Study On The Anti-apoptotic Effects Of Erythropoietin And Its Derivatives On Myocardium Of Epileptic Rats And The Mechanism

Carbamylated EPO(CEPO) is a derivative of erythropoietin(EPO) by subjecting it to carbamylation.Unlike EPO,it does not stimulate erythropoiesis,but effectively protects tissue from injury. A number of signaling pathways,such as Jak2/STAT5 signaling,PI3K signaling,protein kinase C and MAPK,have been implicated in EPO-induced cardioprotection. Since CEPO signals only through EPOR-βcR,the mechanism of CEPO-induced cardiorotection may be different from that of EPO. As an important signal transducing pathway,the Janus kinase/signal transducer and activator of transcription(JAK/STAT) pathway is activated by various cytokines and growth factors.The JAK/STAT pathway plays an important role in growth,proliferation,differentiation,inflammation and apoptosis of cells.More and more studies have shown that the JAK/STAT pathway have important role in pathogenesis of cardiac disease.EPO can activate the STAT family members STAT1, STAT3, and STAT5 rapidly. Although STAT1, STAT3,and STAT5 have been implicated in the regulation of apoptosis, only STAT5 so far has been demonstrated in the anti-apoptotic signaling of EPO.Epilepsy is a common chronic disease in nervous system,which is often accompanied with complications in course.In the complications,arrhythmia and cardiac arrest are the most harmful,many patients'sudden cardiac death has no reason.In this situation,more and more scholars are carrying out the research on brain-heart syndrome. Whether epilepsy can cause definite changes in cardiovascular function is unclear,and which way such changes is mediated in is also unclear.So the effort to identify such mechanism and to give effective intervention to cerebrocardiac syndrome will have significant meaning. The apoptosis of cardiomyocyte was detected with terminal deoxynucleotidyl transferase-mediated dUDP-nick end labeling (TUNEL) method. Immunohistochemistry was used to detect caspase-3 and bcl-xl. The mRNA expression of JAK2 and STAT5 was detected by in situ hybridization.The study applied epilepsy rat model.At time of 0h,2h,6h,12h,24h after epilepsy,rats were put to death and heart tissues were obtained.To esplore the following questions: First, What is the effect of epilepsy on cardiovascular activity?if epilepsy can increase the apoptosis of cardiac myocyte,lead to myocardial injury ?Second, if CEPO and EPO can decrease the apoptosis rate of cardiac myocyte, lessen myocardial injury with epilepsy ?Third, if CEPO and EPO can affect the expression of caspase3 and bcl-xl?Forth,what is the mechanism of cardioprotection ?And if cardioprotection by the activation of JAK2/STAT5 pathway ?The main results were as followed:1.The increase of blood pressure and heart rate in epilepsy-group is more significant than control group(P<0.05).2.Apoptosis of all rats at different time after epilepsy:There were positive cells in epilepsy 0h group by chance.After 2 hours apoptosis cells began increasing,and nucleus of TUNEL positive apoptosis cells were brown.Then they increased gradually,and at 24 hours it reach the peak(P<0.01).The rate of apoptosis reduced gradually in EPO and CEPO group,and the difference is much significant at 24 hours compared with epilepsy group(P<0.01). The rate of apoptosis increased in AG490 group, and the difference is significant at 24 hours compared with ethanol group(P<0.01).3.Caspase-3 bcl-xl protein expression:There was weakly positive expression of caspase-3 in cardiomyocyte of epilepsy 0h group. After 2 hours the expression began increasing.Then they increased gradually,and at 24 hours it reach the peak(P<0.01).The expression reduced gradually in EPO and CEPO group,and the difference is much significant at 24 hours compared with epilepsy group(P<0.01). The expression of caspase-3 increased in AG490 group, and the difference is significant at 24 hours compared with ethanol group(P<0.01). The expression of bcl-xl increased gradually in EPO and CEPO group,and the difference is much significant at 24 hours compared with epilepsy group(P<0.01). The expression of bcl-xl reduced in AG490 group, and the difference is significant at 24 hours compared with ethanol group(P<0.05).4.JAK2/STAT5 mRNA expression: There was weakly positive expression of JAK2/STAT5a mRNA in cardiomyocyte of epilepsy 0h group. After 2 hours the expression began increasing.Then they increased gradually,and at 24 hours it reach the peak(P<0.01).The expression increased gradually in EPO group,and the difference is much significant at 24 hours compared with epilepsy group(P<0.01,P<0.05).The expression of JAK2/STAT5a mRNA reduced in AG490 group, and the difference is significant at 24 hours compared with ethanol group(P<0.01,P<0.05). There were no significant difference in the JAK2/STAT5a mRNA expression of CEPO group compared with epilepsy group.The main conclusions were as followed:1.Epilepsy can alter cardiovascular activity acutely,lead to the increase of the apoptosis of cardiac myocyte. 2.The apoptotic rate increase with the prolongation of epileptic time.3.EPO and CEPO can affect the expression of caspase3 and bcl-xl,decrease the apoptotic rate of cardiac myocyte. EPO and CEPO can decrease the apoptosis induced by epilepsy with no significance.4.EPO can activate the pathway of JAK2/STAT5.The machanism of anti-apoptosis may be related to the pathway, down-regulating of the expression of caspase3, up-regulating of the expression of bcl-xl. CEPO have no effect on the pathway of JAK2/STAT5.The machanism of anti-apoptosis may be related to other pathway, down-regulating of the expression of caspase3, up-regulating of the expression of bcl-xl.Backing up all our datas,we found that EPO and CEPO can reduce the apoptosis rate of cardiac myocyte.They may be an effective drug in the therapy of epilepsy and other heart disease in the future.