Overexpression of Regucalcin Suppresses Cell Response for TNF‐α or TGF‐β1 in Rat Kidney Proximal Tubular Epithelial NRK52E Cells

The regulatory role of regucalcin, a regulatory protein in intracellular signaling, on cell responses for TNF‐alpha or TGF‐beta1 was investigated using rat kidney proximal tubular epithelial NRK52E cells overexpressing regucalcin. NRK52E cells (wild type) and stable regucalcin‐transfected cells (transfectant) with subconfluency were cultured for 24–72 h in medium without bovine serum containing either vehicle, TNF‐alpha (0.1 or 1.0 ng/ml of medium), or TGF‐beta1 (1.0 or 5.0 ng/ml). Culture with TNF‐alpha or TGF‐beta1 caused a significant decrease in the number of wild‐type cells. This decrease was significantly prevented in transfectants overexpressing regucalcin. DNA fragments of adherent wild‐type cells was induced with TNF‐alpha or TGF‐beta1. This was significantly suppressed in transfectants. TNF‐alpha caused a significant increase in NO synthase activity in wild‐type cells. This effect was not seen in transfectants. TGF‐beta1 did not cause a significant increase in NO synthase activity in both cell types. TNF‐alpha or TGF‐beta1 caused a remarkable increase in alpha‐smooth muscle actin in wild‐type cells. This increase was significantly prevented in transfectants. The expression of Smad 2 or NF‐kapperB mRNAs was significantly increased in transfectants as compared with that of wild‐type cells. NF‐kapperB mRNA expression in wild‐type cells was significantly increased with TNF‐alpha. Smad 2 mRNA expression was significantly enhanced in wild‐type cells cultured with TGF‐beta1. These increases were not significantly enhanced in transfectants. Overexpression of regucalcin had suppressive effects on cell responses which are mediated through intracellular signaling pathways of TNF‐alpha or TGF‐beta1 in kidney NRK52E cells.