The Effects Of LncRNA ENST00000584923 Down-Regulated By LLDT-8 On The Function Of RA-FLS And Its Mechanism

Objective1.To research hot spots and future trends of Tripterygium wilfordii in the treatment of RA by constructing knowledge map.2.To investigate the function of lncRNA ENST00000584923 in RA-FLS in vitro.3.To study the effect and mechanism of RA-FLS function by means of lncRNA,and to further explore the pathogenesis of RA and the therapeutic effect of LLDT-8 on RA.Method1.English literatures on the treatment of RA by Tripterygium wilfordii were retrieved by Web of Science,and Chinese literatures on the treatment of RA by CNKI were retrieved.Cite Space software was used to construct the knowledge map of the treatment of RA by using “author”,“keywords”,“institution” as nodes.2.RA-FLS cell was cultured in vitro and sh RNA lentiviral vector was constructed to interfere with the expression of lncRNA ENST00000584923 in RA-FLS cell.CCK8 experiment,clone formation experiment,transwell invasion experiment,cell scratch experiment,Annexin V/ PI double staining method,ELISA,q RT-PCR and Western Blot were used to determine the function of RA-FLS and the expression of related inflammatory factors and proteins.3.Interference and overexpression of lncRNA ENST00000584923 expression in RA-FLS,adding LLDT-8 stimulation,using CCK8,Transwell invasion,Annexin V/ PI double staining method,ELISA,q RT-PCR,Western Blot measures the function of RA-FLS and related inflammatory factors and protein expression.4.Fluorescence in situ hybridization was used to detect the cellular localization of lncRNA ENST00000584923;RNA-seq was used to detect the m RNA expression profile of RA-FLS cell after respectively determine the interference of LncRNA and LLDT-8 treatment.ELISA、q RT-PCR and Western Blot was used to analyze verified the screened genes and related pathways..Results1.According to the screening criteria,362 English literatures and 1473 Chinese literatures related to the treatment of RA by Tripterygium wilfordii were obtained.The number of literatures on the treatment of RA by Tripterygium wilfordii fluctuates year by year and China is the country with the most published literatures.Author collaboration network formed a research team with Yao Xueming,Huang Ying,Ma Wukai;Lin Na;Jiang Quan as the core;AIPING LU,KAMAL D MOUDGIL,etc.as the core,and further analysis found that the author’s teams are closely connected,but the teams are less connected.Institution collaboration network found that China Academy of Chinese Medical Sciences,Nanjing and Beijing University of Traditional Chinese Medicine have a relatively large number of publications in both Chinese and English.Further analysis reveals that the research is mainly based on universities of Chinese medicine and affiliated hospitals,the inter-organizations are not closely connected.Main research content of Chinese literature is tripterygium glycosides,triptolide,signaling pathways,molecular mechanism;the main research contents of English literature are Celastrol,Wilfordii hook f,Cell,inflammation.The research content at home and abroad mainly focused on the mechanism and effective components of Tripterygium wilfordii in RA treatment.The mechanism is mainly the study of signal pathway,and the active component is mainly concentrated in the study of triptolide.The effective ingredients,mechanism of action,curative effect of Tripterygium wilfordii and the combination of Tripterygium wilfordii and western medicine to treat RA are still the focus of domestic and foreign research.2.According to our previous study,lncRNA ENST00000584923 was up-regulated in RA synovial tissue.Interference with the expression of lncRNA ENST00000584923 could inhibit the proliferation,reduce the clone formation,inhibit the function of invasion,promote the apoptosis,reduce the expression of inflammatory cytokines IL-1and TGF-β1,decrease the expression of Cyclin D1 and increase the expression of pro-apoptotic protein Bax of RA-FLS.3.LLDT-8 can inhibit the cell proliferation of RA-FLS by interfering with the expression of lncRNA ENST00000584923,promote the apoptosis of RA-FLS,inhibit the secretion of IL-1 and TGF-β1,inhibit the expression of Cyclin D1 of RA-FLS,and promote the expression of Bax protein.4.LncRNA ENST00000584923 was distributed in both cytoplasm and nucleus.After the interference of lncRNA ENST00000584923 expression and LLDT-8 treatment,25 common ra-FLs differentially expressed genes were detected.The main functions of co-expression of DEGs are mainly in inflammatory response,immune response,etc.,and the pathways are mainly concentrated in cytokine-cytokine receptor interaction pathway,NOD-like receptor signaling pathway,and rheumatoid arthritis signaling pathway.It was verified that MMP1,c-Jun and p-c-Jun were differentially expressed in the two groups.Conclusion1.At present,the research on the active ingredients of Tripterygium wilfordii and its mechanism in treating RA is a hot research topic.2.Interference with lncRNA ENST00000584923 in RA-FLS cells can inhibit cell proliferation,invasion,promote cell apoptosis,reduce the expression of cytokines IL-1,TGF-β1,and reduce cyclin D1 and promote the expression of Bax.3.LLDT-8 can further affect the RA-FLS cell function that interferes with lncRNA ENST00000584923.Further inhibit cell proliferation,promote cell apoptosis,inhibit the expression of IL-1,TGF-β1,inhibit cyclin D1,and promote the expression of Bax.4.LLDT-8 may play a role by interfering with the further action of lncRNA ENST00000584923 on c-Jun/MMP1.