The Effect Of EPO On The Cardiac Structure And Function Of Mouse After Myocardial Infarction

Backgroud:Myocardial Infarction(MI) is a kind of disease that myocardium is under ischemiahypoxia for long time and eventually lead to myocardial injury or death. Until 2020, the globe mortality rate of cardiovascular disease will increase by 50% and MI will be the main factor that lead to death and bring a heavy burdens on the global health care.With the application of various medical technology, the mortality rate of MI has been obviously declined. However, after the MI, a lot of ischemic or necrotic cardiomyocytes will be replaced by scar. The scar tissue which will affect the normal systolic and diastolic function and eventually lead to heart failure at last. Drug therapy, interventional therapy and coronary artery bypass graft can just restore regional myocardial blood supply, but cannot solve the problem of the loss of cardiomyocytes and decreased cardiac function after MI.Erythropoietin(EPO) is a kind of glycoprotein hormones secreted by kidney, which is widely used in the treatment of anemia clinically. The role of EPO in cardiovascular disease has been found that EPO has the role of anti-apoptosis, anti-inflammation, antioxidant, promoting stem cell migration, proliferation and differentiation and inducing angiogenesis. Therefore, it can be concluded that in some degree EPO can protect the damaged myocardium. However, systemic administration of EPO will bring many side effects such as fever, diarrhea, vomiting, asthma, feeling change and upper respiratory tract infection and even high blood pressure, epilepsy and thrombosis, etc. As a consequence, the route of EPO administration is essential.In summary, in order to observe the curative effect of EPO to MI, a new MI model was built and conducted EPO intramyocardial injection post MI 3 days. Changes of cardiac structure and function and apoptosis of cardiomyocytes were observed dynamically after MI 1, 2, 4 weeks. Objective:(1)Build the new method to isolate cardiomyocytes from the adult mouse heart.(2)Observe the dynamical effect of EPO on the cardiac structure and function and apoptosisin cardiomyocytes after MI. Methods:(1)The improvement isolation method of adult mice ventricular cardiomyocytes. The original surgical method was used to retrograde catheterization in vivo for the C57 adult male mouse; The heart was washed by Langendorff perfusion and after fully enzymatic digestion by enzyme solution. The heart was torn and blowed into the cell suspension. The cell suspension was planted on the cell culture plate.(2)A new mouse AMI model. After anesthesia, the mice was fixed in supine position, cut the left chest wall a micro oblique incision, blunt separate the muscle, squeeze the heart out of the chest; Ligature the left main coronary artery, reset the heart quickly after the apex turn gray, push air out of the chest, close muscle and skin by pouched suture. The AMI pseudo operation is through the operation line under the left main coronary artery without ligation.(3)EPO or PBS injection in myocardium. At the 3rd day of MI, open the chest of mouse, expose the heart, inject 20 ul EPO or PBS at the border zone; divide the mouse into 5 groups: sham, sham + EPO, sham + PBS, MI + EPO, MI + PBS. After 7, 14, 28 days, the heave of heart and body, the myocardial infract size, the collagen deposion and the dynamic change of cardiac function.(4)Observe the effect of EPO on the myocardial apoptosis after MI 7, 14, 28 days. Compare the myocardial apoptosis rate of the two groups by TUNEL staining. Results:(1)Isolation of AVCMS: Under microscrope, freshly isolated mouse ventricular myocytes are in long rod shapeand clear cell lines. Based on rapid in vivo aorta intubation, 6.1± 0.27×106 AVCMS were collected by each mouse heartand the survive rate is 58.4±10.92 % after the cell culture for 24 hours.(2)The effect of EPO on myocardial infarction: Compared with MI + PBS group, after 1, 2, 4 weeks, a higher survive rate, a smaller myocardial infract area and a lower degree of myocardial fibrosis existed in MI+EPO group. The cardiac funtion is in MI+EPO group better than that in the control group.(3)The effect of EPO on the myocardial apoptosis: Compared with MI + PBS group, at the 1st week post MI, the TUNEL staining showed that EPO can obviously decline the myocardial apoptosis rate. Conclusion:(1)The in vivo adult mouse cardiomyocytes isolation method is better than the method in vitro with a shorter time, more the number of cells, higher cells viability.(2)Erythropoietin can significantly improve the cardiac structure and function after myocardial infarction, and reduce the rate of cell apoptosis.