Over-represented Pathways Research Articles

Polycystic Ovary Syndrome and Ferroptosis: Following Ariadne's Thread.

Background: Recent literature suggests that ferroptosis (FPT) may be a key player in polycystic ovary syndrome (PCOS) pathogenesis, but the underlying mechanism(s) remain(s) unclear. Aim: Therefore, herein, we made an effort to reproduce the molecular signature of the syndrome by including FPT and exploring novel drug targets for PCOS. Methods: (a) Our previously constructed PCOS interactions molecular network was extended with the addition of FPT-associated genes (interaction score above 0.7) and (b) gene set enrichment analysis was performed so as to detect over-represented KEGG pathways. Results: The updated interactome includes 140 molecules, 20 of which are predicted/novel, with an interaction score of 7.3, and 12 major hubs. Moreover, we identified 16 over-represented KEGG pathways, with FPT being the most overexpressed pathway. The FPT subnetwork is connected with the PCOS network through KDM1A. Conclusions: FPT cell death is involved in PCOS development, as its major hub TP53 was shown to be the most important hub in the whole PCOS interactome, hence representing a prioritized drug target.

The lung proteome in HIV-associated obstructive lung disease (OLD)

RationaleObstructive lung disease is increasingly common among persons living with HIV (PLWH). There are currently no validated biomarkers that identify individuals at risk of developing obstructive lung disease (OLD) and specific mechanisms contributing to HIV-associated OLD remain elusive, independent of smoking.ObjectivesWe sought to identify biomarkers and biological pathways associated with OLD using a broad proteomic approach.MethodsWe performed tandem mass tagging and mass spectrometry (MS) analysis on bronchoalveolar lavage fluid (BALF) samples from persons living with HIV with OLD (n=26) and without OLD (n=26). We combined untargeted MS with a targeted SomaScan aptamer-based approach. We used Pearson correlation tests to identify associations between each protein and lung function (FEV1pp). We adjusted for multiple comparisons using a false discovery rate (FDR) adjustment. Significant proteins were entered into a pathway overrepresentation analysis. Protein-driven endotypes were constructed using K-means clustering.Measurements and main resultsWe identified over 3800 proteins by MS and identified 254 proteins that correlated with FEV1pp when we combined the MS and SomaScan proteomes when adjusting for smoking status. Pathway analysis revealed cell adhesion molecules as significant.ConclusionsProtein expression differs in the lung of PWH and decreased lung function (FEV1pp). Pathway analysis reveals cell adhesion molecules having potentially important roles in this process.

Tick extracellular vesicles undermine epidermal wound healing during hematophagy.

Wound healing has been extensively studied through the lens of inflammatory disorders and cancer, but limited attention has been given to hematophagy and arthropod-borne diseases. Hematophagous ectoparasites, including ticks, subvert the wound healing response to maintain prolonged attachment and facilitate blood-feeding. Here, we unveil a strategy by which extracellular vesicles (EVs) ensure blood-feeding and arthropod survival in three medically relevant tick species. We demonstrate through single cell RNA sequencing and murine genetics that wildtype animals infested with EV-deficient Ixodes scapularis display a unique population of keratinocytes with an overrepresentation of pathways connected to wound healing. Tick feeding affected keratinocyte proliferation in a density-dependent manner, which relied on EVs and dendritic epidermal T cells (DETCs). This occurrence was linked to phosphoinositide 3-kinase activity, keratinocyte growth factor (KGF) and transforming growth factor β (TGF-β) levels. Collectively, we uncovered a strategy employed by a blood-feeding arthropod that impairs the integrity of the epithelial barrier, contributing to ectoparasite fitness.

PSIII-2 Exploring differential gene expression and pathways associated with pregnancy outcomes in beef heifers

