Pathogenesis Of RA Research Articles

YTHDC1 Regulates the Migration, Invasion, Proliferation, and Apoptosis of Rheumatoid Fibroblast-Like Synoviocytes

BackgroundRheumatoid arthritis (RA), a chronic autoimmune condition, is characterized by persistent synovial inflammation, bone degradation, and progressive joint deterioration. Despite considerable research efforts, the precise molecular mechanism underlying RA remains elusive. This investigation aims to elucidate the potential role and molecular mechanism of N6-methyladenosine (m6A) methylation regulators in the pathogenesis of RA.MethodsIn this study, we employed bioinformatics tools to elucidate the association between RA and m6A modifications, aiming to identify potential biological markers. We extracted datasets GSE12021, GSE55235, and GSE55457 from the Gene Expression Omnibus (GEO) database for comprehensive analysis. Utilizing differential expression analysis, protein-protein interaction (PPI) analysis, and single-cell sequencing techniques, we identified pivotal hub genes implicated in the pathogenesis of RA. Subsequently, we assessed the correlation between these hub genes and the pathogenesis of RA using Gene Set Enrichment Analysis (GSEA). Both in vivo and in vitro experiments were performed to confirm the expression and functional roles of the identified key hub gene in RA.ResultsDifferential expression analysis, PPI analysis, and single-cell analysis identified three key hub genes (YTHDC1, YTHDC2, and YTHDF2) associated with RA. GSEA results further revealed that these genes are enriched in pathways associated with inflammatory responses. Subsequent correlation analysis demonstrated a significant negative correlation between YTHDC1 expression and CD8+ T cell levels. Notably, the gene and protein expression levels of YTHDC1 and YTHDF2 were significantly reduced in the synovial tissue of RA patients. Furthermore, silencing YTHDC1 in fibroblast-like synoviocytes (FLSs) significantly inhibited their migration, invasion, proliferation, and induced apoptosis.ConclusionYTHDC1 may potentially be involved in the pathogenesis of RA through its regulation of migration, invasion, proliferation, and apoptosis in FLSs.

N-(4-methoxyphenyl) quinoline-8-sulfonamide reduces the inflammatory response of fibroblast-like synoviocytes by targeting receptor (calcitonin) activity modifying protein 1 in rheumatoid arthritis

BackgroundRheumatoid arthritis (RA) is characterized by chronic inflammation of the synovium of joints. Fibroblast-like synoviocytes (FLS) play an important role in RA pathogenesis. We aimed to investigate the effect of N-(4-methoxyphenyl) quinoline-8-sulfonamide (QS-3g) on the inflammatory response of FLS and explore the potential underlying mechanisms. MethodsWe screened and found that QS-3g exhibits the best anti-inflammatory response against FLS using the CCK8 assay. To investigate the therapeutic effects of QS-3g on K/BxN STA mice, we used H&E staining, immunofluorescence, immunohistochemistry, micro-CT, and other techniques. Additional investigations, including RNA-seq, molecular docking, and CETSA, revealed that QS-3g binds to RAMP1. ResultsAmong a series of 8-quinoline sulfonyl amide derivatives, QS-3g reduced the inflammatory response in TNF-α stimulated FLS, such as the release of interleukin (IL)-1β and IL-6. H&E staining and micro-CT showed that QS-3g inhibited synovial hypertrophy, inflammatory cell infiltration, and bone destruction. RNA-seq and CETSA analyses revealed the targeted inhibition of RAMP1 by QS-3g. Inhibition of RAMP1 expression could reduce IL-6 and IL-1β levels. Compared with RAMP1-si, combined administration of QS-3g and RAMP1-si reduced the TNF-α-induced inflammation in TNF-α stimulated FLS without statistically significant differences. Finally, the results of in vitro experiments showed that QS-3g could restore the balance of Gαi/Gαs by inhibiting Gαi and activating Gαs and up-regulate the expression of cAMP protein, thus inhibiting the RAMP1-mediated inflammatory response in FLS. ConclusionQS-3g could inhibit RAMP1 activity and mediate the Gαs/Gαi–cAMP pathway to reduce FLS inflammatory response. Therefore, QS-3g may serve as a novel anti-inflammatory compound for treating RA.

