4-Hydroxyhexenal Is a Potent Inducer of the Mitochondrial Permeability Transition

Mitochondria undergo at least two types of structural alteration in response to various physiological and pathophysiological stimuli. One type is nonreversible and is associated with mitochondrial lysis. The second is reversible and appears to be associated with calcium-mediated activation of a specific inner mitochondrial membrane channel. The mechanisms underlying the induction of this second alteration, termed a mitochondrial permeability transition (PT), have been the subject of a great deal of recent research. Using rat liver mitochondria, our data demonstrate that calcium-mediated PT induction can be affected by the lipid peroxidation byproducts 4-hydroxynonenal and 4-hydroxyhexenal (HHE). 4-Hydroxynonenal appears inactive at concentrations <1 uM but displays both stimulatory and inhibitory effects as part of a biphasic dose response between approximately 1 and 200 uM. In contrast, HHE consistently enhances calcium-mediated induction of the PT, even at femtomolar concentrations. The exquisite specificity and sensitivity of HHE led to further studies to examine the nature of this induction. Studies showing that HHE-mediated induction could be prevented by cyclosporin A confirmed PT involvement. Further studies showed that induction was dependent on both calcium and electron transport chain function. Pretreatment of the HHE with glutathione also prevented PT induction, but simultaneous addition of the thiol reagents dithiothreitol or glutathione, which often prevents PT induction, was ineffective, attesting to the effectiveness of HHE as an inducer. Together, these data provide a possible mechanistic explanation for the previously observed effects of lipid peroxidation on PT induction. Mitochondria undergo at least two types of structural alteration in response to various physiological and pathophysiological stimuli. One type is nonreversible and is associated with mitochondrial lysis. The second is reversible and appears to be associated with calcium-mediated activation of a specific inner mitochondrial membrane channel. The mechanisms underlying the induction of this second alteration, termed a mitochondrial permeability transition (PT), have been the subject of a great deal of recent research. Using rat liver mitochondria, our data demonstrate that calcium-mediated PT induction can be affected by the lipid peroxidation byproducts 4-hydroxynonenal and 4-hydroxyhexenal (HHE). 4-Hydroxynonenal appears inactive at concentrations <1 uM but displays both stimulatory and inhibitory effects as part of a biphasic dose response between approximately 1 and 200 uM. In contrast, HHE consistently enhances calcium-mediated induction of the PT, even at femtomolar concentrations. The exquisite specificity and sensitivity of HHE led to further studies to examine the nature of this induction. Studies showing that HHE-mediated induction could be prevented by cyclosporin A confirmed PT involvement. Further studies showed that induction was dependent on both calcium and electron transport chain function. Pretreatment of the HHE with glutathione also prevented PT induction, but simultaneous addition of the thiol reagents dithiothreitol or glutathione, which often prevents PT induction, was ineffective, attesting to the effectiveness of HHE as an inducer. Together, these data provide a possible mechanistic explanation for the previously observed effects of lipid peroxidation on PT induction.