HTK基因对脐静脉内皮细胞相关基因表达的影响

The Change of Human Umbilical Vein Endothelial Cells Affected by Recombinant Adeno-associated Virus-mediated Kallikrein Gene

目的 利用已经构建好的腺相关病毒-人组织激肽释放酶基因重组体(rAAV-HTK),感 染体外培养的人脐静脉内皮细胞(HUVEC),观察HUVEC细胞中内皮型一氧化氮合酶(eNOS)、凋亡 蛋白酶(caspase-3)、内皮素-1(ET-1)、血管内皮生长因子(VEGF)、内皮素B1受体(ETR-B1)以及缓激 肽B1受体、缓激肽B2受体的mRNA表达量变化情况,探讨利用rAAV-HTK治疗高血压、缺血性心脏 病的可行性。方法 将已经构建好的rAAV-HTK重组质粒感染体外培养的HUVEC细胞。应用半定 量RT-PCR方法检测感染rAAV-HTK前后HUVEC中eNOS、caspase-3、ET-1、VEGF、ETR-B1以及缓激 肽B1受体、缓激肽B2受体的mRNA表达量变化情况。结果 转染有rAAV-HTK的HUVEC细胞内 其细胞内eNOS的mRNA合成量增加,caspase-3的mRNA表达量降低,VEGF、ET-1、ETR-B1、缓激肽 B1受体、缓激肽B2受体的mRNA表达量没有变化。结论 rAAV-HTK重组体感染HUVEC细胞能够 使HUVEC细胞内eNOS的mRNA表达量增高,caspase-3的mRNA表达量减低,提示HTK能够改善内 皮细胞功能,转HTK能够应用于高血压等血管内皮功能异常的心血管疾病的基因治疗。

Objective To evaluate the functional changes of HUVEC cells which are infected with constructed rAAV-HTK. The feasibility of treating hypertension and ischemic heart disease with rAAV-HTK was discussed. Methods HUVEC cells were infected by constructed rAAV-HTK. Measure the variance of mRNA quantity of eNOS, caspase-3, ET-1, VEGF, ETR-B, Bradykinin B1 receptor and Bradykinin B2 receptor in the HUVEC cells before and after infection by semiquantitative RT-PCR. Results The mRNA quantity of eNOS in the infected HUVEC cells was higher than the control, but the mRNA quantity of caspase-3 was lower, and there was no obvious difference in the expression of VEGF、ET-1、ETR-B、Bradykinin B1 receptor and Bradykinin B2 receptor. Conclusions The expression level of eNOS mRNA was increased in the rAAV-HTK infected HUVEC cells, while the expression level of caspase-3 mRNA was decreased, which implies the HTK can enhance endothelial cells dysfunction. These beneficial effects show the potential implication for the gene therapy in treating cardiovascular diseases with HTK gene caused by blood vessel endothelium dysfunction, such as hypertension.