Functional effects of transforming growth factor beta (TGFβ) on adhesive properties of porcine trophectoderm

Abstract

In pigs, expression and amounts of biologically active beta transforming growth factors (TGFβs) at the conceptus-maternal interface increase significantly as conceptuses elongate and begin the implantation process. Prior to their activation, secreted TGFβs are non-covalently associated with their respective, isoform-specific latency-associated peptides (LAPs) which contain the Arg-Gly-Asp (RGD) amino acid sequence that serves as a ligand for numerous integrins. Objectives of this study were to determine if TGFβ1 increases production of fibronectin by porcine trophectoderm, if porcine trophectoderm adheres specifically to fibronectin and LAP, and if functional interactions between porcine trophectoderm and the two TGFβ-associated proteins, fibronectin and LAP, are integrin-mediated. Porcine trophectoderm cells (pTr2) were cultured in presence of TGFβ1, LAP, or pan-neutralizing anti-TGFβ antibody; TGFβ specifically increased (p<0.05) fibronectin mRNA levels, as determined by Northern and slot blot analyses. Immunofluorescence microscopy demonstrated a TGFβ-induced increase in fibronectin in pTr2 cells. In dispersed cell adhesion assays, adhesion of pTr2 cells to fibronectin was inhibited by an RGD-containing peptide (p<0.05) and pTr2 cells attached to recombinant LAP but not to an LAP mutant which contained an RGE sequence rather than the RGD site (p<0.05). Fibronectin- and LAP-coated microbeads induced integrin activation at apical surfaces of both trophectoderm and uterine luminal epithelial cells, as indicated by aggregation and transmembrane accumulation of talin detected with immunofluorescence microscopy. Cell surface biotinylation and immunoprecipitation revealed integrin subunits αv and β1 on apical membranes of pTr2 cells. These results suggest multiple effects of TGFβ at the porcine conceptus-maternal interface, including integrin-mediated conceptus-maternal communication through LAP. Copyright © 2005 by The Endocrine Society.