Sphingosine 1-phosphate attenuates peroxide-induced apoptosis in HaCaT cells culturedin vitro

Abstract

Sphingosine 1-phosphate (S1P) is a sphingolipid mediator that elicits a wide array of physiological responses in various types of mammalian cells. Among the numerous biological activities elicited by S1P is protection from apoptotic cell death, which seems to take place through the cell-surface S1P receptor and the downstream phosphoinositide 3'-OH kinase (PI3-K)/Akt pathway. It is unclear whether and how S1P protects human keratinocytes from hydrogen peroxide (H2 O2 )-induced apoptosis. We investigated the effects of S1P on apoptotic cell death in HaCaT cells, spontaneously immortalized human keratinocytes. HaCaT cells were treated with hydrogen peroxide (H2 O2 ) 1-2mmol/L as an inducer of apoptosis. Cellular apoptosis was assessed with terminal dUTP nick-end labelling (TUNEL), WST-8 and immunoblot assays. In WST-8 and TUNEL assays, S1P pretreatment (1μmol/L for 30min) attenuated H2 O2 -induced cell death. Promotion of the cleavage of caspase-3 by H2 O2 was markedly attenuated when cells had been preincubated with S1P. S1P markedly potentiated phosphorylation (activation) of Akt in the presence of H2 O2 . Wortmannin, a selective inhibitor of the PI3-K/Akt pathway, significantly suppressed S1P-induced attenuation of caspase-3 cleavage promoted by H2 O2 . S1P, a sphingolipid mediator, attenuates H2 O2 -induced apoptosis of HaCaT cells, by promoting phosphorylation of the Akt pathway.