Accumulation of endogenous adenosine improves cardiomyocyte metabolism via epigenetic reprogramming in an ischemia-reperfusion model.

Adenosine kinase (ADK) plays the major role in cardiac adenosine metabolism, so that inhibition of ADK increases myocardial adenosine levels. While the cardioprotective actions of extracellular adenosine against ischemia/reperfusion (I/R) are well-established, the role of cellular adenosine in protection against I/R remains unknown. Here we investigated the role of cellular adenosine in epigenetic regulation on cardiomyocyte gene expression, glucose metabolism and tolerance to I/R. Evans blue/TTC staining and echocardiography were used to assess the extent of I/R injury in mice. Glucose metabolism was evaluated by positron emission tomography and computed tomography (PET/CT). Methylated DNA immunoprecipitation (MeDIP) and bisulfite sequencing PCR (BSP) were used to evaluate DNA methylation. Lentiviral/adenovirus transduction was used to overexpress DNMT1, and the OSI-906 was administered to inhibit IGF-1. Cardiomyocyte-specific ADK/IGF-1-knockout mice were used for mechanistic experiments.Cardiomyocyte-specific ADK knockout enhanced glucose metabolism and ameliorated myocardial I/R injury in vivo. Mechanistically, ADK deletion caused cellular adenosine accumulation, decreased DNA methyltransferase 1 (DNMT1) expression and caused hypomethylation of multiple metabolic genes, including insulin growth factor 1 (IGF-1). DNMT1 overexpression abrogated these beneficial effects by enhancing apoptosis and decreasing IGF-1 expression. Inhibition of IGF-1 signaling with OSI-906 or genetic knocking down of IGF-1 also abrogated the cardioprotective effects of ADK knockout, revealing the therapeutic potential of increasing IGF-1 expression in attenuating myocardial I/R injury. In conclusion, the present study demonstrated that cardiomyocyte ADK deletion ameliorates myocardial I/R injury via epigenetic upregulation of IGF-1 expression via the cardiomyocyte adenosine/DNMT1/IGF-1 axis.

Mitochondrial aldehyde dehydrogenase-2 deficiency compromises therapeutic effect of ALDH bright cell on peripheral ischemia.

The autologous ALDH bright (ALDHbr) cell therapy for ischemic injury is clinically safe and effective, while the underlying mechanism remains elusive. Here, we demonstrated that the glycolysis dominant metabolism of ALDHbr cells is permissive to restore blood flow in an ischemic hind limb model compared with bone marrow mononuclear cells (BMNCs). PCR array analysis showed overtly elevated Aldh2 expression of ALDHbr cells following hypoxic challenge. Notably, ALDHbr cells therapy induced blood flow recovery in this model was reduced in case of ALDH2 deficiency. Moreover, significantly reduced glycolysis flux and increased reactive oxygen species (ROS) levels were detected in ALDHbr cell from Aldh2-/- mice. Compromised effect on blood flow recovery was also noticed post transplanting the human ALDHbr cell from ALDH2 deficient patients (GA or AA genotypes) in this ischemic hindlimb mice model. Taken together, our findings illustrate the indispensable role of ALDH2 in maintaining glycolysis dominant metabolism of ALDHbr cell and advocate that patient's Aldh2 genotype is a prerequisite for the efficacy of ALDHbr cell therapy for peripheral ischemia.

Regulation of p53 by jagged1 contributes to angiotensin II-induced impairment of myocardial angiogenesis.

Angiotensin II (AngII) is a major contributor to the development of heart failure, however, the molecular and cellular mechanisms still remain elucidative. Inadequate angiogenesis in myocardium leads to transition from cardiac hypertrophy to dysfunction, this study was therefore conducted to examine the effects of AngII on myocardial angiogenesis and the underlying mechanisms. AngII treatment significantly impaired angiogenetic responses, which were determined by counting the capillaries either in matrigel formed by cultured cardiac microvascular endothelial cells (CMVECs) or in myocardium of mice and by measuring the in vitro and in vivo production of VEGF proteins, and stimulated accumulation and phosphorylation of cytosolic p53 which led to increases in phosphorylated p53 and decreases of hypoxia inducible factor (Hif-1) in nucleus. All of these cellular and molecular events induced by AngII in CEMCs and hearts of mice were largely reduced by a p53 inhibitor, pifithrin-α (PFT-α). Interestingly, AngII stimulated the upregulation of Jagged1, a ligand of Notch, but it didn't affect the expression of Delta-like 4 (Dll-4), another ligand of Notch. Inhibition of p53 by PFT-α partly abolished this effect of AngII. Further experiments showed that knockdown ofJagged1 by addition of siRNA to cultured CMVECs dramatically declined AngII-stimulated accumulation and phosphorylation of p53 in cytosol, upregulation of phosphorylated p53 and downregulation of Hif-1 expression in nucleus, decrease of VEGF production and impairment of capillary-like tube formation by the cells. Our data collectively suggest that AngII impairs myocardial angiogenetic responses through p53-dependent downregulation of Hif-1 which is regulated by Jagged1/Notch1 signaling.

AngiotensinII preconditioning promotes angiogenesis in vitro via ERKs phosphorylation.

AngiotensinII (AngII) is involved in not only the formation of cardiac hypertrophy but also the development of cardiac remodeling both of which are associated with myocardial angiogenesis. This study was therefore performed to clarify the effects of AngII on the formation of vasculatures by cultured cardiac microvascular endothelial cells (CMVECs) after a long-period stimulation with or without the AngII preconditioning. Incubation with AngII for 18 hrs significantly impaired the formation of capillary-like tubes comparing to that without AngII. CMVECs with AngII pretreatment for 5 and 10 min formed more capillary-like tubes than those without AngII pretreatment, suggesting that preconditioning with AngII at a lower dose for a short period could prevent the further damage of CMVECs by a higher concentration of AngII. Moreover, AngII (10(-7) M) stimulation for 5 and 10 min significantly induced the increase in extracellular signal-regulated protein kinases (ERKs) phosphorylation, and an ERKs inhibitor, PD98059, abrogated the increase in the formation of capillary-like tubes induced by the AngII-pretreatment. In conclusion, preconditioning with a lower concentration of AngII for a short period prevents the subsequent impairment of CMVECs by a higher dose of AngII, at least in part, through the increase in ERKs phosphorylation.