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.
Highlights
Recent evidence indicates that the vascular endothelial growth factor (VEGF) plays a role in human knee osteoarthritis (OA) pathology
Recent reports have implicated TGFβ signaling in osteoarthritic synovial tissue (SYT) in OA progression because TGFβ levels are elevated in the synovial fibroblasts (SFs) of OA patients [15, 16]
Our preliminary experiments indicated that 10 ng/ml human recombinant TGFβ significantly increased VEGF mRNA expression compared to 1 ng/ml hrTGFβ and culture medium (Supplementary Figure 2). erefore, 10 ng/ml hrTGFβ was used for stimulation experiments
Summary
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). E 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. ALK5 (SB505124) and TAK1 (5Z-oxozeaenol) inhibitors completely suppressed TGFβ-induced VEGF mRNA expression and VEGF protein production. Both SB505124 and 5Z-oxozeaenol suppressed SMAD2 and p38 phosphorylation. E p38 inhibitor (SB203580) partially inhibited TGFβ-mediated VEGF mRNA and VEGF protein production. TGFβ-mediated regulation of VEGF expression and VEGF protein production in the SYT of OA patients occurs through both the canonical and noncanonical pathway
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