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Renal ischemia/reperfusion and ATP depletion/repletion in LLC-PK[sub 1] cells result in phosphorylation of FKHR and FKHRL1. Background. Cell death and survival pathways are critical determinants of epithelial cell fate after ischemia. Forkhead proteins have been implicated in the regulation of cellular survival. Methods and Results. We have found that none of the forkhead family of proteins, FKHR, is phosphorylated after ischemia/reperfusion in the rat kidney. The time course of phosphorylation is similar to the time course of activation of the forkhead protein kinase Akt/protein kinase B (PKB), with maximal phosphorylation at 24 to 48 hours postreperfusion when the process of regeneration peaks. Extracellular signal-regulated kinase (ERK)1/2 activation has also been implicated as prosurvival in the injured kidney. ERK1/2 were phosphorylated in postischemic kidneys at 5, 30, and 90 minutes of reperfusion, with phosphorylation decreased by 24 and 48 hours. Immunocytochemical analysis revealed increased phospho-ERK1/2 in the thick ascending limb and isolated cells of the S3 segment, which have lost apical actin staining. To understand the relationship between forkhead phosphorylation, Akt, and ERK1/2, an in vitro model of injury was employed. After 40 minutes of chemical anoxia followed by dextrose addition for 20 minutes to replete adenosine triphosphate (ATP) levels, FKHR and FKHRL1 are phosphorylated. The levels of phospho-Akt are increased for at least 120 minutes after dextrose addition with a maximum at 20 minutes. Phosphorylation of Akt, FKHR, and FKHRL1 are phosphatidylinositol 3-kinase (PI 3-kinase) dependent since phosphorylation is reduced by the PI 3-kinase inhibitors, wortmannin, or LY294002. Inhibition of mitogen-activated protein kinase (MAPK)/ERK kinase (MEK1/2), the upstream activator of ERK1/2, has no effect on forkhead protein phosphorylation after chemical anoxia/dextrose addition. Conclusion. We conclude that PI 3-kinase and Akt are activated after renal ischemia/reperfusion and that Akt phosphorylation leads to phosphorylation of FKHR and FKHRL1, which may affect epithelial cell fate in acute renal failure. [ABSTRACT FROM AUTHOR]