Activin A, B, and AB Increase Human Trophoblast Cell Invasion by Up-regulating N-Cadherin

Abstract

CONTEXT: Inhibin β-subunits and activin receptors are expressed by human trophoblast cells. Although activin A has been shown to enhance human trophoblast cell invasion, whether two additional activin isoforms, activin B and AB, exert similar effects remains unknown. Moreover, whether the expression of mesenchymal adhesion molecule neural cadherin (N-cadherin) is essential for this proinvasive effect of activin has yet to be determined. OBJECTIVE: To examine the effects of all three activin isoforms on human trophoblast cell invasion and the involvement of N-cadherin. DESIGN: HTR8/SVneo immortalized extravillous cytotrophoblast cells and primary cultures of human extravillous cytotrophoblast cells were used as study models. Small interfering RNA-mediated knockdown approaches were used to investigate the molecular determinants of activin-mediated functions. SETTING: An academic research center. MAIN OUTCOME MEASURES: Reverse transcription-quantitative real-time PCR and Western blot analysis were used to examine mRNA and protein levels, respectively. Cell invasiveness was assessed by Matrigel-coated transwell assays. RESULTS: All three activin isoforms produced comparable increases in HTR8/SVneo cell invasion as well as N-cadherin expression. In addition, the up-regulatory effect of activin isoforms on N-cadherin was confirmed in primary cultures of human trophoblast cells. Interestingly, small interfering RNA-mediated down-regulation of N-cadherin attenuated basal and activin-induced invasion of both HTR8/SVneo and primary trophoblast cells. All three activin isoforms induced equivalent phosphorylation of SMAD2 and SMAD3. Importantly, activin-stimulated cell invasion, up-regulation of N-cadherin, as well as activation of SMAD2/SMAD3 were abolished by the TGF-β type I receptor inhibitor SB431542 in HTR8/SVneo cells. Furthermore, knockdown of SMAD2/3 or common SMAD4 abolished the stimulatory effects of all three activin isoforms on N-cadherin expression. CONCLUSION: Activin A, B, and AB produce comparable increases in human trophoblast cell invasion by up-regulating N-cadherin expression in a SMAD2/3-SMAD4-dependent manner.