Abstract Heifer fertility is a key factor in the success of beef production systems, as the reproductive performance of a heifer correlates with her stayability in the breeding herd. However, despite being reproductively mature and physically healthy, 5 to 15% of these heifers fail to conceive within a designated breeding season. The underlying physiological mechanisms preventing these heifers from achieving pregnancy remain elusive. Therefore, this study aimed to determine differentially expressed genes (DEGs) and biological pathways associated with pregnancy outcomes in beef heifers. Angus-Simmental crossbred heifers were subjected to an estrus synchronization and fixed-time artificial insemination (AI) program (7-d CO-Synch + CIDR) and blood samples were collected at the time of AI for isolation of peripheral white blood cells (PWBC). After pregnancy check, heifers were classified as pregnant (P) and non-pregnant (NP). Based on that, 6 animals per group were randomly selected for total RNA isolation from the PWBC samples, and genome-wide expression was measured through RNA sequencing. The RNA-Seq data were analyzed using an in-house bioinformatics pipeline, with filtered reads aligned to the Bos taurus reference genome using STAR. DEGs were identified using DESeq2, followed by functional analysis using ShinyGO. Our approach unveiled 230 DEGs, comprising 102 up and 128 downregulated genes in the NP group (P-value ≤ 0.05 and |log2FC| > 0.5). Upregulated genes included those from the cytochrome P450 superfamily (CYP27B1 and CYP2U1). The genes SCML2, USIG2, and FBP1 were upregulated and have been associated with immune response and increased PWBC counts. This may be related to the over-representation of biological processes involved in cell division based on the upregulated genes. KEGG pathway over-representation analysis of DEGs retrieved allograft rejection, graft-versus-host disease, type I Diabetes Mellitus, insulin signaling pathway, and transcriptional misregulation of cancer. These pathways included genes such as BOLA-DQB, PRKAR1B, CALML4 and JUP, which were reported as downregulated during times of inflammatory response. Another downregulated gene was GZMB, which is needed for proper cell targeting during immune response. This finding may provide a link between immune function and early pregnancy failure. However, the relationship between the immune system and pregnancy warrants further investigation. Our results shed light on the molecular mechanisms underlying reproductive performance in beef heifers and support previous studies associating immune function with pregnancy outcomes. These differentially expressed genes may be further explored as potential avenues for enhancing fertility in beef production systems.

Peritoneal infection after colorectal cancer surgery induces substantial alterations in postoperative protein levels: an exploratory study

PurposePeritoneal infection, due to anastomotic leakage, after resection for colorectal cancer have been shown to associate with increased cancer recurrence and mortality, as well as cardiovascsular morbidity. Alterations in circulating protein levels could help shed light on the underlying mechanisms, prompting this exploratory study of 64 patients operated for colorectal cancer with anastomosis.MethodsThirty-two cases who suffered a postoperative peritoneal infection were matched with 32 controls who had a complication-free postoperative stay. Proteins in serum samples at their first postoperative visit and at one year after surgery were analysed using proximity extension assays and enzyme-linked immunosorbent assays. Multivariate projection methods, adjusted for multiple testing, were used to compare levels between groups, and enrichment and network analyses were performed.ResultsSeventy-seven proteins, out of 270 tested, were differentially expressed at a median sampling time of 41 days postoperatively. These proteins were all normalised one year after surgery. Many of the differentially expressed top hub proteins have known involvement in cancer progression, survival, invasiveness and metastasis. Over-represented pathways were related to cardiomyopathy, cell-adhesion, extracellular matrix, phosphatidylinositol-3-kinase/Akt (PI3K-Akt) and transforming growth factor beta (TGF-β) signaling.ConclusionThese affected proteins and pathways could provide clues as to why patients with peritoneal infection might suffer increased cancer recurrence, mortality and cardiovascular morbidity.

Impact of vaccination with the Anaplasma phagocytophilum MSP4 chimeric antigen on gene expression in the rabbit host

There are currently no vaccines available to prevent and control of Anaplasma phagocytophilum, an intracellular bacterial pathogen transmitted by ticks that occurs in many regions of the world and causes disease in a wide range of domestic and wild hosts, including humans. Vaccines induce long-lasting immunity and could prevent or reduce transmission of this pathogen.Understanding how vaccines induce a protective response can be difficult due to the complexity of the immune system, which operates at many levels throughout the organism. New perspectives in vaccinology, based on systems biology approaches, integrate many scientific disciplines to fully understand the biological responses to vaccination, where a transcriptomic approach could reveal relevant information of the host immune system, allowing profiling for rational design of vaccine formulations, administration, and potential protection.In the present study we report the gene expression profiles by RNA-seq followed by functional analysis using whole blood samples from rabbits immunized with a recombinant chimeric protein containing peptides from the MSP4 protein of A. phagocytophilum, which showed satisfactory results in terms of potential protection. Transcriptomic analysis revealed differential expression of 720 genes, with 346 genes upregulated and 374 genes downregulated. Overrepresentation of biological and metabolic pathways correlated with immune response, protein signaling, cytoskeleton organization and protein synthesis were found. These changes in gene expression could provide a complete and unique picture of the biological response to the epitope candidate vaccine against A. phagocytophilum in the host.

IL-17 is associated with disease severity and targetable inflammatory processes in heart failure.

