The selective NADPH oxidase inhibitor apocynin has potential prophylactic effects on melamine-related nephrolithiasis in vitro and in vivo.

Abstract

The objective of this study is to examine the effects of apocynin on melamine-cyanuric acid mixture (MCM)-induced nephrolithiasis in vitro and in vivo. For the in vitro experiments, changes in oxidative stress (OS) markers and the expression of osteopontin (OPN) and phospho-p38 (p-p38) were measured to assess the effects of apocynin treatment after MCM-induced crystallization in HK-2 cells, a human renal epithelial-derived cell line. For in vivo studies, the potential effects of apocynin in preventing and treating nephrolithiasis were analyzed with a MCM-induced nephrolithiasis rat model, and urea and creatinine levels were measured. Urinary 8-IP (a product of lipid peroxidation) and malondialdehyde levels and superoxide dismutase activity were assessed in the kidneys as markers of renal OS. The kidneys were removed, weighed, and subjected to histopathological examination. The urolithiasis-associated proteins p-p38 and OPN were evaluated by immunohistochemistry and Western blotting. Apocynin treatment prevented the MCM-induced changes in OS and in OPN and p-p38 expression in HK-2 cells. For in vivo experiments, the expression of OS markers, renal OPN, and p-p38 increased after MCM administration, and these increases were diminished by apocynin. In addition, apocynin prevented MCM-induced renal crystallization. Moreover, prophylactic apocynin treatment reduced MCM-induced nephrotoxicity. After therapeutic apocynin treatment in nephrolithic rats, OS decreased, but the other indicators did not improve significantly. Prophylactic apocynin administration reduced renal melamine-related-crystal deposition, potentially by modulating OS and thereby decreasing p-p38 and OPN expression.