RA-related Genes Research Articles

Amelioration of melittin on adjuvant-induced rheumatoid arthritis: Integrated transcriptome and metabolome

BackgroundRheumatoid arthritis (RA) is a chronic autoimmune disease lacking a definitive cure. Although conventional treatments such as dexamethasone and methotrexate are prevalent, their usage is constrained by potential adverse effects. Melittin (MLT) has emerged as a promising natural anti-rheumatic drug; however, studies focusing on the role of MLT in modulating the expression and metabolism of RA-related genes are scarce. MethodArthritis was induced in rats using Complete Freund's Adjuvant (CFA), followed by MLT injections for treatment. Post-treatment, the inflammatory status of each group was assessed, and the mechanistic underpinnings of MLT's ameliorative effects on RA were elucidated through transcriptomic and metabolomic analyses. Additionally, this study conducted qRT-PCR validation of key therapeutic genes and characterized the molecular docking interactions of MLT with key receptor proteins (TNF-α and IL-1β) using the AutoDock Vina software. ResultMLT significantly diminished redness and swelling in affected joints, ameliorated inflammatory cell infiltration, and mitigated joint damage. Integration of transcriptomic and metabolomic data revealed that MLT predominantly regulated the transcription levels of pathways and genes related to cytokines and immune responses, and the metabolic biomarkers of Sphingomyelin, fatty acid, and flavonoid. qRT-PCR confirmed MLT's downregulation of inflammation-related genes such as Il6, Jak2, Stat3, and Ptx3. Molecular docking simulations demonstrated the stable binding of MLT to TNF-α and IL-1β. ConclusionMLT demonstrated significant efficacy in alleviating RA. This study provides a comprehensive summary of MLT's impact on gene expression and metabolic processes associated with RA.

Identification of Hedyotisdiffusa Willd-specific mRNA–miRNA–lncRNA network in rheumatoid arthritis based on network pharmacology, bioinformatics analysis, and experimental verification

Hedyotisdiffusa Willd (HDW) possesses heat-clearing, detoxification, anti-cancer, and anti-inflammatory properties. However, its effects on rheumatoid arthritis (RA) remain under-researched. In this study, we identified potential targets of HDW and collected differentially expressed genes of RA from the GEO dataset GSE77298, leading to the construction of a drug-component-target-disease regulatory network. The intersecting genes underwent GO and KEGG analysis. A PPI protein interaction network was established in the STRING database. Through LASSO, RF, and SVM-RFE algorithms, we identified the core gene MMP9. Subsequent analyses, including ROC, GSEA enrichment, and immune cell infiltration, correlated core genes with RA. mRNA–miRNA–lncRNA regulatory networks were predicted using databases like TargetScan, miRTarBase, miRWalk, starBase, lncBase, and the GEO dataset GSE122616. Experimental verification in RA-FLS cells confirmed HDW’s regulatory impact on core genes and their ceRNA expression. We obtained 11 main active ingredients of HDW and 180 corresponding targets, 2150 RA-related genes, and 36 drug-disease intersection targets. The PPI network diagram and three machine learning methods screened to obtain MMP9, and further analysis showed that MMP9 had high diagnostic significance and was significantly correlated with the main infiltrated immune cells, and the molecular docking verification also showed that MMP9 and the main active components of HDW were well combined. Next, we predicted 6 miRNAs and 314 lncRNAs acting on MMP9, and two ceRNA regulatory axes were obtained according to the screening. Cellular assays indicated HDW inhibits RA-FLS cell proliferation and MMP9 protein expression dose-dependently, suggesting HDW might influence RA’s progression by regulating the MMP9/miR-204-5p/MIAT axis. This innovative analytical thinking provides guidance and reference for the future research on the ceRNA mechanism of traditional Chinese medicine in the treatment of RA.

