Derivation and Comprehensive Analysis of Aging Patterns in Patients with Bladder Cancer

Background. Aging is an essential risk factor for cancer. However, aging-related genes (ARGs) have not been comprehensively analyzed in bladder cancer (BC). Therefore, the study is aimed at derivating a risk stratification system for BC patients based on ARGs. Methods. Public databases were used to acquire ARGs sets, transcriptome files, and clinical data. The “limma” package was then used to screen for differential ARGs while also using univariate Cox regression analysis to explore for prognostic ARGs. The “ConsensusClusterPlus” package was used to perform aging patterns in BC patients based on the above prognostic ARGs. Subsequently, aging patterns were investigated in survival prediction, mutation landscape, immunotherapy, immunological checkpoints, and immune microenvironment. We likewise utilized gene enrichment analysis to explore the biological functions that were behind the findings. To construct a risk signature and nonogram for prognostic prediction, we used LASSO and Cox regression analysis based on differential genes in aging patterns. In addition, we plotted a nomogram and validate the accuracy of the risk signature in GEO and TCGA cohorts. We explored the possible biological mechanism using GSEA analysis and preliminarily identified a hub gene using PPI network. Finally, we validated the expression of hub gene in BC cell lines. Results. We screened 84 downregulated ARGs, 74 upregulated ARGs, and 32 prognostic ARGs in the human aging genome resource. The aging patterns based on prognostic genes had excellent survival prediction ( p < 0.001 ) and discriminatory ability in 405 BC patients. In addition, we found no significant differences in aging patterns in mutation analysis, which were all characterized by TP53, TTN, and KMT2D mutations. It is worth noting that cluster B in the aging patterns has a better response to immunotherapy and a more active immune microenvironment ( p < 0.05 ). In addition, gene enrichment analysis showed that aging patterns may be related to biological processes such as Staphylococcus aureus infection, phagosome, and cytokine-cytokine receptor interaction. Subsequently, we constructed a risk signature based on 16 differential genes from different aging patterns and had good survival prediction ability in both GEO and TCGA cohort. Specifically, survival analysis revealed a significantly shorter survival time in the high-risk group than in the low-risk group (TCGA and GEO, p < 0.001 ). In addition, AUC values in the ROC analysis predicted 1, 3, and 5 years in TCGA cohort that are 0.713, 0.714, and 0.738, respectively. AUC values predicted 1, 3, and 5 years in GEO cohort that are 0.606, 0.663, and 0.718, respectively. There is no doubt that risk score was an independent prognostic factor from results of multivariate Cox regression analysis in BC patients ( p < 0.001 ). There were also significant differences in immune cell infiltration, immune checkpoint, and immune score between the two groups ( p < 0.05 ), but it should not be ignored that the correlation with the HLA expression was weak. Finally, we identified and validated CLIC3 as a hub gene that may be involved in the Wnt signaling pathway, etc. Conclusion. We provided robust evidences that aging patterns based on ARGs can guide targeted therapy and survival prediction in BC patients.

Bioinformatics Analysis Predicts ceRNA Regulatory Axes Related to Melanoma Pathogenesis

Background Melanoma is a highly malignant skin tumour, with an incidence and mortality rates accounting for approximately 5% and >75% of all tumours, respectively. In the present study, we aimed to screen mRNAs, microRNAs, and circRNAs related to the pathogenesis of melanoma via bioinformatics methods. Materials and methods The microarray data correlating with melanoma were obtained from the GEO database, and the differentially expressed genes between melanomatissues and non-melanoma tissues were screened using GEO2R. Moreover, the Hub genes were screened using the STRING database, Cytoscape software, and GEPIA database. The DAVID database was used for gene enrichment analysis. Thereafter, the ceRNA network diagram was constructed using the starBase database and was analysed for survival in order to predict the molecular markers for diagnosis and treatment of skin cutaneous melanoma; the most probable ceRNA mechanism was screened via pan-cancer analysis. Furthermore, the ceRNA network was verified using the GEPIA and UALCAN databases. Results In total, 266, 20, and 10 differentially expressed mRNAs, microRNAs, and circRNA10 were screened. Using Cytoscape software, 59 Hub and 31 key genes were screened, followed by construction of the ceRNA and ceRNA-gene enrichment analysis networks. Conclusion Clinical verification revealed that SIPA1L1 could regulate the expression of DSC3 by sponge adsorption of has-miR-106b and has-miR-20b, and thereby affect the pathogenesis and progression of skin cutaneous melanoma.

