TIMP-1对绒毛外滋养细胞侵袭迁移和分泌VEGF水平影响研究

Effects of TIMP-1 on Invasion Migration and Secretion of VEGF Level in Villous Trophoblast Cells

目的:研究基质金属蛋白酶组织抑制因子1(TIMP-1)对绒毛外滋养细胞侵袭、迁移和分泌血管内皮生长因子(VEGF)水平的影响。方法:缺氧复氧处理绒毛外滋养细胞HTR8/SVneo,以荧光定量PCR和Western blot方法检测细胞内TIMP-1的表达变化。在HTR8/SVneo细胞中转染TIMP-1siRNA,经缺氧复氧处理以后,以荧光定量PCR和Western blot方法检测细胞内TIMP-1的表达变化。Transwell小室检测敲低TIMP-1对缺氧复氧HTR8/SVneo细胞迁移和侵袭能力的影响,Western blot法检测基质金属蛋白酶-9(MMP-9)蛋白水平,ELISA法检测培养液上清中VEGF、可溶性fms样酪氨酸激酶1(s Flt-1)、可溶性内皮因子(s Eng)含量。结果:缺氧复氧处理后的HTR8/SVneo细胞中TIMP-1mRNA和蛋白水平均高于未经缺氧复氧处理的细胞(P<0.05)。在细胞中转染TIMP-1 siRNA可以降低缺氧复氧细胞中TIMP-1的表达。缺氧复氧处理后的HTR8/SVneo细胞侵袭和迁移能力下降,细胞中MMP-9蛋白水平降低,细胞分泌的VEGF减少,s Flt-1、s Eng增多,与未经缺氧复氧处理的细胞比较,差异有统计学意义(P<0.05)。下调TIMP-1可以促进缺氧复氧条件下HTR8/SVneo细胞侵袭和迁移,促进细胞中MMP-9的表达,诱导细胞分泌VEGF,减少细胞分泌s Flt-1、s Eng。结论:下调TIMP-1促进缺氧复氧条件下绒毛外滋养细胞侵袭和迁移,促进细胞分泌VEGF。

Objective： To study the effect of TIMP-1 on the invasion,migration and secretion of VEGF in the villous trophoblast cells. Methods： Anoxic reoxygenation was used to treat HTR8/SVneo in the villous trophoblastic cells; the expression of TIMP-1 in cells was detected by fluorescence quantitative PCR and Western blot,transfection of TIMP-1 siRNA in HTR8/SVneo cells,after the anoxia and reoxygenation treatment,the expression of TIMP-1 in cells was detected by fluorescence quantitative PCR and Western blot; the effects of low TIMP-1 on the migration and invasion of anoxic reoxygenated HTR8/SVneo cells were detected in the Transwell compartment; Western blot assay was used to detect the level of MMP-9 protein,the content of VEGF,s Flt-1 and s Eng in the supernatant of the culture liquid was detected by ELISA. Results： The level of TIMP-1 mRNA and protein in HTR8/SVneo cells after anoxic reoxygenation was higher than that of cells without anoxic reoxygenation（P 0.05）. The transfection of TIMP-1 siRNA in cells can reduce the expression of TIMP-1 in the hypoxic and reoxygenation cells. The invasion and migration of HTR8/SVneo cells decreased after anoxic reoxygenation,the level of MMP-9 protein in the cells decreased,the cell secretion of VEGF decreased,and the number of s Flt-1 and s Eng increased. Compared with the cells without anoxic reoxygenation,the difference was statistically significant（P 0.05）. Down regulation of TIMP-1 can promote the invasion and migration of HTR8/SVneo cells under hypoxia and reoxygenation,promote the expression of MMP-9 in cells,Inducing cells to secrete VEGF,decrease the secretion of s Flt-1 and s Eng in cells. Conclusion： Down regulation of TIMP-1 promoted the invasion and migration of villous trophoblastic cells under hypoxia and reoxygenation,and promoted the secretion of VEGF by cells.