CAJ | 학술논문

目的：探讨中期因子( midkine, MK)在人乳腺癌MDA-MB-231细胞体外血管生成中的作用及其机制。方法采用shR-NA干扰技术降低MDA-MB-231细胞MK表达,应用Western blot技术检测肿瘤细胞中内皮蛋白C受体( endothelial protein C receptor, EPCR)的表达；干扰MK和EPCR表达或通过抗体阻断活化蛋白酶激活受体1(protease-activated receptor 1, PAR1)作用后,制备肿瘤条件培养基作用于人脐静脉内皮细胞( human umbilical vein endothelial cells, HUVECs),通过CCK-8试剂盒检测HUVECs增殖、Transwell小室检测迁移以及Matrigel表面培养检测脉管形成能力。结果干扰MK表达后,EPCR表达随之降低。干扰MK和EPCR低表达后,HUVECs增殖、迁移及脉管形成能力均低于对照组(P<0．05),EPCR干扰组低于MK干扰组(P<0．05)。应用抗PAR1作用后,HUVECs增殖、迁移及脉管形成能力低于对照组和EPCR干扰组(P<0．05)。结论 MK可通过EPCR/PAR1通路促进乳腺癌MDA-MB-231细胞体外血管生成。
목적：탐토중기인자( midkine, MK)재인유선암MDA-MB-231세포체외혈관생성중적작용급기궤제。방법채용shR-NA간우기술강저MDA-MB-231세포MK표체,응용Western blot기술검측종류세포중내피단백C수체( endothelial protein C receptor, EPCR)적표체；간우MK화EPCR표체혹통과항체조단활화단백매격활수체1(protease-activated receptor 1, PAR1)작용후,제비종류조건배양기작용우인제정맥내피세포( human umbilical vein endothelial cells, HUVECs),통과CCK-8시제합검측HUVECs증식、Transwell소실검측천이이급Matrigel표면배양검측맥관형성능력。결과간우MK표체후,EPCR표체수지강저。간우MK화EPCR저표체후,HUVECs증식、천이급맥관형성능력균저우대조조(P<0．05),EPCR간우조저우MK간우조(P<0．05)。응용항PAR1작용후,HUVECs증식、천이급맥관형성능력저우대조조화EPCR간우조(P<0．05)。결론 MK가통과EPCR/PAR1통로촉진유선암MDA-MB-231세포체외혈관생성。
Purpose To observe the effects of midkine ( MK) on human breast cancer cell line MDA-MB-231 angiogenesis in vitro, and to explore its mechanism. Method shRNA interference was performed to silence the expression of MK in MDA-MB-231 cells, and Western blot was used to identify the expression of MK and EPCR. After MK and EPCR knockdown, or treated with anti protease-activated receptor 1 (PAR1) antibody, the culture medium of MDA-MB-231 cells were collected and the conditioned medium were pre-pared. Human umbilical vein endothelial cells ( HUVECs) were cultured with conditioned medium, and the endothelial cells prolifera-tion was detected by CCK-8 assay, cell migration was detected by transwell method, vasculogenic activity was assessed by Matrigel-based tube formation assay. Results After knockdown of MK, the protein level of EPCR was decreased in MDA-MB-231 cells. Com-pared with control, knockdown of MK and EPCR decreased the proliferation, migration and angiogenesis ability of HUVECs significant-ly (P<0. 05), and the effect of EPCR knockdown group was stronger than MK knockdown group (P<0. 05). After treated with anti-PAR1 antibody, the proliferation, migration and angiogenesis ability of HUVECs were decreased compared with control and EPCR knockdown group (P<0. 05). Conclusion MK promotes human breast cancer cell line MDA-MB-231 angiogenesis through EPCR /PAR1 signaling pathway in vitro.