Genome-wide association study reveals susceptibility loci for self-reported headache in a large community-based Asian population

Background The genetic substrate for headache in the general population has not been identified in Asians. We investigated susceptible genetic variants for self-reported headache in a large community-based Asian population. Methods We conducted a genome-wide association study in participants recruited from a community-based cohort to identify the genetic variants associated with headache in Taiwanese. All participants received a structured questionnaire for self-reported headache. A total of 2084 patients with “self-reported headache” and 11,822 age- and sex-matched controls were enrolled. Gene enrichment analysis using the Genotype-Tissue Expression version 6 database was performed to explore the potential function of the identified variants. Results We identified two novel loci, rs10493859 in TGFBR3 and rs13312779 in FGF23, that are functionally relevant to vascular function and migraine to be significantly associated with self-reported headache after adjusting age, sex and top 10 principal components ( p = 8.53 × 10−11 and p = 1.07 × 10−8, respectively). Gene enrichment analysis for genes with GWAS suggestive significance ( p < 10−6) demonstrated that the expression of these genes was significantly enriched in the artery ( p = 8.18 × 10−4) and adipose tissue ( p = 8.95 × 10−4). Conclusion Our results suggest that vascular dysfunction might play important roles in the pathogenesis of self-reported headache in Asian populations.

Cluster analysis of transcriptomic datasets to identify endotypes of Idiopathic Pulmonary Fibrosis

Rationale: Considerable clinical heterogeneity in Idiopathic Pulmonary Fibrosis (IPF) suggests the existence of multiple disease endotypes. Identifying these endotypes could allow for a biomarker-driven personalised medicine approach in IPF. Objectives: To improve our understanding of the pathogenesis of IPF by identifying clinically distinct groups of patients with IPF that could represent distinct disease endotypes. Methods: We co-normalised, pooled and clustered three publicly available blood transcriptomic datasets (total 220 IPF cases). We compared clinical traits across clusters and used gene enrichment analysis to identify biological pathways and processes that were over-represented among the genes that were differentially expressed across clusters. A gene-based classifier was developed and validated using three additional independent datasets (total 194 IPF cases). Measurements and main results: We identified three clusters of IPF patients with statistically significant differences in lung function (P=0.009) and mortality (P=0.009) between groups. Gene enrichment analysis implicated dysregulation of mitochondrial homeostasis, apoptosis, cell cycle and innate and adaptive immunity in the pathogenesis underlying these groups. We developed and validated a 13-gene cluster classifier that predicted mortality in IPF (high-risk clusters vs low-risk cluster: hazard ratio= 4.25, 95% confidence interval= [2.14, 8.46], P=3.7×10-5). Conclusions: We have identified blood gene expression signatures capable of discerning groups of IPF patients with significant differences in survival. These clusters could be representative of distinct pathophysiological states, which would support the theory of multiple endotypes of IPF. Although more work must be done to confirm the existence of these endotypes, our classifier could be a useful tool in patient stratification and outcome prediction in IPF.

Differentially Expressed Circular Non-coding RNAs in Atherosclerotic Aortic Vessels and Their Potential Functions in Endothelial Injury

