Potential role of HOXD9 in synoviocyte proliferation

Objective. To investigate the role of HOXD9 in the proliferation activity o f cultured synoviocytes as well as the mechanisms that regulate HOXD9 trans cription. Methods. Synoviocytes from patients with rheumatoid arthritis (RA) and oste oarthritis (OA) were transfected with HOX-D9 complementary DNA to establish stable transformants that overexpressed HOXD9. HOXD9 expression was detect ed by Western blotting with anti-HOXD9 antibody. The growth properties of t he transformants were investigated by proliferation and colony formation as says. The expression of basic fibroblast growth factor (bFGF), tumor necros is factor alpha (TNF alpha), interieukin-1 beta, c-Fos, and c-Myc was exami ned by Western blotting. Transcriptional regulation of HOXD9 was examined b y transient cotransfection. Results. HOXD9 protein was highly expressed in RA synoviocytes, but there w as no expression in OA synoviocytes. HOXD9 transfection induced stable HOXD 9 protein expression in synoviocytes and showed an increased proliferation rate under both normal and serum-starved conditions, as well as an enhanced capacity to proliferate anchorage independently to form colonies in soft a gar cultures, compared with control transfectants. Higher levels of bFGF an d c-Fos were detected in HOXD9 transformants than in controls. Transient co transfection assays of NIH3T3 fibroblasts and synoviocytes showed that HOXD 9 activated the luciferase reporter construct containing the highly conserv ed region (HCR), an autoregulatory element of HOXD9 promoter. This activati on was significantly increased by bFGF, suppressed by TNF alpha, and unchan ged by transforming growth factor beta in synoviocytes. Human T lymphotropi c virus type I tax also activated the luciferase reporter construct contain ing the HCR and had a synergistic effect with HOXD9 on HCR promoter activat ion. Conclusion. Our data suggest that HOXD9 plays a potential role in synovial proliferation. In addition, they suggest that the involvement of HOXD9 in t he regulation of cellular growth might be mediated, at least in part, by up -regulation of growth-related factors such as bFGF and c-Fos and/or might r esult from increased transcription activity by its regulators.