TYROSINE KINASE CELL SIGNALING PATHWAYS OF RAT MESANGIAL CELLS IN 3-DIMENSIONAL CULTURES - RESPONSE TO FETAL BOVINE SERUM AND PLATELET-DERIVED GROWTH FACTOR-BB

Cells grown in 3-dimensional collagen gels adopt a nonproliferative, c ontractile phenotype which is more characteristic of cells in vivo tha n cells grown in 2-dimensional culture. The floating collagen gel cont raction assay is a well-defined system used to study cell-extracellula r matrix interactions grown in 3-dimensional culture, Although the cel l biology of this system is well defined, the cell signalling associat ed with gel contraction has not been well characterized. in this study we demonstrate that fetal bovine (FBS) and platelet-derived growth fa ctor (PDGF)-induced mesangial cell-collagen gel contraction is associa ted with increased tyrosine phosphorylation of a number of proteins in cluding focal adhesion kinase (FAK) and the 42-kDa isoform of MAPK (ER K2). FBS-induced gel contraction is not affected by the presence of th e MEK inhibitor PD098059. Low concentrations of PDGF-BB (10 ng/ml) ind uce gel contraction; however, at higher PDGF-BB concentrations (80 ng/ ml) gel contraction is not observed. PDGF-BB-induced gel contraction a s well as tyrosine phosphorylation of FAK are inhibited in the presenc e of the PI-3 kinase inhibitor wortmanin, Minimal autophosphorylation of the PDGF-beta receptor is observed under 3-dimensional culture cond itions following PDGF-BB stimulation; however, when mesangial cells gr own in 2-dimensional culture are exposed to PDGF-BB , the PDGF-beta re ceptor was prominently phosphorylated, We conclude that induction of c ollagen gel contraction by FBS and PDGF-BB is associated with tyrosine kinase phosphorylation and that these responses differ substantially from what occurs in 2-dimensional cultures in the presence of the same agonists. (C) 1998 Academic Press.