Heart failure (HF) is recognized as an inflammatory disease in which cytokines play an important role. In animal HF models, interleukin-17A (IL-17) has been linked to deterioration of cardiac function and fibrosis, whereas knock-out of IL-17 showed beneficial cardiac effects. However, there is limited evidence of IL-17 involvement in patients with HF. This study aims to investigate the clinical characteristics, outcomes, and pathophysiological processes associated with circulating IL-17 concentrations in patients with HF. IL-17 was measured by ELISA in 2082 patients diagnosed with HF along with 363 circulating proteins using proximity extension assay technology for differential expression and pathway analysis. Data were validated in an independent cohort of 1737 patients with HF. Patients with elevated IL-17 concentrations had more severe HF, as reflected by more frequent current or previous hospitalizations for HF, higher New York Heart Association functional class (NYHA) and higher levels of N-terminal pro-B-type natriuretic peptide (NT-proBNP). High IL-17 concentrations were independently associated with an increased risk of hospitalization for HF and mortality. In both cohorts, the most strongly up-regulated proteins in patients with high IL-17 were fibroblast growth factor 21 (FGF-21), interleukin-6 (IL-6), C-X-C motif chemokine ligand 13 (CXCL13), tumour necrosis factor receptor superfamily member 6B (TNFRSF6B) and interleukin-1 receptor antagonist (IL-1RA). Pathway over-representation analysis showed increased activity of pathways related to lymphocyte-mediated immunity, leukocyte activation and regulation of the immune response. In patients with HF, elevated IL-17 concentrations indicate more severe HF and increased activity of inflammatory processes known to be involved in the pathophysiology of HF. IL-17 might hold potential for identifying and targeting inflammation in HF.

Gene co-expression network analysis for porcine intramuscular fatty acid composition

In pigs, meat quality depends markedly on the fatty acid (FA) content and composition of the intramuscular fat, which is partly determined by the gene expression in this tissue. The aim of this work was to identify the link between muscle gene expression and its FA composition. In an (Iberian × Duroc) × Duroc backcrossed pig population, we identified modules of co-expressed genes, and correlation analyses were performed for each of them versus the phenotypes, finding four relevant modules. Two of the modules were positively correlated with saturated FAs (SFAs) and monounsaturated FAs (MUFAs), while negatively correlated with polyunsaturated FAs (PUFAs) and the omega-6/omega-3 ratio. The gene-enrichment analysis showed that these modules had over-representation of pathways related with the biosynthesis of unsaturated FAs, the Peroxisome proliferator-activated receptor signalling pathway and FA elongation. The two other relevant modules were positively correlated with PUFA and the n-6/n-3 ratio, but negatively correlated with SFA and MUFA. In this case, they had an over-representation of pathways related with fatty and amino acid degradation, and with oxidative phosphorylation. Using a graphical Gaussian model, we inferred a network of connections between the genes within each module. The first module had 52 genes with 87 connections, and the most connected genes were ADIPOQ, which is related with FA oxidation, and ELOVL6 and FABP4, both involved in FA metabolism. The second module showed 196 genes connected by 263 edges, being FN1 and MAP3K11 the most connected genes. On the other hand, the third module had 161 genes connected by 251 edges and ATG13 was the top neighbouring gene, while the fourth module had 224 genes and 655 connections, and its most connected genes were related with mitochondrial pathways. Overall, this work successfully identified relevant muscle gene networks and modules linked with FA composition, providing further insights on how the physiology of the pigs influences FA composition.

Plasma proteins associated with plant-based diets: Results from the Atherosclerosis Risk in Communities (ARIC) study and Framingham Heart Study (FHS)

Background & AimsPlant-based diets are associated with a lower risk of chronic diseases. Large-scale proteomics can identify objective biomarkers of plant-based diets, and can improve our understanding of the pathways that link plant-based diets to health outcomes. This study investigated the plasma proteome of four different plant-based diets [overall plant-based diet (PDI), provegetarian diet, healthful plant-based diet (hPDI), and unhealthful plant-based diet (uPDI)] in the Atherosclerosis Risk in Communities (ARIC) Study and replicated the findings in the Framingham Heart Study (FHS) Offspring cohort. MethodsARIC Study participants at visit 3 (1993-1995) with completed food frequency questionnaire (FFQ) data and proteomics data were divided into internal discovery (n=7,690) and replication (n=2,543) data sets. Multivariable linear regression was used to examine associations between plant-based diet indices (PDIs) and 4,955 individual proteins in the discovery sample. Then, proteins that were internally replicated in the ARIC Study were tested for external replication in FHS (n=1,358). Pathway overrepresentation analysis was conducted for diet-related proteins. C-statistics were used to predict if the proteins improved prediction of plant-based diet indices beyond participant characteristics. ResultsIn ARIC discovery, a total of 837 diet-protein associations (PDI=233; provegetarian=182; hPDI=406; uPDI=16) were observed at false discovery rate (FDR) <0.05. Of these, 453 diet-protein associations (PDI=132; provegetarian=104; hPDI=208; uPDI=13) were internally replicated. In FHS, 167/453 diet-protein associations were available for external replication, of which 8 proteins (PDI=1; provegetarian=0; hPDI=8; uPDI=0) replicated. Complement and coagulation cascades, cell adhesion molecules, and retinol metabolism were over-represented. C-C motif chemokine 25 for PDI and 8 proteins for hPDI modestly but significantly improved the prediction of these indices individually and collectively (P value for difference in C-statistics<0.05 for all tests) ConclusionsUsing large-scale proteomics, we identified potential candidate biomarkers of plant-based diets, and pathways that may partially explain the associations between plant-based diets and chronic conditions.

