The Effect And Mechanism Of Chronic Hypoxia On Hypothermic Cardioprotection During Cardiopulmonary Bypass

Background:Cardiac arrest is required to facilitate the surgical procedures during cardiac surgery,and therefore cardiopulmonary bypass(CPB)is necessary and myoeardial lsehemim-reperfusion injury is inevitable.In clinical practice,there is a group of patients suffering from chronic hypoxia(CH),such as patients with cyanotic congenital heart disease(CCHD),chronic obstructive pulmonary disease(COPD)and obstructive sleep apnea(OSA)as well as individuals who live at high altitudes.Accumulating evidence showed that these patients are at high risk for severe CPB-related myocardial injury following cardiac surgery.Objective:It is well known that low temperature confers cardioprotection by reducing metabolic activity and oxygen demands and hypothermia is a commonly used strategy to protect the heart against myocardial injury during CPB in open-heart surgery.However,hypothermia i5 associated with some disadvantages at the cellular level.To counter its negative effects,cold shock proteins are activated by hypothermia.However,the role of cold shock proteins in hypothermic cardioprotection is unknown and the effect of chronic hypoxia on cardioprotective effects of hypothermia has not been clearly elucidated.This study will investigate the underlying mechanism that links hypoxia with compromised hypothermic cardioprotection during CPB and explore its potential translational values.Methods:Using rat models of CH and animal models of CPB,as well as proteomic analysis,we sought to evaluate the effect of chronic hypoxia on hypothermic cardioprotection during CPB and identify the most relevant protein in this process.Furthermore,we used genetically modified rats and in vitro cell experiments to validate the central role of the identified protein and explore its upstream regulators and downstream effectors.Finally,we modified the formula of cardioplegic solution based on the identified target and observed if it could enhance the cardioprotective effects for chronically hypoxic myocardium.Results:We established a novel rat model to completely mimic CPB in clinical practice,in which we adopted ice-slush and cold cardioplegic solutions to protect the arrested heart.Then,we established a rat model of CH(10%O2;4 weeks)and subjected them to CPB.Compared to the normoxic control,the postoperative cardiac function decreased significantly in CH rats.Tunel staining showed that CH rats had more apoptotic cardiomyocytes than normoxic rats.Moreover,CH rats showed significantly increased level of oxidative stress and decreased level of ATP in cardiomyocytes during CPB.In addition,compared with normoxic rats,postoperative serum cardiac enzymes were significantly increased in CH rats.Cardiac proteome analysis detected a significantly decreased expression of Cold-inducible RNA-binding protein(CIRBP)in CH rats during CPB.By using bisulfite sequencing and chromatin immunoprecipitation followed by quantitative PCR(ChBP-qPCR),we found CH caused hypermethylation of the GC box at the Cirbp promoter region.The methylation inhibits SP1 binding to the GC box,resulting in CIRBP depression and its failure to respond to cold stress.We observed similar molecular changes in myocardial specimens from CCHD patients and high-altitude-dwelling patients.Furthermore,we developed Cirbp-knockout(Cirbp-/-)and Cirbp transgenic(Cirbp-Tg)rats and found hypothermic cardioprotection was attenuated in Cirbp-/-rats but enhanced in Cirbp-Tg rats,presenting as increased rates of cardiac apoptosis,higher concentration of myocardial enzyme and worse cardiac function after CPB in Cirbp-/-rats,and decreased rates of cardiac apoptosis,lower concentration of myocardial enzyme and better cardiac function after CPB in Cirbp-Tg rats.Proteomics analysis revealed that the cardiac ubiquinone(CoQ10)biosynthesis pathway was down-regulated during CPB in Cirbp-/-rats.Western blotting showed that biosynthesis monooxygenase COQ6(COQ6)and ubiquinone biosynthesis protein COQ9(COQ9)were significantly decreased during CPB in cardiac tissues harvested from Cirbp-/-rats.RNA pull-down and RNA immunoprecipitation(RNA-IP)analysis showed that CIRBP could bind to the mRNAs of Coq6 and Coq9.Moreover,luciferase assay and polysome profiling analysis showed that CIRBP could post-transcriptionally modulates the expression of COQ6 and COQ9 by facilitating their translations.Mass spectrum analysis showed that cardiac CoQ10 contents during CPB were significantly decreased in the Cirbp-/-rat.Consequently,its cardiac oxidative stress was aggravated and ATP production was impaired,leading to increased myocardial injury during CPB.Finally,after adding CoQ10 to cardioplegic solution,significant improvement of cardioprotection was achieved in CH rats,but its effect was limited in normoxic rats.Conclusions:hypothermia up-regulates Cirbp to mediate the hypothermic cardioprotection through promoting the biosynthesis of CoQ10 in the heart,but chronic hypoxia-induced hypermethylation in Cirbp promoter region affects its response to hypothermia,which attenuates cardioprotective effects of hypothermia during CPB.This finding suggests individualized cardioprotection strategies based on hypoxic states are necessary for CH patients undergoing open-heart surgery.