Myocyte Apoptosis And Progressive Cardiac Remodeling Following Coronary Microthrombosis In Rats

OBJECTIVESThe aim of this study was to develop a new model of coronary artery microthrombosis (CAM) in rats, to explore the mechanism of myocyte loss at acute and chronic phases following CAM, to evaluate the association between myocyte loss and cardiac dysfunction, and to investigate the effect of irbesartan(IB) on myocyte loss and cardiac remodeling. METHODS1 0mg/kg of sodium laurate (SL) was injected into aorta root during clamping the ascending aorta in order to create a model of CAM. Sixty-six rats were divided into five groups: acute CAM group(n=18), acute control group(n-18), chronic CAM group(n=10), chronic CAM plus IB intervention group(n=10), chronic control group(n=10). HE-staining was used to assess the endothelial damage and platlet-related microthrombus in the coronary microcirculation, and Nagar-Olsen's staining to evaluate myocardial ischemia in acute phase. A microscopy incorporated with an image analysis software (ImagePro-4) was employed to calculate the myocyte apoptosis rate in sections with TUNEL-staining or immunohischemical method for caspase-3, and to determine the myocyte nuclear density in sections with HE-staining. Western-blotting was conducted to detect the expression of activated caspase-3 in myocardial tissue, and serial echocardiography was performed to monitoring the alterations of cardiac function during chronic phase of CAM. RESULTS1. Acute Phase: One and a half hours after injection of SL, HE-staining sections revealed that endothelial damage (10.17 2.33%) and platelet-fibrin microthrombi (9.42 2.02%) formed in the coronary microcirculation in all animals of acute CAM group, without any changes in the control animals (p<0.01 each); Nagar-Olsen'sstaining showed that (7.20 2.43)% of myocytes experienced ischemia, without any alterations in the control group (p<0.01); Western blotting found that activated caspase-3 expression significantly increased in acute CAM group; Myocyte apoptosis rate significantly increased (TUNEL-staining, 2.19 0.67% vs 0.10 0.04%, p<0.01; Activated caspase-3 Immunohistochemistry, 2.85 0.67% vs 0.06 0.02%,p<0.01).2. Chronic Phase: Myocyte apoptosis rates in the groups of chronic CAM, chronic CAM plus IB intervention and chronic controls were 0.47 0.10%, 0.21 0.09% and 0.08 0.02%, respectively with TUNEL-staining (p<0.01 each); 0.49 0.14%, 0.24 0.11% and 0.04 0.02%, respectively with the activated caspase-3 Immunohistochemistry (p<0.01 each); Myocyte nuclear density was 2041 363, 2491 222 and 3067 254 nuclear/mm2, respectively (p<0.01 each); LVEDD was 9.25 0.59, 6.97 0.51 and 6.30 0.18mm, respectively (p<0.01 each); LVESD was 7.40 0.41, 4.69 0.74 and 3.49 0.22mm, respectively (p<0.01 each); EF was 38.7 3.4%, 60.1 8.9% and 74.8 3.5%, respectively (p<0.01 each).3. Cardiac function and myocyte loss: LVEDD was highly correlated to myocyte nuclear density(r = -0.793, p<0.01) and to myocyte apoptosis rate(r = 0.842, p<0.01); EF was highly correlated to the myocyte nuclear density(r = 0.816, p<0.01) and to myocyte apoptosis rate(r =- 0.797, p<0.01). CONCLUSIONS1. A new rat model of CAM is created by injection of sodium laurate.2. Myocyte apoptosis rate increases both in acute and chronic stages of CAM, which may be the main mechanism of the myocyte loss after CAM.3. Progressive myocyte apoptosis leads to myocyte loss and probably to progressive cardiac remodeling and dysfunction in chronic stages of CAM.4. IB may reduce myocyte loss and improve cardiac function after CAM.