Aberrant overexpression of the autoantigen protein vimentin promotes Th17 cell differentiation and autoimmune arthritis via activation of STAT3 signaling

Vimentin contributes to the positioning and function of organelles, cell migration, adhesion, and division. However, secreted vimentin accumulates on the cell surface (Mor-Vaknin et al., 2003; Ramos et al., 2020 [1,2]) where it acts as a coreceptor for viral infection and as an autoantigen in inflammatory and autoimmune diseases. The roles of vimentin in Th17 cells were examined in mice with knockdown of vimentin. We also examined whether STAT3 is required for vimentin expression.Vimentin expression was significantly increased in Th17 cells through STAT3 activation, and vimentin+ IL-17+ T cells were markedly increased in the joint and spleen tissues of CIA mice. The arthritis score and expression levels of proinflammatory cytokines were significantly decreased in CIA mice treated with vimentin shRNA vector.In this study, we demonstrated that vimentin is significantly expressed in Th17 cells through STAT3 activation. Our results provide new insights into the role of vimentin in Th17 cells and the complex pathogenesis of RA.

Intra-Articular Delivery of an AAV-Anti-TNF-α Vector Alleviates the Progress of Arthritis in a RA Mouse Model.

Rheumatoid arthritis (RA) is a chronic, inflammatory autoimmune disease marked by joint destruction and functional impairment. Tumor necrosis factor (TNF) plays a critical role in RA pathogenesis. Although TNF-targeting drugs are clinically effective, their need for frequent and long-term administration often results in poor patient adherence and suboptimal outcomes. This study developed a gene therapy approach using engineered adeno-associated virus (AAV) vectors to deliver an anti-TNF agent directly into the joint cavity of RA animal models. Animals receiving this therapy demonstrated sustained improvement in clinical scores, inflammatory markers, and joint tissue health. Immunofluorescence staining revealed that AAV vectors could transduce various cell types, including T cells, type A synoviocytes, and dendritic cells. Our results indicate that a single administration of this gene therapy provided long-term efficacy. This suggests that AAV-mediated anti-TNF gene therapy can offer prolonged relief from clinical symptoms and reduce inflammatory damage in a mouse model of RA. This innovative approach presents a promising new therapy with significant clinical prospects to treat patients with RA.

Identification of diagnostic biomarkers of rheumatoid arthritis based on machine learning-assisted comprehensive bioinformatics and its correlation with immune cells

BackgroundRheumatoid arthritis (RA) is a chronic systemic autoimmune disease characterized by inflammatory cell infiltration, which can lead to chronic disability, joint destruction and loss of function. At present, the pathogenesis of RA is still unclear. The purpose of this study is to explore the potential biomarkers and immune molecular mechanisms of rheumatoid arthritis through machine learning-assisted bioinformatics analysis, in order to provide reference for the early diagnosis and treatment of RA disease. MethodsRA gene chips were screened from the public gene GEO database, and batch correction of different groups of RA gene chips was performed using Strawberry Perl. DEGs were obtained using the limma package of R software, and functional enrichment analysis such as gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), disease ontology (DO), and gene set (GSEA) were performed. Three machine learning methods, least absolute shrinkage and selection operator regression (LASSO), support vector machine recursive feature elimination (SVM-RFE) and random forest tree (Random Forest), were used to identify potential biomarkers of RA. The validation group data set was used to verify and further confirm its expression and diagnostic value. In addition, CIBERSORT algorithm was used to evaluate the infiltration of immune cells in RA and control samples, and the correlation between confirmed RA diagnostic biomarkers and immune cells was analyzed. ResultsThrough feature screening, 79 key DEGs were obtained, mainly involving virus response, Parkinson's pathway, dermatitis and cell junction components. A total of 29 hub genes were screened by LASSO regression, 34 hub genes were screened by SVM-RFE, and 39 hub genes were screened by Random Forest. Combined with the three algorithms, a total of 12 hub genes were obtained. Through the expression and diagnostic value verification in the validation group data set, 7 genes that can be used as diagnostic biomarkers for RA were preliminarily confirmed. At the same time, the correlation analysis of immune cells found that γδT cells, CD4+ memory activated T cells, activated dendritic cells and other immune cells were positively correlated with multiple RA diagnostic biomarkers, CD4+ naive T cells, regulatory T cells and other immune cells were negatively correlated with multiple RA diagnostic biomarkers. ConclusionsThe results of novel characteristic gene analysis of RA showed that KYNU, EVI2A, CD52, C1QB, BATF, AIM2 and NDC80 had good diagnostic and clinical value for the diagnosis of RA, and were closely related to immune cells. Therefore, these seven DEGs may become new diagnostic markers and immunotherapy markers for RA.

