptake in pulmonary hypertension.

Alterations in metabolism and bioenergetics are hypothesized in the mechanisms leading to pulmonary vascular remodeling and heart failure in pulmonary hypertension (PH). To test this, we performed metabolomic analyses on 30 PH individuals and 12 controls. Furthermore, using 2-[18F]fluoro-2-deoxy-D-glucose positron emission tomography, we dichotomized PH patients into metabolic phenotypes of high and low right ventricle (RV) glucose uptake and followed them longitudinally. In support of metabolic alterations in PH and its progression, the high RV glucose group had higher RV systolic pressure (p < 0.001), worse RV function as measured by RV fractional area change and peak global longitudinal strain (both p < 0.05) and may be associated with poorer outcomes (33% death or transplantation in the high glucose RV uptake group compared to 7% in the low RV glucose uptake group at five years follow-up, log-ranked p = 0.07). Pathway enrichment analysis identified key metabolic pathways including fructose catabolism, arginine-nitric oxide metabolism, tricarboxylic acid cycle, and ketones metabolism. Integrative human protein-protein interactome network analysis of metabolomic and transcriptomic data identified key pathobiological pathways: arginine biosynthesis, tricarboxylic acid cycle, purine metabolism, hypoxia-inducible factor 1, and apelin signaling. These findings identify a PH metabolomic endophenotype, and for the first time link this to disease severity and outcomes.