Abstract
BackgroundNerve growth factor (NGF) contributes to pain in knee osteoarthritis (KOA) patients. Transforming growth factor-beta (TGF-β) stimulates NGF expression in chondrocytes from KOA patients. However, the correlation between synovial TGF-β and NGF levels has not been sufficiently studied in human KOA patients. Further, the mechanism governing NGF regulation by TGF-β in synovial cells is unclear.MethodsDuring total knee arthroplasty, we extracted the synovial tissue (SYT) of 107 subjects with unilateral Kellgren/Lawrence grade 3–4 KOA confirmed by radiography. We examined the distribution of TGF-β and NGF using immunohistochemistry, and analyzed the relationship between NGF and TGFB mRNA levels. Cultured synovial cells extracted from SYT were exposed to culture medium (control), human recombinant TGF-β (rhTGF-β), rhTGF-β + ALK5 inhibitor SB505124, rhTGF-β + transforming growth factor activating kinase 1 (TAK1) inhibitor (5Z)-7-oxozeaenol, or rhTGF-β + p38 inhibitor SB203580 for 30 min, 6 h and 24 h. NGF mRNA expressed by the cultured cells and NGF protein levels in the cell supernatant were detected by real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. Phosphorylation of p38 was evaluated by western blotting.ResultsNGF mRNA levels were positively correlated with those of TGFB. Cells expressing TGF-β and NGF protein were observed in the lining layer of SYT. TGF-β stimulated increased NGF mRNA expression and NGF protein production. The ALK5 inhibitor completely suppressed the TGF-β-mediated increase in NGF expression and NGF production in synovial cells. ALK5, TAK1 and p38 inhibitors inhibited the TGF-β-induced phosphorylation of p38, and TAK1 and p38 inhibitors partially inhibited the TGF-β-mediated increase in NGF expression and NGF production in synovial cells.ConclusionTGF-β regulates NGF production via the TGF-β/ALK5 signaling pathway in osteoarthritic synovium. This effect may partially occur through inhibition of the TAK1/p38 pathway in the SYT of KOA patients.
Highlights
Nerve growth factor (NGF) contributes to pain in knee osteoarthritis (KOA) patients
Localization of NGF and transforming growth factor (TGF)-β protein in the Synovial tissue (SYT) of KOA patients As qPCR analysis detected a correlation between NGF and TGFB mRNA expression, immunohistochemical analysis was performed to investigate the localization of NGF and TGF-β in SYT
Effect of an ALK5 inhibitor on TGF-β-induced NGF expression and NGF production in synovial cells Flow cytometry demonstrated that cultured synovial cells were primarily made up of CD45-CD90+ fibroblasts (86.3 ± 0.5%, mean ± standard error (SE); Additional file 1: Figure S1A) and some CD45 + CD14+ macrophages (8.6 ± 0.6%; Additional file 1: Figure S1B)
Summary
Nerve growth factor (NGF) contributes to pain in knee osteoarthritis (KOA) patients. Transforming growth factor-beta (TGF-β) stimulates NGF expression in chondrocytes from KOA patients. The correlation between synovial TGF-β and NGF levels has not been sufficiently studied in human KOA patients. Nerve growth factor (NGF) is a neurotrophin that modulates nociception It is elevated in chronic pain conditions, leading to increased perception of pain [3]. Numerous studies have shown that NGF is regulated by inflammatory cytokines, including tumor necrosis factor (TNF)-α and interleukin (IL)-1β, in mouse and human articular chondrocytes, synovial fibroblasts, and synovial macrophages in vitro [8,9,10,11]. Synovial NGF regulation under non- or moderate synovial inflammatory states may play an important role in OA, in the late stages of the disease. Synovial NGF regulation under nonor moderate-inflammatory states is not fully understood
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