Dexpramipexole Attenuates Myocardial Ischemia/reperfusion Injury Through Upregulation Of Mitophagy By Targeting PINK1 And Parkin

BackgroundAcute myocardial infarction presenting as ST-segment elevation(STEMI)is the result of abrupt occlusion of an epicardial coronary artery,which is associated with significant morbidity and mortality.Timely restoration of blood flow to the ischemic myocardium to limit infarct size,improve long-term myocardial function,and reduce mortality has become the standard treatment for patients.However,a large body of experimental and clinical evidence supports the notion that reperfusion induces additional damage to the myocardium,known as reperfusion injury.Myocardial ischemia/reperfusion(I/R)injury is a complex phenomenon involving many players,all contributing to the final damage inflicted on the heart.The mechanisms underlying I/R-induced injury remain incompletely explored;however,increasing evidence indicates that modulation of mitophagy is a novel therapeutic strategy in myocardial I/R injury.Mitophagy is a type of autophagy that occurs in the mitochondria,and it plays a critical role in the selective removal of damaged or unwanted mitochondria to avoid cell death.Mitophagy is induced in mammalian cells characteristically by the PINK1-Parkin pathway.The proteins PINK1/Parkin are selectively recruited to dysfunctional mitochondria with a low membrane potential,where it subsequently mediates the degradation of impaired mitochondria by autophagosomes.It has been found that PINK1/Parkin-mediated mitophagy is also induced by hypoxia and helps to restore cellular homeostasis by degrading defective mitochondria.Dexpramipexole(DPX),is the non-dopaminergic R(+)enantiomer of the high-affinity dopamine agonist and Parkinson's disease therapeutic pramipexole.Due to the non-high-affinity property of DPX,its use avoids the side effects caused by dopaminergic agonists.It has been reported that DPX can increase cells' energy production efficiency,including neurons,at concentrations that constrain large-conductance mitochondrial currents.Research has demonstrated that currents located at the inner mitochondrial membrane are suppressed by DPX and are also inhibited by ATP.Previous studies have reported that DPX reduces the production of reactive oxygen species(ROS),attenuates the activation of apoptotic pathways,and increases cell survival.Thus,it is plausible that DPX may promote mitophagy and participate in ischemia/reperfusion injury.ObjectivesThis article assesses the significance of DPX in acute myocardial infarction,and whether DPX can promote mitophagy via PINK1 and Parkin translocation to mitochondria induced by ischemic/reperfusion injury in vivo and in vitro.Methods and ResultsNeonatal rat cardiomyocytes(NRCs)were isolated from newborn(1 to 2 day old)Sprague-Dawley rats.Cells were subjected to hypoxia followed by reoxygenation.Hypoxia/reoxygenation(H/R)treatments were used to mimic an in vivo I/R model.Cells were washed with PBS and incubated in PBS then transferred to in a hypoxic chamber(Modular Incubator Chamber)in a humidified atmosphere that equilibrated with 95%N2 and 5%Co2 at 37? for 3h.Reoxygenation was then exchanged with normoxic complete medium for 3h.DPX treatments were administered 24 and 1 h before hypoxia induction.Results:DPX up to 100 ?M did not cause cellular cytotoxicity.PINK1 protein levels were reduced after hypoxia or hypoxia followed by 3h of reoxygenation.DPX pretreatment inhibited the reduction of PINK1.Treatment with 10 ?M DPX increased survival of cardiomyocyte cells undergoing H/R challenge.LDH leakage increased in the H/R group compared with the control group but was significantly decreased by DPX treatment.TUNEL assay for the apoptosis index was significantly decreased by DPX treatment.The activity of cleaved caspase-3 was inhibited by DPX.These results suggest that DPX alleviates H/R injury.Male adult C57BL/6 mice(20-25 g)were subjected to myocardial ischemia followed by reperfusion.Mice were occluded the left anterior descending coronary artery by snaring with a vinyl tube through ligature to subjected to 45 min of ischemia,then released the ligature for 24 h of reperfusion.DPX reduced the I/R-induced myocardial infarct size.DPX displayed partial but significantly restored cardiac function.DPX significantly reduced TUNEL-positive myocytes in ischemic heart.DPX was resistant to I/R-induced myocardial necrosis,as evidenced by reductions in serum creatine kinase(CK),lactate dehydrogenase(LDH)release and cardiac-specific isoform of troponin-I(cTnI).These results suggest that the protective effect of DPX is attributable to its inhibitory effect on I/R-induced myocardial necrosis.We then investigated the molecular mechanism underlying the protective effects of DPX.Results indicate that DPX exerts protective effects against H/R-induced mitochondrial dysfunction.DPX enhances mitophagy in vivo and vitro.PINK1 takes part in the agitating effects of DPX on myocardial H/R injury.DPX activates mitophagy by enhancing translocation of Parkin to the mitochondria.Knock-down of PINK1 and Parkin by specific siRNAs reversed dexpramipexole-induced inhibition of myocardial ischemic reperfusion injury.These results suggested that DPX regulates mitophagy by activating PINK1 and Parkin.Conclusions1.DPX pretreatment protects against myocardial I/R injury in vivo;2.DPX pretreatment attenuates H/R injury in vitro;3.We firstly identified DPX ameliorates myocardial ischemic reperfusion injury through upregulation of mitophagy by targeting both PINK1 and Parkin-dependent mechanisms.