Tetrandrine Protects Hearts Against Ischemia‐Reperfusion Injury Via The Inhibition of p66Shc Phosphorylation And Mitochondrial Translocation

Abstract

Introduction Increased Mitochondrial (Mito) reactive oxygen species (ROS) generation is a major cause of cardiac ischemia/reperfusion (I/R) injury. p66 Shc is an adaptor protein that regulates multiple pathways governing Mito redox state and ROS generation. Tetrandrine (TET), a bisbenzylisoquinoline alkaloid that is extracted from the root of Stephania tetrandra S. Moore, has been used to treat cardiovascular diseases and hypertension. Here we investigate whether Tet exerts cardioprotection in I/R injury and its underlying mechanism mechanisms. Here we studied the effects of TET on phosphorylation and Mito translocation of p66 Shc, Mito oxidative stress/damage, cardiac function and infarct size in mouse heart model of I/R injury. Methods: Hearts (n = 14 each) isolated from C57BL/6 mice were perfused in the Langendorf mode. After 30‐min equilibration, the heart was treated with TET or vehicle for 10 min, then subjected to global, no‐flow normothermic ischemia for 30 min followed by reperfusion with the corresponding treatment for 120 min. Cardiac function and myocardial energy metabolism were measured with 31 P‐NMR spectroscopy continuously and simultaneously. Effluent was collected to detect the release of lactate dehydrogenase. At the end of the reperfusion, half of the hearts were stained with 2,3,5‐triphenyltetrazolium chloride to evaluate infarct size, whereas the other half were freeze‐clamped and whole protein lysate, cytosolic and Mito fractions were prepared from the clamped tissues to assess the phosphorylation at ser36 and Mito translocation of p66 SHC. Mito H2O2, malondialdehyde (MDA), and protein carbonyls were measured to assess ROS generation and oxidative damage to lipids and proteins, respectively. Results 3 μM TET significantly reduced the heart infarct size and improved cardiac functional recovery after I/R injury. Consistently, TET reversed I/R‐induced increases in H2O2, MDA, and protein carbonyls. Interestingly, 3 μM TET significantly decreased the I/R‐induced phosphorylation of p66 Shc at serine 36 and its translocation to mitochondria. Conclusions: Our data indicate that TET protects the heart against myocardial I/R injury by preventing Mito oxidative damage via mechanisms that are likely to suppressing p66 Shc phosphorylation and Mito accumulation. Targeting p66 Shc phosphorylation and translocation may serve as new therapeutic strategies for I/R injury.