Identification of shared disease marker genes and underlying mechanisms between major depression and rheumatoid arthritis

Both depression and rheumatoid arthritis (RA) have a very high comorbidity rate. A bilateral association is estimated to increase the mutual risk and the common denominator is inflammation being observed in both diseases. Previous studies have mainly focused on assessing peripheral blood's inflammatory and pro-inflammatory cytokines levels.We aimed to extend insights into the molecular mechanisms of depression based on hub RA genes. To do so, we prioritized RA-related genes using in-silico tools. We then investigated whether RA-related genes undergo altered expression in patients with unipolar and bipolar depression without a concurrent RA diagnosis and any exponents of active inflammation. In addition, we selected a homogeneous group of patients treated with lithium (Li), which has immunomodulatory properties.The study was performed on patients with bipolar depression (BD, n = 45; Li, n = 20), unipolar depression (UD, n = 27), and healthy controls (HC, n = 22) of both sexes. To identify DEGs in peripheral blood mononuclear cells (PBMCs), we used the SurePrint G3 Microarray and GeneSpring software.We selected a list of 180 hub genes whose altered expression we analyzed using the expression microarray results. In the entire study group, we identified altered expression of 93 of the 180 genes, including 35 down-regulated (OPRM1 gene with highest FC > 3) and 58 up-regulated (TLR4 gene with highest FC > 3). In UD patients, we observed maximally up-regulated expression of the TEK gene (FC > 3), and in BD of the CXCL8 gene (FC > 5). On the other hand, in lithium-treated patients, the gene with the most reduced expression was the TRPV1 gene.The study proved that depression and RA are produced by a partially shared “inflammatory interactome” in which the opioid and angiogenesis pathways are important.

Epigenome-wide methylation haplotype association analysis identified HLA-DRB1, HLA-DRB5 and HLA-DQB1 as risk factors for rheumatoid arthritis.

The aim of this study was to compare nonrandom associations between physically adjacent single methylation polymorphism loci among rheumatoid arthritis (RA) and normal subjects for investigating RA-risk methylation haplotypes (meplotype). With 354 ACPA-positive RA patients and 335 normal controls selected from a case-control study based on Swedish population, we conducted the first RA epigenome-wide meplotype association study using our software EWAS2.0, mainly including (i) converted the β value to methylation genotype (menotype) data, (ii) identified methylation disequilibrium (MD) block, (iii) calculated frequent of each meplotypes in MD block and performed case-control association test and (iv) screened for RA-risk meplotypes by odd ratio (OR) and p-values. Ultimately, 545 meplotypes on 334 MD blocks were identified significantly associated with RA (p-value<.05). These meplotypes were mapped to 329 candidate genes related to RA. Subsequently, combined with gene optimization, eight RA-risk meplotypes were identified on three risk genes: HLA-DRB1, HLA-DRB5 and HLA-DQB1. Our results reported the relationship between DNA methylation pattern on HLA-DQB1 and the risk of RA for the first time, demonstrating the co-demethylation of 'cg22984282' and 'cg13423887' on HLA-DQB1 gene (meplotype UU, p-value=2.90E-6, OR=1.68, 95% CI=[1.35, 2.10]) may increase the risk of RA. Our results demonstrates the potential of methylation haplotype analysis to identify RA-related genes from a new perspective and its applicability to the study of other disease.

Network pharmacology analysis of the pharmacological mechanism of Artemisia lavandulaefolia DC. in rheumatoid arthritis

ObjectiveThe traditional She medicine is a notable type of traditional Chinese medicine, which has been applied for a long history despite the lack of sufficient mechanistic understanding. Our study revealed the possible molecular mechanism of sesquiterpene 5α-Hydroxycostic acid, active ingredient of traditional She medicine Artemisia lavandulaefolia DC., in the treatment of rheumatoid arthritis (RA). MethodsRA-fibroblast like synoviocytes (RA-FLSs) were treated with 5α-Hydroxycostic acid, Anthemidin, and methotrexate (MTX). CCK-8 and ELISA were used to measure the resultant viability of RA-FLSs and to quantify pro-inflammatory cytokines. Target genes of 5α-Hydroxycostic acid and Anthemidin, RA-related differentially expressed genes, and RA-related genes were retrieved by bioinformatics analyses, results of which were further intersected to identify candidate genes. The protein-protein interaction network was constructed to develop the pharmacophore model. The molecular docking was simulated to determine the core target androgen receptor (AR) for subsequent molecular mechanism investigation in vitro. ResultsThe 5ɑ-Hydroxycostic acid, Anthemidin, or MTX of different concentrations inhibited the viability of RA-FLSs, and downregulated the levels of proinflammatory cytokines. The pharmacophore model and molecular docking of 10 candidate targets with 5α-Hydroxycostic acid were successfully established. In vitro experiments provided evidence confirming that 5α-Hydroxycostic acid elevated AR expression to inhibit inflammatory responses of RA-FLSs and degradation of extracellular matrix. ConclusionTherefore, this study reveals the active ingredient sesquiterpene 5α-Hydroxycostic acid of traditional She medicine Artemisia lavandulaefolia DC., and illustrates potential molecular mechanism in RA treatment by upregulating AR expression. This study is the first to report the effect of the active ingredient sesquiterpenes in traditional She medicine A.lavandulaefolia DC on RA and elucidate the underlying molecular mechanism associated with up-regulated AR expression. This study provides new insights into the mechanistic understanding of traditional She medicine in the treatment of RA.

