Vascular Endothelial Growth Factor Is Regulated by the Canonical and Noncanonical Transforming Growth Factor-βPathway in Synovial Fibroblasts Derived from Osteoarthritis Patients

Abstract

Background Previous studies suggest the presence of an association of vascular endothelial growth factor (VEGF) with osteoarthritis (OA) severity and pain in patients with knee OA. VEGF expression in human synovial fibroblasts (SFs) is induced by transforming growth factor-beta (TGFβ). However, the signaling pathway governing TGFβ-mediated regulation of VEGF in SFs has not been identified. Methods OA patients who underwent total knee arthroplasty had their synovial tissue (SYT) extracted and the constituent SFs cultured. The cells were stimulated with culture medium (control), human recombinant TGFβ (hrTGFβ), hrTGFβ + ALK5 inhibitor SB505124, hrTGFβ + transforming growth factor activating kinase 1 (TAK1) inhibitor (5Z)-7-oxozeaenol, or hrTGFβ + p38 inhibitor SB203580 for 6 h. VEGF mRNA expression in SFs was examined using real-time polymerase chain reaction and VEGF protein production in the cell supernatant was examined using enzyme-linked immunosorbent assay. Additionally, phosphorylated levels of SMAD2 and p38 were examined using western blotting. Results ALK5 (SB505124) and TAK1 (5Z-oxozeaenol) inhibitors completely suppressed TGFβ-induced VEGF mRNA expression and VEGF protein production. Both SB505124 and 5Z-oxozeaenol also suppressed SMAD2 and p38 phosphorylation. The p38 inhibitor (SB203580) partially inhibited TGFβ-mediated VEGF mRNA and VEGF protein production. Conclusion TGFβ-mediated regulation of VEGF expression and VEGF protein production in the SYT of OA patients occurs through both the canonical and noncanonical pathway.

The inflammatory speech of fibroblasts

“…249 VEGF expression is induced in OA synoviocytes by inflammatory and growth factors including IL-1β, 250 PGE2, 251 and TGFβ. 252 Increased vascularization permits additional immune cell infiltration that further contributes to synovial fibrosis via inflammatory mediators. Overall, this suggests that OA-FLS are potent contributors to their inflammatory microenvironment and contribute to OA synovial fibrosis in autocrine and paracrine fashions.…”

“…Although VEGF is detected in macrophages and endothelial cells, the majority of VEGF is expressed by FLS 249 . VEGF expression is induced in OA synoviocytes by inflammatory and growth factors including IL‐1β, 250 PGE2, 251 and TGF‐β 252 . Increased vascularization permits additional immune cell infiltration that further contributes to synovial fibrosis via inflammatory mediators.…”

Identification of Transcription Factors Responsible for a Transforming Growth Factor-β-Driven Hypertrophy-like Phenotype in Human Osteoarthritic Chondrocytes

“…COL10A1 is considered as the standard marker and RUNX2 as the master transcription factor of chondrocyte hypertrophy [ 2 , 34 , 93 , 94 , 95 , 96 , 97 ]. IHH is a key regulator of endochondral ossification and is mainly produced and secreted by pre-hypertrophic chondrocytes [ 98 , 99 , 100 ], while VEGFA is mainly found in the late hypertrophic phase, correlating with osteoblast formation [ 95 , 101 , 102 , 103 ]. Lastly, TGF-β increased COL1A1 expression, which is associated with dedifferentiation towards fibrotic chondrocytes.…”

Adult fibroblasts use aggresomes only in distinct cell-states

“…Further, TAK1 has many targets outside of p38 MAPKs which could additionally play a role in aggresome formation such as c-Jun N-terminal kinases (JNKs) and Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-kB) (Totzke, Scarneo et al 2020). Lastly, although TAK-715 and 5Z-7-oxozeaenol are recognized as inhibitors of p38α and p38β MAPKs or TAK1 respectively (Ninomiya-Tsuji, Kajino et al 2003, Miwatashi, Arikawa et al 2005, Verkaar, van der Doelen et al 2011, Takano, Uchida et al 2019), both of these drugs are known to have off-target effects. Therefore, it is possible that the effects we observe from these drugs on aggresome formation is through their effect on other proteins.…”