氧化型低密度脂蛋白对骨髓内皮前体细胞增殖、凋亡及NO分泌功能的影响论文.doc

氧化型低密度脂蛋白对骨髓内皮前体细胞增殖、凋亡及NO分泌功能的影响论文.doc氧化型低密度脂蛋白对骨髓内皮前体细胞增殖、凋亡及NO分泌功能的影响论文
宗小娟,王海昌,周祥,程康【关键词】内皮祖细胞
Effect of oxidized lo bone marroarro bone marrog/L; group C, 10 mg/L; group D, 20 mg/L) edium to detect the effect of oxLDL on EPCs. MTT assay ultiplication ability of EPCs. Floetry easured in the cell culture medium by nitrate reductase method to find the injury of oxLDL on cells and cell functions. RESULTS: pared oted the apoptosis of EPCs (group B, ±; group C, ±; group D, ±) (n=6), and damaged the NO secreting function of EPCs. These effects age the NO secreting function and promote the apoptosis of the cells. It had been seen that EPCs ay be some relationship betg/L,C组10 mg/L,D组20 mg/L)作用于EPCs,,小鼠flk1、山羊属CD133(Santa Cruz,美国)、鼠CD31, FITC羊抗小鼠(IgG)、FITC羊抗兔(IgG)、罗丹明兔抗羊(IgG)(Zymed,美国), NO, TBA试剂盒(南京建成,中国),LDL(Sigma,美国), 6孔培养板(Nunclon,丹麦); Elite ESP流式细胞仪(美国COULTER公司),Olympus Tokyo CK相差显微镜(奥林巴斯),DG3022A型酶联免疫检测仪(南京华东电子管厂),UV2450分光光度计(日本SHIMADZU公司),MRC1024激光共聚焦显微镜(美国BioRad公司).

g/L浓度苯巴比妥1 mL/kg麻醉后,左侧髂后棘局部常规消毒铺巾,20 g/L利多卡因5 mL局麻,沿髂后棘行骨穿,有落空感后接肝素化的20 mL注射器用力抽吸. 抽出的细胞悬液经等体积DMEM混合重悬后,小心叠加于等体积淋巴细胞分离液上,保持界面清晰,2000 r/min, 离心20 min,吸取中间云雾层细胞,5倍体积DMEM重悬,1500 r/min, 10 min,洗涤两次. 末次离心后,弃上清,加入含100 mL/L胎牛血清的DMEM重悬. 取微量细胞悬液与等体积2 g/L台盼蓝染液混合,进行细胞活力计数,死细胞被染成蓝色,活细胞不着色,活细胞数占98%以上. 细胞接种于培养瓶中,由于成纤维细胞贴壁较快,多于24 h内贴壁,故取24 h未贴壁细胞,经含100 mL/L胎牛血清、VEGF(10 μg/L)