Low-Intensity Physical Exercise Decreases Inflammation and Joint Damage in the Preclinical Phase of a Rheumatoid Arthritis Murine Model

Lifestyle modifications in preclinical Rheumatoid Arthritis (RA) could delay the ongoing pathogenic immune processes and potentially prevent its onset. Physical exercise (PE) benefits RA patients; however, its impact in reducing the risk of developing RA has scarcely been studied. The objective was to describe the effects of low-intensity PE applied at the disease's preclinical phase on the joints of DBA/1 mice with collagen-induced arthritis (CIA). Twelve mice with CIA were randomly distributed into two groups: the CIA-Ex group, which undertook treadmill PE, and the CIA-NoEx, which was not exercised. The effects of PE were evaluated through clinical, histological, transcriptomics, and immunodetection analyses in the mice's hind paws. The CIA-Ex group showed lower joint inflammation and damage and a decreased expression of RA-related genes (Tnf Il2, Il10, Il12a, IL23a, and Tgfb1) and signaling pathways (Cytokines, Chemokines, JAK-STAT, MAPK, NF-kappa B, TNF, and TGF-beta). TNF-α expression was decreased by PE in the inflamed joints. Low-intensity PE in pre-arthritic CIA reduced the severity through joint down-expression of proinflammatory genes and proteins. Knowledge on the underlying mechanisms of PE in preclinical arthritis and its impact on reducing the risk of developing RA is still needed.

Integrated Network Pharmacology and Experimental Validation Approach to Investigate the Mechanisms of Stigmasterol in the Treatment of Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory disease of the joints associated with systemic comorbidities. Sinomenium acutum is regarded as an effective traditional Chinese medicine (TCM) for the treatment of RA. Based on network pharmacology and Gene Expression Omnibus (GEO) database, 33 RA-related differentially-expressed genes (DEGs) targeting active compounds of Sinomenium acutum were initially screened in our investigation. Gene Ontology (GO) and Kyoto encyclopaedia of genes and genome (KEGG) analyses found the important involvement of these DEGs in osteoclast differentiation, and finally 5 core DEGs, including NCF4, NFKB1, CYBA, IL-1β and NCF1 were determined through protein-protein interaction (PPI) network. We also identified the related active component of Sinomenium acutum include Stigmasterol. Finally, in order to experimentally verify these results, a rat model of collagen-induced arthritis (CIA) was established, and subsequently treated with Stigmasterol solution. Similar to the healing effect of Indomethacin, Stigmasterol was observed to reduce the levels of inflammatory factors (IL-6 and IL-1β) and osteoclast differentiation-related factors (RANKL, ACP5 and Cathepsin K), which can also reduce the arthritis index score and alleviate the degree of pathological injury of rat ankle joints. The predictions and experimental data uncover the involvement of Stigmasterol, an active component of Sinomenium acutum, in regulation of osteoclast differentiation, exerting great medicinal potential in the treatment of RA.

NFIL3 and its immunoregulatory role in rheumatoid arthritis patients.

Nuclear-factor, interleukin 3 regulated (NFIL3) is an immune regulator that plays an essential role in autoimmune diseases. However, the relationship between rheumatoid arthritis (RA) and NFIL3 remains largely unknown. In this study, we examined NFIL3 expression in RA patients and its potential molecular mechanisms in RA. Increased NFIL3 expression levels were identified in peripheral blood mononuclear cells (PBMCs) from 62 initially diagnosed RA patients and 75 healthy controls (HCs) by quantitative real-time PCR (qRT-PCR). No correlation between NFIL3 and disease activity was observed. In addition, NFIL3 expression was significantly upregulated in RA synovial tissues analyzed in the Gene Expression Omnibus (GEO) dataset (GSE89408). Then, we classified synovial tissues into NFIL3-high (≥75%) and NFIL3-low (≤25%) groups according to NFIL3 expression levels. Four hundred five differentially expressed genes (DEGs) between the NFIL3-high and NFIL3-low groups were screened out using the "limma" R package. Enrichment analysis showed that most of the enriched genes were primarily involved in the TNF signaling pathway via NFκB, IL-17 signaling pathway, and rheumatoid arthritis pathways. Then, 10 genes (IL6, IL1β, CXCL8, CCL2, PTGS2, MMP3, MMP1, FOS, SPP1, and ADIPOQ) were identified as hub genes, and most of them play a key role in RA. Positive correlations between the hub genes and NFIL3 were revealed by qRT-PCR in RA PBMCs. An NFIL3-related protein-protein interaction (PPI) network was constructed using the STRING database, and four clusters (mainly participating in the inflammatory response, lipid metabolism process, extracellular matrix organization, and circadian rhythm) were constructed with MCODE in Cytoscape. Furthermore, 29 DEGs overlapped with RA-related genes from the RADB database and were mainly enriched in IL-17 signaling pathways. Thus, our study revealed the elevated expression of NFIL3 in both RA peripheral blood and synovial tissues, and the high expression of NFIL3 correlated with the abnormal inflammatory cytokines and inflammatory responses, which potentially contributed to RA progression.

