京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第21960号 / 医博第4502号 / 新制||医||1037(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 平井 豊博, 教授 松原 和夫, 教授 湊谷 謙司 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

京都大学 / 新制・論文博士 / 博士(医学) / 乙第13401号 / 論医博第2225号 / 新制||医||1051(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 長船 健二, 教授 渡邊 直樹, 教授 江藤 浩之 / 学位規則第4条第2項該当 / Doctor of Medical Science / Kyoto University / DFAM

Background: Histone deacetylases (HDACs) play a critical role in modulating myocardial protection and cardiomyocyte survivals. However, Specific HDAC isoforms in mediating myocardial ischemia/reperfusion injury remain currently unknown. We used cardiomyocyte-specific overexpression of active HDAC4 to determine the functional role of activated HDAC4 in regulating myocardial ischemia and reperfusion in isovolumetric perfused hearts. Methods: In this study, we created myocyte-specific active HDAC4 transgenic mice to examine the functional role of active HDAC4 in mediating myocardial I/R injury. Ventricular function was determined in the isovolumetric heart, and infarct size was determined using tetrazolium chloride staining. Results: Myocyte-specific overexpressing activated HDAC4 in mice promoted myocardial I/R injury, as indicated by the increases in infarct size and reduction of ventricular functional recovery following I/R injury. Notably, active HDAC4 overexpression led to an increase in LC-3 and active caspase 3 and decrease in SOD-1 in myocardium. Delivery of chemical HDAC inhibitor attenuated the detrimental effects of active HDAC4 on I/R injury, revealing the pivotal role of active HDAC4 in response to myocardial I/R injury. Conclusions: Taken together, these findings are the first to define that activated HDAC4 as a crucial regulator for myocardial ischemia and reperfusion injury.

No description available.

Background: Erythropoietin (EPO) can reduce myocardial ischemia/reperfusion (I/R) injury. However, the cellular mechanisms have not been elucidated entirely. The present study was to investigate whether transcription factor GATA-4 could be involved in EPO-induced cardioprotection when it was administered after ischemia, immediately before reperfusion. Methods and results: Male Balb/c mice treated with or without EPO were subjected to ischemia (45 min) followed by reperfusion (4 h). TTC staining showed that the infarct size in EPO-treated mice was significantly reduced compared with untreated I/R mice (P < 0.05). Echocardiography examination suggested that EPO administration significantly improved cardiac function following I/R. TUNEL assay indicated that EPO treatment decreased apoptosis. EPO administration also significantly increased the level of nuclear GATA-4 phosphorylation in the myocardium which was positively correlated with the reduction of myocardial infarction. In vitro hypoxia/re-oxygenation study showed that EPO treatment increased the levels of phospho-GATA-4 and decreased cardiomyocyte apoptosis. More significantly, blocking GATA-4 by transfection of a dominant-negative form of GATA-4 (dnGATA-4) abolished EPO-induced cardioprotective effects. Conclusion: EPO administration after ischemia, just before reperfusion induced cardioprotection and stimulated GATA-4 phosphorylation. Activation of GATA-4 may be one of the mechanisms by which EPO induced protection against myocardial I/R injury.

Inhibitors of apoptosis proteins (IAPs) are key intrinsic regulators of caspases-3 and -7. During ischemia, IAP-2 is upregulated dramatically, while the other IAPs show little or no change. To test whether IAP-2 prevents cardiac apoptosis and injury following ischemia/reperfusion, we generated a line of transgenic mice that carried a mouse IAP-2 transgene. High levels of mouse IAP-2 transcripts and 70 kDa IAP-2 were expressed in the hearts of transgenic mice, whereas IAP-1 and XIAP levels remained the same. Immunohistochemical studies revealed more intense staining of IAP-2 in the myocytes of transgenic mouse hearts. To assess the role of IAP-2 in I/R injury, the transgenic mice were subjected to ligation of the left descending anterior coronary artery ligation followed by reperfusion. The infarct sizes, expressed as the percentage of the area at risk, were significantly smaller in the transgenic mice than in the non-transgenic mice (30 ± 2% vs. 44 ± 2%, respectively, P < 0.05). This protection was accompanied by a decrease of the serum level of troponin I in the transgenic mice. IAP-2 transgenic hearts had significantly fewer TUNEL-positive cardiac cells, which indicated an attenuation of apoptosis. Our results demonstrate that overexpression of IAP-2 renders the heart more resistant to apoptosis and I/R injury.

