PMID- 32373221 OWN - NLM STAT- MEDLINE DCOM- 20210726 LR - 20210726 IS - 1838-7640 (Electronic) IS - 1838-7640 (Linking) VI - 10 IP - 12 DP - 2020 TI - Identification of a novel microRNA-141-3p/Forkhead box C1/beta-catenin axis associated with rheumatoid arthritis synovial fibroblast function in vivo and in vitro. PG - 5412-5434 LID - 10.7150/thno.45214 [doi] AB - Rationale: Rheumatoid arthritis (RA) is a prototype of inflammatory arthritis in which synovial fibroblasts (SFs) play key roles in cartilage and bone destruction through tumor-like proliferation, migration, invasion and inflammation. This study aimed to research forkhead box protein C1 (FoxC1) and microRNA (miR)-141-3p, which modulate pathological changes in the synovial membrane, to find possible strategies for treating RA. Methods: FoxC1, beta-catenin and miR-141-3p gene expression in synovial tissues and SFs was quantified by real-time PCR; FoxC1 and beta-catenin protein levels were evaluated by immunohistochemistry, immunofluorescence, and Western blotting. We transiently transfected human SFs with FoxC1 and beta-catenin overexpression and silencing vectors and assessed proliferation, migration, invasion and inflammation by cell function and enzyme-linked immunosorbent assays. We also assessed downstream signaling activation using immunofluorescence, real-time PCR and Western blotting. Double luciferase, coimmunoprecipitation and chromatin immunoprecipitation assays were used to verify miR-141-3p, FoxC1 and beta-catenin gene and protein combinations. Finally, the therapeutic effects of FoxC1 silencing and miR-141-3p overexpression were evaluated in type II collagen-induced arthritis (CIA) rats. Results: We found that FoxC1 expression was significantly upregulated in synovium and SFs in both RA patients and rats with collagen-induced arthritis (CIA). FoxC1 overexpression increased beta-catenin messenger RNA (mRNA) and protein levels and upregulated cyclin D1, c-Myc, fibronectin and matrix metalloproteinase 3 (MMP3) mRNA and protein expression in RA SFs (RASFs). In contrast, FoxC1 knockdown reduced beta-catenin mRNA and protein levels as well as cyclin D1, c-Myc, and fibronectin mRNA and protein levels in RASFs. Furthermore, altering FoxC1 expression did not significantly change GSK3beta and pGSK3beta levels. FoxC1 overexpression promoted proliferation, migration, invasion and proinflammatory cytokine (interleukin (IL)-1beta, IL-6, and tumor necrosis factor-alpha (TNF-alpha)) production and reduced anti-inflammatory cytokine (IL-10) levels in RASFs. FoxC1 bound to the beta-catenin promoter, and beta-catenin mediated the FoxC1-induced pathological changes. We also observed downregulated microRNA (miR)-141-3p expression in SFs from both RA patients and CIA rats and further found that miR-141-3p bound to the FoxC1 3'UTR and suppressed FoxC1 expression. Intra-ankle miR-141-3p agomir or FoxC1-specific siRNA injection hindered CIA development in rats. Conclusions: FoxC1 and miR-141-3p participate in RA pathogenesis by mediating inflammation and SF proliferation, migration, and invasion and thus could be novel targets for RA therapy as a nonimmunosuppressive approach. CI - (c) The author(s). FAU - Wang, Jun AU - Wang J AD - Department of Orthopaedics, The First Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Wang, Yin AU - Wang Y AD - Department of Plastic Surgery, The Fourth Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Zhang, Hui AU - Zhang H AD - Department of Orthopaedics, The First Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Chang, Jun AU - Chang J AD - Department of Orthopaedics, The Fourth Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Lu, Ming AU - Lu M AD - Department of Orthopaedics, The First Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Gao, Weilu AU - Gao W AD - Department of Orthopaedics, The First Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Liu, Wendong AU - Liu W AD - Department of Radiology, The First Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Li, Yetian AU - Li Y AD - Department of Orthopaedics, The First Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Yin, Li AU - Yin L AD - Department of Orthopaedics, The First Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Wang, Xiaohe AU - Wang X AD - Department of Orthopaedics, The First Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Wang, Yuejun AU - Wang Y AD - Department of Pathology, The Fourth Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Gao, Mengru AU - Gao M AD - Department of Pathology, The Fourth Affiliated Hospital of Anhui Medical University, Anhui, China. FAU - Yin, Zongsheng AU - Yin Z AD - Department of Orthopaedics, The First Affiliated Hospital of Anhui Medical University, Anhui, China. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20200406 PL - Australia TA - Theranostics JT - Theranostics JID - 101552395 RN - 0 (CTNNB1 protein, human) RN - 0 (FOXC1 protein, human) RN - 0 (Forkhead Transcription Factors) RN - 0 (MIRN141 microRNA, human) RN - 0 (MicroRNAs) RN - 0 (beta Catenin) SB - IM MH - Animals MH - Arthritis, Rheumatoid/genetics/*metabolism MH - Blotting, Western MH - Cell Proliferation/genetics/physiology MH - Cells, Cultured MH - Female MH - Fibroblasts/*cytology/*metabolism MH - Forkhead Transcription Factors/genetics/*metabolism MH - Humans MH - Immunohistochemistry MH - MicroRNAs/genetics/*metabolism MH - Rats MH - Real-Time Polymerase Chain Reaction MH - beta Catenin/*metabolism PMC - PMC7196314 OTO - NOTNLM OT - FoxC1 OT - collagen-induced arthritis OT - miR-141-3p OT - rheumatoid arthritis OT - synovial fibroblasts COIS- Competing Interests: The authors have declared that no competing interest exists. EDAT- 2020/05/07 06:00 MHDA- 2021/07/27 06:00 PMCR- 2020/01/01 CRDT- 2020/05/07 06:00 PHST- 2020/02/22 00:00 [received] PHST- 2020/03/20 00:00 [accepted] PHST- 2020/05/07 06:00 [entrez] PHST- 2020/05/07 06:00 [pubmed] PHST- 2021/07/27 06:00 [medline] PHST- 2020/01/01 00:00 [pmc-release] AID - thnov10p5412 [pii] AID - 10.7150/thno.45214 [doi] PST - epublish SO - Theranostics. 2020 Apr 6;10(12):5412-5434. doi: 10.7150/thno.45214. eCollection 2020.