Background: Circular non-coding RNA (circRNA) has a variety of biological functions. However, the expression profile and potential effects of circRNA on atherosclerosis (AS) and vascular endothelial injury have not been fully elucidated. This study aims to identify the differentially expressed circRNAs in atherosclerotic aortic vessels and predict their potential functions in endothelial injury.Method: ApoE-/- mice were fed with high-fat diet for 12 weeks to induce AS. Atherosclerotic plaques were evaluated by H&amp;E and Masson staining and immunohistochemistry; differentially expressed circRNAs were detected by Arraystar Circular RNA Microarray and verified by RT-PCR; the potential target mircoRNAs of circRNAs were predicted by miRanda, Tarbase, Targetscan and their expression changes were verified by RT-PCR; the potential target genes of mircoRNAs were predicted by Targetscan and verified by Western blot; the signaling pathways that they might annotate or regulate and their potential functions in vascular endothelial injury were predicted by gene enrichment analysis.Results: Fifty two circRNAs were up-regulated more than twice and 47 circRNAs were down-regulated more than 1.5 times in AS aortic vessels. Mmmu_circRNA_36781 and 37699 were up-regulated both in AS aortic vessels and H2O2-treated mouse aortic endothelial cells (MAECs). The expression of miR-30d-3p and miR-140-3p, the target microRNA of circRNA_37699 and circRNA_36781, were downregulated both in AS vessels and H2O2-treated MAECs. On the contrary, MKK6 and TP53RK, the potential target gene of miR-140-3p and miR-30d-3p, were upregulated both in AS aortic roots and H2O2-treated MAECs. Besides, gene enrichment analysis showed that MAPK and PI3K-AKT signaling pathway were the most potential signaling pathways regulated by the differentially expressed circRNAs in atherosclerosis.Conclusions: Mmu_circRNA_36781 (circRNA ABCA1) and 37699 (circRNA KHDRBS1) were significantly up-regulated in AS aortic vessels and H2O2-treated MAECs. They have potential regulatory effects on atherosclerosis and vascular endothelial injury by targeting miR-30d-3p-TP53RK and miR-140-3p-MKK6 axis and their downstream signaling pathways.

Network pharmacology-based elucidation of the molecular mechanism underlying the anti-migraine effect of Asari Radix et Rhizoma

Purpose: To determine the molecular mechanism involved in the anti-migraine effect of Asari Radix et Rhizoma (ARR) using network pharmacology. Methods: The compounds present in ARR were identified through information retrieval from literature and public databases, and were screened based on absorption, distribution, metabolism, excretion and toxicity. Target genes related to the selected compounds and migraine were identified or predicted from public databases. Hub genes in ARR against migraine were identified through analysis of interactions in overlapping genes between compounds and migraine target genes, based on STRING database. Gene enrichment analysis of overlapping genes was performed using Database for Annotation, Visualization and Integrated Discovery. Results: A total of 138 compounds were selected as potential bioactive compounds in ARR. Target genes related to the selected compounds (611 genes) and migraine (278 genes) were obtained, including 71 overlapping genes. The hub genes in the anti-migraine effect of ARR were BDNF, IL6, COMT, APP and TNF. Gene enrichment analysis showed the top 10 biological processes or pathways involved in the mechanism of anti-migraine action of ARR. The tissue source of the overlapping genes was not limited to the brain. The results from gene enrichment analysis revealed that the effect of ARR on migraine was holistic, which is characteristic of traditional Chinese medicines. Conclusion: Network pharmacology has been used to decipher the molecular mechanism involved in the action of ARR against migraine. The results provide a scientific basis for the clinical effect of ARR on migraine.

Draft Genome of the Korean smelt Hypomesus nipponensis and its transcriptomic responses to heat stress in the liver and muscle

