Effects Of Low-molecular Weight Heparin On MMP-2,TIMP-2 Expression And Invasiveness Of Vitro Cytotrophoblast Cells

Objective:To investigate the hypothesis that low-molecular weight heparin(LMWH) might regulate in vitro cytotrophoblast invasiveness and production of metalloproteinases-2(MMP-2),tissue inhibitor of metalloproteinas-2(TIMP-2).Methods:Chorionic villi tissue of normal 6-8 weeks pregnancy was obtained under asepsis condition. Trophoblast cells were collected by trypsin-collagenase digestion and Percoll gradient centrifugation. The cytotrophoblast cells were cultured 24h and divided into four groups according to the concentration of LMWH adding into the medium, which include 0 IU·mL-1,0.1 IU·mL-1,1 IU·mL-1,10 IU·mL-1. The determination of MMP-2 and TIMP-2 in cell culture supernatants were performed using ELISA. Cytotrophoblast invasiveness was determined by Transwell chamber.Results:1. After 24h culture, the concentration of MMP-2 in cell culture supernatants were (69.3±24.4) pg/ml in group 0.1 IU·mL-1, (111.754±19.5) pg/ml in group 1.0 IU·mL-1(78.488±18.9) pg/ml in group 10 IU·mL-1, which were all higer than (45.288±20.8) pg/ml in control group significantly (P<0.05). At 1 IU·mL-1, LMWH greatly enhanced the expression of MMP-2 (P<0.01).2. With the increasing concentrations of LMWH, the levels of TIMP-2 was decreased after intervention with LMWH. At 1 IU·mL-1 and 10 IU·mL-1, LMWH induced a significant decrease in TIMP-2 expression(P<0.05).There was no significant difference between group 1 IU·mL-1 and group 10 IU·mL-1(P>0.05).3.In matrigel invasion assay, after 24h culture,the number of the trans-membrance cells in different groups, including (33±5) in group 0.1 IU·mL-1,(47±9) in group 1.0 IU·mL-1, (30±4) in group 10 IU·mL-1, were all higher than (25±5) in control group with statistical significance(P<0.05). At 1 IU·mL-1, LMWH greatly enhanced cytotrophoblast invasiveness (P<0.01).Conclusions:LMWH might regulate in vitro cytotrophoblast invasiveness by influencing the expression of MMP-2 and TIMP-2 of cytotrophoblast cells.Significance:This results inform us that LMWH might regulate in vitro cytotrophoblast invasiveness by influencing the expression of MMP-2 and TIMP-2 of cytotrophoblast cells. And The effects were influenced by different concentrations of LMWH. This study will help us to understand the mechanism of the treatment of recurrent abortion with LMWH clinically.