Constitutive expression and modulation of the functional thrombin receptor in the human kidney

Abstract

Thrombin exerts procoagulant effects and has also many cellular effects mediated by cell surface receptors. A functional thrombin receptor from human platelets has been cloned and sequenced. In the present study, by reverse transcription and polymerase chain reaction, using specific primers designed from the thrombin receptor cDNA sequence, we show that the mRNA encoding for this receptor can be amplified from freshly isolated human glomeruli obtained by microdissection of normal kidney cortex. By immunohistochemistry using a specific monoclonal antibody, ATAP2, directed against the extracellular N-terminus of this receptor, we find that this functional thrombin receptor is constitutively expressed in the normal human kidney. The three glomerular cell types, endothelial, mesangial, and epithelial cells, were positively stained, as were the endothelial cells of renal arteries, arterioles, venules, and peritubular capillaries. Occasionally, interstitial cells and smooth muscle cells in the media of renal arteries were also stained. Proximal and distal tubular cells were not stained. By in situ hybridization, using a digoxigenin-labeled cDNA probe specific for thrombin receptor, the thrombin receptor mRNA was found to have the same distribution as the thrombin receptor protein detected by immunohistochemistry. A lighter staining of glomerular endocapillary cells was observed in cases of thrombotic microangiopathy and extracapillary glomerulonephritis, two renal diseases associated with in situ thrombin generation and fibrin formation. In one case of thrombotic microangiopathy, we observed an increase in thrombin receptor mRNA. This suggests that thrombin receptor protein is not always correlated with thrombin receptor mRNA level. Internalization and degradation of thrombin receptor protein have been demonstrated in vitro and could also occur after activation in vivo. This is the first demonstration of the constitutive expression of the functional thrombin receptor in the human kidney. These results suggest that thrombin may exert glomerular and vascular effects within the kidney in normal and in pathological conditions.