O-098 The transcriptome of endometrial organoids of fertile and subfertile women exposed to seminal plasma of fertile men

Abstract Study question What is the effect of seminal plasma (SP) from fertile males on the transcriptome of endometrial epithelial organoids of fertile and subfertile women? Summary answer Transcriptomic changes in fertile and subfertile endometrial organoids are observed upon treatment with SP, related to hormone- and immune-regulation, reproduction and cellular organisation. What is known already Clinical studies have demonstrated the potential beneficial effect of intrauterine SP (liquid fraction of semen) insemination during assisted reproductive technology (ART), resulting in higher implantation- and pregnancy rates. However, the molecular mechanism by which SP exerts its function remains unclear. SP components such as TGF-β1 or IL-8 have the potential capacity of conditioning the uterus for pregnancy. SP has shown to induce significant transcriptional changes in 2D endometrial epithelial cells. 3D self-organising human endometrial organoids have been established over the last years, mimicking a more in vivo-like physiology, and therefore may better represent the interaction of the endometrium with SP. Study design, size, duration Endometrial tissue biopsies from fertile (n = 5) and subfertile (n = 5; subfertile defined as women not having reached clinical pregnancy after at least 12 months of regular unprotected intercourse) were used to establish endometrial epithelial organoids. Subsequently, SP from fertile males (n = 5) was collected and pooled. Endometrial organoids were cultured in complex media supplemented with 1 µM 17β-estradiol (E2) and incubated with or without (control) 1% SP for 6 h. Participants/materials, setting, methods RNA sequencing was performed using NextSeq600 (Illumina). Significant DEGs upon SP exposure were identified using DESeq2 (Fold Change (FC) of ≥ 1.5 or ≤ -1.5). Eight selected DEGs were validated by qPCR in one cell line. Principal Component Analysis (PCA) was performed to compare endometrium donor correlation. Gene Set Enrichment Analysis (GSEA; v4.3.2) was then used to identify overrepresented pathways and gene ontology (GO) gene sets (P &amp;lt; 0.05; False Discovery Rate (FDR) &amp;lt; 0.25). Main results and the role of chance High heterogeneity between donors was observed, regardless of their fertility phenotype. PCA analysis indicated clustering based on donor rather than SP treatment or phenotype. Moreover, the number of significantly DEGs ranged from 629 to 2332. Treatment with SP resulted in identification of 105 significant DEGs (p &amp;lt; 0.05) among all donors (fertile and subfertile) from which 80 DEGs were upregulated and 25 DEGs were downregulated. Several of these DEGs were also reported to be affected by SP in other 2D epithelial cultures studies. Interesting DEGs include progesterone-associated endometrial protein (PAEP; log2FC 2.0 ± 1.0), interleukin-6 (IL6; log2FC 4.5 ± 1.5), insulin-like growth factor-binding protein 3 (IGFBP3; log2FC 1.1 ± 0.2) and mucin 6 (MUC6; log2FC 3.7 ± 1.6). GSEA analysis identified gene set enrichment of pathways and biological processes related to hormone- and immune regulation, reproduction, and cellular organisation in all endometrial organoids treated with SP. Preliminary RNAseq validation with qPCR showed comparable expression patterns for 8 genes (e.g. PAEP and IL6). Moreover, expression of these genes was uniform when using a distinct SP batch (n = 6; fertile males). No clear differential response between fertile and subfertile organoids upon SP exposure was observed. Limitations, reasons for caution The apical-inside organisation of the organoids (i.e. SP interaction from the basolateral side) should be taken into account, since in vivo interaction will be apically. Moreover, high heterogeneity between donors was observed between and within the fertility phenotypes. Therefore, conclusions based on fertility phenotypes should be drawn with caution. Wider implications of the findings Understanding the effect of SP on the endometrium will shed light on male-female reproductive tract crosstalk, which is omitted during ART. Future research should focus on identifying key regulatory SP molecules and functional studies should include apical SP interaction. Potentially providing therapeutic options for improving success rates in ART. Trial registration number Not applicable

