Effects of Matrix Proteins on the Expression of Matrix Metalloproteinase-2, -9, and -14 and Tissue Inhibitors of Metalloproteinases in Human Cytotrophoblast Cells During the First Trimester1

Abstract

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.

Animal models of implantation

“…Possible ligands for the integrins on the maternal surface include osteopontin, laminin, thrombospondin, and perlecan (Kimber & Spanswick 2000), and on the blastocyst, the oncofetal fibronectin, a protein restricted only to trophoblasts (Feinberg et al 1991, Craven et al 2000. In addition, in vitro studies have shown that a v b3 may also interact and activate specific matrix metalloproteinases in the extracellular matrix of the uterus (Xu et al 2001), thereby implicating integrins in the transition between attachment and epithelial cell penetration.…”

IGF2, IGF binding protein 1, and matrix metalloproteinases 2 and 9 in implantation-stage endometrium following immunoneutralization of vascular endothelial growth factor in the rhesus monkey

“…As the cells lose their proliferative ability, they invade the uterine wall by expressing specific matrix metalloproteinases (MMPs; Xu et al 2001, Staun-Ram et al 2004. It has been demonstrated that VEGF and MMPs 2 and 9 may comprise a functional network regulating the invasive phenotype in a variety of cell types including trophoblast cells (Anteby et al 2004, Hollborn et al 2007.…”

DNA Methylation Changes in Sera of Women in Early Pregnancy Are Similar to Those in Advanced Breast Cancer Patients

“…We also asked whether such an invasion-specific methylation pattern in serum shows differences between women with normal pregnancies and women developing preeclampsia-a disease known to be accompanied by a disturbed invasion process in the first trimester (6 ). We chose a panel of three genes (CDH1, TIMP-3, and PTGS-2) known to play key roles in the invasion process of tumor cells (17)(18)(19), which are often regulated by promoter hypermethylation (20 -22 ), or in the invasion process of trophoblast cells (2,(23)(24)(25). We also chose one gene (BLT1) involved in the regulation of immune response and regulated by promoter hypermethylation (26 ).…”

“…From our point of view, the most important finding is the similarity between pregnant women and metastasized breast cancer patients in the methylation changes in genes that are known to be involved in metastasis and tumor cell invasion (17-19, 27, 28 ) or even in the invasion process of trophoblast cells (2,(23)(24)(25) and in the regulation of the immune response (26 ). As sera of those cancer patients without evidence of metastasis at the time of diagnosis lacked the methylation changes found in advanced breast cancer and pregnancy (Table 1), we speculate that the observed methylation pattern reflects DNA release from invasive cells, specifically trophoblast cells and tumor cells.…”