Pond smelt (Hypomesus nipponensis) is a cold-freshwater fish species as a winter economic resource of aquaculture in South Korea. Due to its high susceptibility to abnormal water temperature from global warming, a large number of smelt die in hot summer. Here, we present the first draft genome of H. nipponensis and transcriptomic changes in molecular mechanisms or intracellular responses under heat stress. We combined Illumina and PacBio sequencing technologies to generate the draft genome of H. nipponensis. Based on the reference genome, we conducted transcriptome analysis of liver and muscle tissues under normal (NT, 5 °C) versus warm (HT, 23 °C) conditions, to identify heat stress-induced genes and gene categories. We observed a total of 1,987 contigs, with N50 of 0.46 Mbp with a largest contig (3.03 Mbp) in the assembled genome. A total number of 20,644 protein coding genes were predicted, and 19,224 genes were functionally annotated: 15,955 genes for Gene Ontology (GO) terms; and 11,560 genes for KEGG Orthology (KO). We conducted the lost and gained genes analysis compared with three species that human, zebrafish and salmon. In the lost genes analysis, we detected smelt lost 4,461 (22.16%), 2,825 (10.62%), and 1,499 (3.09%) genes compare with above three species, respectively. In the gained genes analysis, we observed smelt gain 1,133 (5.49%), 1,670 (8.09%), and 229 (1.11%) genes compare with above species, respectively. From transcriptome analysis, a total of 297 and 331 differentially expressed genes (DEGs) with False discovery rate (FDR) &lt; 0.05 were identified in the liver and muscle tissues, respectively. Gene enrichment analysis of DEGs indicates that up-regulated genes were significantly enriched for lipid biosynthetic process (GO : 0008610, P &lt; 0.001) and regulation of apoptotic process (GO : 0042981, P &lt; 0.01), and down-regulated genes by immune responses such as myeloid cell differentiation (GO : 0030099, P &lt; 0.001) in the liver under heat stress. In muscle tissue, up-regulated genes were enriched for hypoxia (GO : 0001666, P &lt; 0.05), transcription regulator activity (GO : 0140110, P &lt; 0.001) and calcium-release channel activity (GO : 0015278, P &lt; 0.01), and down-regulated genes for nicotinamide nucleotide biosynthetic process (GO : 0019359, P &lt; 0.01). The results of KEGG pathway analysis were similar to that of gene enrichment analysis. The draft genome and transcriptomic of H. nipponensis will be used as a useful genetic resource for functional and evolutionary studies. Our findings will improve understanding of the molecular mechanisms and heat responses and will be useful for predicting survival of the smelt and its closely related species under global warming.

Divergent transcriptomic profiles in skeletal muscle of diabetics with and without heart failure

Funding Acknowledgements Type of funding sources: None. Background Patients with type 2 diabetes mellitus (DM) that have coexistent heart failure (HF) have exacerbated symptoms and prognosis, however beside cardiac dysfunction the mechanisms governing these features are incompletely understood. Evidence indicates abnormalities in the periphery could contribute to this worse clinical phenotype, including a role for skeletal muscle whereby disturbances in the transcriptome could disrupt muscle homeostasis/repair to offer a novel therapeutic approach. Purpose Is the skeletal muscle transcriptome distinguishable between DM patients with and without HF? Methods DM patients without (n = 11) or with HF with reduced left ventricular ejection fraction (LVEF) (n = 16) were included. Muscle biopsies were collected from the pectoralis major during pacemaker implantation. Following RNA extraction and cDNA synthesis, non-bias RNA sequencing (RNAseq) was performed (Cambridge Genomic Services, UK) followed by targeted RT-PCR gene expression of relevant targets. DESeq2 identified differentially expressed genes (DEGs) with a false discovery rate (p &lt; 0.05). Gene enrichment analysis was performed with clusterProfiler v3.16.0 to interrogate the gene ontology database, while pathway analysis was conducted using ReactomePA v1.32.0 to interrogate the Reactome database, using an adjusted p value. Values of p &lt; 0.05 were accepted as significant. Results Groups were not different (p &gt; 0.05) for age (74 ± 11 vs. 66 ± 10 years), BMI (31 ± 7 vs 29 ± 6), sex (n = 2 females per group), or HbA1c (56 ± 10 vs. 57 ± 8 mmol/mol), although LVEF was lower in the group with HF (27 ± 8 vs. 54 ± 2%; p &lt; 0.05). Of the 19,544 genes analysed, RNAseq identified 53 DEGs between DM patients with and without HF, with several relevant targets related to myofiber homeostasis such as autophagy (RUBCN), protein synthesis (DGKζ), and inflammation/apoptosis (TLE1). Follow-up RT-PCR analysis confirmed a trend towards upregulation of the autophagy-related machinery p62 (p = 0.043) and BNIP3 (p = 0.085) in the HF group, but not ubiquitin-proteasome (MuRF1, MAFbx; p &gt; 0.05). Gene-enrichment analysis of DEGs identified 7 overrepresented terms (P &lt; 0.05), including lipid metabolism/signalling alongside epigenetic modifications related to histone deacetylases (HDAC6/10). Furthermore, pathway analysis identified 4 terms (p &lt; 0.05) related to NOTCH signalling and phosphatidyl inositol-bisphosphate (PIP2) hydrolysis thus indicating alterations to muscle repair and lipid signalling respectively. Conclusion(s): This study confirms that DM patients with and without HF demonstrate distinct skeletal muscle transcriptome profiles. Key differences related to skeletal muscle myogenesis, autophagy, epigenetic regulation, and lipid signalling were identified that could form part of important therapeutic targets. Whether these underlying muscle transcriptome differences contribute to poorer clinical outcomes in DM patients with HF remains to be determined.