To determine the mechanism by which AMP 579, an adenosine A1/A2 agonist, administered at reperfusion protects ischemic myocardium, buffer-perfused rabbit hearts were subjected to 30 min of global ischemia and 2 h of reperfusion. AMP 579 (500 nM) was included in the reperfusate for the first 70 min. Average left ventricular diastolic pressure during reperfusion in hearts receiving AMP 579 was lower than that in control hearts (17.9 ± 2.4 vs. 39.0 ± 6.5 mm Hg, p < 0.05), indicating attenuation of contracture. Left ventricular developed pressure and coronary flow during reperfusion were also significantly improved with AMP 579 treatment. AMP 579's anti-contracture effect was blocked by the adenosine A2-receptor antagonist 8-(3-chlorostyryl)caffeine (CSC), but not by the A1 antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). CSC, but not DPCPX, also blocked AMP 579's ability to preserve developed pressure and coronary flow in these hearts. AMP 579 significantly reduced infarction in isolated hearts subjected to regional ischemia. The anti-infarct effect again was abolished by CSC but not by DPCPX. Finally, we tested whether 5′-(N-ethylcarboxamido)adenosine (NECA), another A1/A2 agonist, also administered for the initial 70 min of reperfusion, could duplicate the anti-infarct effect of AMP 579. One-hundred-nanomolar NECA duplicated the protection, but neither 50 nM CGS21680, a selective A2 agonist, nor 100 μM adenosine was protective. Therefore, AMP 579 given at reperfusion reduces contracture and infarction. Anti-contracture and anti-infarct effects require the adenosine A2, but not the A1, receptor suggesting that prevention of contracture and tissue salvage are mechanistically related. Not all A2 agonists were able to duplicate the anti-infarct effect, suggesting something unique about AMP579.

Generation of free radicals upon reperfusion has been cited as one of the major causes of ischaemia/reperfusion injury. The following series of experiments was designed to study the effect of manganese superoxide dismutase (MnSOD) overexpression in transgenic mice on ischemia/reperfusion injury. A species of 1.4 kb human MnSOD mRNA was expressed, and a 325% increase in MnSOD activity was detected in the hearts of transgenic mice with no changes in the other antioxidant enzymes or heat shock proteins. Immunocytochemical study indicated an increased labeling of MnSOD mainly in the heart mitochondria of the transgenic mice. When these hearts were perfused as Langendorff preparations for 45 min after 35 min of global ischemia, the functional recovery of the hearts, expressed as heart rate x left ventricular developed pressure, was 52 ± 4% in the transgenic hearts as compared to 31 ± 4% in the non-transgenic hearts. This protection was accompanied by a significant decrease in lactate dehydrogenase release from the transgenic hearts. Overexpression of MnSOD limited the infarct size in vivo in a left coronary artery ligation model. Our results demonstrate that overexpression of MnSOD renders the heart more resistant to ischemia/reperfusion injury.

PURPOSE: Previous work suggests that Dihydroorotate dehydrogenase (DHODH) inhibition via teriflunomide (TERI) may provide protection in multiple disease models. To date, little is known about the effect of TERI on the heart. This study was performed to assess the potential effects of TERI on cardiac ischemia reperfusion injury. METHODS: Male and female rat hearts were subjected to global ischemia (25 min) and reperfusion (120 min) on a Langendorff apparatus. Hearts were given either DMSO (VEH) or teriflunomide (TERI) for 5 min prior to induction of ischemia and during the reperfusion period. Left ventricular pressure, ECG, coronary flow, and infarct size were determined using established methods. Mitochondrial respiration was assessed via respirometry. RESULTS: Perfusion of hearts with TERI led to no acute effects in any values measured across 500 pM-50 nM doses. However, following ischemia-reperfusion injury, we found that 50 nM TERI-treated hearts had an increase in myocardial infarction (p < 0.001). In 50 nM TERI-treated hearts, we also observed a marked increase in the severity of contracture (p < 0.001) at an earlier time-point (p = 0.004), as well as reductions in coronary flow (p = 0.037), left ventricular pressure development (p = 0.025), and the rate-pressure product (p = 0.008). No differences in mitochondrial respiration were observed with 50 nM TERI treatment (p = 0.24-0.87). CONCLUSION: This study suggests that treatment with TERI leads to more negative outcomes following cardiac ischemia reperfusion, and administration of TERI to at-risk populations should receive special considerations.

No description available.