Phototherapy Alters the Plasma Metabolite Profile in Infants Born Preterm with Hyperbilirubinemia

To investigate the effects of gestational age (GA) and phototherapy on the plasma metabolite profile of preterm infants with neonatal hyperbilirubinemia (NHB). From a cohort of prospectively enrolled infants born preterm (n=92), plasma samples of very preterm (VPT; GA, 28+0 to 31+6weeks, n=27) and moderate/late preterm (M/LPT; GA, 32+0 to 35+6weeks, n=33) infants requiring phototherapy for NHB were collected prior to the initiation of phototherapy and 24hours after starting phototherapy. An additional sample was collected 48hours after starting phototherapy in a randomly selected subset (n=30; VPT n=15; M/LPT n=15). Metabolite profiles were determined using ultraperformance liquid chromatography tandem mass spectroscopy. Two-way ANCOVA was used to identify metabolites that differed between GA groups and timepoints after adjusting for total serum bilirubin levels (false discovery rate q-value < 0.05). Top impacted pathways were identified using pathway over-representation analysis. Phototherapy was initiated at lower total serum bilirubin (mean±SD mg/dL) levels in VPT compared with M/LPT infants (7.3±1.4 vs 9.9±1.9, P<.01). We identified 664 metabolites that were significant for a phototherapy effect, 191 metabolites significant for GA, and 46 metabolites significant for GA×phototherapy interaction (false discovery rate q-value < 0.05). Longer duration phototherapy had a larger mean effect size (24hours postphototherapy: d=0.36; 48hours postphototherapy: d=0.43). Top pathways affected by phototherapy included membrane lipid metabolism, one-carbon metabolism, creatine biosynthesis, and oligodendrocyte differentiation. Phototherapy alters the plasma metabolite profile more than GA in preterm infants with NHB, affecting pathways related to lipid and one-carbon metabolism, energy biosynthesis, and oligodendrocyte differentiation.

Integrative Bioinformatics Approaches to Uncover Hub Genes and Pathways Involved in Cardiovascular Diseases.

Cardiovascular diseases (CVD) represent a significant global health challenge resulting from a complex interplay of genetic, environmental, and lifestyle factors. However, the molecular pathways and genetic factors involved in the onset and progression of CVDs remain incompletely understood. Here, we performed an integrative bioinformatic analysis to highlight specific genes and signaling pathways implicated in the pathogenesis of 80 CVDs. Differentially expressed genes (DEGs) were identified through the integrated analysis of microarray and GWAS datasets. Then, hub genes were identified after gene ontology functional annotation analysis and protein-protein internet (PPI) analysis. In addition, pathways were identified through KEGG and gene ontology enrichment analyses. A total of 821 hub genes related to 80 CVDs were identified, including 135 common and frequent CVD-associated genes. TNF, IL6, VEGFA, and TGFB.1 genes were the central core genes expressed in 50% or more of CVDs, confirming that the inflammation is a key pathological feature of CVDs. Analysis of hub genes by KEGG enrichment revealed predominant enrichment in 201 KEGG pathways, of which the AGE-RAGE signaling pathway in diabetic complications was identified as the common key KEGG implicated in 62 CVDs. In addition, the outcomes showed an overrepresentation in pathways categorized under human diseases, particularly in the subcategories of infectious diseases and cancers, which may be common risk factors for CVDs. In conclusion, this powerful approach for in silico fine-mapping of genes and pathways allowed the identification of determinant hubs genes and pathways implicated in the pathogenesis of CVDs which could be employed in developing more targeted and effective interventions for preventing, diagnosing, and treating CVDs. The function of these hub genes in CVDs needs further exploration to elucidate their biological characteristics.

