Maleate injection causes dose dependent injury in proximal tubular cells. This study sought to better define underlying pathogenic mechanisms, and to test whether maleate toxicity recapitulates critical components of the hypoxic/ischemic- renal injury cascade. CD-1 mice were injected with maleate or used as a source for proximal tubule segments (PTS) for in vitro studies. Maleate induced dose dependent PTS injury [LDH release, ATP reductions, non esterified fatty acid (NEFA) accumulation]. These changes were partially dependent on maleate metabolism (protection conferred by succinate / acetoacetate: metabolic inhibitors). Maleate toxicity reproduced critical characteristics of the hypoxia/ATP depletion- induced injury cascade: 1) Glutathione (GSH) conferred protection, but due to its glycine, not cysteine (antioxidant), content; 2) ATP reductions reflected decreased production, not NaK-ATPase driven increased consumption; 3) cell death was completely blocked by extracellular acidosis (pH6.6); 4) intracellular Ca²⁺ chelation (BAPTA) mitigated cell death; 5) maleate and hypoxia each caused plasma membrane cholesterol shedding; in both instances, this was completely suppressed by glycine; 6) maleate + hypoxia caused neither additive NEFA accumulation nor LDH release, suggesting shared pathogenic pathways; and 7) maleate, like ischemia, induced renal cortical cholesterol loading: increased HMG CoA reductase (HMGCR) activity (statin inhibitable), increased HMGCR mRNA levels, and increased RNA polymerase II recruitment to the HMGCR locus (chromatin immunoprecipitation, ChIP, assay) were involved. These results further define critical determinants of maleate nephrotoxicity, and suggest that it can serve as a useful adjunct for studies of ischemia / ATP depletion- induced, proximal tubule specific, cell death.