Involvement of activation of NADPH oxidase and extracellular signal-regulated kinase (ERK) in renal cell injury induced by zinc.
Zinc is employed as a supplement; however, zinc-related nephropathy is not generally known. In this study, we investigated zinc-induced renal cell injury using a pig kidney-derived cultured renal epithelial cell line, LLC-PK(1), with proximal kidney tubule-like features, and examined the involvement of free radicals and extracellular signal-regulated kinase (ERK) in the cell injury. The LLC-PK(1) cells showed early uptake of zinc (30 microM), and the release of lactate dehydrogenase (LDH), an index of cell injury, was observed 24 hr after uptake. Three hours after zinc exposure, generation of reactive oxygen species (ROS) was increased. An antioxidant, N, N'-diphenyl-p-phenylenediamine (DPPD), inhibited a zinc-related increase in ROS generation and zinc-induced renal cell injury. An NADPH oxidase inhibitor, diphenyleneiodonium (DPI), inhibited a zinc-related increase in ROS generation and cell injury. We investigated translocation from the cytosol fraction of the p67(phox) subunit, which is involved in the activation of NADPH oxidase, to the membrane fraction, and translocation was induced 3 hr after zinc exposure. We examined the involvement of ERK1/2 in the deterioration of zinc-induced renal cell injury, and the association between ERK1/2 and an increase in ROS generation. Six hours after zinc exposure, the activation (phosphorylation) of ERK1/2 was observed. An antioxidant, DPPD, inhibited the zinc-related activation of ERK1/2. An MAPK/ERK kinase (MEK1/2) inhibitor, U0126, almost completely inhibited zinc-related cell injury (the release of LDH), but did not influence ROS generation. These results suggest that early intracellular uptake of zinc by LLC-PK(1) cells causes the activation of NADPH oxidase, and that ROS generation by the activation of the enzyme leads to the deterioration of renal cell injury via the activation of ERK1/2.
Matsunaga
Yoshiko
Y
Division of Pharmacology, Osaka University of Pharmaceutical Sciences, Nasahara, Takatsuki, Japan.
Kawai
Yoshiko
Y
Kohda
Yuka
Y
Gemba
Munekazu
M
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D004789:Enzyme Activation; D004847:Epithelial Cells / Q000187:drug effects / Q000473:pathology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000502:physiology*; D007668:Kidney / Q000187:drug effects* / Q000473:pathology; D019255:NADPH Oxidases / Q000502:physiology*; D010655:Phenylenediamines / Q000494:pharmacology; D017382:Reactive Oxygen Species; D013552:Swine; D015032:Zinc / Q000378:metabolism / Q000633:toxicity*
D000975:Antioxidants; D010655:Phenylenediamines; D017382:Reactive Oxygen Species; C004845:N,N'-diphenyl-4-phenylenediamine; D019255:NADPH Oxidases; D048049:Extracellular Signal-Regulated MAP Kinases; D015032:Zinc