Indoxyl sulfate enhances p53-TGF-ß1-Smad3 pathway in proximal tubular cells.

BACKGROUND/AIM: Indoxyl sulfate-induced activation of nuclear factor (NF)-ĸB promotes transforming growth factor (TGF)-ß1 in human proximal tubular cells (HK-2 cells). The present study aimed to elucidate the cross talk among indoxyl sulfate, p53 and TGF-ß1-Smad3 signaling in proximal tubular cells. METHODS: The effects of indoxyl sulfate on the expression of TGF-ß1, Smad3, and α-smooth muscle actin (α-SMA) were determined using HK-2 cells. As for in vivo experiments the following animals were used: Dahl salt-resistant normotensive rats (DN) and indoxyl sulfate-administered Dahl salt-resistant normotensive rats (DN+IS). RESULTS: Both indoxyl sulfate and nutlin-3, a specific p53 inducer, stimulated TGF-ß1 expression, which was suppressed by pifithrin-α, p-nitro, a p53 inhibitor. Further, indoxyl sulfate stimulated TGF-ß1-induced expression of α-SMA by enhancing Smad3 expression and TGF-ß1-induced Smad3 phosphorylation. Indoxyl sulfate induced phosphorylation of extracellular signal-regulated kinase (ERK). U0126, an inhibitor of ERK pathway, prevented indoxyl sulfate-induced upregulation of Smad3 expression. Immunohistochemistry demonstrated that TGF-ß1 and Smad3 were localized in renal tubular cells, and that indoxyl sulfate increased the TGF-ß1 and Smad3-positive area in the kidney. CONCLUSION: Indoxyl sulfate stimulates p53-induced TGF-ß1 expression and TGF-ß1-induced α-SMA expression in proximal tubular cells. Indoxyl sulfate-induced Smad3 accelerates TGF-ß1-induced α-SMA expression through ERK activation. Thus, indoxyl sulfate enhances p53-TGF-ß1-Smad3 pathway in proximal tubular cells.