Abstract
BACKGROUND: Calcium accumulation and fibrotic activities are principal mechanisms for calcific aortic valve disease (CAVD). Active complement products are observed in human stenotic aortic valves. Runt-related transcription factor 2 (Runx-2) is involved in tissue calcification. We hypothesized that complement upregulates Runx-2 to induce profibrogenic change in human aortic valve interstitial cells (AVICs). METHODS: AVICs were isolated from 6 normal and 6 CAVD donor valves. Cells were treated with complement cocktails. Profibrogenic activities and associated signaling molecules were analyzed by Western blot assay and collagen staining. RESULTS: Complement time and dose dependently enhanced profibrogenic activities in AVICs, and complement exposure also induced total collagen deposition in AVICs. Complement-induced profibrogenic responses were associated with increased Runx-2 expression and phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2). Genetic silencing of Runx-2 decreased both matrix metalloproteinase 9 (MMP-9) and collagen I levels. Pharmacological inhibition of ERK1/2 decreased complement-mediated MMP-9, collagen I, and Runx-2 expression as well as total collagen deposition in human AVICs. Further, treating AVICs with heat-deactivated complement resulted in reduced MMP-9, collagen I, and Runx-2 levels compared with active complement treatment. CONCLUSIONS: Complement induced profibrogenic activities in AVICs by activation of ERK1/2-mediated Runx-2 signaling pathways. This study demonstrates a potential role for complement-mediated CAVD pathogenesis, establishing a possible therapeutic target to limit CAVD progression.