Construction and comprehensive analysis of the biological network related to rheumatoid arthritis-related interstitial lung disease.

Interstitial lung disease (ILD) is a severe manifestation of rheumatoid arthritis (RA), which is characterized by low survival time post-diagnosis. Thus, it is important to explore the role of gene regulation related with ILD. Constructed a RA-ILD-related long chain noncoding RNA - messenger RNA (lncRNA-mRNA) network (ILD-LMN), based on ILD- and RA-related genes. We analyzed the topological properties of the resulting network. The results for network modularization and functional analysis showed that ILD-LMN performed basic and specific functions in ILD pathology. Furthermore, differential expression and correlation analysis of hub nodes revealed highly correlated competitive endogenous RNA regulatory relationships with important roles in pathological regulation. Following this, statistical analysis of disease-related single nucleotide polymorphisms (SNPs) in hub lncRNAs revealed that some of transcription factor-related SNPs were significantly associated with the expression of lncRNA. In fact, these SNPs exhibited significant differential expression in disease and normal samples. These results suggest that ILD-LMN has important implications in the study of disease. Altogether, the study of RA- and ILD-related lncRNA and genes on the basis of biological network would assist in providing better treatment opportunities for ILD patients. Additionally, it would promote further research on treatment of the disease.

Anti-angiogenic effect of YuXueBi tablet in experimental rheumatoid arthritis by suppressing LOX/Ras/Raf-1 signaling

Ethnopharmacological relevanceA Chinese patent medicine derived from a classical traditional Chinese medicine formula, Yu-Xue-Bi tablet (YXB) is widely used in the clinic to treat rheumatoid arthritis (RA). During the progression of RA, angiogenesis plays a central role in fostering the production of inflammatory cells, leading to synovial hyperplasia and bone destruction. However, whether YXB attenuates the angiogenesis during RA progression remains to be defined. Aim of the studyWe aimed to evaluate the anti-angiogenic activity of YXB and explore its mechanism of action in collagen-induced arthritis (CIA) rats and VEGF-induced HUVECs. Materials and methodsTranscriptional regulatory network analysis and a network pharmacology approach were employed to explore mechanism of YXB in RA angiogenesis. The antiarthritic effect of YXB was evaluated by determining the arthritis incidence, and score, and by micro-CT analysis. The anti-angiogenic effect of YXB in vivo was assessed by histological and immunohistochemical analyses. The anti-angiogenic effect of YXB in vitro was assessed by wound healing, Transwell migration, Transwell invasion, and tube formation assays. Western-blotting and immunohistochemical analysis were employed to explore the molecular mechanisms of YXB. ResultsYXB reduced disease severity and ameliorated pathological features in CIA rats. YXB markedly decreased bone destruction and synovial angiogenesis. Consistently, we also demonstrated that YXB effectively suppressed angiogenesis marker CD31 and VEGF expression. In vitro, YXB effectively inhibited HUVEC migration, invasion, and tube formation. Following the identification of transcriptional expression profiles, “YXB putative targets–known RA-related genes–genes associated with the therapeutic effect of YXB” interaction network was constructed and analyzed. After that, the LOX/Ras/Raf-1 signaling axis, which is involved in RA angiogenesis, was identified as one of the candidate mechanisms of YXB against RA. Experimentally, YXB dose-dependently decreased the expression levels of LOX, Ras, and Raf-1, as well as the phosphorylation of MEK and ERK in CIA rats, and these effects were better than the inhibitory effects of methotrexate (MTX), an FDA approved drug used for some autoimmune diseases such as RA. In addition, YXB may function as a potent angiogenesis inhibitor and significantly suppress the VEGF-induced activation of LOX/Ras/Raf-1 signaling in vitro. ConclusionsWe provide evidence that YXB may decrease the disease severity of RA and reduce bone erosion by suppressing angiogenesis via inhibition of LOX/Ras/Raf-1 signaling.

