靶向血管内皮生长因子受体2的sKDR抑制血管内皮细胞生长和血管生成的研究  

Study on the inhibitory effect of sKDR of the targeting vascular endothelial growth factor receptor 2 on the growth of vascular endothelial cells and angiogenesis

作　　者：郑红淑[1] 王秀岩[1] 李然伟[1] 冯树强[1] 李春艳[1] 徐文翠[1] ZHENG Hong-shu;WANG Xiu-yan;LI Ran-we(Department of Urology,the Second Hospital of Jilin University,Changchun 130041,China)

机构地区：[1]吉林大学第二医院泌尿外科,吉林长春130041

出　　处：《中国实验诊断学》2020年第7期1187-1191,共5页Chinese Journal of Laboratory Diagnosis

摘　　要：目的研究可溶性血管内皮生长因子受体2(sKDR)对人脐静脉内皮细胞(HUVEC)及血管形成的抑制作用。方法利用RT-PCR方法提取KDR胞外血管内皮生长因子(VEGF)结合区编码基因,构建真核分泌型表达质粒PQE40-KDR,即可溶性血管内皮生长因子受体sKDR,脂质体法转染HUVEC,经G418抗性压力筛选稳定转染细胞株。ELISA法检测细胞上清VEGF结合活性,筛选稳定表达sKDR的HUVEC。RT-PCR检测细胞KDR蛋白表达水平,比色法和鸡胚尿囊膜(CAM)试验分别测定sKDR对HUVEC增殖及其对CAM血管生成的影响。结果稳定表达sKDR的HUVEC KDRmRNA表达水平明显高于空质粒对照组,细胞增殖水平下降,新生血管数目减少。提示sKDR可抑制KDR刺激的HUVEC增殖,并遏制KDR诱导的CAM血管生成。结论 sKDR具有与KDR结合的生物学功能,KDR有望成为基因抗血管形成治疗恶性实体肿瘤的新靶点。Objective To study the inhibitory effect of soluble vascular endothelial growth factor receptor 2(sKDR)on human umbilical vein endothelial cells(HUVEC)and angiogenesis.Methods The RT-PCR method was used to extract the encoding gene of KDR extracellular vascular endothelial growth factor(VEGF)binding region,and the secretory expression plasmid PQE40-KDR was constructed.The soluble vascular endothelial growth factor receptor sKDR,liposome method transfected to HUVEC,and stable transfected cell lines by G418 resistance pressure screening.ELISA assay was used to detect the VEGF binding activity of cell supernatants and to screen HUVEC with stable expression of sKDR.RT-PCR was used to detect the expression level of KDR protein,and the effects of sKDR on HUVEC proliferation and CAM angiogenesis were detected by colorimetry and chick allantoic membrane(CAM)test respectively.Results The expression level of HUVEC KDRmRNA in stable sKDR expression was significantly higher than that in empty plasmid control group.The proliferation level decreased and the number of neovascularization decreased.It is suggested that sKDR can inhibit the proliferation of KDR stimulated HUVEC and inhibit KDR induced CAM angiogenesis.Conclusion SKDR has biological functions associated with KDR.KDR is expected to become a new target for gene therapy against malignant solid tumors.

关 键 词：血管内皮生长因子受体2 人脐静脉内皮细胞 鸡胚尿囊膜 免疫组织化学 

分 类 号：Q78[生物学—分子生物学]