Toll-like receptor 2 ligand mediates the upregulation of angiogenic factor, vascular endothelial growth factor and interleukin-8/CXCL8 in human rheumatoid synovial fibroblasts.
ABSTRACT Rheumatoid arthritis (RA) is characterized by infiltrations of inflammatory cells accompanied by neovascularization in the joint. We hypothesized that cell activation via the toll-like receptor (TLR) may be involved in the induction of angiogenic molecules, which are relevant to the pathogenesis of RA. RA fibroblast like synoviocytes (FLS) were stimulated with TLR-2 ligand bacterial peptidoglycan (PGN), TLR-4 ligand lipopolysaccharide (LPS) and various cytokines. Vascular endothelial growth factor (VEGF) and IL-8 were measured by ELISA in culture supernatants; mRNA levels were assessed by RT-PCR and real time PCR. The levels of TLR-2, VEGF and IL-8 were analyzed by dual immunohistochemistry in RA synovium and compared with osteoarthritis (OA). Regulation of MyD88, IRAK4, IRAK1, IRAK-M and TRAF-6 mRNA expression levels by PGN were analyzed by RT-PCR. Phosphorylation of I kappa B alpha was evaluated by western blotting. Levels of VEGF and IL-8 were upregulated in culture supernatants of RA FLS stimulated with PGN, similar to the levels of IL-1beta and IL-17 stimulation. Neutralization of TLR-2 with a blocking monoclonal antibody significantly reduced both VEGF and IL-8 levels (P<0.05), which reflected the functional relevance of TLR-2 activation to the induction of VEGF and IL-8 production. Downstream intracellular signaling following TLR-2 stimulation involved MyD88-IRAK-4-TRAF-6 pathways, resulting in NF-kappaB activation. Thus, TLR-2 activation in RA FLS by microbial constituents could be involved in the induction of VEGF and IL-8 and thereby promote inflammation either directly or via angiogenesis. This possibly contributes to the perpetuation of synovitis in patients with RA.
02/2012; , ISBN: 978-953-51-0063-8
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ABSTRACT: Innate immune and inflammatory responses have been implicated in myocardial ischemia/reperfusion (I/R) injury. However, the mechanisms by which innate immunity and inflammatory response are involved in myocardial I/R have not been elucidated completely. Recent studies highlight the role of Toll-like receptors (TLRs) in the induction of innate immune and inflammatory responses. Growing evidence has demonstrated that TLRs play a critical role in myocardial I/R injury. Specifically, deficiency of TLR4 protects the myocardium from ischemic injury, whereas modulation of TLR2 induces cardioprotection against ischemic insult. Importantly, cardioprotection induced by modulation of TLRs involves activation of the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway, suggesting that there is a crosstalk between TLRs and PI3K/Akt signaling pathways. In addition, TLRs also associate with other coreceptors, such as macrophage scavenger receptors in the recognition of their ligands. TLRs are also involved in the induction of angiogenesis, modulation of stem cell function, and expression of microRNA, which are currently important topic areas in myocardial I/R. Understanding how TLRs contribute to myocardial I/R injury could provide basic scientific knowledge for the development of new therapeutic approaches for the treatment and management of patients with heart attack.Antioxidants & Redox Signaling 11/2010; 15(7):1875-93. · 8.20 Impact Factor
Article: Arthritis is associated with T-cell-induced upregulation of Toll-like receptor 3 on synovial fibroblasts.[show abstract] [hide abstract]
ABSTRACT: Toll-like receptors (TLRs) are likely to play crucial roles in the pathogenesis of rheumatoid arthritis (RA). The aim of this study was to determine the key TLRs in synovium and explore their roles in the activation of fibroblast-like synoviocytes (FLSs) mediated by T cells in arthritis. Pristane-induced arthritis (PIA) was established by subcutaneous injection with pristane at the base of the rat's tail. TLR expression in synovium from PIA rats was detected at different time points by performing real-time PCR. Polyinosinic:polycytidylic acid (poly(I:C)) was intra-articularly administrated to PIA rats, and arthritis was monitored macroscopically and microscopically. Synovial TLR3 was detected by immunohistochemical staining. Rat FLSs were stimulated with pristane-primed T cells or pristane-primed, T-cell conditioned medium. The intervention of TLR3 in FLSs was achieved by specific short-hairpin RNA (shRNA) or an antibody. The migration ability of FLSs was measured by using the scratch test, and gene expression was detected by using real-time PCR. FLSs from RA patients were stimulated with various cytokines and TLR ligands, and TLR3 expression was detected by performing real-time PCR. In addition, with different concentrations of poly(I:C) stimulation, TLR3 expression of FLSs from RA patients and patients with osteoarthritis (OA) was compared. Synovium TLR3 displayed early and persistent overexpression in PIA rats. TLR3 was expressed in FLSs, and local treatment with poly(I:C) synergistically aggravated the arthritis. Rat FLSs co-cultured with pristane-primed T cells showed strengthened migration ability and significant upregulation of TLR3, IFN-β, IL-6 and matrix metalloproteinase 3 (MMP3) expression, which could also be induced by pristane-primed, T-cell conditioned medium. The upregulation of cytokines and MMPs was blocked by shRNA or TLR3 antibodies. In RA FLSs with cytokine or TLR ligand stimulation, TLR3 expression exhibited remarkable upregulation. Furthermore, RA FLSs showed higher reactivity than OA FLSs to poly(I:C). TLR3 in the synovium of PIA rats was overexpressed, and activation of the TLR3 signaling pathway could aggravate this arthritis. The induction of TLR3 in FLSs resulted from T cell-derived inflammatory stimulation and could further mediate FLS activation in arthritis. We conclude that TLR3 upregulation of FLSs activated by T cells results in articular inflammation.Arthritis research & therapy 06/2011; 13(3):R103. · 4.27 Impact Factor