Xin-Sheng Deng1, Xianzhong Meng1, David Fullerton1, Matthew Stone2, James Jaggers3. 1. Cardiothoracic Surgery, University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045 USA. 2. Cardiothoracic Surgery, University of Colorado, Children's Hospital Colorado, Aurora, Colorado 80045 USA. 3. Cardiothoracic Surgery, University of Colorado, Children's Hospital Colorado, Aurora, Colorado 80045 USA. Electronic address: james.jaggers@childrenscolorado.org.
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 up-regulate Runx-2 to induce pro-fibrogenic 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. Pro-fibrogenic activities and associated signaling molecules were analyzed by Western blot assay and collagen staining. RESULTS: Complement time and dose dependently enhanced pro-fibrogenic activities in AVICs, and complement exposure also induced total collagen deposition in AVICs. Complement induced pro-fibrogenic responses were associated with increased Runx-2 expression and phosphorylation of ERK1/2. Genetic silencing of Runx-2 decreased both 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 to active complement treatment. CONCLUSIONS: Complement induced pro-fibrogenic 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.
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 up-regulate Runx-2 to induce pro-fibrogenic 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. Pro-fibrogenic activities and associated signaling molecules were analyzed by Western blot assay and collagen staining. RESULTS: Complement time and dose dependently enhanced pro-fibrogenic activities in AVICs, and complement exposure also induced total collagen deposition in AVICs. Complement induced pro-fibrogenic responses were associated with increased Runx-2 expression and phosphorylation of ERK1/2. Genetic silencing of Runx-2 decreased both 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 to active complement treatment. CONCLUSIONS: Complement induced pro-fibrogenic 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.
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