Lycium barbarum polysaccharide modulates gut microbiota to alleviate rheumatoid arthritis in a rat model

Rheumatoid arthritis (RA) seriously impairs the quality of life of sufferers. It has been shown that Lycium barbarum polysaccharide (LBP), a natural active indigestible ingredient with medicinal and edible functions, can effectively relieve RA, however, whether this effect is related to gut microbiota is not known. This study aimed to explore the RA alleviating mechanism of LBP mediated by gut microbiota using a collagen-induced arthritis rat model. The results showed that LBP significantly changed the gut microflora structure accompanied with the RA alleviation. Specifically, a LBP intervention reduced the relative abundance of Lachnospiraceae_NK4A136_group and uncultured_bacterium_f_Ruminococcaceae and significantly increased the abundance of Romboutsia, Lactobacillus, Dubosiella and Faecalibaculum. The mRNA contents of several colonic epithelial genes including Dpep3, Gstm6, Slc27a2, Col11a2, Sycp2, SNORA22, Tnni1, Gpnmb, Mypn and Acsl6, which are potentially associated to RA, were down-regulated due to the DNA hypermethylation, possibly caused by the elevating content of a bacterial metabolite S-adenosyl methionine (SAM). In conclusion, our current study suggests that LBP alleviated RA by reshaping the composition of intestinal microflora which may generate SAM, inducing DNA hypermethylation of RA-related genes in the host intestinal epithelium and subsequently reducing their expression.

Analysis and Experimental Validation of Rheumatoid Arthritis Innate Immunity Gene CYFIP2 and Pan-Cancer.

Rheumatoid arthritis (RA) is a chronic, heterogeneous autoimmune disease. Its high disability rate has a serious impact on society and individuals, but there is still a lack of effective and reliable diagnostic markers and therapeutic targets for RA. In this study, we integrated RA patient information from three GEO databases for differential gene expression analysis. Additionally, we also obtained pan-cancer-related genes from the TCGA and GTEx databases. For RA-related differential genes, we performed functional enrichment analysis and constructed a weighted gene co-expression network (WGCNA). Then, we obtained 490 key genes by intersecting the significant module genes selected by WGCNA and the differential genes. After using the RanddomForest, SVM-REF, and LASSO three algorithms to analyze these key genes and take the intersection, based on the four core genes (BTN3A2, CYFIP2, ST8SIA1, and TYMS) that we found, we constructed an RA diagnosis. The nomogram model showed good reliability and validity after evaluation, and the ROC curves of the four genes showed that these four genes played an important role in the pathogenesis of RA. After further gene correlation analysis, immune infiltration analysis, and mouse gene expression validation, we finally selected CYFIP2 as the cut-in gene for pan-cancer analysis. The results of the pan-cancer analysis showed that CYFIP2 was closely related to the prognosis of patients with various tumors, the degree of immune cell infiltration, as well as TMB, MSI, and other indicators, suggesting that this gene may be a potential intervention target for human diseases including RA and tumors.

Therapeutic mechanism of emodin for treatment of rheumatoid arthritis: a network pharmacology-based analysis

To investigate the therapeutic mechanism of emodin in the treatment of rheumatoid arthritis (RA) using a network pharmacology-based method and validate this mechanism in a fibroblast-like synovial cell line. The PubChem, Targetnet, SwissTargetPrediction, Genecards, OMIM, and DisGeNET databases were searched to obtain emodin targets and RA-related genes. A protein-protein interaction (PPI) network was constructed, and GO and KEGG pathway enrichment analyses were carried out to analyze the intersection genes. AutoDock4.2.6 software was used to simulate molecular docking between emodin and its candidate targets. In a cultured fibroblast-like synovial cell line (MH7A), the effects of different concentrations of emodin on proliferation of tumor necrosis factor-α (TNF-α)-induced cells were investigated using CCK-8 assay, cell scratch experiment and flow cytometry; the changes in the expressions of nuclear factor-κB (NF-κB) pathway proteins were detected using Western blotting, and the mRNA expressions of the hub genes were examined with RT-qPCR. We identified 32 intersection genes of emodin and RA, and the key targets including CAPS3, ESR1, and MAPK14 involved mainly the NF-κB signaling pathway. Cell scratch experiment and flow cytometry demonstrated a strong inhibitory effect of emodin on MH7A cell proliferation. Treatment with TNF-α significantly increased the cellular expressions of the NF-κB pathway proteins, which were obviously lowered by treatment with 80 μmol/L emodin. The results of RT-qPCR showed that TNF-α treatment obviously up-regulated the expressions of the hub genes COX2 and P38MAPK, and emodin treatment significantly down-regulated the expressions of MAPK and PTGS2 and up-regulated the expression of CASP3. The therapeutic effect of emodin on RA is mediated mainly through regulation of cell proliferation, apoptosis, and the NF-κB pathway.

