重组福安泰-03抑制人脐静脉血管内皮细胞的迁移和增殖  

Recombinant Fuantai-03 Inhibits the Migration and Proliferation in Human Umbilical Vein Endothelial Cells

作　　者：黄来珍[1] 马润娣[1] 于立坚[1] 张永平[1] 苏伟明[1] 廖铭能[1] 于廷曦[1,2] 

机构地区：[1]广东海洋大学海洋药物研究与开发重点实验室.湛江524025 [2]Department of Surgery , Department of Pathology ,University of Maryland School of Medicine and Baltimore Veterans Affairs Medical Center, Baltimore, MD 21201, USA

出　　处：《中国生物化学与分子生物学报》2010年第6期561-567,共7页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家高技术研究发展计划海洋技术领域专题(863计划,No.2007AA092422);国家自然科学基金(No.30271493);广东省自然科学基金重点项目(No.021386);广东省海洋与渔业局科技兴海重大项目(No.A200099B01)~~

摘　　要：研究重组福安泰-03(recombinant Fuantai-03,rFAT-03)对人脐静脉血管内皮细胞(human umbilical vein endothelial cells,HUVECs)迁移和增殖的影响.聚碳酸酯膜小室趋化运动模型(transwell model)检测rFAT-03对HUVECs迁移能力的影响;MTT法检测rFAT-03对HUVECs和多种人癌细胞生长的影响;荧光显微镜观察rFAT-03作用下HUVECs的形态变化;膜联蛋白V-异硫氰酸荧光素(annexin V-fluorescein isothiocyanate,annexinV-FITC)双染检测rFAT-03对HUVECs早期凋亡的影响;流式细胞术分析rFAT-03对HUVECs周期及凋亡的影响;Western印迹检查rFAT-03对HUVECs血管内皮细胞生长因子(VEGF)、Bax和Bcl-2表达的影响.结果显示,rFAT-03明显抑制HUVECs细胞的迁移和增殖,其抑制效果与剂量和作用时间相关.0.20mg/mL恩度(endostar),0.10、0.20mg/mLrFAT-03作用HUVECs24h,对细胞迁移的抑制率分别为32.0%、32.6%、57.1%(P<0.01).0.20mg/mL恩度,0.05、0.10、0.20mg/mLrFAT-03作用HUVECs72h,其对细胞生长的抑制率分别为40.9%、63.7%、69.3%、87.0%.但rFAT-03对多种人癌细胞株的生长却无明显影响.rFAT-03处理HUVECs24h,细胞的早期凋亡率增加(P<0.05).rFAT-03阻滞HUVECs于G0/G1期,并呈现典型的凋亡峰.终浓度为0.05、0.10、0.20mg/mLrFAT-03作用于HUVECs24h,G0/G1期细胞指数分别为63.4%、67.5%和75.7%(对照组为62.1%),凋亡指数分别为4.2%、8.5%和10.3%.rFAT-03下调HUVECs的VEGF和抑调亡基因Bcl-2的表达,上调促凋亡基因Bax的表达,其效果与剂量相关.结果提示,rFAT-03明显抑制HUVECs细胞的迁移和增殖,诱导其凋亡,它的这些作用与其下调VEGF、Bcl-2表达,上调Bax表达密切相关.The present study was conducted to investigate the effects of recombinant Fuantai-03 （rFAT03）on the migration and proliferation in human umbilical vein endothelial cells （HUVECs）.Migration assay was performed using a Transwell model with polycarbonate membrane;cell growth inhibitory effect was measured by MTT assay;apoptotic induction of HUVECs by rFAT-03 was determined by fluorescence microscopy and flow cytometry;and Western blot analysis was performed for examining expressions of vascular endothelial growth factor （VEGF）,Bcl-2 and bax.rFAT-03 obviously inhibited the migration and proliferation of HUVECs in a dose-and time-dependent manner.The doses 0.20 mg /mL endostar,0.10 and 0.20 mg /mL rFAT-03 decreased the percentage of migrating HUVECs by 32.0%,32.6%,and 57.1% （P 0.01）,respectively.The doses 0.20 mg /mL endostar and 0.05,0.10,0.20 mg /mL rFAT-03 displayed growth inhibitory activity against HUVECs with inhibition rates of 40.9%,63.7%,69.3% and 87.0% for 72 hours,respectively,but did not show significant effects on several human malignant tumor cell lines.rFAT-03-treated HUVECs showed typical morphologic and cellular evidence of apoptosis.The expressions of VEGF and Bcl-2 in the rFAT-03-treated HUVECs were evidently downregulated,and expression of bax was obviously up-regulated.These findings provide evidence that rFAT03 significantly inhibits the migration and proliferation of HUVECs and induces apoptosis,and the effects of rFAT-03 might result from the down-regulation of expressions of VEGF and Bcl-2 and up-regulation of expression of bax in the rFAT-03-treated HUVECs.

关 键 词：重组福安泰-03 人脐静脉内皮细胞 细胞周期 细胞凋亡 血管内皮细胞生长因子 

分 类 号：Q331[生物学—遗传学]