Matrix metalloproteinases (MMPs) are implicated in the degradation of extracellular matrix; they play important roles in the invasion of the trophoblast cell into the maternal endometrium during placentation. Previous studies have concentrated on comparison of MMP expression in trophoblast cells between the first and third trimester. But the dynamic expression of MMPs during the first trimester has not been reported. In the present study, the expression of MMP-2, -9, and -14 (membrane-type MMP-1) and the production of tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2) by cultured human cytotrophoblast cells from 6 to 11 wk of gestation were investigated. The cells were cultured under serum-free conditions. There was no MMP-9 secretion by the cells at Week 6, but from Week 7 to 11 the MMP-9 secretion increased gradually. Week 11 cells secreted more than 10-fold as much MMP-9 (167.7 +/- 18.8 ng/ml) as Week 7 (14.7 +/- 3.9 ng/ml) cultures. However, MMP-2 production declined from Week 6 to Week 11, and the production at Week 11 (32.3 +/- 8.1 ng/ml) was about one sixth that at Week 6 (205.7 +/- 27.2 ng/ml). The expression of mRNA transcripts for MMP-2 and MMP-9 correlated with enzyme secretion; we did not detect any MMP-9 mRNA signal in 20 microg total RNA extracted from cultured cells at Weeks 6, 7, and 8 of pregnancy, but a signal was apparent in Weeks 9 and 11. MMP-2 mRNA was expressed throughout the 6- to 11-wk period and exhibited a remarkable decline during this period. MMP-14 mRNA transcripts remained relatively stable from 6 to 11 wk. Significantly more TIMP-1 (P < 0.01) was detected in Week 9 (87.5 +/- 15.0 ng/ml) and Week 11 (169.1 +/- 30.2 ng/ml) media compared to Week 6 media (23.5 +/- 4.8 ng/ml), but we did not detect any TIMP-2 in the media of the tested cells. This study demonstrated that first-trimester human cytotrophoblast cells were able to produce abundant laminin, fibronectin, and vitronectin. However, we did not observe detectable secretion of collagen I and collagen IV. These data indicated that human trophoblast-derived MMPs and their inhibitors are intrinsically and developmentally regulated. The same cytotrophoblast cells that produced MMPs could also secrete various substrates for these enzymes.

The activity of matrix metalloproteinases (MMPs) specifies the ability of the trophoblast cell to degrade extracellular matrix (ECM) substrates. Usually the process of normal human placentation involves a coordinated interaction between the fetal-derived trophoblast cells and their microenvironment in the uterus. In this study, the effects of ECM proteins on the expression of MMP-2, -9, and -14 (membrane-type MMP-1); and the production of tissue inhibitors of metalloproteinase (TIMP) types -1, -2, and -3 have been investigated. Cytotrophoblast cells at 9 or 10 wk of gestation were cultured on various ECM coated dishes under serum-free conditions. Gelatin zymography analysis showed that cells grown on fibronectin (FN), laminin (LN), and vitronectin (VN) secreted more MMP-9 (about 1.5- to 3-fold more) than cells cultured on collagen I (Col I), whereas the secretion of MMP-9 by cells cultured on collagen IV (Col IV) was only half that by the cells on Col I. Northern Blot analysis gave the same results as zymography, indicating that expression of the MMP-9 gene in cytotrophoblast cells can be affected by matrix proteins. There was no significant difference in the expression of MMP-2 either at protein or mRNA levels among the cells cultured on the different matrix substrates. The expression of MMP-14 was regulated in a manner similar to that of MMP-2. Using ELISA, we detected higher levels of TIMP-1 in the culture medium of cells grown on VN, LN, and FN compared with that grown on Col I. But the expression of TIMP-3 mRNA was remarkably inhibited by VN, and ECM proteins had no effect on TIMP-1 and TIMP-2 mRNA expression. It was also observed that cultured cytotrophoblast cells expressed the corresponding receptors for the tested matrix proteins, such as integrins alpha(1), alpha(5), alpha(6), beta(1), and beta(4). Furthermore, the adhesiveness of cytotrophoblast cells on Col I, Col IV, FN, and LN was increased by 62%, 45%, 21%, and 22%, respectively, when compared with adhesiveness on VN. Isolated cytotrophoblast cells remained stationary when cultured on dishes coated with Col I and Col IV, but they assumed a more motile morphology and aggregated into a network when cultured on LN and VN. These data indicate that human trophoblast cells interact with their microenvironment to control their behavior and function.

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