Protective Effects of N-Acetylcysteine Amide (NACA) on Gentamicin-Induced Apoptosis in LLC-PK1 Cells

Abstract

Aim: Apoptosis plays a critical role in the pathogenesis of gentamicin (Gen)-induced nephrotoxicity. However, the underlying molecular mechanisms still remain unclear. In this study, we addressed the role of p38 mitogen-activated protein kinase (MAPK)/inducible nitric oxide synthase (iNOS) signaling pathway in Gen-induced nephrotoxicity and evaluated the protective effect of the free-radical scavenger N-acetylcysteine amide (NACA). Methods: Pig kidney epithelial cells (LLC-PK1) cells were exposed to Gen for variable times and doses. Cytotoxicity was assessed by morphology and by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Protein expression was assessed by Western blotting. Results: Exposure to Gen-induced apoptosis in a dose-dependent and time-dependent manner was assessed by DNA content analysis and poly ADP ribose polymerase (PARP) cleavage. Gen caused increased phosphorylation of p38 MAPK and induction of iNOS. This was accompanied by a significant upregulation of Bax and nuclear factor κB (NF-κB) and a downregulation of Bcl-2 expression. Pretreatment with SB203580, aminoguanidine (AG), and NACA inhibited apoptosis. Furthermore, pretreatment with SB203580 and NACA not only attenuated the pro-apoptotic effect of Gen, but also significantly reversed its effects on p38 MAPK phosphorylation and iNOS induction. The Gen-induced effects on Bcl-2, Bax, and NF-κB expression were also reversed by SB203580, AG, and NACA. Conclusion: In conclusion, NACA can attenuate Gen-induced apoptotic injury in LLC-PK1 cells through inhibiting p38 MAPK/iNOS signaling pathway.

• apoptosis
• gentamicin-induced nephrotoxicity
• N-acetylcysteine amide
• p38 mitogen-activated protein kinase
• inducible nitric oxide synthase

ACKNOWLEDGMENT

We thank Dr. Glenn Goldstein for providing N-acetylcysteine amide.

Declaration of interest: The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.