Tetsuro Marunouchi

Simultaneous loss of phospholipase Cδ1 and phospholipase Cδ3 causes cardiomyocyte apoptosis and cardiomyopathy.

Phospholipase C (PLC) is a key enzyme in phosphoinositide turnover. Among 13 PLC isozymes, PLCδ1 and PLCδ3 share high sequence homology and similar tissue distribution, and are expected to have functional redundancy in many tissues. We previously reported that the simultaneous loss of PLCδ1 and PLCδ3 caused embryonic lethality because of excessive apoptosis and impaired vascularization of the placenta. Prenatal death of PLCδ1/PLCδ3 double-knockout mice hampered our investigation of the roles of these genes in adult animals. Here, we generated PLCδ1/PLCδ3 double-knockout mice that expressed PLCδ1 in extra-embryonic tissues (cDKO mice) to escape embryonic lethality. The cDKO mice were born at the expected Mendelian ratio, which indicated that the simultaneous loss of PLCδ1 and PLCδ3 in the embryo proper did not impair embryonic development. However, half of the cDKO mice died prematurely. In addition, the surviving cDKO mice spontaneously showed cardiac abnormalities, such as increased heart weight/tibial length ratios, impaired cardiac function, cardiac fibrosis, dilation, and hypertrophy. Predating these abnormalities, excessive apoptosis of their cardiomyocytes was observed. In addition, siRNA-mediated simultaneous silencing of PLCδ1 and PLCδ3 increased apoptosis in differentiated-H9c2 cardiomyoblasts. Activation of Akt and protein kinase C (PKC) θ was impaired in the hearts of the cDKO mice. siRNA-mediated simultaneous silencing of PLCδ1 and PLCδ3 also decreased activated Akt and PKCθ in differentiated-H9c2 cardiomyoblasts. These results indicate that PLCδ1 and PLCδ3 are required for cardiomyocyte survival and normal cardiac function.

Cell Death in the Cardiac Myocyte

Loss of cardiac myocytes plays a critical role in the pathogenesis of cardiovascular disorders. A decrease in the number of cardiac myocytes in cardiac diseases results in sustained, irreversible contractile failure of myocardium. Therefore prevention of cardiac cell death is a potential therapeutic strategy for various heart diseases. It is well accepted that three types of phenomena such as apoptosis, necrosis, and autophagy may be involved in myocardial cell death. Apoptosis is a highly regulated process that is promoted via death receptor pathway in the plasma membrane or via mitochondrial pathway. Necrosis is induced via mitochondrial swelling, cell rupture, and subsequent inflammation. Autophagy is a cell survival mechanism that involves degradation and recycling of cytoplasmic components. As compared with the other two mechanisms, autophagy may mediate cell death under specific conditions. These three types of cell death in the myocardium are discussed in this article.

Protective effect of geranylgeranylacetone via enhanced induction of HSPB1 and HSPB8 in mitochondria of the failing heart following myocardial infarction in rats

The mechanisms underlying mitochondrial impairment in the failing heart are not yet clear. In a previous study, we found that the levels of small heat shock proteins (HSP) such as mitochondrial HSPB1 and HSPB8 in the failing heart following myocardial infarction were decreased. In the present study, to verify the hypothesis that mitochondrial dysfunction in the failing heart is associated with alterations in mitochondrial small heat shock proteins, we examined the effects of geranylgeranylacetone, a heat shock protein inducer, on the cardiac mitochondrial function after myocardial infarction. When hemodynamic parameters of rats with myocardial infarction were measured at the 8th (8W) week after coronary artery ligation (CAL), the 8W-CAL showed signs of chronic heart failure concomitant with a reduced mitochondrial oxygen consumption rate. HSPB1 and HSPB8 contents in the mitochondrial fraction prepared from the failing heart were decreased, suggesting that an attenuation of mitochondrial translocation of HSPB1 and HSPB8 had led to an impairment of mitochondrial energy-producing ability. Geranylgeranylacetone treatment from the 2nd to 8th week after myocardial infarction attenuated the reduction in mitochondrial HSPB1 and HSPB8 contents. Furthermore, the mitochondrial energy-producing ability and cardiac pump function were preserved by orally administered geranylgeranylacetone during the development of heart failure. These results suggest that the induction of small heat shock proteins in the infarcted heart by geranylgeranylacetone treatment contributed to the preservation of mitochondrial function, leading to an improvement of cardiac contractile function.

