HIF-1-Dependent IL-6 Activation in Articular Chondrocytes Initiating Synovitis in Femoral Head Ischemic Osteonecrosis

Background: Ischemic osteonecrosis of the femoral head in children is associated with chronic hip synovitis and increased levels of the pro-inflammatory cytokine interleukin-6 (IL-6) in the synovial fluid due to unknown mechanisms. The purpose of this study was to investigate hypoxia-inducible factor-1 (HIF-1) activation as a molecular mechanism linking the induction of ischemic osteonecrosis to IL-6 production and the initiation of hip synovitis. Methods: Ischemic osteonecrosis was surgically induced in the right femoral head of 6 piglets. A histologic score, synovial fluid volume, and IL-6 level were used to assess hip synovitis. IL-6 immunostaining of articular cartilage and synovial tissue was performed as well. To study the role of HIF-1 in IL-6 activation, in vitro experiments using an HIF-1α activator (deferoxamine) and inhibitor (HIF-1 small interfering-RNA [siRNA]) were carried out. Synovial cell responses to hypoxic chondrocyte-conditioned media with and without an IL-6 receptor blocker (tocilizumab) were assessed on the basis of IL-1β and tumor necrosis factor-alpha (TNF-α) gene expressions and with a synovial cell-proliferation assay. Results: Induction of ischemic osteonecrosis produced hip synovitis and increased IL-6 levels in the synovial fluid. Immunostaining and protein analysis demonstrated articular chondrocytes as a source of increased IL-6 production. When articular chondrocytes were cultured under hypoxic conditions, significantly increased HIF-1α and IL-6 expressions were observed. Under hypoxic culture conditions, IL-6 gene expression was significantly increased by HIF-1α activation using deferoxamine and inhibited by HIF-1α inhibition using HIF-1 siRNA. Synovial cells exposed to hypoxic chondrocyte-conditioned medium showed significant increases in IL-1β and TNF-α gene expressions and cell proliferation, which were inhibited by the IL-6 receptor blocker tocilizumab. Conclusions: Induction of ischemic osteonecrosis results in IL-6 production in the articular cartilage through an HIF-1-dependent pathway. IL-6 produced by hypoxic articular chondrocytes stimulates inflammatory cytokine responses in synovial cells, which were significantly decreased by tocilizumab. Clinical Relevance: This study provides new insight into the inherent relationship between the induction of ischemia and the initiation of hip synovitis following ischemic osteonecrosis and suggests a potential therapeutic target in the treatment of the synovitis.