랫드 심근 허혈/재관류 손상 동물모델에서 좌심실 이완기능 장애 완화에 대한 메트포르민 또는 사파논A의 심장보호 효과 = Cardioprotective effects of metformin or sappanone A on alleviating left ventricular diastolic dysfucntion in a rat myocardial I/R injury model

다국어 초록 (Multilingual Abstract)

The increased incidence of myocardial infarction (MI) has emerged globally as a leading cause of cardiovascular morbidity and mortality. In the development of new drugs, the evaluation of cardiac function is important factor in the withdrawal of drug candidates. In addition, as life expectancy increases, the prevalence of metabolic syndrome is gradually increasing, and metabolic syndromes such as diabetes, abdominal obesity, dyslipidemia, and hypertension are the main causes of cardiovascular disease. The aim of this study was to establish a rat myocardial ischemia and reperfusion (I/R) injury model similar to human acute MI and investigate cardiac function commonly referred to as left ventricular (LV) function including LV systolic and diastolic function using echocardiographic evaluation.
Male Sprague-Dawley rats, 8-9-week-old, were ligated and reperfused the left anterior descending coronary artery (LAD) for 30 minutes under ventilation. Echocardiography was performed under anesthesia after the induction of MI according to the experimental plan. Among the echocardiographic indices, EF and FS represent the left ventricular systolic function in tree-dimensional and two-dimensional meanings respectively. To assess the left ventricular diastolic function, the values of peak velocity of the transmitral flow at early filling (E), the early diastolic mitral annular velocity (E’) at the medial mitral annulus, and E/E’ ratio were measured. In the preliminary study, the EF and FS values of rats with MI were significantly decreased compared with those of the sham group at 2 hours and on day 1 (P<0.01). However, the significantly decreased E’ and increased E/E’ values of MI group were shown compared with those of sham group during the entire experimental period. These results indicate that, unlike left ventricular systolic function, left ventricular diastolic function does not recover at the early phase of rat myocardial I/R injury model. Moreover, this acute myocardial infarction animal model and echocardiographic assessment of cardiac function show the importance of the left ventricular diastolic function, which is currently highlighted in patients with heart failure in human. In this study, in a rat myocardial I/R injury model, we can evaluate the efficacy study of candidates for new drug development especially using echocardiographic evaluation of left ventricular diastolic function, and applied metformin and sappanone A.
Metformin, dimethyl biguanide as a first-line treatment for type Ⅱ diabetes, significantly improved left ventricular systolic function including EF and FS values on days 3 and 7 after MI surgery. In addition, metformin resulted in recovery of left ventricular diastolic function in the early phase of rat ischemic reperfusion injury in this model. It was also possible to confirm the cardioprotective effect of metformin in histopathological results, in line with echocardiographic results.
Sappanone A, isolated from heartwood Caesalpinia sappan, homoisoflavanone, has proven anti-inflammatory effects with cells and LPS-induced bone loss mouse model. Sapannone A administration significantly attenuated left ventricular systolic and diastolic dysfunction in a rat myocardial I/R injury model in the early phase of MI. Also, myocardial infarct size, serum cardiac marker assay, histopathological assessment of inflammatory cells infiltration of rat hearts, transcriptome analysis of rat left ventricles showed that 50 mg/kg sappanone A has a cardioprotective effect.
Therefore, this rat myocardial I/R injury model using multimodal assessment of cardiac function including echocardiographic evaluation of left ventricular systolic and diastolic function, serum cardiac marker, myocardial infarct size, histopathologic evaluation, and mRNA sequencing can be widely used in translational research and in the development of new heart failure-related drugs.

목차 (Table of Contents)

• Contents
• Abstract i
• Contents iii
• List of Tables vi
• List of Figures vii
• Abbreviations ix
• Introduction 1
• Chapter Ⅰ. Preliminary study 4
• 1. Materials and Methods 4
• 1.1. Animals 4
• 1.2. Study design 4
• 1.3. Rat myocardial ischemia/reperfusion injury model 5
• 1.4. Echocardiographic analysis 7
• 1.5. Statistical analysis 7
• 2. Results 9
• 2.1. Echocardiographic results 9
• 3. Discussion 12
• Chapter Ⅱ. Efficacy study of metformin 14
• 1. Materials and Methods 14
• 1.1. Animals 14
• 1.2. Study design 14
• 1.3. Rat myocardial ischemia/reperfusion injury model 16
• 1.4. Echocardiographic analysis 16
• 1.5. Myocardial infarct size 16
• 1.6. Histopathological analysis 17
• 1.7. mRNA sequencing and data analysis 18
• 1.8. Identification of differentially expressed genes (DEGs) 18
• 1.9. Statistical analysis 19
• 2. Results 20
• 2.1. Body weight and heart changes 20
• 2.2. Echocardiographic results 22
• 2.3. Myocardial infarct size 26
• 2.4. Histopathological results 28
• 2.5. mRNA sequencing and data analysis 33
• 2.6. Identification of differentially expressed genes (DEGs) 101
• 3. Discussion 103
• Chapter Ⅲ. Efficacy study of sappanone A 106
• 1. Materials and Methods 106
• 1.1. Animals 106
• 1.2. Study design 106
• 1.3. Rat myocardial ischemia/reperfusion injury model 108
• 1.4. Echocardiographic analysis 108
• 1.5. Myocardial infarct size 108
• 1.6. Serum chemistry of cardiac marker 109
• 1.7. Histopathological analysis 109
• 1.8. mRNA sequencing and data analysis 110
• 1.9. Identification of differentially expressed genes (DEGs) 110
• 1.10. RT-PCR 111
• 1.11. Statistical analysis 112
• 2. Results 113
• 2.1. Echocardiographic results 113
• 2.2. Myocardial infarct size 117
• 2.3. Serum chemistry of cardiac markers 119
• 2.4. Histopathological results 121
• 2.5. mRNA sequencing and data analysis 127
• 2.6. Identification of differentially expressed genes (DEGs) 137
• 2.7. RT-PCR analysis 139
• 3. Discussion 143
• References 146
• Summary 157
• 국문 요약 (Summary in Korean) 159