#1980 Past peritonitis episodes are linked to decreased peritoneal Immune Cell Function and Associated with Chronic Increased Peritoneal Inflammation

Abstract Background and Aims Chronic peritoneal inflammation and infectious complications occur in a significant proportion of PD patients. A relationship between peritoneal IL-6 levels and local immunocompetence is suspected. To analyse local immunocompetence, an ex-vivo stimulation assay of PD effluent was developed. The aim of this study was to analyse peritoneal inflammation and immunocompetence in a PD cohort in relation to clinical outcomes. Method Prospectively, 245 routinely performed 4-hour PETs (143 patients, 2679 patient-months) of patients exclusively receiving 2. generation PD fluids (two-chamber, pH neutral, low amount of glucose degradation products) were analysed. An ex-vivo cell stimulation protocol of the fresh effluents was performed of all PETs. Peritoneal immune cell function was analysed using ex-vivo Toll-like receptor (LPS/Pam3Cys) stimulated IL-6 release as an index of immunocompetence. The molecular effects of effluent components and the addition of alanyl-glutamine to PD fluid were analysed in heterologous cell stimulation experiments and using an effluent proteomics and metabolomics approach. Results IL-6 in the effluent showed no association with various patient characteristics (incl. RRF, underlying disease, peritonitis episodes, time on PD). A significant association of peritoneal IL-6 with solute transport rates and systemic markers of uremic inflammation was found. IL-6 in effluent (HR 2.54) and stimulated 1 h effluent (HR 0.34) was predictive of subsequent peritonitis episodes. An inverse association was observed with ex-vivo stimulated IL-6 release from peritoneal cells (P = .02), most pronounced in patients with a history of peritonitis (P = .008). Patients with above median elevated IL-6 levels showed reduced peritoneal immune cell function. The results were validated in an independent multi-centre PD cohort (n = 48). A proteomics (n = 17) and targeted metabolomics (patients n = 134, 1004 metabolites) analysis of PD effluents show an overrepresentation of apoptosis and neutrophil degranulation pathways in patients with elevated effluent IL-6 levels and reduced response mechanisms to pathogens when compared to effluents from patients with low effluent IL-6 levels. Heterologous donor-PBMC stimulation (4 Healthy donors) with 4h PET PD-effluents of 20 patients showed reduced immunocompetence with elevated IL-6 (P = .038) and improved immunocompetence when 8 mM alanyl-glutamine were added to the PD fluid during the dwell. These results could also be validated in a post-hoc analysis of a RCT testing alanyl-glutamine addition to PD fluid for 8 weeks in PD patients. Conclusion This is the first longitudinal study to show the relationship between peritoneal inflammation and impaired immune function of peritoneal cells, possibly predisposing to infectious complications. The possible underlying mechanistic link may represent the biological correlate (“immune-paralysis”) for PD patients at increased risk for recurrent episodes of peritonitis.

The Effect of Ovariectomy and Estradiol Substitution on the Metabolic Parameters and Transcriptomic Profile of Adipose Tissue in a Prediabetic Model.

Menopause brings about profound physiological changes, including the acceleration of insulin resistance and other abnormalities, in which adipose tissue can play a significant role. This study analyzed the effect of ovariectomy and estradiol substitution on the metabolic parameters and transcriptomic profile of adipose tissue in prediabetic females of hereditary hypertriglyceridemic rats (HHTgs). The HHTgs underwent ovariectomy (OVX) or sham surgery (SHAM), and half of the OVX group received 17β-estradiol (OVX+E2) post-surgery. Ovariectomy resulted in weight gain, an impaired glucose tolerance, ectopic triglyceride (TG) deposition, and insulin resistance exemplified by impaired glycogenesis and lipogenesis. Estradiol alleviated some of the disorders associated with ovariectomy; in particular, it improved insulin sensitivity and reduced TG deposition. A transcriptomic analysis of perimetrial adipose tissue revealed 809 differentially expressed transcripts in the OVX vs. SHAM groups, mostly pertaining to the regulation of lipid and glucose metabolism, and oxidative stress. Estradiol substitution affected 1049 transcripts with overrepresentation in the signaling pathways of lipid metabolism. The principal component and hierarchical clustering analyses of transcriptome shifts corroborated the metabolic data, showing a closer resemblance between the OVX+E2 and SHAM groups compared to the OVX group. Changes in the adipose tissue transcriptome may contribute to metabolic abnormalities accompanying ovariectomy-induced menopause in HHTg females. Estradiol substitution may partially mitigate some of these disorders.

Analysis of MicroRNA Cargo in Circulating Extracellular Vesicles from HIV-Infected Individuals with Pulmonary Hypertension.

