HYPOXIA INDUCES CHANGES IN PHOSPHOLIPASE A(2) IN RAT PROXIMAL TUBULES- EVIDENCE FOR MULTIPLE FORMS

Increased free fatty release during hypoxia is believed to contribute to cell injury. This phenomenon is likely to be mediated through activ ation of specific isoforms of phospholipase A(2) (PLA(2)). In this stu dy, PLA(2) enzymatic activity was measured in cell-free extracts prepa red from rat renal proximal tubules. Both soluble and membrane-associa ted PLA(2) activity were detected. All PLA(2) activity detected during normoxia was Ca2+ dependent. Fractionation of cytosolic extracts by g el filtration revealed three peaks of PLA(2) activity. Exposure of tub ules to hypoxia resulted in stable activation of soluble PLA(2) activi ty, which correlated with disappearance of the highest molecular mass form (>100 kDa) and appearance of a low-molecular-mass form (similar t o 15 kDa) of PLA(2). Hypoxia also resulted in release of a low-molecul ar-mass form of PLA(2) into the extracellular medium. Pretreatment of tubules with glycine before hypoxia blocked this release of PLA(2) but not activation of soluble FLAP activity. These studies provide direct evidence for PLA(2) activation during hypoxia and suggest that multip le mechanisms regulate free fatty acid release associated with hypoxic injury.