Identification of critical genes and metabolic pathways in rheumatoid arthritis and osteoporosis toward drug repurposing

BackgroundRheumatoid arthritis (RA) and osteoporosis (OP) are considered to be complex diseases. In recent studies, a positive association between RA and OP has been reported triggering growing research interest. This study aims to investigate the drugs related to critical genes in RA and OP, using bioinformatics approaches, toward drug repurposing. MethodRA and OP genes were identified. The RA–OP PPI network was constructed and analyzed using the STRING and Cytoscape, respectively. Hub genes and modules were extracted and enriched Gene Ontology, through the WebGestalt and g:Profiler. The identification of the drugs related to critical genes using the DGIDB, and extracted the miRNAs using miRWalk and miRNet. ResultsBy network clustering, five significant modules were obtained that have important roles in the immune system. IL6, TNF, IL1B, STAT3, TGFB1, TP53, HIF1A, CCL2, IL10, and MMP9 were found as the top 10 hub genes in the RA-OP network. Hub genes were shown to have implications in inflammatory response, significant functions in cytokine receptor binding, and localized mostly in extracellular space. By investigating the drugs related to hub genes, 16 drugs were identified as repurposing candidate drugs. The 10 drugs included Hydroxychloroquine, Infliximab, Adalimumab, Etanercept, Certolizumab, Cyclosporine, Diacerein, Gevokizumab, Canakinumab, and Olokizumab proposed for OP. Also, six drugs including Pirfenidone, Pentoxifylline, Vadimezan, Rilonacept, Metelimumab, and Siltuximab have important roles in inflammatory control and were proposed for both RA and OP. ConclusionsThe results of the present study can provide novel insights into the pathogenesis and treatment of RA and OP.

Proinflammatory cytokines in rheumatoid arthritis: dynamics against therapy with Janus kinase and interleukin 6 inhibitors

Increased production of proinflammatory cytokines in serum and synovial fluid plays an important role in the pathogenesis of RA. JAK inhibitors and bDMARD are aimed at suppressing various pathological reactions caused by them.The aim of the study. To determine the effect of therapy with JAK and IL-6 inhibitors on the concentration of proinflammatory cytokines in RA patients in real clinical practice.Materials and methods. The study included 30 patients with a reliable diagnosis of RA, advanced stage of disease, with moderate or high RA activity and ineffectiveness of previous therapy with csDMARD or bDMARD for at least 6 months. 10 patients received TOFA at a dose of 5 mg twice daily 10 received UPA at a dose of 15 mg once daily and 10 were on OKZ therapy at a dose of 64 mg subcutaneously every 4 weeks. Studies were performed before treatment, after 3 and 6 months of therapy. The levels of IL-1β, IL-6, IL-17A, IL-17F, IL-23, IL-31, IL-33, INF-γ, TNF-α in serum were investigated using multiplex xMAR technology on Bio-PlexTM 200 System analyser (BIO-RAD, USA).Results. In all groups of patients after 3 and 6 months from the start of therapy, there was a significant decrease in the RA activity index compared to baseline values. The concentration of IL-1β, IL-17A, IL-17F and IL-23 did not change significantly during treatment with any of the drugs. IL-6 values on TOFA background significantly decreased after 3 and 6 months of follow-up compared to the baseline level. UPA therapy had no effect on IL-6 level during the whole observation period, and against the background of OKZ application its values significantly increased after 3 months, and after 6 months – decreased, remaining higher than the initial values. The concentration of IL-31 after 3 months of TOFA treatment significantly decreased (respectively: 6.95 (3.85; 17.72) pg/ml and 3.00 (1.50; 3.85) pg/ml, p<0.05), and after 6 months – increased, but remained lower than baseline (5.09 (3.85; 6.33) pg/ml, p<0.05). IL-33 level on the background of UPA decreased and after 6 months was significantly lower than baseline (1.11 (0.86; 3.95) pg/ ml; 1.05 (0.37; 3.95) and 0.37 (0.12; 1.23) pg/ml, p<0.05). The concentration of INF-γ after 3 and 6 months of TOFA administration decreased significantly compared to the start of therapy (2.05 (1.48; 3.19) pg/ml; 0.99 (0.49; 2.05) pg/ml and 0.99 (0.49; 2.62) pg/ml, p<0.05). Treatment with OKZ resulted in increased TNF-α levels after 6 months compared to baseline values of 0.79 (0.41; 0.98) pg/ml and 1.23 (0.67; 2.06) pg/ml, p<0.05.Conclusions. The use of TOFA, UPA and OKZ in RA patients has a positive effect on disease activity, but has different effects on the level of proinflammatory cytokines in serum.