Effects of metabotropic glutamate mGlu5 receptor antagonist on tyrosine phosphorylation of NMDA receptor subunits and cell death in the hippocampus after brain ischemia in rats

Tyrosine phosphorylation of the N-methyl-D-aspartate (NMDA) receptor appears to be associated with the regulation of the receptors ion channel. This study focused on the effect of a metabotropic glutamate mGlu5 receptor antagonist on tyrosine phosphorylation of NMDA receptor subunits and cell death in the hippocampal CA1 region after transient global ischemia and sought to explore their mechanisms. Pretreatment with the mGlu5 receptor antagonist reduced cell death in the hippocampal CA1 region on day 3 after the transient ischemia. Transient ischemia increased the tyrosine phosphorylation of NMDA receptor subunits, which are a major target of Src family tyrosine kinases. Therefore, we investigated the effect of the antagonist on tyrosine phosphorylation of the NMDA receptor subunits after transient ischemia. Tyrosine phosphorylation of the NR2A subunit, but not that of the NR2B one, was inhibited by the mGlu5 receptor antagonist. The administration of the antagonist also attenuated the increase in the amount of active form of Src after the reperfusion. We further demonstrated that the administration of a Src-family kinase inhibitor prevented cell death in the hippocampal CA1 region and attenuated the increase in the tyrosine phosphorylation of the NMDA receptor subunits after the reperfusion. These findings suggest that mGlu5 receptor in the hippocampal CA1 region after transient ischemia is involved in the activation of Src and subsequent tyrosine phosphorylation of NMDA receptor subunits, which actions may contribute to alterations of properties of the NMDA receptor and may be related to pathogenic events leading to neuronal cell death.

Effects of 2-Octynyladenosine (YT-146) on Mitochondrial Function in Ischemic/Reperfused Rat Hearts

This study investigated the effects of an adenosine receptor agonist, 2-octynyladenosine (YT-146), on mitochondrial function in ischemic and ischemic/reperfused hearts. Isolated rat hearts were perfused in the Langendorff manner with a constant flow rate, and exposed to 30 min of ischemia followed by 60 min of reperfusion. Preischemic treatment with YT-146 significantly improved postischemic recovery of left ventricular developed pressure. The high-energy phosphate content in reperfused hearts treated with YT-146 was also more greatly restored than in untreated hearts. YT-146 treatment attenuated the Na(+) content of a mitochondria-enriched fraction, but not the myocardial Na(+) content, at the end of ischemia. These results suggest that preischemic YT-146 treatment preserves the energy-producing ability of mitochondria during ischemia in the Na(+)-accumulated myocardium. YT-146 also attenuated both the sodium lactate-induced decrease in mitochondrial energy-producing ability and the increase in mitochondrial Na(+) concentration in the myocardial skinned fibers. YT-146 may attenuate Na(+) influx to myocardial mitochondria in ischemic cardiac cells, resulting in both preservation of the ability of mitochondria to produce energy and enhancement of the contractile recovery in reperfused hearts. Our findings suggest that the cardioprotective effects of YT-146 against ischemia/reperfusion injury are at least partially due to the preservation of mitochondrial function in the ischemic myocardium.

Effects of cardiosphere-derived cell transplantation on cardiac mitochondrial oxygen consumption after myocardial infarction in rats

BACKGROUND It is postulated that impaired mitochondrial energy-producing ability may lead to the development of chronic heart failure following an acute myocardial infarction. In this study, the effects of transplantation of cardiosphere-derived cells (CDCs) into the viable cardiac tissue after a myocardial infarction on the cardiac mitochondrial oxygen consumption rate (OCR) were examined. METHODS CDCs isolated from adult rat cardiac tissue fragments were cultured. Myocardial infarction was induced by ligation of the left ventricular coronary artery in rats. Immediately after coronary artery ligation (CAL), approximately 1 million CDCs were injected into the viable myocardium around the infarct area. RESULTS Eight weeks after CAL, animals without transplantation showed signs of heart failure such as impaired cardiac pump function. Furthermore, the mitochondrial OCR of the viable cardiac tissue in rats with heart failure was reduced. In contrast, the cardiac pump function and mitochondrial OCR were preserved without a reduction in the infarct size in the animals with transplantation of CDCs. CONCLUSIONS These results suggest that the transplantation of CDCs into the infarcted rat heart contributes to a preservation of mitochondrial function, leading to an improvement of cardiac contractile function without regeneration of cardiac tissues.