摘要
目的:观察不同剂量的血小板源生长因子BB刺激对血管平滑肌细胞生长率的影响及血管平滑肌细胞不同增殖程度下p27和p57蛋白表达量的变化。方法:实验于1998-09/2000-05在南方医科大学附属南方医院心内科实验室完成。①选用七八周雄性SD大鼠80只,体外分离SD大鼠主动脉中层平滑肌,贴壁法培养平滑肌细胞,选用2～4代培养的细胞。②胰酶消化的血管平滑肌细胞,浓度为1×108L-1,分别接种于6孔板上,无血清的培养基静止48h,分别加入血小板源生长因子BB10和20μg/L,无血清的培养基为对照组,分别在刺激1,3,5d后,计算不同剂量的血小板源生长因子BB刺激下的血管平滑肌细胞数量,每组重复4次,取平均值。③在用血小板源生长因子BB10和20μg/L刺激6,12,24h后收集细胞,用免疫印迹分析法检测细胞中p27和p57蛋白表达量。结果:①血管平滑肌细胞数量:血小板源生长因子BB刺激1和3及5d后血小板源生长因子BB10和20μg/L组明显高于对照组(t=4.06～21.76,P<0.05～0.01)。②血管平滑肌细胞中p27蛋白表达量(A值):血小板源生长因子BB刺激12和24h后血小板源生长因子BB10和20μg/L组明显低于对照组(P<0.05,0.01),血小板源生长因子BB20μg/L组明显低于血小板源生长因子BB10μg/L组(P<0.01)。p57蛋白表达量(A值):血小板源生长因子BB刺激24h后血小板源生长因子BB10和20μg/L组明显高于对照组(P<0.01),血小板源生长因子BB20μg/L组明显高于血小板源生长因子BB10μg/L组(P<0.01)。结论:①在血小板源生长因子BB刺激的血管平滑肌细胞增殖过程中,p27和p57蛋白表达量变化不同。②在增殖的血管平滑肌细胞中随刺激因子作用时间的延长,表达量逐渐下降,而p57蛋白表达水平随血管平滑肌细胞的增殖而增加,p27蛋白表达量的变化可能是决定血管平滑肌细胞增殖的关键因素。
AIM： To observe the influence of platelet-derived growth factor-BB（PDGF-BB） of different dosage on the growth rate of the cultured vascular smooth muscle cells, and the changes of the p27 and p57 protein expressions in vascular smooth muscle cells under different proliferation severity. METHODS： The experiment was carried out in the cardiological laboratory of Nafang Hospital affiliated to Southern Medical University from September 1998 to May 2000. ① Eighty male SD rats were used, their aorta media smooth muscles were isolated and cultured by means of attachment, and cells of passages 4-6 were selected. ② The vascular smooth muscle cells were digested by pancreatin, inoculated in 6-well plate. After serum withdrawal for 48 hours to induce quiescence, 10 and 20 μg/L PDGF-BB were added into serum-free medium respectively. The serum-free group was treated as comparison. The cells were harvested after stimulated for 1, 3 and 5 days. The cell numbers were calculated. All the steps were repeated for 4 times, and the average value was used. ③ The quiescenced vascular smooth muscle cells were stimulated by 10 and 20 μg/L PDGF-BB respectively, and harvested after 6,12,24 hours. The expressions of protein p27and p57 were examined by Western blot. RESULTS： ①Number of vascular smooth muscle cells： After stimulated by PDGF-BB for 1, 3 and 5 days, the numbers of vascular smooth muscle cells in the PDGF-BB 10 and 20 μg/L groups were more than that in the control group （t=-4.06-21.76, P 〈 0.05-0.01）, ② The expression amount （A value） of p27 in vascular smooth muscle cells： After stimulated by PDGF-BB for 12 and 24 hours, the expression of p27 was highly decreased in the PDGF-BB 10 and 20 μg/L groups as compared with that in the control group （P 〈 0.05,0.01）. The expression of protein p27 was lowest in the PDGF-BB 20 μg/L group （P 〈 0.01）. The expression amount （A value） of protein p57 in vascular smooth muscle cells： After stimulated by PDGF-BB for 24 hours, the expressio
出处
《中国临床康复》
CSCD
北大核心
2005年第38期20-22,共3页
Chinese Journal of Clinical Rehabilitation
