Studies Of Proteomics Of Adenosine A (1) Receptor Agonist-Induced Delayed Myocardial Protection In Rabbits

Objectives: (1) To investigate whether the adenosine A1 receptoragonist 2-chloro-N~6-cyclopentyladenosine(CCPA)could induce the latephase of preconditioning against myocardial ischemia-reperfusion(I/R)injury in rabbits. (2) To investigate the changes of myocardial proteinexpression profiles 24h after CCPA pretreatment, and to search for theproteins probably involved in the late phase of preconditioning inducedby CCPA with proteomics techniques. (3) To identified some differentprotents with western blotting according the above results.Methods: The experiment consisted of three parts: 1.Rabbits weredivided into four groups including group CCPA,group NS,group I/R andgroup sham.CCPA was infused into rabbits in group CCPA,while thesame volume of normal saline(NS)was given those in group NS on day 1.Any treatment was not done in rabbits in groupI/R and sham. 24 hoursafter CCPA or NS prtreatment,rabbits in group CCPA,NS and I/R weresubjected to 30 min left anterior descending coronary artery(LAD)occlusions and were reperfused for 2h. Rabbits in group sham did notsubjecte LAD occulsion and reperfusion. Myocardial infarct size and theconcentrations of cTnI,CK,CK-MB were determined, the tissue and cellinjury of myocardium was examined with optical and electronmicroscope. 2.Proteomic analysis of myocardium of CCPA pretreatment. The left ventricle tissues of CCPA(group CCPA) orNS(group NS)preconditioned rabbits were sampled for proteomicanalysis. 3. According the above results,some protents were identifiedwith western blotting.Results: 1.The concentration of CK, CK-MB, cTn I and themyocardial infarct size were decreased significantly in group CCPA thanthose in group I/R and NS (P＜0.05). The tissue and cell injury ofmyocardium examined with optical and electron microscope in groupCCPA was decreased compared with the group I/R and NS. 2. Theresults revealed that 55 protein spots with significant differentiationexpression were screened. Those protein spots included 25overexpression spots and 16 spots down-regulation in group CCPAagainst group NS, 14 spots only expression in group CCPA. In this study,we selected 20 protein spots to further study by MALDI-TOF-MS andgot 17 peptide mass fingerprints. Furthermore, we identified 17 proteinsfrom those 17 peptide mass fingerprints by using Mascot and Expasybioinformatics software. Those proteins included stress protein (such asαB crystallin), metabolism-associated protein (D-aspartateO-methyltransferase) and signal transduction pathway-related proteins(phosphor- inositide-3-kinase) and so on. 3. we further detected theexpression ofαB crystallin (232 spot) by using immunobltting (WesternBlot). The results suggested thatαB crystallin was up-regulated by the 100μg/kg CCPA pretreatment.Conclusions: 1 Adenosine A1 receptor agonist 2-chloro-N~6-cyclopentyladenosine(CCPA)could induce the late phase ofpreconditioning in rabbits heart. 2. CCPA pretreatment 24h before canresult in the changes of myocardial protein expression profiles.Thedifferential proteins identified by MALDI-TOF-MS might be involved inthe delayed cardioprotection induced by CCPA. It would be helpful forclarifying the mechanisms of CCPA-mediated the second windowprotection against myocardial injury.