Association of circulating micro-RNAs (miR-21-5p and miR-142-3p (and tumor necrosis factor-α with Rheumatoid Arthritis

Micro-RNAs (miRNAs) become a promising biomarker for diagnosis due to their stability and reproducibility and could be help to find a new effective treatment. This study is designed to evaluate the diagnostic utility and clinical significance of circulating micro-RNAs (miR-21-5p and miR-142-3p (and tumor necrosis factor-α (TNF-α) for patient with RA and to assess their correlation with disease activity. This case control study involved sixty patients and 30 apparently healthy subjects as control group. The diagnosis was done depending on the classification criteria of American college of rheumatology-European league against rheumatism (ACR- EULAR). Expression levels of circulating miRNA were determined by two step reversed transcription polymerase chain reactions (RT- PCR). The concentrations of TNF-α, anti-citrullinated protein antibodies(ACPA) and C- reactive protein(CRP) were estimated by Enzyme linked immune-sorbent assay technique (ELISA). The expression level of circulating miRNA- 21- 5p and miRNA - 142- 3p was significantly elevated (P <0.01) in patients group as well as the levels of blood ESR, serum CRP, ACPA, and TNF-α compared to controls. There are significant differences (P < 0.01) in miRNA- 21- 5p levels between different patients groups, while the expression of miR-142-3p show non-significant difference (p>0.05). It is found that miRNA- 21-5p expression levels are significantly correlated with RA activity. Both miR-21-5p and miR-142-3p have significant positive correlation with levels of CRP, ACPA, and TNF-α. The level of TNF-α have significant positive correlation with both of CRP and ACPA also with the DAS-28 and CDAI. In conclusion, circulating miR-21- 5p and miRNA - 142- 3p may be associated to RA pathogenesis due to their positive correlation with inflammatory cytokine TNF-α, CRP and ACPA, therefore these miRNAs might be considered new targets for RA treatment, and alterations in their expression could be used for diagnosis and monitoring the disease activity and treatment response.

Association of Rheumatoid Arthritis with Glucose-6-Phosphate Dehydrogenase Deficiency: Results from a Case-Control Study.

Carriers of G6PD deficiency were at an increased risk of RA. This finding opens new windows to better understanding the RA pathogenesis.

Bibliometric analysis of the top 100 most cited articles on Th17/Treg balance and rheumatoid arthritis

ObjectiveRheumatoid arthritis (RA) is an autoimmune disease. The role of Th17/Treg balance in RA pathogenesis has been increasingly emphasized. In this study, bibliometric and visualization analyses of the top 100 most cited articles on Th17/Treg balance in the field of RA were conducted. MethodsBy searching the Web of Science Core Collection database, the top 100 most cited articles of related studies were included, and the authors, countries, institutions, journals, keywords and other information were extracted for analysis using VOSviewer software. ResultsThe top 100 most cited papers had a total of 7185 citations, with an average citation frequency of 72 (range 21–730). All of them were published between 2011 and 2022. The most influential paper, with 730 citations, was written by “Komatsu, Noriko” in 2014 and published in NATURE MEDICINE. The author with the highest output was “Cho, Mi-La” (n = 24). China was the country with the highest number of publications (n = 42). Catholic University of Korea was the institution with the highest number of publications (n = 24). ARTHRITIS AND RHEUMATISM (n = 7), ARTHRITIS & RHEUMATOLOGY (n = 7) and INTERNATIONAL IMMUNOPHARMACOLOGY (n = 7) were the journals that published the most literature. “Expression” (cytokines and transcription factors, etc) and “differentiation” (T cells, Treg cells, and Th17 cells) were the themes of the research. “Mechanisms”, “gut microbiota”, “STAT3”, “interleukin-6”, “synovial fibroblasts” were the hot spots of research in recent years. ConclusionsFor the first time, the top 100 most cited articles were analyzed using bibliometric methods. We aimed to grasp the current development and research trends of RA and Th17/Treg-related studies. It is hoped that this study will provide direction and support for future research.

