Effect Of MSX2, Msh Homeobox 2, In The Invasion Of Placental Trophoblast Cells

Background: Normal implantation depends on appropriate trophoblast growth and invasion. Inadequate trophoblast invasion results in pregnancy-related disorders, such as early miscarriage and pre-eclampsia, which are dangerous to both the mother and fetus. MSX2(Msh Homeobox 2), a member of the MSX family of homeobox proteins, plays a significant role in the proliferation and differentiation of various cells and tissues, including ectodermal organs, teeth, and chondrocytes. Recently, MSX2 was also found to play important roles in the invasion of cancer cells into adjacent tissues via the epithelial-mesenchymal transition(EMT). However, the role of MSX2 in trophoblastic invasion during placental development has yet to be explored.Methods: 1. immunohistochemistry was used to detect the expression of human placental trophoblast cells in different trimesters. 2. Detection the expression level in four kinds of trophoblastic cells(HTR8/SVneo, JEG-3, JAR, and BeWo cells) 3. MSX2 overexpression plasmid or MSX2-specific siRNA transfected into HTR8/SVneo cells to up or down the expression of MSX2 protein, then useing of cell invasion assay(Matrigel invasion assay) to detect MSX2 Effect of trophoblast invasion ability. The supernatant was collected and the enzymatic activities of matrix metalloproteinases(MMP-2 and MMP-9) in HTR8/SVneo cells were analyzed by gelatin zymography. 4. We examined the effects of different MSX2 expression levels on the expression levels of β-catenin, E-cadherin and vimentin, to elucidate the role of MSX2 during the EMT process. 5. We compared the differential expression patterns of MSX2 in pre-eclampsic and normal placental villi to investigate the role of MSX2 in PE pathogenesis.Results: In the present study, we detected MSX2 expression in cytotrophoblast, syncytiotrophoblast, and extravillous cytotrophoblast cells of third trimester human placentas via immunohistochemistry analysis. Furthermore, we found that the in vitro invasive ability of HTR8/SVneo cells was enhanced by exogenous overexpression of MSX2(271±45vs. 443±75,P<0.05), and that this effect was accompanied by increased protein expression of MMP-2, vimentin, and β-catenin(P<0.05). Conversely, treatment of HTR8/SVneo cells with MSX2-specific siRNAs resulted in decreased protein expression of MMP-2, vimentin, and β-catenin, and reduced invasion levels in a Matrigel invasion test(P<0.05). Notably, however, treatment with the MSX2 overexpression plasmid and the MSX2 siRNAs had no effect on the mRNA expression levels of β-catenin(P>0.05). Meanwhile, overexpression of MSX2 and treatment with the MSX2-specific siRNA resulted in decreased and increased E-cadherin expression(P<0.05), respectively, in JEG-3 cells. Lastly, the protein expression levels of MSX2 were significantly lower in human pre-eclamptic placental villi than in the matched control placentas.Conclusion: Collectively, our results suggest that MSX2 may induce human trophoblast cell invasion, and that modulation of abnormal MSX2 expression could be utilized to treat pre-eclampsia.