Involvement of Raf-1/MEK/ERK1/2 signaling pathway in zinc-induced injury in rat renal cortical slices.
Zinc is an essential nutrient that can also be toxic. We have previously reported that zinc-related renal toxicity is due, in part, to free radical generation in the renal epithelial cell line, LLC-PK(1) cells. We have also shown that an MEK1/2 inhibitor, U0126, markedly inhibits zinc-induced renal cell injury. In this study, we investigated the role of an upstream MEK/ERK pathway, Raf-1 kinase pathway, and the transcription factor and ERK substrate Elk-1, in rat renal cortical slices exposed to zinc. Immediately after preparing slices from rat renal cortex, the slices were incubated in medium containing Raf-1 and MEK inhibitors. ERK1/2 and Elk-1 activation were determined by Western blot analysis for phosphorylated ERK (pERK) 1/2 and phosphorylated Elk-1 (pElk-1) in nuclear fractions prepared from slices exposed to zinc. Zinc caused not only increases in 4-hydroxynonenal (4-HNE) modified protein and lipid peroxidation, as an index of oxidant stress, and decreases in PAH accumulation, as that of renal cell injury in the slices. Zinc also induced a rapid increase in ERK/Elk-1 activity accompanied by increased expressions of pERK and pElk-1 in the nuclear fraction. A Raf-1 kinase inhibitor and an MEK1/2 inhibitor U0126 significantly attenuated zinc-induced decreases PAH accumulation in the slices. The Raf-1 kinase inhibitor and U0126 also suppressed ERK1/2 activation in nuclear fractions prepared from slices treated with zinc. The present results suggest that a Raf-1/MEK/ERK1/2 pathway and the ERK substrate Elk-1 are involved in free radical-induced injury in rat renal cortical slices exposed to zinc.
Kohda
Yuka
Y
Division of Pharmacology, Osaka University of Pharmaceutical Sciences, 4-21-1 Nasahara, Takatsuki, Osaka 569-1094, Japan. kohda@gly.oups.ac.jp
Matsunaga
Yoshiko
Y
Shiota
Ryugo
R
Satoh
Tomohiko
T
Kishi
Yuko
Y
Kawai
Yoshiko
Y
Gemba
Munekazu
M
D000447:Aldehydes / Q000378:metabolism; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000502:physiology*; D007672:Kidney Cortex / Q000187:drug effects* / Q000378:metabolism; D015227:Lipid Peroxidation / Q000187:drug effects; D048369:MAP Kinase Kinase 1 / Q000502:physiology*; D048370:MAP Kinase Kinase 2 / Q000502:physiology*; D008297:Male; D010655:Phenylenediamines / Q000494:pharmacology; D010766:Phosphorylation; D019908:Proto-Oncogene Proteins c-raf / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology*; D015032:Zinc / Q000633:toxicity*
D000447:Aldehydes; D000975:Antioxidants; D010655:Phenylenediamines; C004845:N,N'-diphenyl-4-phenylenediamine; D019908:Proto-Oncogene Proteins c-raf; D048049:Extracellular Signal-Regulated MAP Kinases; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; D015032:Zinc; C027576:4-hydroxy-2-nonenal