Computation-Based Discovery of Potential Targets for Rheumatoid Arthritis and Related Molecular Screening and Mechanism Analysis of Traditional Chinese Medicine.

This study is aimed at screening potential therapeutic ingredients in traditional Chinese medicine (TCM) and identifying the key rheumatoid arthritis (RA) targets using computational simulations. Data for TCM-active ingredients with clear pharmacological effects were collected. Absorption, distribution, metabolism, excretion, and toxicity were evaluated. Potential RA targets were identified using the Gene Expression Omnibus (GEO) database, protein–protein interaction network, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses and potential TCM ingredients using AutoDock Vina. To examine the mechanisms underlying small molecules, target prediction, Gene Ontology, KEGG, and network modeling analyses were conducted; the effects were verified in rat synovial cells using cell proliferation assay. The activities of tumor necrosis factor TNF-α and IL-1β and alterations in cellular target protein levels were detected by ELISA and Western blotting, respectively. In total, data for 432 TCM active ingredients with clear pharmacological effects were obtained. Five critical RA-related genes were identified; CCL5 and CXCL10 were selected for molecular docking. Target prediction and network-based proximity analysis showed that dioscin could modulate 22 known RA clinical targets. Dioscin, asiaticoside, and ginsenoside Re could effectively inhibit in vitro cell proliferation and secretion of TNF-α and IL-1β in RA rat synovial cells. Using bioinformatics and computer-aided drug design, the potential small anti-RA molecules and their mechanisms of action were comprehensively identified. Dioscin could significantly inhibit proliferation and induce apoptosis in RA rat synovial cells by reducing TNF-α and IL-1β secretion and inhibiting abnormal CCL5, CXCL10, CXCR2, and IL2 expression.

PCBP1 is associated with rheumatoid arthritis by affecting RNA products of genes involved in immune response in Th1 cells

Rheumatoid arthritis (RA) is an autoimmune disease characterized by persistent synovitis, in which T helper 1 (Th1) can promote the development of a pro-inflammatory microenvironment. Poly(rC)-binding protein 1 (PCBP1) has been identified as a promising biomarker of RA, while its molecular mechanisms in RA development are unknown. As a canonical RNA binding protein, we propose that PCBP1 could play roles in RA by affecting both expression and alternative splicing levels in Th1 cells. Here, microarray datasets (GSE15573 and GSE23561), including 102 peripheral blood mononuclear cell samples from 39 RA patients and 63 controls, were used to evaluate the PCBP1 expression changes in RA patients. High throughput sequencing data (GSE84702) of iron driven pathogenesis in Th1 cells were downloaded and reanalyzed, including two Pcbp1 deficiency samples and two control samples in Th1 cells. In addition, CLIP-seq data of PCBP1 in Jurkat T cells was also analyzed to investigate the regulatory mechanisms of PCBP1. We found PCBP1 were down-regulated in RA specimens compared with control. The result of differentially expressed genes (DEGs) showed that Pcbp1 silencing in Th1 cells affected the expression of genes involved in immune response pathway. Alternative splicing analysis also revealed that PCBP1-regulated alternative splicing genes (RASGs) were enriched in TNF-a/NF-κB signaling pathway, T cell activation, T cell differentiation and T cell differentiation associated immune response pathways, which were highly associated with RA. DEGs and RASGs by Pcbp1 deficiency in mice were validated in PBMCs specimens of RA patients by RT-qPCR. Investigation of the CLIP-seq data revealed PCBP1 preferred to bind to 3′UTR and intron regions. PCBP1-bound genes were also significantly associated with RASGs, identifying 102 overlapped genes of these two gene sets. These genes were significantly enriched in several immune response related pathways, including myeloid cell differentiation and positive regulation of NF-κB transcription factor activity. Two RA-related genes, PML and IRAK1, were screened from the above immune related pathways. These results together support our hypothesis that PCBP1 can regulate the expression of genes involved in immune response pathway, and can bind to and regulate the alternative splicing of immune response related genes in immune T cells, and ultimately participate in the molecular mechanism of RA, providing new research ideas and directions for clinical diagnosis and treatment.

