双层共培养模型中内皮细胞对平滑肌细胞增殖能力-第三军医大学学报.doc

编号：200808068 论著 文章编号： 双层共培养模型中缝隙连接在内皮细胞调节平滑肌细胞增殖能力中的作用 宋明宝，于学军，朱光旭，赵 刚，于世勇，董红梅，康华利，黄 岚 （第三军医大学新桥医院全军心血管病研究所，重庆 400037） 摘 要：目的 探讨在双层共培养体系中缝隙连接在血管内皮细胞（endothelial cell, EC）调节血管平滑肌细胞(smooth muscle cell, SMC)增殖能力中的作用。方法 本实验采用植块法分别培养大鼠主动脉内皮及平滑肌细胞，并将2种细胞分别接种至transwell insert膜的两侧以建立内皮细胞与平滑肌细胞双层共培养模型，并用透射电镜观察2种细胞在transwell insert膜两侧生长情况。用3H-TdR掺入法观察双层共培养模型中平滑肌细胞增殖情况，并观察缝隙连接阻断剂18α-甘草次酸（18α-GA）对平滑肌细胞增殖能力的影响。结果 在共培养第1天，EC/SMC组与SMC/SMC组的平滑肌细胞3H-TdR掺入率无统计学差异[(10900±1320) cpm/well vs.（10430±1200） cpm/well, P＞0.05]。第2天，EC/SMC组中平滑肌细胞3H-TdR的掺入率有低于SMC/SMC组[(17200±1734 )cpm/well vs. （20900±1659 ）cpm/well ]的趋势,但两者差异未达到统计学标准。共培养第3天时，EC/SMC组平滑肌细胞3H-TdR掺入率显著低于SMC/SMC组，两者相比有显著性差异[(25800±1984 )cpm/well vs.（ 33070±3569） cpm/well, P＜0.05]。在用18α-GA预处理24h后，EC/SMC共培养３ｄ后平滑肌细胞3H-TdR的掺入率明显高于同期未处理EC/SMC组，两者相比有显著性差异［(30500±2650)cpm/well vs. （25800±1984 ）cpm/well, P＜0.05]。结论 　双层共培养模型中内皮细胞通过缝隙连接可抑制血管平滑肌细胞的增殖能力。 关键词：内皮细胞；平滑肌细胞；缝隙连接；共培养；血管 中图法分类号： 文献标识码：A Role of gap junctions on interactin between endothelial cells and smooth muscle cell in a coculture system SONG Ming-bao, YU Xue-jun, ZHU Guang-xu,, ZHAO Gang，YU Shi-yong, DONG Hong-mei, KANG Hua-li，HUANG Lan (Institute of Cardiovascular Diseases of PLA, Xinqiao Hospital, Third Military Medical University, Chongqing 400037) Abstract: Objective To explore the Role of gap junctions on interactin between endothelial cells and smooth muscle cell in a coculture system. Methods and Materials Rat aortic endothelial cells (RAECs) and smooth muscle cells (SMCs) were cultured by explanted rat aortic wall tissue and were identified with cell immunofluorescence staining for vWF or smooth muscle α-actin. A coculture system was made by seeding ECs and SMCs on the both side of transwell insert with millipores respectively and observed with transmission electron microscope. Then, the proliferation of SMCs was evaluated by 3H-TdR incorporation. Meanwhile, 18α-glycyrrhetinic acid (