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The cellular response to stress involves the recruitment and coordination of molecular signaling pathways that prevent cell death. D-dopachrome tautomerase (DDT) is an enzyme that lacks physiologic substrates in mammalian cells, but shares partial sequence and structural homology with macrophage migration inhibitory factor (MIF). Here, we observed that DDT is highly expressed in murine cardiomyocytes and secreted by the heart after ischemic stress. Antibody-dependent neutralization of secreted DDT exacerbated both ischemia-induced cardiac contractile dysfunction and necrosis. We generated cardiomyocyte-specific DDT knockout mice (Myh6-Cre [Ddtf.sup.fl/fl]), which demonstrated normal baseline cardiac size and function, but had an impaired physiologic response to ischemia-reperfusion. Hearts from Myh6-Cre [Ddtf.sup.fl/fl] mice exhibited more necrosis and LV contractile dysfunction than control hearts after coronary artery ligation and reperfusion. Furthermore, treatment with DDT protected isolated hearts against injury and contractile dysfunction after ischemia-reperfusion. The protective effect of DDT required activation of the metabolic stress enzyme AMP-activated protein kinase (AMPK), which was mediated by a CD74/CaMKK2-dependent mechanism. Together, our data indicate that cardiomyocyte secretion of DDT has important autocrine/paracrine effects during ischemia-reperfusion that protect the heart against injury., Introduction Myocardial ischemia resulting from coronary arterial occlusion leads to myocardial necrosis, arrhythmias, contractile dysfunction, and heart failure. Cellular adaptation to ischemic stress requires the coordination of cellular metabolic pathways [...]

Myocardial hypertrophy is an adaptation to increased hemodynamic demands. An increase in heart tissue must be matched by a corresponding expansion of the coronary vasculature to maintain and adequate supply [...]

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine that also modulates physiologic cell signaling pathways. MIF is expressed in cardiomyocytes and limits cardiac injury by enhancing AMPK activity during ischemia. Reperfusion injury is mediated in part by activation of the stress kinase JNK, but whether MIF modulates JNK in this setting is unknown. We examined the role of MIF in regulating JNK activation and cardiac injury during experimental ischemia/reperfusion in mouse hearts. Isolated perfused [Mif.sup.[-/-]] hearts had greater contractile dysfunction, necrosis, and JNK activation than WT hearts, with increased upstream MAPK kinase 4 phosphorylation, following ischemia/reperfusion. These effects were reversed if recombinant MIF was present during reperfusion, indicating that MIF deficiency during reperfusion exacerbated injury. Activated JNK acts in a proapoptotic manner by regulating BCL2-associated agonist of cell death (BAD) phosphorylation, and this effect was accentuated in [Mif.sup.[-/-]] hearts after ischemia/reperfusion. Similar detrimental effects of MIF deficiency were observed in vivo following coronary occlusion and reperfusion in [Mif.sup.[-/-]] mice. Importantly, excess JNK activation also was observed after hypoxia-reoxygenation in human fibroblasts homozygous for the MIF allele with the lowest level of promoter activity. These data indicate that endogenous MIF inhibits JNK pathway activation during reperfusion and protects the heart from injury. These findings have clinical implications for patients with the low-expression MIF allele., Introduction Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine that is released from immune cells (1) and is involved in inflammatory diseases, including rheumatoid arthritis (1), atherosclerosis (2), and [...]

A study was conducted to evaluate the benefits of coronary artery disease (CAD) screening in identifying those high-risk patients with type 2 diabetes and also whether it improved their outcomes. Results indicated that cardiac event rates were low in diabetic patients and also that CAD screening did not really impact the outcomes.

Understanding cellular response to environmental stress has broad implications for human disease. AMP-activated protein kinase (AMPK) orchestrates the regulation of energy-generating and-consuming pathways, and protects the heart against ischaemic injury [...]

Glucose uptake in the heart is mediated by specific glucose transporters (GLUTs) present on cardiomyocyte cell surface membranes. Metabolic stress and insulin both increase glucose transport by stimulating the translocation of glucose transporters from intracellular storage vesicles to the cell surface. Isolated perfused transgenic mouse hearts are commonly used to investigate the molecular regulation of heart metabolism; however, current methods to quantify cell surface glucose transporter content in intact mouse hearts are limited. Therefore, we developed a novel technique to directly assess the cell surface content of the cardiomyocyte glucose transporter GLUT4 in perfused mouse hearts, using a cell surface impermeant biotinylated his-glucose photolabeling reagent (bio-LC-ATB-BGPA). Bio-LC-ATB-BGPA was infused through the aorta and cross-linked to cell surface GLUTs. Bio-LCATB-BGPA-labeled GLUT4 was recovered from cardiac membranes by streptavidin isolation and quantified by immunoblotting. Bio-LCATB-BGPA-labeling of GLUT4 was saturable and competitively inhibited by D-glucose. Stimulation of glucose uptake by insulin in the perfused heart was associated with parallel increases in bio-LC-ATB-BGPA-labeling of cell surface GLUT4. Bio-LC-ATB-BGPA also labeled cell surface GLUT1 in the perfused heart. Thus, photolabeling provides a novel approach to assess cell surface glucose transporter content in the isolated perfused mouse heart and may prove useful to investigate the mechanisms through which insulin, ischemia, and other stimuli regulate glucose metabolism in the heart and other perfused organs. glucose transport; perfused mouse heart; photolabeling; glucose transporter 4 doi:10.1152/ajpendo.00170.2006

