Long-term treatment with TGFβ1 impairs mechanotransduction in bovine aortic endothelial cells

Background and purpose: Vascular endothelial cells play a role in the physiological response to mechanical stress. Transforming growth factor β1 (TGFβ1) induces morphological changes in endothelial cells, and this may alter their mechanosensitive responses. The aim of this study was to examine the effects of TGFβ1 on hypotonic stress (HTS)-induced responses in bovine aortic endothelial cells (BAECs). Experimental approach: Cultured BAECs were treated with 3 ng ml-1 TGFβ1 for 24 h (24h-TGFβ1) or 7 days (7d-TGFβ1). Cytosolic actin fibres were stained with rhodamine-phalloidin. Intracellular Ca2+ concentration was measured using fura2. Tyrosine phosphorylation and RhoA expression were assessed by Western blotting. Expression of RhoA mRNA was assessed by real-time PCR. Key results: BAECs developed pseudopod-like processes within 24 h and showed a fibroblast-like appearance after 7 days. HTS induced Ca2+ transients via endogenous ATP release in both control and 24h-TGFβ1 BAECs but not in 7d-TGFβ1 BAECs. We have previously shown that HTS-induced ATP release is mediated by sequential activation of RhoA and tyrosine kinases. The basal amount of membrane-bound RhoA was significantly lower in 7d-TGFβ1 than in 24h-TGFβ1 or control BAECs. HTS increased the membrane-bound RhoA to the same fractional level in 24h-TGFβ1 and control BAECs, but its net maximal amount was significantly lower in 7d-TGFβ1. HTS-induced downstream signals of RhoA activation, i.e. the tyrosine phosphorylation of FAK and paxillin, were markedly suppressed in 7d-TGFβ1 BAECs. Conclusions and Implications: These results indicate that long-term treatment with TGFβ1 does not impair mechanoreception in BAECs but impairs mechanotransduction by affecting RhoA membrane translocation. British Journal of Pharmacology (2007) 150, 424–433. doi:10.1038/sj.bjp.0707123