Protection by the Chinese prescription Wen-Pi-Tang against renal tubular LLC-PK1 cell damage induced by 3-morpholinosydnonimine.

Abstract

We investigated the effects of Wen-Pi-Tang extract on the protective mechanisms of renal tubular LLC-PK1 cells, as renal tubular cells are the most vulnerable renal tissue to oxidative stress. Exposure to 800 microM 3-morpholinosydnonimine (SIN-1) resulted in a marked increase in cellular peroxynitrite (ONOO-), which converted nonfluorescent dihydrorhodamine 123 to fluorescent rhodamine 123, a detectable probe for the long-lived ONOO-. In addition, it resulted in apoptotic cell death, assessed by a DNA fragmentation assay. However, treatment with Wen-Pi-Tang extract, at concentrations of 50 and 100 microg mL(-1) together with SIN-1 protected renal tubular cells against ONOO- through scavenging ONOO- and inhibiting apoptotic cell death in a dose-dependent manner. Moreover, treatment with Wen-Pi-Tang extract both before and after exposure to SIN-1 was also protective: it reduced cellular ONOO- levels, increased cell viability and decreased the DNA fragmentation rate. These results suggest that Wen-Pi-Tang would have protective activity against ONOO- -induced renal tubular injury through the inhibition of ONOO- production and apoptotic cell death by both preventing and treating renal injury. Furthermore, morphological characteristics of apoptosis were observed in SIN-1 treated tubular cells, while the addition of Wen-Pi-Tang extract with SIN-1 attenuated these morphological changes. ONOO- generated by SIN-1 also disturbed the cell cycle by decreasing the cellular G2/M phase ratio, while Wen-Pi-Tang extract regulated the cell cycle by G2/M phase arrest.