The risk of developing pulmonary hypertension (PH) in people living with HIV is at least 300-fold higher than in the general population, and illicit drug use further potentiates the development of HIV-associated PH. The relevance of extracellular vesicles (EVs) containing both coding as well as non-coding RNAs in PH secondary to HIV infection and drug abuse is yet to be explored. We here compared the miRNA cargo of plasma-derived EVs from HIV-infected stimulant users with (HIV + Stimulants + PH) and without PH (HIV + Stimulants) using small RNA sequencing. The data were compared with 12 PH datasets available in the GEO database to identify potential candidate gene targets for differentially altered miRNAs using the following functional analysis tools: ingenuity pathway analysis (IPA), over-representation analysis (ORA), and gene set enrichment analysis (GSEA). MiRNAs involved in promoting cell proliferation and inhibition of intrinsic apoptotic signaling pathways were among the top upregulated miRNAs identified in EVs from the HIV + Stimulants + PH group compared to the HIV + Stimulants group. Alternatively, the downregulated miRNAs in the HIV + Stimulants + PH group suggested an association with the negative regulation of smooth muscle cell proliferation, IL-2 mediated signaling, and transmembrane receptor protein tyrosine kinase signaling pathways. The validation of significantly differentially expressed miRNAs in an independent set of HIV-infected (cocaine users and nondrug users) with and without PH confirmed the upregulation of miR-32-5p, 92-b-3p, and 301a-3p positively regulating cellular proliferation and downregulation of miR-5571, -4670 negatively regulating smooth muscle proliferation in EVs from HIV-PH patients. This increase in miR-301a-3p and decrease in miR-4670 were negatively correlated with the CD4 count and FEV1/FVC ratio, and positively correlated with viral load. Collectively, this data suggest the association of alterations in the miRNA cargo of circulating EVs with HIV-PH.

Evaluation of Toll-like Receptor 4 (TLR4) Involvement in Human Atrial Fibrillation: A Computational Study.

In the present study, we have explored the involvement of Toll-like Receptor 4 (TLR4) in atrial fibrillation (AF), by using a meta-analysis of publicly available human transcriptomic data. The meta-analysis revealed 565 upregulated and 267 downregulated differentially expressed genes associated with AF. Pathway enrichment analysis highlighted a significant overrepresentation in immune-related pathways for the upregulated genes. A significant overlap between AF differentially expressed genes and TLR4-modulated genes was also identified, suggesting the potential role of TLR4 in AF-related transcriptional changes. Additionally, the analysis of other Toll-like receptors (TLRs) revealed a significant association with TLR2 and TLR3 in AF-related gene expression patterns. The examination of MYD88 and TICAM1, genes associated with TLR4 signalling pathways, indicated a significant yet nonspecific enrichment of AF differentially expressed genes. In summary, this study offers novel insights into the molecular aspects of AF, suggesting a pathophysiological role of TLR4 and other TLRs. By targeting these specific receptors, new treatments might be designed to better manage AF, offering hope for improved outcomes in affected patients.

Elucidating Cancer Subtypes by Using the Relationship between DNA Methylation and Gene Expression.

Advancements in the field of next generation sequencing (NGS) have generated vast amounts of data for the same set of subjects. The challenge that arises is how to combine and reconcile results from different omics studies, such as epigenome and transcriptome, to improve the classification of disease subtypes. In this study, we introduce sCClust (sparse canonical correlation analysis with clustering), a technique to combine high-dimensional omics data using sparse canonical correlation analysis (sCCA), such that the correlation between datasets is maximized. This stage is followed by clustering the integrated data in a lower-dimensional space. We apply sCClust to gene expression and DNA methylation data for three cancer genomics datasets from the Cancer Genome Atlas (TCGA) to distinguish between underlying subtypes. We evaluate the identified subtypes using Kaplan-Meier plots and hazard ratio analysis on the three types of cancer-GBM (glioblastoma multiform), lung cancer and colon cancer. Comparison with subtypes identified by both single- and multi-omics studies implies improved clinical association. We also perform pathway over-representation analysis in order to identify up-regulated and down-regulated genes as tentative drug targets. The main goal of the paper is twofold: the integration of epigenomic and transcriptomic datasets followed by elucidating subtypes in the latent space. The significance of this study lies in the enhanced categorization of cancer data, which is crucial to precision medicine.

Functional transcriptome analysis revealed upregulation of MAPK-SMAD signalling pathways in chronic heat stress in crossbred cattle.

