Regenerative and proinflammatory effects of thrombin on human proximal tubular cells

Interstitial fibrin deposition is a common histologic feature of tubulointe rstitial diseases, which suggests that the coagulation system is activated. Thrombin, generated during the activation of the coagulation cascade, is a powerful activating factor for different cell types. Although proximal tub ular cells are potential targets for this coagulation factor, no informatio n is available on the effect of thrombin on these cells. Thus, the expressi on of protease-activated receptor-1 (PAR-1), the main thrombin receptor, wa s investigated in human proximal tubular cells (hPTC) in vivo and in vitro. A diffuse expression of PAR-1 was observed by immunohistochemistry along t he basolateral membrane of PTC in normal human kidney. This observation was confirmed in vitro in cultured hPTC. Because tubular damage and monocyte i nfiltration are two hallmarks of tubulointerstitial injury, the effect of t hrombin on DNA synthesis and monocyte chemotactic peptide-1 (MCP-1) gene an d protein expression was evaluated in cultured hPTC. Thrombin induced a sig nificant and dose-dependent increase in thymidine uptake and a striking upr egulation of MCP-1 mRNA expression and protein release into the supernatant . Although PAR-1 is a G protein-coupled receptor, its activation in hPTC, a s in other cell systems, resulted in a transient increase in cellular level s of tyrosine-phosphorylated proteins. An increased level of tyrosine-phosp horylated c-src suggested the activation of this cytoplasmic tyrosine kinas e in response to thrombin and its potential role in thrombin-induced protei n-tyrosine phosphorylation. Interestingly, thrombin-induced DNA synthesis a nd MCP-1 gene expression were completely blocked by genistein, a specific t yrosine kinase inhibitor, but not by its inactive analogue daidzein, demons trating a central role for tyrosine kinase activation in the thrombin effec ts on hPTC. Moreover, the specific src inhibitor PP1 abolished the thrombin effect on DNA. synthesis. In conclusion, thrombin might represent a powerf ul regenerative and proinflammatory stimulus for hPTC in acute and chronic tubulointerstitial diseases.