Orphan Nuclear Receptor NR4A2 Is Constitutively Expressed in Cartilage and Upregulated in Inflamed Synovium From hTNF-Alpha Transgenic Mice.

Orphan nuclear receptor 4A2 (NR4A2/Nurr1) is a constitutively active transcription factor with potential roles in the onset and progression of inflammatory arthropathies. NR4A2 is overexpressed in synovium and cartilage from individuals with rheumatoid arthritis (RA), psoriatic arthritis, and osteoarthritis. This study documents the expression and tissue localization of NR4A2 and upstream regulator nuclear factor kappa B (NF-κB) in the human tumor necrosis factor-alpha (hTNF-α) transgenic mouse model of RA. Since TNF-α is a potent inducer of NR4A2 in vitro, we hypothesized that NR4A2 would also be upregulated and active during disease progression in this model. Expression levels of NR4A2, related receptors NR4A1 (Nur77) and 3 (NOR1), and NF-κB1 transcripts were quantified by RT-qPCR in hTNF-α and wild-type joints at three stages of disease. The protein distribution of NR4A2 and NF-κB subunit RelA (p65) was analyzed by quantitative immunohistochemistry. Global gene expression of 88 RA-related genes was also screened and compared between groups. Consistent with previous reports on the hTNF-α model, transgenic mice exhibited significant weight loss and severely swollen paws by 19 weeks of age compared to age-matched wild-type controls. NR4A1-3 and NF-κB1 were constitutively expressed at disease onset and in healthy joints. NF-κB1 transcript levels increased 2-fold in hTNF-α paws with established disease (12 weeks), followed by a 2-fold increase in NR4A2 at the late disease stage (19 weeks). NR4A2 and RelA proteins were overexpressed in inflamed synovium prior to symptoms of arthritis, suggesting that gene expression changes documented in whole paws were largely driven by elevated expression in diseased synovium. Broader screening of RA-related genes by RT-qPCR identified several differentially expressed genes in hTNF-α joints including those encoding inflammatory cytokines and chemokines, matrix-degrading enzymes and inhibitors, cell surface receptors, intracellular signaling proteins and transcription factors. Consensus binding sites for NR4A receptors and NF-κB1 were enriched in the promoters of differentially expressed genes suggesting central roles for these transcription factors in this model. This study is the first comprehensive analysis of NR4A2 in an animal model of RA and validates the hTNF-α model for testing of small molecules and genetic strategies targeting this transcription factor.

Integrative pharmacology powers the detection of active components and mechanism underlying Wang Bi granules in rheumatoid arthritis

In China, Wang Bi Granule (WBG)2, composed of 16 herbal and 1 animal-based compounds, is used for clinical treatment of the "Wang Bi" syndrome, commonly referred to as later rheumatoid arthritis (RA) in modern medicine. It is also used in the treatment of ankylosing spondylitis, tuberculous arthritis, and Kashin-Beck disease, which are characterized by joint pain and swelling deformation. However, its pharmacological mechanisms remain unknown. We aimed to characterize the chemical components in WBG and examine the underlying mechanism for RA treatment using integrative pharmacological strategy, including chemical composition detection, efficacy evaluation, and mechanism exploration. We employed UPLC-QTOF-MS/MS to describe the chemical profile of WBG. TNF-α-stimulated RAW264.7 cells were used to simulate the inflammatory processes in RA and evaluate the anti-inflammatory effects of WBG. Network pharmacology was used to determine the mechanism underlying WBG action in RA. A total of 278 chemical components were identified or tentatively characterized. The water extract of WBG improved the imbalance in inflammation in TNF-α-stimulated RAW264.7 cells by regulating 179 differential genes. 55 key active constituents were obtained based on the interactions among "components" targets, RA-related genes, and differential genes (WBG vs TNF-α group) which may ameliorate RA by regulating 161 hub genes primarily involved in inflammation-related pathways. The present study, for the first time, employed integrative pharmacology to characterize the chemical profile of WBG and elucidate its mechanism of action against RA through an inflammation-immune regulatory system.

