Role of ceramide synthase in oxidant injury to renal tubular epithelial cells.
Ceramide has been implicated to play an important role in the cell signaling pathway involved in apoptosis. Most studies that have used the apoptotic model of cellular injury have suggested that enhanced ceramide generation is the result of the breakdown of sphingomyelin by sphingomyelinases. However, the role of ceramide synthase in enhanced ceramide generation in response to oxidant stress has not been previously examined in any tissue. Hydrogen peroxide (H(2)O(2)) (1 mM) resulted in a rapid increase in ceramide generation (as measured by in vitro diacylglycerol kinase assay) in LLC-PK1 cells. The intracellular ceramide level was significantly increased at 5 min after exposure of cells to H(2)O(2) and thereafter continuously increased up to 60 min. H(2)O(2) also resulted in a rapid increase (within 5 min) in ceramide synthase activity (as measured by incorporation of [(14)C] from the labeled palmytoyl-CoA into dihydroceramide) in microsomes. In contrast, the exposure of cells to H(2)O(2) did not result in any significant change in sphingomyelin content or acid or neutral sphingomyelinase activity. An increase in ceramide production induced by H(2)O(2) preceded any evidence of DNA damage and cell death. The specific inhibitor of ceramide synthase, fumonisin B1 (50 microM), was able to suppress H(2)O(2)-induced ceramide generation and provided a marked protection against H(2)O(2)-induced DNA strand breaks, DNA fragmentation, and cell death. Taken together, these data provide the first evidence that H(2)O(2) is a regulator of ceramide synthase rather than sphingomyelinases and that ceramide synthase-dependent ceramide generation plays a key role in DNA damage and cell death in oxidant stress to renal tubular epithelial cells.
Ueda
Norishi
N
University of Arkansas for Medical Sciences, Division of Nephrology, and Central Arkansas Veterans Healthcare System, Little Rock, Arkansas.
Camargo
Simone M R
SMR
University of Arkansas for Medical Sciences, Division of Nephrology, and Central Arkansas Veterans Healthcare System, Little Rock, Arkansas.
Hong
Xiaoman
X
University of Arkansas for Medical Sciences, Division of Nephrology, and Central Arkansas Veterans Healthcare System, Little Rock, Arkansas.
Basnakian
Alexei G
AG
University of Arkansas for Medical Sciences, Division of Nephrology, and Central Arkansas Veterans Healthcare System, Little Rock, Arkansas.
Walker
Patrick D
PD
University of Arkansas for Medical Sciences, Division of Nephrology, and Central Arkansas Veterans Healthcare System, Little Rock, Arkansas.
Shah
Sudhir V
SV
University of Arkansas for Medical Sciences, Division of Nephrology, and Central Arkansas Veterans Healthcare System, Little Rock, Arkansas.
D000818:Animals; D002264:Carboxylic Acids / Q000494:pharmacology; D016923:Cell Death / Q000187:drug effects; D002518:Ceramides / Q000096:biosynthesis; D004249:DNA Damage / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000502:physiology; D037341:Fumonisins; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D007684:Kidney Tubules / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D018374:LLC-PK1 Cells; D016877:Oxidants / Q000494:pharmacology*; D010088:Oxidoreductases / Q000037:antagonists & inhibitors / Q000502:physiology*; D013108:Sphingomyelin Phosphodiesterase / Q000378:metabolism; D013109:Sphingomyelins / Q000378:metabolism; D013552:Swine
D002264:Carboxylic Acids; D002518:Ceramides; D004791:Enzyme Inhibitors; D037341:Fumonisins; D016877:Oxidants; D013109:Sphingomyelins; C056933:fumonisin B1; D006861:Hydrogen Peroxide; D010088:Oxidoreductases; C108138:dihydroceramide desaturase; D013108:Sphingomyelin Phosphodiesterase