Xiaowugui decoction alleviates experimental rheumatoid arthritis by suppressing Rab5a-mediated TLR4 internalization in macrophages

BackgroundRheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by exacerbated synovial inflammation and joint destruction. Recent studies suggest toll-like receptor 4 (TLR4) internalization facilitate inflammatory response of macrophage. The role of TLR4 internalization in the pathogenesis of RA is unknown. PurposeTo investigate the role and mechanism of TLR4 internalization in macrophage inflammatory response of RA and explore whether TLR4 internalization mediates the anti-arthritic effect of Xiaowugui (XWG) decoction, a patented herbal formula used in China. MethodsThe co-expression of TLR4 and the internalization marker, early endosome antigen 1 (EEA1), in the synovial samples of RA patients and joint tissue of collagen-induced arthritis (CIA) mice, were evaluated using immunofluorescence. The effect of Rab5a-mediated early internalization of TLR4 on the activation induced by lipopolysaccharide (LPS) in RAW264.7 cells was investigated using small interfering RNAs that act against Rab5a. CIA was induced in Rab5a−/− mice to evaluate the role of Rab5a in vivo. The disease progression and expression of Rab5a and TLR4 in the joint tissue were evaluated in CIA mice treated with XWG. Inflammatory factors production, TLR4 internalization, and activation of downstream signaling pathways were examined in RAW264.7 cells treated with XWG in vitro. ResultsThe co-expression and co-localization of TLR4 and EEA1 were elevated in the synovial samples of RA patients and joint tissue of CIA mice. Pharmaceutical inhibition of TLR4 internalization reduced macrophages inflammatory responses induced by LPS. The co-expression and co-localization of Rab5a and TLR4 were significantly increased in macrophages treated with LPS. Silencing Rab5a reduced LPS-induced TLR4 internalization, inflammatory factors production, and phosphorylation of Jun N-terminal kinases (JNK) and p65. Genetic deletion of Rab5a inhibited TLR4 internalization and the development of arthritis in vivo. The co-expression of TLR4 and Rab5a was also elevated in the synovial samples of RA patients. XWG treatment of mice with CIA alleviated arthritis and reduced the co-expression of Rab5a and TLR4 in the joint tissue. XWG treatment of macrophage inhibited LPS-induced IL-6 and TNF-α production, co-expression of Rab5a and TLR4, and phosphorylation of JNK and p65. ConclusionsOur findings highlight the pathogenic role of TLR4 internalization in patients with RA and identify a novel Rab5a-dependent internalization pathway that promotes macrophage inflammatory response. XWG treatment demonstrated outstanding therapeutic effects in experimental arthritis, and targeting the Rab5a-mediated internalization of TLR4 may be the main underlying mechanism.

Elderly-onset rheumatoid arthritis — clinical findings and treatment features (systematic literature review)

Background. Elderly-onset rheumatoid arthritis ­(EORA) is characterized by the disease onset after 60 years. Gi­ven the importance of the global increase in the proportion of older people in the population and the potential consequences, the problem of diagnosis and treatment of EORA is quite relevant. Purpose: to analyze the current literature data on the peculiarities of the clinical picture, differential diagnosis, and treatment of patients with EORA, taking into account comorbidity. Materials and methods. An analytical review of literature data was conducted using information analysis of the databases PubMed, Web of Science, Scopus, and Google Scholar for the period 2013–2023, but it did not exclude key works that were published earlier, using the keywords “elderly-onset rheumatoid arthritis”, “rheumatoid arthritis”, “diagnosis”, “treatment”, “prognosis”. Results. A review of the lite­rature demonstrated that while previous studies have mainly argued that EORA is a milder form of the disease with a favorable prognosis, recent studies have shown greater disease activity and severity, as well as worse clinical, functional, and radiological outcomes. Despite recent advances in the understanding of RA pathogenesis and new treatment strategies, there is still controversy regarding the management of EORA patients. Clinical practice shows that most patients with EORA are prescribed purely nonsteroidal anti-inflammatory drugs (NSAID) and glucocorticoids (GC) without the addition of disease-modifying antirheumatic drugs (DMARD). Real-world data have demonstrated that the treatment of elderly patients with RA is often unsatisfactory due to concerns about the possible side effects of DMARD, the presence of comorbidities, polypharmacy, and cognitive dysfunction in elderly patients. Conclusions. EORA presents a unique clinical profile, such patients require individualized treatment strategies, mandatory addition or switch to DMARD based on disease activity, comorbidities, and safety considerations to optimize treatment outcomes and minimize GC and NSAIDs intake, thereby improving the quality of treatment of elderly patients.