Animal geneticists and breeders have the impending challenge of enhancing the resilience of Indian livestock to heat stress through better selection strategies. Climate change's impact on livestock is more intense in tropical countries like India where dairy cattle crossbreeds are more sensitive to heat stress. The main reason for this study was to find the missing relative changes in transcript levels in thermo-neutral and heat stress conditions in crossbred cattle through whole-transcriptome analysis of RNA-Seq data. Differentially expressed genes (DEGs) identified based on the minimum log twofold change value and false discovery rate 0.05 revealed 468 up-regulated genes and 2273 down-regulated significant genes. Functional annotation and pathway analysis of these significant DEGs were compared based on Gene Ontology (Biological process), Kyoto Encyclopedia of Genes and Genome (KEGG), and Reactome pathways using g: Profiler, ShinyGO v0.76, and iDEP.951 web tools. On finding network visualization, the most over-represented and correlated pathways were neuronal and sensory organ development, calcium signalling pathway, Mitogen-activated protein kinase (MAPK) and Smad signalling pathway, Ras-proximate-1, or Ras-related protein 1 (Rap 1) signalling pathway, apoptosis, and oxidative stress. Similarly, down-regulated genes were most expressed in mRNA processing, immune system, B-cell receptor signalling pathway, Nucleotide oligomerization domain(NOD)-like receptors (NLRs) signalling pathway and nonsense-mediated decay (NMD) pathway. The heat stress-responsive genes identified in this study will facilitate our understanding of the molecular basis for climate resilience and heat tolerance in Indian dairy crossbreeds.

Age- and sex- divergent translatomic responses of the mouse retinal pigmented epithelium

Aging is the main risk factor for age-related macular degeneration (AMD), a retinal neurodegenerative disease that leads to irreversible blindness, particularly in people over 60 years old. Retinal pigmented epithelium (RPE) atrophy is an AMD hallmark. Genome-wide chromatin accessibility, DNA methylation, and gene expression studies of AMD and control RPE demonstrate epigenomic/transcriptomic changes occur during AMD onset and progression. However, mechanisms by which molecular alterations of normal aging impair RPE function and contribute to AMD pathogenesis are unclear.Here, we specifically interrogate the RPE translatome with advanced age and across sexes in a novel RPE reporter mouse model. We find differential age- and sex- associated transcript expression with overrepresentation of pathways related to inflammation in the RPE. Concordant with impaired RPE function, the phenotypic changes in the aged translatome suggest that aged RPE becomes immunologically active, in both males and females, with some sex-specific signatures, which supports the need for sex representation for in vivo studies.

OA01 Cluster medicine: the role of genetics for identifying shared biological pathways between rheumatoid arthritis and common comorbidities for targeting treatments

Abstract Background/Aims Rheumatoid arthritis (RA) is a chronic inflammatory disorder associated with significant pain and disability, as well as increased risk of comorbidities. These comorbidities, such as coronary artery disease, stroke, heart failure and type 2 diabetes, often form predictable clusters highlighting the importance of common disease mechanisms. Identifying these clusters, and the underlying shared mechanisms, is key to the concept of cluster medicine which aims to take a more systemic approach to treating comorbidity. It is hypothesised that inflammatory processes in RA promote the pathogenesis of these comorbid diseases, which typically occur at an earlier stage of life than in the general population. However, few studies have investigated the degree of shared genetic susceptibility implicated across these traits. The present study aimed to identify and characterise pleiotropic loci and shared biological pathways involved in the pathogenesis of RA and cardiometabolic disorders. Methods European-only GWAS summary statistics were collected for RA (seropositive, seronegative, and overall) and seven cardiometabolic traits, including ischemic stroke, any stroke, heart failure, coronary artery disease, hypertension, atrial fibrillation, and type 2 diabetes. Cross-trait linkage disequilibrium score regression was employed to estimate the degree of genetic correlation (rg) between all pairs of traits. Subset-based meta-analysis was performed using the R package ASSET to identify pleiotropic single nucleotide polymorphisms (SNPs). Pleiotropic SNPs were assigned to genes based on positional and eQTL mapping. Prioritised genes were tested for over-representation in functional categories and biological pathways in gene sets obtained from MsigDB and WikiPathways using the hypergeometric test loci. Gene mapping and enrichment analyses were conducted in FUMA. Results RA showed significant positive genetic correlation with all cardiometabolic traits, with the seronegative subtype showing greater correlation than the seropositive subtype. Correlations between RA and cardiometabolic phenotypes ranged from 0.11 - 0.38, the strongest of which was between seronegative RA and heart failure (rg = 0.38, P = 2.6e-07). A total of 67 lead SNPs were identified as potentially pleiotropic. Pleiotropic SNPs were mapped to genes including IL6R, PTPN11, ATXN2, PLCL1 and SOX6. Pathway enrichment analyses revealed that mapped genes were primarily involved in inflammatory signalling pathways, involving interleukins, TNF-α and interferon (gamma and alpha) response. Conclusion This study provides evidence for pleiotropy across RA and cardiometabolic traits, whereby inherited genetic variants predispose to aberrant cytokine signalling, thus conferring risk to multiple diseases where inflammation is a key driver of pathogenesis. Disclosure M. Stoves: None. M. Soomro: None. S. Zhao: None. D. Plant: None. A. Barton: None. J. Bowes: None.