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目的:探讨人内皮型一氧化氮合酶(heNOS)基因转染犬内皮祖细胞(EPCs)的有效方法。方法:分别用5型腺病毒和pEGFP-N1质粒作为载体携带heNOS,对体外定向培养扩增的骨髓来源的犬EPCs进行体外转染,然后对转染heNOS基因后的EPCs进行鉴定及功能检测,观察heNOS基因的功能表达情况。结果:heNOS转染犬EPCs48h后,酶联免疫吸附试验(ELISA)检测到5型腺病毒转染组EPCs的eNOS表达量(2091.67±172.49pg/ml)较pEGFP-N1质粒转染组(173.67±36.76pg/ml)和未转染组(158.00±30.91pg/ml)均显著增多(P<0.01);硝酸还原酶法测定5型腺病毒转染组细胞培养上清中的NO含量(49.5±5.2μmol/L)较质粒转染组(38.5±7.1μmol/L)和未转染组(39.7±7.2μmol/L)均显著增多(P<0.01)。结论:5型腺病毒载体介导的heNOS基因能够成功地转染犬EPCs,并能在EPCs中有效表达。
Objective: To explore the method of transfection of endothelial nitric oxide synthase (henos) gene into EPCs in dogs. Methods: HeNOS were transfected with adenovirus type 5 and plasmid pEGFP-N1 respectively, and EPCs were transfected into bone marrow-derived EPCs in vitro. Then the EPCs were identified and their function was detected. Observed the function of heNOS gene expression. Results: The expression of eNOS in EPCs of type 5 adenovirus transfection group (2091.67 ± 172.49pg / ml) was significantly higher than that of pEGFP-N1 plasmid transfection group (173.67 ± 36.76pg / ml) and non-transfected group (158.00 ± 30.91pg / ml) (P <0.01). Nitric acid reductase assay was used to determine NO content in cell culture supernatant of type 5 adenovirus transfection group (49.5 ± 5.2μmol / L) were significantly increased compared with the control group (38.5 ± 7.1μmol / L) and the untransfected group (39.7 ± 7.2μmol / L) (P <0.01). CONCLUSION: The heNOS gene mediated by adenovirus type 5 can be successfully transfected into canine EPCs and can be efficiently expressed in EPCs.