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.

Rapamycin suppresses rheumatoid arthritis fibroblast synovial cell proliferation and induces apoptosis via the AKT/mTORC1 pathway

AbstractBackgroundRheumatoid arthritis (RA) is an autoimmune disease characterized by the overproliferation of synovial fibroblast‐like synoviocytes (FLSs) in the lining, leading to chronic inflammation and progressive joint damage. RA pathogenesis involves lymphocyte infiltration, increased synovial cell proliferation, and impaired cell death. This study investigated the effects of rapamycin on RA‐FLSs and explored the underlying molecular mechanisms.MethodsThe optimal drug concentration and time for rapamycin administration were determined using a cell counting kit‐8 assay. The concentration of rapamycin varied from 1 to 25 nmol/L. The mRNA expression levels of protein kinase B (AKT), mammalian target of rapamycin (mTOR), B cell lymphoma 2 (Bcl‐2), and Bcl‐2‐associated X protein (Bax) were quantified by real‐time polymerase chain reaction. Protein expression (p‐AKT, AKT, p‐mTOR, mTOR, pS6 kinase [S6K], S6K, p‐4E‐binding protein 1 [4EBP1], 4EBP1, Bcl‐2, Bax, caspase 3, caspase 9, cyclin‐dependent kinase 2 [CDK2], and CD1) were detected by Western blotting analysis.ResultsRapamycin suppresses RA‐FLS proliferation and induces apoptosis in a dose‐dependent manner. Rapamycin significantly elevated the protein expression of Bax (p &lt; 0.01), caspase 3 (p &lt; 0.05), and caspase 9 (p &lt; 0.001), and downregulated Bcl‐2 expression (p &lt; 0.01). Rapamycin increased Bax mRNA expression (p &lt; 0.01) and decreased Bcl‐2 (p &lt; 0.05), AKT (p &lt; 0.05), and mTOR (p &lt; 0.05) expression. Additionally, there was a marked reduction in the expression levels of p‐AKT(Ser473) (p &lt; 0.01), p‐mTOR (Ser2448) (p &lt; 0.01), p‐S6K1 (p &lt; 0.01), and p‐4EBP1, CDK2, and CD1 (p &lt; 0.01).ConclusionsRapamycin effectively inhibited proliferation and induced apoptosis of RA‐FLSs by targeting the AKT/mTORC1 pathway. These findings underscore the potential of rapamycin as a therapeutic candidate for addressing the dysregulated proliferation and inflammatory characteristics of RA. Further clinical investigations are required to validate its application in the management of RA.

Targeting TNF-α-induced expression of TTR and RAGE in rheumatoid arthritis: Apigenin's mediated therapeutic approach

BackgroundRheumatoid arthritis (RA) is a chronic inflammatory disease induced by TNF-α, which increases fibroblast-like synoviocytes inflammation, resulting in cartilage destruction. The current work sought to comprehend the pathophysiological importance of TNF-α stimulation on differential protein expression and their regulation by apigenin using in-vitro and in-vivo models of RA. MethodsThe human RA synovial fibroblast cells were stimulated with or without TNF-α (10 ng/ml) and treated with 40 μM apigenin. In-silico, in-vitro and in-vivo studies were performed to confirm the pathophysiological significance of apigenin on pro-inflammatory cytokines and on differential expression of TTR and RAGE proteins. ResultsTNF-α induced inflammatory response in synoviocytes revealed higher levels of IL-6, IL-1β, and TNF-α cytokines and upregulated differential expression of TTR and RAGE. In-silico results demonstrated that apigenin has a binding affinity towards TNF-α, indicating its potential effect in the inflammatory process. Both in-vitro and in-vivo results obtained by Western Blot analysis suggested that apigenin reduced the level of p65 (p = 0.005), TTR (p = 0.002), and RAGE (p = 0.020). ConclusionThe findings of this study suggested that TNF-α promotes the differential expression of pro-inflammatory cytokines, TTR, and RAGE via NF-kB pathways activation. Anti-inflammatory effect of apigenin impedes TNF-α mediated dysregulation or expression associated with RA pathogenesis.

