| ObjectivesTo create a model of chronic hibernating myocardium (CHM) in rabbits, to explore the changes of myocyte and the interstitial collagen after CHM, and to determine whether these alterations affect the cardiac function, and to further observe the effects of losartan on ventricular remodeling.MethodsA left anterior descending (LAD) coronary artery stenosis was created and maintained for 4 weeks to create a chronic hibernating myocardium (CHM) model in rabbits. Thirty-six rabbits were assigned to the following three groups(12 rabbits per group): CHM for 4 weeks(CHM group), low-dose of losartan intervention group for 4 weeks(LT1 group, 10mg.kg-1.d-1), high-dose of losartan intervention group for 4 weeks (LT2 group, 30mg.kg-1.d-1); and 12 sham-operated rabbits served as controls (SO group).A microscopic imaging system (Image-Pro Plus, Olympus) was used to assess the Interstitial collagen volume fraction (ICVF) in myocardial sections with picrosirius-red staining, and a polarized light microscopy to quantitatively analysis the changes in the type and the proportion of collagen fibers, to semi-quantitatively score the proportion of collagen I to collagenⅢ(PI/III). The expression of MMP-2,9 and TIMP-2 was assessed by immunohistochemistry and western bloting. The myocyte apoptosis rate (Rapo) was calculated with TUNEL-staining. And echocardiography was performed to measure left ventricular end-systolic and end-diastolic diameter (LVESD, LVEDD), and left ventricular short-axis fraction shortening(LVFS) and ejection fraction(LVEF). Results1. Validation of CHM modelThe animal model of CHM was induced successfully in 12 rabbits of the CHM group. The average stenosis of LAD was 84.90%±4.24%. Echocardiography revealed 38 segments with abnormal wall motion of the left ventricle, 27 of which showed a biphasic response, i.e. an improvement followed by a deterioration with dobutamine dosage increasing during DSE. Compared with the measurements of sham group, LVEF and LVFS were greatly reduced in CHM group (P<0.01). Histological examination with HE-staining demonstrated disarrangement of myofibers, degeneration of cardiomyocytes, without necrosis in CHM group.2. Changes of interstitial collagen after CHM and effects of Losartan 28 days after operation: compared with the sham group, ICVF was significantly increased(P<0.01)in CHM group; compared with CHM group, ICVF was significantly decreased(P<0.01,each)in LT1 group and LT2 group ,and the change were more remarkable in LT2 group, compared to LT1 group(P<0.05).3. Changes of collagen fiber types after CHM and effects of Losartan 28 days after operation: compared with sham group, Picrosirius-red staining and polarization microscopy revealed a large number of collagen fibrils of type I that showed either red or yellow intense birefringence in the myocardial interstitium, with a small amount of typeⅢcollagen fibers characterized by green thin fibril with weak greenish birefringence, with a significant increase of PI/III (P < 0.01) in CHM group; compared with CHM group, the amount of type I collagen fibrils were markedly reduced with a significant decrease of PI/III (P<0.01,each) in the losartan intervention groups, and the change were more remarkable in LT2 group compared to LT1 group (P<0.05).4. Changes of MMPs and inhibiters after CHM and effects of Losartan 28 days after operation: compared with sham group, the expression of MMP-2 and MMP-9 were greatly increased(P<0.01)in CHM group, while TIMP-2 were greatly decreased(P< 0.01)compared with CHM group, the expression of MMP-2 and MMP-9 were significantly decreased(P<0.01,each), while TIMP-2 were significantly increased(P<0.01,each)in the losartan intervention groups, and the change were more remarkable in LT2 group than in LT1 group (P<0.05).5. Changes of myocyte apoptosis after CHM and effects of Losartan 28 days after operation: compared with sham group, myocyte Rapo was markedly increased (P<0.01)in CHM group; compared with CHM group, myocyte Rapo was significantly decreased(P<0.01,each)in the losartan intervention groups, and the change were more remarkable in LT2 group than in LT1 group (P<0.05).6. Changes of cardiac function after CHM and effects of Losartan 28 days after operation: compared with sham group, LVEF and LVFS were significantly reduced in CHM group (P <0.01), compared with CHM group, LVEF and LVFS were higher (P<0.01,each) in the losartan intervention groups ,and the changes were more remarkable in LT2 group than in LT1 group (P<0.05).Conclusions1. A rabbit model of CHM is successfully established through creating a left anterior descending coronary artery stenosis.2. CHM underwent interstitial collagen proliferation and myocyte apoptosis, leading to ventricular remodeling and ventricular functional impairment.3. Losartan intervention reduces myocyte apoptosis and interstitial collagen proliferation, and these beneficial effects ultimately translate to improve ventricular function. |
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