Ab0074 rapamycin inhibits il-1β induced ra synovial fibroblast activation, an effect associated with alteration in their metabolic profile
Annals of the Rheumatic Diseases(2022)
摘要
BackgroundRheumatoid Arthritis (RA) is a common autoimmune disease characterized by systemic polyarthritis affecting the joints, most notably of the hands and feet. A fundamental feature of RA is inflammation within the synovial joint due to neo-angiogenesis which facilitates an influx of immune cells and release of pro-inflammatory mediators.ObjectivesIn this study we investigate the effect of metabolic reprogramming on IL-1β regulation of stromal cell activation and invasiveness in RA.MethodsPrimary RA synovial fibroblasts (RAFLS) and human umbilical vein endothelial cells (HUVEC) were cultured with IL-1 β (2 ng/mL) alone or in combination with the metabolic inhibitor rapamycin (100nM). Pro-inflammatory cytokines IL-6, MCP-1 and Rantes were quantified by real-time PCR and ELISA. Cellular adhesion and network formation were quantified by adhesion binding assays and Matrigel invasion assays. pS6 (a surrogate marker of the mTOR pathway) was quantified by flow cytometry. Cellular bioenergetics was assessed using the Seahorse-XFe-technology and key glycolytic genes (HK2, PKM2, G6DP) were quantified by real-time PCR. YAP was measured by Western-Blot.ResultsIL-1β significantly induced secretion of IL-6 (p<0.05), MCP-1 (p<0.01) and Rantes (p<0.01) from RAFLS. IL-1β induced leukocyte adhesion to RAFLS (p=0.062) and HUVEC (p=0.051), in addition to an increase RAFLS and HUVEC network formation. This was accompanied by a change in the cellular bioenergetic profile of cells, where IL-1β increased the ECAR/OCR ratio in favour of glycolysis for RAFLS (p<0.05) and HUVEC, and induced the % frequency in the cell surface expression of pS6 on RAFLS. Rapamycin inhibited IL-1β -induced leukocyte adhesion and network formation in RAFLS (p<0.05; p<0.05 respectively) and HUVEC (p<0.05). Rapamycin inhibited IL-1β -induced Rantes secretion (p<0.05), no effect observed for IL-6 and MCP-1. Rapamycin also inhibited IL-1b-induce YAP protein expression which is involved in FLS invasive capacity through cytoskeletal rearrangement. This was accompanied by a shift in the metabolic profile from a glycolytic/energetic state back towards a more quiescent state.ConclusionRapamycin inhibits IL-1β -induced pro-inflammatory mechanisms in key stromal cells involved in the pathogenesis of RA. Targeting metabolism may lead to new potential therapeutic or adjuvant strategies, particularly for those patients who have sub-optimal responses to current treatments.References[1]Kim EK, Min HK, Lee SY, Kim DS, Ryu JG, Na HS, Jung KA, Choi JW, Park SH, Cho ML. Metformin rescues rapamycin-induced mitochondrial dysfunction and attenuates rheumatoid arthritis with metabolic syndrome. Arthritis Res Ther. 2020 Apr 10;22(1):77. doi: 10.1186/s13075-020-02174-3. PMID: 32276645; PMCID: PMC7149912.Disclosure of InterestsNone declared
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