Current Pathogenesis and Treatment of Autoimmune Diseases

Nowadays, autoimmune diseases(AD) still torment many people and they always suffer from severe symptoms and complications. However, the pathogenesis of AD is still unclear and needs more research. Also, there are shortages of treatments which can totally cure AD. This research mainly focuses on current findings of possible pathogenesis and treatments for AD by discussing and analyzing two of most famous AD: rheumatoid arthritis and systemic lupus erythematosus (SLE). This essay discusses pathogenesis of RA and SLE from three aspects: genes, immune systems and environmental factors, and then concludes current medications and therapies as well as their drawbacks. This study finds that susceptible genes, abnormal immune responses and environmental stimulation can synergistically lead to autoimmune diseases, and existing treatments are mainly composed of symptomatic treatments and modifying treatments. But each medication has its inevitable side effects. Furthermore, more research should focus on systematic treatments with fewer drawbacks in order to truly cure AD.

PIM Kinases as Potential Biomarkers and Therapeutic Targets in Inflammatory Arthritides.

The Proviral Integration site for the Moloney murine leukemia virus (PIM)-1 kinase and its family members (PIM-2 and PIM-3) regulate several cellular functions including survival, proliferation, and apoptosis. Recent studies showed their involvement in the pathogenesis of rheumatoid arthritis RA, while no studies are available on psoriatic arthritis (PsA) and axial spondyloarthritis (axSpA). The main objective of this study is to assess the expression of PIM kinases in inflammatory arthritides, their correlation with proinflammatory cytokines, and their variation after treatment with biologic disease-modifying anti-rheumatic drugs or JAK inhibitors. We evaluated PIM-1, -2, and -3 expression at the gene and protein level, respectively, in the peripheral blood mononuclear cells and serum of patients with RA, PsA, axSpA, and healthy individuals (CTR). All the samples showed expression of PIM-1, -2, and -3 kinases both at the gene and protein level. PIM-1 was the most expressed protein, PIM-3 the least. PIM kinase levels differed between controls and disease groups, with reduced PIM-1 protein and increased PIM-3 protein in all disease samples compared to controls. No difference was found in the expression of these molecules between the three different pathologies. PIM levels were not modified after 6 months of therapy. In conclusion, our preliminary data suggest a deregulation of the PIM pathway in inflammatory arthritides. In-depth studies on the role of PIM kinases in this field are warranted.

Pathologically expanded peripheral CD4+PD‐1+Foxp3− T‐cell subset promotes B‐cell hyperactivity in patients with rheumatoid arthritis

AbstractBackgroundRheumatoid arthritis (RA) is an autoimmune disease characterized by destructive polyarthritis, and abnormal T–B‐cell interactions may contribute to its pathogenesis. This study aimed to investigate the characteristics and roles of CD4+ programmed death 1 (PD‐1)+Foxp3− T cells in relation to the B‐cell response in patients with RA.MethodsThis study included 155 patients with RA and 36 age‐ and sex‐matched healthy controls (HCs) from the Second Hospital of Dalian Medical University in China. Flow cytometry was used to assess the proportion and properties of peripheral CD4+PD‐1+Foxp3+ T cells, including their proliferation, activation, cytokine production, and capacity to induce B‐cell differentiation.ResultsThe proportion of CD4+PD‐1+Foxp3− T cells was increased in patients with RA compared with HCs ([10.78 ± 0.60]% vs. [5.67 ± 0.40]%, p &lt; 0.001), and this was positively associated with the B‐cell response. Compared with CD4+PD‐1+Foxp3+ T cells, CD4+PD‐1+Foxp3− T cells from patients with RA exhibited increased expression of Ki67 ([6.52 ± 0.41]% vs. [3.87 ± 0.42]%, p &lt; 0.01) and activation markers, produced higher levels of cytokines, and showed enhanced B‐cell differentiation. Furthermore, anti‐interleukin‐6R antagonists decreased the proportion, activation, and cytokine production of CD4+PD‐1+Foxp3− T cells in vitro. The frequency of type 2 CD4+PD‐1+Foxp3− T cells was significantly higher in patients with RA than that in HCs ([37.27 ± 1.43]% vs. [29.05 ± 1.30]%, p &lt; 0.05).ConclusionsPeripherally expanded CD4+PD‐1+Foxp3− T cells in patients with RA, which induced B‐cell hyperactivity, may be inclined toward type 2 helper T cells. Our findings revealed a novel T‐cell subset that contributes to B‐cell hyperactivity in the pathogenesis of RA.

