| Objective:To study the protection of noninvasive delayed limb ischemic preconditioning (NDLIP) against myocardial ischemia reperfusion (I/R) injury in streptozotocin (STZ)-induced diabetic rats.Method:Healthy male SD rats were administered once intravenous injection of2%STZ solution at a dose of50mg/kg to induce acute diabetes mellitus.Then the diabetic rats were assigned randomly into four groups after4weeks feeding.(1) Sham group (n=16):rats were threaded a silk suture under the left coronary artery anterior descending (LAD) and laid up throughout the experiment.(2) I/R group (n=24):rats were subjected to30min LAD occlusion followed by120min reperfusion.(3) early myocardial ischemic preconditioning (EMIP) group (n=24):rats were subjected to three episodes of5min LAD occlusion followed by5min reperfusion before30min ischemia and120min reperfusion.(4) NDLIP group (n=24) rats were subjected to three episodes of5min ischemia of left hind limb followed by5min reperfusion daily for three consecutive days to induced NDLIP. On the forth day,30min ischemia followed by120min reperfusion of LAD was performed. Delayed protection of NDLIP were evaluated according to the changes of ECG. injury and necrosis of myocardial cell, mitochondria apoptosis pathway, blood vessel endothelial function. Changes of arrhythmia incidence during LAD30min/120reperfusion were monitored. Myocardial infarct size was determined based on2,3,5-triphenyltetrazodium chloride staining. Activity of creatine kinase-MB type isoenzyme (CK-MB) in serum was determined by immunological inhibition method. Myocardial apoptosis was detected using the TUNEL method. Expression of mitochondria apoptosis-associated protein Bcl-2, Bax, Caspase-3was measured by Western blotting technique. Serum concentrations of cardiac troponin Ⅰ (cTnI), endothelin-1(ET-1), nitric oxide (NO) were detected by ELISA method.Results:1. Effects of NDLIP on myocardial cell injury and necrosis induced by myocardial I/R injuryCompared with I/R group, the incidence of ventricular premature contraction (VPC) were decreased50%and37.5%vs100%(P<0.01), ventricular tachycardia (VT) decreased43.75%(P<0.01) and25%(P<0.05) vs62.5%, ventricular fibrillation (VF) decreased25%(P<0.01) and18.75%(P<0.05) vs25%in the EMIP group and NDLIP group during the ischemia period, respectively; the amplitude of increase in the activity of CK-MB and serum content of cTnI were reduced (CK-MB:2657±501U/L and3607±808U/L vs6420±714U/L, P<0.01; cTnl:1.26±0.17ng/L vs2.13±0.22ng/L, P<0.01,1.90±0.12ng/L vs2.13±0.22ng/L, no statistical difference, respectively); myocardial infarct size was diminished (IS/AAR%:18.88±6.96%and27.47±4.86%vs48.06±6.26%, respectively, P<0.01) in the EMIP group and NDLIP group at the end of reperfusion.2. Effects of NDLIP on mitochondria apoptosis induced by myocardial I/R injuryCompared with I/R group, apoptosis of cadiocytes was decreased (TUNEL:33.60±6.52%vs51.39±9.85%, P<0.05,40.96±9.18%vs51.39±9.85%, no statistical difference, respectively); expression of apoptosis-associated protein Bcl-2was upregulated (0.81±0.36and0.75±0.33vs0.70±0.28, no statistical difference), Bax and Caspase-3were downregulated (Bax:1.62±0.56vs1.88±0.52, P<0.05,1.77±0.67vs1.88±0.52, no statistical difference; Caspase-3:0.88±0.29vs1.28±0.30, P<0.05,1.02±0.11vs1.28±0.30no statistical difference, respectively) in EMIP and NDLIP group.3. Effects of NDLIP on blood vessel endothelial function before and after myocardial I/R injuryBefore30min ischemia, there is no change of serum concentration of ET-1, NO in EMIP and NDLIP groups compared with I/R group. After120min reperfusion the increase of content of ET-1was reduced (122.33±11.59ng/ml vs166.73±24.12ng/ml, P<0.01,157.31±12.04ng/ml vs166.73±24.12ng/ml, no statistical difference, respectively); the concentration of NO was preserved (8.49±1.19μmol/L vs5.37±1.20μmol/L, P<0.01,6.39±1.08μmol/L vs5.37±1.20μmol/L, no statistical difference); the increase in ET-1/NO was diminished (14.51±1.67and24.87±2.31vs32.64±9.48,P<0.01and P<0.05,respectively).Conclusion:1. NDLIP decreased the incidence of severe ventricular arrhythmia during ischemia and the release of CK-MB, diminished the infarct size, but no work in the release of cTnI. The delayed protection at anti-injury and necrosis of NDLIP was inferior to EMIP.2. There is no significant delayed protection at decreasing apoptotic index, upregulating the expression of Bcl-2, downregulating the expression of Bax and Caspase-3in NDLIP group. This could be associated with the prolonged duration of DM.3. NDLIP failed to increase the basic serum concentration of NO, while there is no significant delayed protection at reliefing the endothelia function, preserving of NO, declining the serum elevation of ET-1. This could be associated with the prolonged duration of DM. |
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