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GW24-e0507 Role of wnt signal pathway in proliferation and collagen secretion of smooth muscle cells from rats aorta induced by AngII
  1. Hua JunYi1,
  2. Xu Yun2,
  3. He Yu-zhou1,
  4. Jiang Xu-hong1,
  5. Pan Zhi-min1,
  6. Ye Wu1
  1. 1The First Affiliated Hospital, ZheJiang University of Traditional Chinese Medicine
  2. 2Medicine Communication and Management Cencer of ZheJiang Province

Abstract

Objectives To evaluate whether Wnt signalling pathway modulates proliferation and collagen secretion of vascular smooth muscle cells and try to find novel gene target of anti-vascular restenosis.

Methods Cultured vascular smooth muscle cells (SMC, A7r5) were devided into seven groups: control group; AngII (10-6 mol/L) group; AngII + atrovastatin (0.1 umol/L) group; AngII + atrovastatin (1 umol/L)group; AngII + atrovastatin (10 umol/L) group; AngII (10-6 mol/L) + SiRNA-dvl-1group; AngII (10-6 mol/L) + SiRNA-dvl-1 + atrovastatin (10 umol/L) group. The SMCs proliferation was determined by MTT assay. Several Wnt genes and proteins werequantitated separatedly by Real-Time quantitative-PCR and Western blot assay.

Results (1) After Dvl-1 Gene was silenced by SiRNA after transcription, proliferation of VSMC induced by AngII was inhibited significantly (AngII + siRNA vs AngII, 0.369 ± 0.105 vs 0.417 ± 0.098, 24h, P < 0.05); Proliferation of VSMC induced by AngII were inhibited significantly by 1umol/L concentrations of atorvastatin in comparison with the control group (0.365 ± 0.077 vs 0.417 ± 0.098, 24h, P < 0.05; 0.321 ± 0.083vs0.434 ± 0.101, 48h, P < 0.05 ; 0.277 ± 0.062 vs 0.425 ± 0.112,72h, P < 0.01), the inhibited effect showed time and concentration dependent manners; (2) High expression of Wnt4, Dvl-1 and ß-catenin mRNA and proteins were induced by AngII, which could be inhibited by atrovastatin in concentration dependent manner (1,10 umol/L vs 0.1 umol/L seperatly, 24 h,wnt4 mRNA:1.188 ± 0.132,0.785 ± 0.246 vs 1.565 ± 0.187, P < 0.05; wnt4 western: 22.73 ± 4.14,15.94 ± 3.38 vs 24.02 ± 5.44, P < 0.05; dvl-1 mRNA:1.254 ± 0.163,1.104 ± 0.086 vs 1.624 ± 0.213, P < 0.05; dvl-1 western: 25.28 ± 4.67,17.57 ± 2.92 vs 27.72 ± 4.29, P < 0.05; ß-catenin mRNA:1.190 ± 0.112,0.986 ± 0.078 vs 1.669 ± 0.194, P < 0.05; ß-catenin western:15.80 ± 3.07,13.67 ± 2.16 vs 28.82 ± 5.10, P < 0.05) and the latter two could be reversed significantly by siRNA-Dvl-1; (3) High expression of collegan I, collegan III mRNA and proteins in VSMC induced by AngII could be inhibited siganificantly by siRNA-Dvl-1 and atrovastatin (1,10 umol/L) (AngII + atv1.0, AngII + atv10 vs AngII, seprately; col1 mRNA 1.660 ± 0.150,1.148 ± 0.091 vs 2.390 ± 0.207, P < 0.05; col1 western 14.02 ± 3.35 vs 16.53 ± 3.86, P > 0.05; 10.12 ± 2.04 vs 16.53 ± 3.86, P < 0.01; col3 mRNA:1.148 ± 0.151, 0.846 ± 0.140 vs 1.552 ± 0.256, P < 0.05; col3 western: 10.23 ± 2.02, 6.35 ± 1.08 vs 18.83 ± 3.45, p < 0.01); (4) After Dvl-1 Gene was silenced by SiRNA, proliferation and collagen secretion of VSMC could be further inhibited by atorvastastin (24 h, AngII + siRNA + atv10 vs AngII + siRNA, MTT: 0.333 ± 0.085 vs 0.369 ± 0.105,p < 0.05; western:col1, 5.86 ± 1.26 vs 8.08 ± 1.88, p < 0.05; col3, 4.18 ± 0.93 vs 6.96 ± 1.14 p < 0.05).

Conclusions 1) Induced by AngII, VSMC proliferate accompanying with Wnt signal moleculars highly expressed; 2) Expression of intracellular wnt signal moleculars reduced significantly when AngII-induced proliferation of VSMC was inhibited by siRNA-Dvl-1 and atrovastatin; 3) Activation of Wnt signal pathway is involved in AngII-induced high expression of collegan genes in VSMC.4)Wnt signal pathway is involved in AngII-induced VSMC proliferation; 4) Besides wnt signal pathway, any other signal pathways enrolled in AngII-induced VSMC proliferation and collagen secretion.

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