Similarity and Potential Relation Between Periimplantitis and Rheumatoid Arthritis on Transcriptomic Level: Results of a Bioinformatics Study.

BackgroundThis bioinformatics study aimed to reveal potential cross-talk genes, related pathways, and transcription factors between periimplantitis and rheumatoid arthritis (RA).MethodsThe datasets GSE33774 (seven periimplantitis and eight control samples) and GSE106090 (six periimplantitis and six control samples) were included from the National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO). A differential expression analysis (p < 0.05 and |logFC (fold change)| ≥ 1) and a functional enrichment analysis (p < 0.05) were performed. Based on this, a protein–protein interaction (PPI) network was constructed by Cytoscape. RA-related genes were extracted from DisGeNET database, and an overlap between periimplantitis-related genes and these RA-related genes was examined to identify potential cross-talk genes. Gene expression was merged between two datasets, and feature selection was performed by Recursive Feature Elimination (RFE) algorithm. For the feature selection cross-talk genes, support vector machine (SVM) models were constructed. The expression of these feature genes was determined from GSE93272 for RA. Finally, a network including cross-talk genes, related pathways, and transcription factors was constructed.ResultsPeriimplantitis datasets included 138 common differentially expressed genes (DEGs) including 101 up- and 37 downregulated DEGs. The PPI interwork of periimplantitis comprised 1,818 nodes and 2,517 edges. The RFE method selected six features, i.e., MERTK, CD14, MAPT, CCR1, C3AR1, and FCGR2B, which had the highest prediction. Out of these feature genes, CD14 and FCGR2B were most highly expressed in periimplantitis and RA. The final activated pathway–gene network contained 181 nodes and 360 edges. Nuclear factor (NF) kappa B signaling pathway and osteoclast differentiation were identified as potentially relevant pathways.ConclusionsThis current study revealed FCGR2B and CD14 as the most relevant potential cross-talk genes between RA and periimplantitis, which suggests a similarity between RA and periimplantitis and can serve as a theoretical basis for future research.

Epilepsy-Associated UBE3A Deficiency Downregulates Retinoic Acid Signalling Pathway.

Ubiquitin-protein ligase E3A (UBE3A) has dual functions as a E3 ubiquitin-protein ligase and coactivator of nuclear hormone receptors. Mutations or deletions of the maternally inherited UBE3A gene cause Angelman syndrome. Here, we performed transcriptome profiling in the hippocampus of Ube3am+/p+ and Ube3am–/p+ mice, and determined that the expression of the retinoic acid (RA) signalling pathway was downregulated in Ube3a-deficient mice compared to WT mice. Furthermore, we demonstrated that UBE3A directly interacts with RARα and may function as a coactivator of the nuclear receptor RARα to participate in the regulation of gene expression. Loss of UBE3A expression caused the downregulation of the expression of RA-related genes, including Erbb4, Dpysl3, Calb1, Pten, and Arhgap5 in Ube3am–/p+ mice brain tissues. This work revealed a new role for UBE3A in regulating retinoic acid (RA) signalling downstream genes and hopefully to shed light on the potential drug target of AS.

Integrative analysis highlighted susceptibility genes for rheumatoid arthritis

Genetic factors underlying susceptibility to rheumatoid arthritis (RA) are largely unknown. The aim of this study was to identify potential genes for RA. We conducted summary statistic data-based Mendelian randomization (SMR) analysis to detect potential causal genes for RA. Further, we performed additional bioinformatics analysis to show the potential relevance of the identified genes to RA. We identified 140 genes that showed causal association with RA. Among these genes, 24 have not been reported to be associated with RA (e.g., IFNAR2, FLOT1, ITPR3, PPP2R3C and SLC35B2). The unreported genes were highly connected with some well-known RA-related genes (e.g., HLA-DQB1, CD226, PTPN22, CD40, IFNGR2, BLK, TRAF1, SYNGR1 and CCR6) that were also found to be causally associated with RA. The identified genes were involved in the significant enriched RA-related biological pathways. We found integrative evidence in support of IFNAR2 as a potential causal gene of RA in SMR, differential expression, weighted gene co-expression network, protein–protein interaction and functional enrichment analyses. The present study highlights a list of potential causal genes for RA. The findings provide new insights into the mechanism underlying known genome-wide associated RA susceptibility loci.