Maleic acid (MA) has been shown to induce
Fanconi syndrome via disturbance of renal energy homeostasis, though the underlying pathomechanism is still under debate. Our study aimed to examine the pathomechanism underlying
maleic acid-induced nephrotoxicity.
Methylmalonic acid (MMA) is structurally similar to MA and accumulates in patients affected with methymalonic aciduria, a defect in the degradation of
branched-chain amino acids, odd-chain
fatty acids and
cholesterol, which is associated with the development of
tubulointerstitial nephritis resulting in
chronic renal failure. We therefore used MMA application as a control experiment in our study and stressed hPTECs with MA and MMA to further validate the specificity of our findings. MMA did not show any toxic effects on proximal tubule cells, whereas
maleic acid induced concentration-dependent and time-dependent cell death shown by increased
lactate dehydrogenase release as well as
ethidium homodimer and
calcein acetoxymethyl ester staining. The toxic effect of MA was blocked by administration of single
amino acids, in particular
L-alanine and
L-glutamate. MA application further resulted in severe impairment of cellular energy homeostasis on the level of glycolysis, respiratory chain, and citric acid cycle resulting in
ATP depletion. As underlying mechanism we could identify disturbance of
calcium homeostasis. MA toxicity was critically dependent on
calcium levels in culture medium and blocked by the extra- and intracellular
calcium chelators EGTA and
BAPTA-AM respectively. Moreover, MA-induced cell death was associated with activation of
calcium-dependent
calpain proteases. In summary, our study shows a comprehensive pathomechanistic concept for MA-induced dysfunction and damage of human proximal tubule cells.