AMP-activated protein kinase (AMPK) plays a key role in modulating cellular metabolic processes. AMPK, a serine-threonine kinase, is a heterotrimeric complex of catalytic [alpha]-subunits and regulatory [beta]- and [gamma]-subunits with multiple isoforms. Mutations in the cardiac [[gamma].sub.2]-isoform have been associated with hypertrophic cardiomyopathy and pre-excitation syndromes. However, physiological regulation of AMPK complexes containing different subunit isoforms is not well defined and is important for an understanding of the function of this signaling pathway in the intact heart. We evaluated the kinase activity associated with heart AMPK complexes containing specific or- and y-subunit isoforms of AMPK in an in vivo rat model of regional ischemia. Left coronary artery occlusion activated the immunoprecipitated [[alpha].sub.1]-isoform (6-fold, P < 0.01) and [[alpha].sub.2]-isoform (9-fold, P < 0.01) in the ischemic left ventricle compared with sham controls. The degree of a-subunit activation depended on the extent of ischemia and paralleled echocardiographic contractile dysfunction. The regulatory [[gamma].sub.1]- and [[gamma].sub.2]-isoforms were expressed in the heart. The [[gamma].sub.1]- and [[gamma].sub.2]-isoforms coimmunoprecipitated with [[alpha].sub.1]- and [[alpha].sub.2]-isoforms in proportion to [alpha]-subunit content, [[gamma].sub.1]-Isoform immunocomplexes accounted for 70% of AMPK activity and AMPK phosphorylation ([Thr.sup.172]) in hearts. Ischemia similarly increased AMPK activity associated with the [[gamma].sub.1]- and [[gamma].sub.2]-isoform complexes threefold (P < 0.01 for each). Thus AMPK catalytic [[alpha].sub.1]-and [[alpha].sub.2]-isoforms are activated by regional ischemia in vivo in the heart, irrespective of the regulatory [[gamma].sub.1]- or [[gamma].sub.2]-isoforms to which they are complexed. Despite the pathophysiological importance of [[gamma].sub.2]-isoform mutations, [[gamma].sub.1]-isoform complexes account for most of the AMPK activity in the ischemic heart. adenosine monophosphate-activated protein kinase; ischemia; subunits; isoforms

Role of the nitric oxide pathway in AMPK-mediated glucose uptake and GLUT4 translocation in heart muscle. Am J Physiol Endocrinol Metab 287: E834-E841, 2004. First published July 20, 2004; doi:10.1152/ajpendo.00234.2004.--AMP-activated protein kinase (AMPK) is a serine-threonine kinase that regulates cellular metabolism and has an essential role in activating glucose transport during hypoxia and ischemia. The mechanisms responsible for AMPK stimulation of glucose transport are uncertain, but may involve interaction with other signaling pathways or direct effects on GLUT vesicular trafficking. One potential downstream mediator of AMPK signaling is the nitric oxide pathway. The aim of this study was to examine the extent to which AMPK mediates glucose transport through activation of the nitric oxide (NO)-signaling pathway in isolated heart muscles. Incubation with 1 mM 5-amino-4-imidazole-1-[beta]-carboxamide ribofuranoside (AICAR) activated AMPK (P < 0.01) and stimulated glucose uptake (P < 0.05) and translocation of the cardiomyocyte glucose transporter GLUT4 to the cell surface (P < 0.05). AICAR treatment increased phosphorylation of endothelial NO synthase (eNOS) ~1.8-fold (P < 0.05). eNOS, but not neuronal NOS, coimmunoprecipitated with both the [[alpha].sub.2] and [[alpha].sub.1] AMPK catalytic subunits in heart muscle. NO donors also increased glucose uptake and GLUT4 translocation (P < 0.05). Inhibition of NOS with [N.sup.[omega]]-nitro-L-arginine and [N.sup.[omega]]-methyl-L-arginine reduced AICAR-stimulated glucose uptake by 21 [+ or -] 3% (P < 0.05) and 25 [+ or -] 4% (P < 0.05), respectively. Inhibition of guanylate cyclase with ODQ and LY-83583 reduced AICAR-stimulated glucose uptake by 31 [+ or -] 4% (P < 0.05) and 22 [+ or -] 3% (P < 0.05), respectively, as well as GLUT4 translocation to the cell surface (P < 0.05). Taken together, these results indicate that activation of the NO-guanylate cyclase pathway contributes to, but is not the sole mediator of, AMPK stimulation of glucose uptake and GLUT4 translocation in heart muscle. nitric oxide synthase; AMP-activated protein kinase; glucose transporter