| Objective: To investigate the effect of erythropoietin(EPO)on myocardial injury after cardiac transplantation and its relationship with PKC pathway.Methods: The model of myocardial ischemia reperfusion injury via isogenic heart transplantation in rat was established,the experiment was divided into two batches(5 days and 10 days),each batch of male adult SD rats were random divided into 5 groups: sham operation group(group A);pure heart transplantation control group(group B);heart transplantation + EPO treatment group(C group);heart transplantation + EPO treatment +PKC inhibitor group(D group);heart transplantation + EPO treatment + PKC agonist group(E group).On 5th day after surgery of the first batche,10 th day after operation of the second batches,the rats were sacrificed and each specimen was stained by HE staining,to observe the tissue structure damage and cell infiltration.By immunohistochemical method the expression of EGFR protein of myocardial tissue was detected by western blot the phosphorylation of PKC and PLC-gamma 1,by special kit to detect MDA,ATP enzyme,GSH-Px and calcium ion level were detected respectively.Results: 1.The results of HE staining: At the 5th day,A group,myocardial structural was integrity,closely arranged,had no abnormal change;B group,the myocardial structure at fifth day had loosely arranged,inflammatory cell infiltration,tissue blood infiltration.the degree of myocardial cell structure at the 10 th day were more disorder osteoporosis,inflammatory infiltration and tissue infiltration of blood more significantly than obvious,and significantly structure damage;group C,at the 5th day a small amount of inflammatory cell infiltration,structure is complete,a small amount of tissue bleeding,at the 10 th day the myocardial tissue tends to be normal,compact structure,no significant cell infiltration;in group D,at the 5th day the structure is complete,myocardial cell gap increased slightly,there were obvious tissue bleeding,at the 10 th day of myocardial structure was observed in disorder,inflammatory cell infiltration;in group E,at the 5th daythe myocardial tissue was a small amount of inflammatory cell infiltration,the structure is complete,at 10 th day of myocardial structural integrity closely,no significant infiltration of inflammatory cells.2.The MDA level of myocardial tissue: At the5 th day,MDA level of A group was significantly lower than that of B,C,D,E group(P<0.05);B group were higher than that of C,D,E group(P<0.05);D group higher than that of C group(P<0.05),E group lower than that of C group(P<0.05).At the 10 th day,A group was lower than that of B and D group(P<0.05),A,C,E groups had no significant difference(P>0.05);C,E group lower than that of B group(P<0.05),no significant difference between B and D groups(P>0.05);D group was lower than that of C group(P<0.05);the 5 days and the 10 days of each corresponding group,in addition to A group(P>0.05)was no significant difference,compared to the other groups were significantly difference(P<0.05),and the result of each group of the 10 days is less than the 5 days groups.3.Comparison of GSH-Px results: At the 5th day,A group higher than that of B group(P<0.05),and lower than the C,D,E group(P<0.05);C group is higher than D but lower than E group(P<0.05);Each group of 10 days,the expression of GSH-Px was similar to groups of 5 days;The corresponding to each group of the two batches,no significant difference except the sham operation group(P>0.05),other groups were significantly different(P<0.05).4.The changes of ATP enzyme: The two batchs were similar,the A group was lower than that of C group and E group(P<0.05),but higher than that of B group(P<0.05),no significant difference between A group and D group(P>0.05);group C was higher than that of D group(P<0.05),but lower than that of group E(P<0.05).5.Immunohistochemical expression of EGFR comparison: The groups of 5 days and 10 days were similar,A group was lower than that of B,C,D and E group(P<0.05);B group was lower than that of C,D,E group(P<0.05);C,D,E group comparison,there was no significant difference(P>0.05);The corresponding to each group of the two batches,no significant difference except the sham operation group and simple transplantation group(P>0.05),other groups were significantly different(P<0.05).6.Western Blot to detect PLC-gamma 1 differential expression: the results of the two methods are consistent,the groups of 5 days and 10 days were similar,A group was lower than that of,B,C,D,E group(P<0.05);B group was lower than that of C,D,E group(P<0.05);C,D,E group compared no significant difference(P>0.05);The corresponding to each group of the two batches,no significant difference except the sham operation group(P>0.05),other groups were significantly different(P<0.05).7.Compare the level of phosphorylated PKC expression in myocardial tissue:(1)at the 5th day,A group was lower than that of C group and E group(P<0.01),higher than that of B group(P<0.05),compared with the D group had no significant difference(P>0.05);B group was significantly lower than that of C,D,E group(P<0.05);C group was higher than that of D group(P<0.05),and lower than that of E group(P<0.05);(2)at the 10 th day,A group was lower than that of C and E group(P<0.05),higher than that of D group(P<0.05),no significant difference compared with B group(P>0.05);B group was lower than that of C group and E group(P<0.05),no significant difference compared with with the D group(P>0.05);C group than that of D group(P<0.05),but lower than that of group E(P<0.05).8.Determination of calcium ion in myocardial tissue: The results of the each groups were no significant difference(P>0.05).Conclusion: EPO has obvious protective effects on myocardial ischemia reperfusion injury after cardiac transplantation,and PKC pathway may be involved in the regulation of EPO on myocardial protection.The activation of PKC pathway contributes to the protective effect. |
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