抗ATPase F1α抗体对人非小细胞肺癌血管内皮细胞的抑制作用

Inhibitory effect of ATPase F1αantibody on human non-smaU cell lung cancer-derived vascular endothelial cells

目的:探讨一种新的抗ATPase F1α抗体,人血管抑制和肿瘤转移相关蛋白(human angiostatin interacting and tumor metastasis involving protein,HAI-TMIP)抗体((MAb3D5AB1,简称GX)对人非小细胞肺癌血管内皮细胞(non-small cell lung cancer-derived vascular endothelial cell,NSCLC-VEC)的抑制作用。方法:从非小细胞肺癌患者手术切除标本分离和原代培养NSCLC-VECs;采用免疫荧光法检测NSCLC-VECs膜上ATPase F1α的表达,RT-PCR分析细胞ATPase FlαmRNA的表达水平;CCK-8法检测GX对NSCLC-VECs增殖的抑制效应,划痕损伤实验检测GX对NSCLC-VECs迁移的抑制作用,FACS检测GX对NSCLC-VECs凋亡的影响。结果:倒置荧光显微镜检测显示,NSCLC-VECs胞膜上有ATPase F1α的显著表达;RT-PCR的结果表明,分别来源于肺腺癌、肺鳞癌的血管内皮细胞中ATPase F1αⅡ肽段基因片段大小为668 bp。CCK-8结果表明,经GX处理,NSCLC-VECs的增殖活力显著低于对照组(P<0.01);划痕损伤实验结果显示,经GX处理的NSCLC-VECs的迁移速度显著低于对照组(P<0.01);FACS结果显示,GX处理后NSCLC-VECs的凋亡率高于对照组(P<0.01)。结论:ATPaseF1α在NSCLC-VECs膜上有较高的表达,GX通过靶向作用于ATPaseF1α抑制NSCLC-VECs的增殖、迁移,并诱导细胞凋亡。

Objective：To investigate the inhibitory effect of human angiostatin interacting and tumor metastasis involving protein （HAI-TMIP） antibody （MAb3DSAB1, GX） ,a new anti-ATPase F1α antibody, against proliferation, migration and apoptosis of non-small cell lung cancer-derived vascular endothelial cells （NSCLC-VECs）. Methods： NSCLC- VECs were isolated and cultured from surgical specimens of NSCLC patients. The membrane expression of ATPase F1α in NSCLC-VECs was observed by immunofluorescence means; RT-PCR was used to analyze the expression of ATPase F1α mRNA. CCK-8 assay, wound-healing assay and fluorescence activated cell sorting （FACS） were used to assess the effects of HAI-TMIP antibody on NSCLC-VECs proliferation, migration and apoptosis. Results： ATPase F1α expression was observed in NSCLC-VECs under inverted fluorescent microscope. RT-PCR showed the fragment length of the ATPase F1αⅡ peptide gene derived from pulmonary adenocarcinoma, squamous carcinoma was 668 bp. It was found that the proliferation and migration of NSCLC-VECs were significantly inhibited in HAI-TMIP antibody treated group compared with those in the control group （P 〈 0.01 ） . FACS result showed that the apoptosis rate of and NSCLC-VECs in HAI-TMIP antibody group was significantly higher than that in the control group （P 〈 0.01 ）. Conclusion： ATPase F1 α is highly expressed on the membrane of NSCLC-VECs. HAI-TMIP antibody can efficiently inhibit the proliferation and migration of NSCLC-VECs and induce apoptasis through interacting with ATPaseF1α.