DiseaseLinc: Disease Enrichment Analysis of Sets of Differentially Expressed LincRNAs

Long intergenic non-coding RNAs (LincRNAs) are long RNAs that do not encode proteins. Functional evidence is lacking for most of them. Their biogenesis is not well-known, but it is thought that many lincRNAs originate from genomic duplication of coding material, resulting in pseudogenes, gene copies that lose their original function and can accumulate mutations. While most pseudogenes eventually stop producing a transcript and become erased by mutations, many of these pseudogene-based lincRNAs keep similarity to the parental gene from which they originated, possibly for functional reasons. For example, they can act as decoys for miRNAs targeting the parental gene. Enrichment analysis of function is a powerful tool to discover the functional effects of a treatment producing differential expression of transcripts. However, in the case of lincRNAs, since their function is not easy to define experimentally, such a tool is lacking. To address this problem, we have developed an enrichment analysis tool that focuses on lincRNAs exploiting their functional association, using as a proxy function that of the parental genes and has a focus on human diseases. The tool is available at: http://cbdm-01.zdv.uni-mainz.de:3838/piyusmor/DiseaseLinc/.

Draft Genome of the Korean smelt (Hypomesus nipponensis) and its transcriptomic responses to heat stress in the liver and muscle

Backgrounds: Hypomesus nipponensis is a cold-freshwater fish species as a winter economic resource of aquaculture in South Korea. Due to its high susceptibility to abnormal water temperature from global warming. Aim of the study: Here, we present the first draft genome of H. nipponensis and transcriptomic changes in molecular mechanisms or intracellular responses under heat stress. Methods: We combined Illumina and PacBio sequencing technologies to generate the draft genome of H. nipponensis. Based on the reference genome, we conducted transcriptome analysis of liver and muscle tissues under normal (NT, 5°C) versus warm (HT, 23°C) conditions, to identify heat stress-induced genes and gene categories. Results: We observed a total of 1,987 contigs, with N50 of 0.46 Gbp with a largest contig (3.03 Mbp) in the assembled genome. A total number of 20,644 protein coding genes were predicted, and 19,224 genes were functionally annotated: 15,955 genes for Gene Ontology (GO) terms; and 11,560 genes for KO (KEGG Orthology). In the missing and gained genes analysis, we detected 4,461 (22.16%), 2,825 (10.62%), and 1,499 (3.09%) genes with coverage less than 10% and for gained genes found from alignment to other species that human, zebrafish and salmon, respectively, we observed 1,133 (5.49%), 1,670 (8.09%), and 229 (1.11%) genes with coverage less than 10% compared with above species, respectively. From transcriptome analysis, a total of 297 and 331 differentially expressed genes (DEGs, adjusted p-value < 0.05) were identified in the liver and muscle tissues, respectively. Gene enrichment analysis of DEGs indicates that up-regulated genes were significantly enriched for lipid biosynthetic process (GO:0008610, P < 0.001) and regulation of apoptotic process (GO:0042981, P < 0.01), and down-regulated genes by immune responses such as myeloid cell differentiation (GO:0030099, P < 0.001) in the liver under heat stress. In muscle tissue, up-regulated genes were enriched for hypoxia (GO:0001666, P < 0.05), transcription regulator activity (GO:0140110, P < 0.001) and calcium-release channel activity (GO:0015278, P < 0.01), and down-regulated genes for nicotinamide nucleotide biosynthetic process (GO:0019359, P < 0.01). The results of KEGG pathway analysis were similar to that of gene enrichment analysis. Conclusion: The draft genome and transcriptomic of H. nipponensis will be used as a useful genetic resource for functional and evolutionary studies. Our findings will improve understanding of the molecular mechanisms and heat responses and be useful for better survival of the smelt and its closely related species under global warming.