Comprehensive Co-Inhibitory Receptor (Co-IR) Expression on T Cells and Soluble Proteins in Rheumatoid Arthritis.

Co-IRs T cells were immunophenotyped employing multicolor flow cytometry, and ELISA was utilized for measuring soluble PD-1, PDL-2, and TIM3. Correlations have been detected across the percentage of T cells expressing Co-IRs (MFI) and different indicators in the blood, including ESR, high-sensitivity CRP (hsCRP), 28 joint disease activity scores (DAS28), and soluble PD-1/PDL-2/TIM3. In RA patients, we recognized elevated levels of PD-1 (CD279), CTLA-4, and TIGIT in CD4+ T cells; TIGIT, HLA-DR, TIM3, and LAG3 in CD8+ T cells; and CD8+CD279+TIM3+, CD8+HLA-DR+CD38+ T cells. The following tests were revealed to be correlated with hsCRP: CD4/CD279 MFI, CD4/CD279%, CD4/TIM3%, CD8/TIM3%, CD8/TIM3 MFI, CD8/LAG3%, and CD8+HLA-DR+CD38+%. CD8/LAG3 and CD8/TIM3 MFIs are linked to ESR. DAS28-ESR and DAS28-CRP exhibited relationships with CD4/CD127 MFI, CD8/CD279%, and CD8/CD127 MFI, respectively. CD4+CD279+TIM3+% was correlated with DAS28-ESR (p = 0.0084, N = 46), DAS28-CRP (p = 0.007, N = 47), and hsCRP (p = 0.002, N = 56), respectively. In the serum of patients with RA, levels of soluble PD-1, PDL-2, and Tim3 were extremely elevated. CD4+ TIM3+% (p = 0.0089, N = 46) and CD8+ TIM3+% (p = 0.0305, N = 46) were correlated with sTIM3 levels; sPD1 levels were correlated with CD4+CD279+% (p < 0.0001, N = 31) and CD3+CD279+% (p = 0.0084, N = 30). Co-IR expressions on CD4+ and CD8+ T cells, as well as soluble PD-1, PDL-2, and TIM3 levels, could function as indicators of disease activity and potentially play crucial roles in the pathogenesis of RA.

Comprehensive analysis revealed the immunoinflammatory targets of rheumatoid arthritis based on intestinal flora, miRNA, transcription factors, and RNA-binding proteins databases, GSEA and GSVA pathway observations, and immunoinfiltration typing

ObjectiveRheumatoid arthritis (RA) is a chronic inflammatory arthritis. This study aimed to identify potential biomarkers and possible pathogenesis of RA using various bioinformatics analysis tools.MethodsThe GMrepo database provided a visual representation of the analysis of intestinal flora. We selected the GSE55235 and GSE55457 datasets from the Gene Expression Omnibus database to identify differentially expressed genes (DEGs) separately. With the intersection of these DEGs with the target genes associated with RA found in the GeneCards database, we obtained the DEGs targeted by RA (DERATGs). Subsequently, Disease Ontology, Gene Ontology, and the Kyoto Encyclopedia of Genes and Genomes were used to analyze DERATGs functionally. Gene Set Enrichment Analysis (GSEA) and Gene Set Variation Analysis (GSVA) were performed on the data from the gene expression matrix. Additionally, the protein-protein interaction network, transcription factor (TF)-targets, target-drug, microRNA (miRNA)-mRNA networks, and RNA-binding proteins (RBPs)-DERATGs correlation analyses were built. The CIBERSORT was used to evaluate the inflammatory immune state. The single-sample GSEA (ssGSEA) algorithm and differential analysis of DERATGs were used among the infiltration degree subtypes.ResultsThere were some correlations between the abundance of gut flora and the prevalence of RA. A total of 54 DERATGs were identified, mainly related to immune and inflammatory responses and immunodeficiency diseases. Through GSEA and GSVA analysis, we found pathway alterations related to metabolic regulations, autoimmune diseases, and immunodeficiency-related disorders. We obtained 20 hub genes and 2 subnetworks. Additionally, we found that 39 TFs, 174 drugs, 2310 miRNAs, and several RBPs were related to DERATGs. Mast, plasma, and naive B cells differed during immune infiltration. We discovered DERATGs’ differences among subtypes using the ssGSEA algorithm and subtype grouping.ConclusionsThe findings of this study could help with RA diagnosis, prognosis, and targeted molecular treatment.