紫杉醇联合骨髓基质干细胞种植体外修复内皮及对血管平滑肌增生的影响  

Effects of paclitaxel combined with bone marrow stromal stem cells implantation on vascular endothelial repair and smooth muscle cells growth in vitro

作　　者：武晓静[1] 黄岚[1] 周骐[2] 宋耀明[1] 李爱民[1] 晋军[1] 于学军[1] 覃军[1] 赵刚[1] 

机构地区：[1]第三军医大学附属新桥医院全军心血管内科中心,重庆400037 [2]第三军医大学附属新桥医院心外科,重庆400037

出　　处：《中华心血管病杂志》2005年第5期464-468,共5页Chinese Journal of Cardiology

基　　金：国家自然科学基金资助项目(30270568)

摘　　要：目的探讨紫杉醇联合骨髓基质干细胞种植体外修复内皮的可行性及对血管平滑肌细胞增生的影响。方法培养兔主动脉内皮、平滑肌和人骨髓基质干细胞,通过细胞共培养将内皮/骨髓基质干细胞接种于下室、平滑肌细胞接种于上室模拟血管内皮修复过程,分别用3H TdR掺入和Westernblot检测紫杉醇(1,10,100nmol/L)干预20min后第10天平滑肌DNA合成和PCNA蛋白表达,用免疫荧光细胞化学法观察与紫杉醇干预内皮共培养的骨髓基质干细胞vWF和Flk1蛋白表达。结果骨髓基质干细胞种植组平滑肌细胞3H TdR掺入和PCNA蛋白光密度相对值均高于融合内皮组(n=6,P<0.05),低于对数内皮组(n=6,P<0.05)。共培养前骨髓基质干细胞不表达vWF和Flk1蛋白,与紫杉醇干预内皮共培养10天时vWF染色阴性,但部分骨髓基质干细胞开始表达Flk1蛋白。结论骨髓基质干细胞种植能部分抑制紫杉醇引起的平滑肌细胞延迟增生,与紫杉醇干预内皮共培养的骨髓基质干细胞有向内皮分化的能力。ObjectiveTo investigate the effects of paclitaxel combined with bone marrow stromal stem cells (MSCs) implantation on inhibiting the smooth muscle cells(SMCs) growth and promoting endothelial repair by developing an endothelial repair model in vitro.MethodsIn a cell coculture system, rabbit endothelial cells (ECs) and human MSCs were seeded in the lower chamber and rabbit SMCs were seeded in the upper chamber. 3H-TdR incorporation and PCNA protein expression were used to evaluate SMCs proliferation at the 10~th day after paclitaxel application (1,10,100 nmol/L; 20 min). Fluorescence immunocytochemistry was employed to observe the Flk-1 and vWF protein expression on MSCs.Results^The SMCs 3H-TdR incorporation of the MSCs implant group was significantly lower than that of the proliferative ECs group(1 nmol/L:12 265±991 vs. 14 505±1013 cpm/well;10 nmol/L:8401±783 vs. ~10 511 ±934 cpm/well;100 nmol/L:5880±569 vs. 7457±768 cpm/well,n=6,P<0.05), but higher than that of the confluent ECs group (1 nmol/L:12 265±991 vs. 8671±642 cpm/well;10 nmol/L:8401±783 vs. 6175±743 cpm/well;100 nmol/L:5880±569 vs. 4423±406 cpm/well,n=6,P<0.05). The expression of SMCs PCNA protein in MSCs implant group was lower than that of the proliferative ECs group(1 nmol/L:0.92±0.06 vs. 1.15±0.07;10 nmol/L:0.97±0.07 vs. 1.07±0.08;100 nmol/L:0.91±0.05 vs. 1.18±0.11,n=6,P< 0.05), but higher than that of the confluent ECs group(1 nmol/L:0.92±0.06 vs. 0.74±0.07;10 nmol/L:0.97±0.07 vs. 0.78±0.06;100 nmol/L:0.91±0.05 vs. 0.71±0.05,n=6,P< 0.05). The MSCs did not express vWF or Flk-1 protein before coculture. Although none cell expressed vWF, some of the MSCs began to express Flk-1 protein after cocultured with mature ECs for 10 days.Conclusion MSCs implantation can partly inhibit the delayed SMCs proliferation. The MSCs cocultured with paclitaxel-treated mature ECs have the ability to differentiate into ECs.

关 键 词：细胞种植 血管平滑肌增生 紫杉醇 ^3H-TDR掺入 体外 血管平滑肌细胞增生 人骨髓基质干细胞 PCNA蛋白表达 Flk-1 Western BLOT检测 细胞接种 主动脉内皮 细胞共培养 DNA合成 细胞化学法 vWF 修复过程 血管内皮 免疫荧光 

分 类 号：R543[医药卫生—心血管疾病]