Omentin Prevents Myocardial Ischemic Injury Through AMP-Activated Protein Kinase- and Akt-Dependent Mechanisms

ObjectivesThis study examined the impact of omentin on myocardial injury in a mouse model of ischemia/reperfusion (I/R) and explored its underlying mechanisms.BackgroundObesity is a major risk factor for ischemic heart disease. Omentin is a circulating adipokine that is down-regulated by obesity.MethodsIn patients who underwent successful reperfusion treatment after acute myocardial infarction, cardiac function and perfusion defect were assessed by using scintigraphic images. Mice were subjected to myocardial ischemia followed by reperfusion.ResultsThis study found that high levels of plasma omentin were associated with improvement of heart damage and function after reperfusion therapy in patients with acute myocardial infarction. Systemic administration of human omentin to mice led to a reduction in myocardial infarct size and apoptosis after I/R, which was accompanied by enhanced phosphorylation of AMP-activated protein kinase (AMPK) and Akt in the ischemic heart. Fat-specific overexpression of human omentin also resulted in reduction of infarct size after I/R. Blockade of AMPK or Akt activity reversed omentin-induced inhibition of myocardial ischemic damage and apoptosis in mice. In cultured cardiomyocytes, omentin suppressed hypoxia/reoxygenation-induced apoptosis, which was blocked by inactivation of AMPK or Akt.ConclusionsOur data indicate that omentin functions as an adipokine that ameliorates acute ischemic injury in the heart by suppressing myocyte apoptosis through both AMPK- and Akt-dependent mechanisms

Echocardiography measurements in wild-type (WT) and cardiac-specific SOCS3-CKO mice during the chronic phase.

<p>Echocardiography was performed at pre-ischemia and from day 1 to day 28 after reperfusion (n = 8 per group). Pooled data for echocardiographic measurements in WT and SOCS3-CKO mice. *<i>P</i> < 0.05 vs. WT mice. FS indicates fractional shortening; LVESD, left ventricular end systolic dimension; LVEDD, left ventricular end diastolic dimension; IVST, interventricular septum thickness; PWT, posterior left ventricular wall thickness.</p

Early and sustained expression of SOCS3 protein during myocardial IRI.

<p>(<b>A</b>) Total cell lysates (TCL) were prepared from the left ventricle of wild-type (WT) and SOCS3-CKO mice at the indicated times after reperfusion and were immunoprecipitated with anti-SOCS3 antibodies. The immunoprecipitates and total cell lysates were subjected to western blotting and probed with antibodies against SOCS3 and GAPDH, respectively. The graphs represent the time course expression of SOCS3 protein in WT mice after reperfusion (left) and the quantitative difference between WT and SOCS3-CKO mice 3 h after reperfusion (right; n = 5 per group). *<i>P</i> < 0.05 vs. WT pre-ischemia. §<i>P</i> < 0.05 vs. WT 3 h after reperfusion. AU indicates arbitrary units; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; IRI, ischemia reperfusion injury. (<b>B</b>) Real-time PCR analysis for SOCS3 mRNA expression in mouse hearts pre-ischemia or 3 h after reperfusion (n = 5 for each group). *<i>P</i> < 0.05 vs. pre-ischemia.</p

Activation of cardioprotective signaling molecules in wild-type (WT) mice during myocardial IRI.

<p>Total cell lysates were prepared from the left ventricle of WT mice at the indicated times after reperfusion. Blots were probed using antibodies against tyrosine-phosphorylated STAT3 (pY-STAT3), STAT3, phosphorylated AKT (p-AKT), AKT, phosphorylated ERK1/2 (p-ERK1/2), and GAPDH. The graphs represent quantitative differences in expression between the ratios of pY-STAT3 to STAT, p-AKT to AKT, and p-ERK1/2 to ERK1/2 (n = 6 per group). *<i>P</i> < 0.05 vs. pre-ischemia, §<i>P</i> < 0.05 vs. 1 h (pY-STAT3 and p-AKT) or 0.3 h (p-ERK1/2) after reperfusion. AU, arbitrary units; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; IRI, ischemia reperfusion injury.</p

(A) Sustained activation of cardioprotective signaling molecules in SOCS3-CKO mice during IRI.

<p>Western blot analysis of total cell lysates prepared from the left ventricle of WT or SOCS3-CKO mice at the indicated times after reperfusion. The blots were probed using antibodies against tyrosine-phosphorylated STAT3 (pY-STAT3), STAT3, phosphorylated AKT (p-AKT), AKT, phosphorylated ERK1/2 (p-ERK1/2), and GAPDH. The graphs represent quantitative differences in the ratios of the expression of pY-STAT3 to STAT3, p-AKT to AKT, and p-ERK1/2 to ERK1/2 (n = 6 per group). *<i>P</i> < 0.05 vs. WT mice. AU, arbitrary units; IRI, ischemia reperfusion injury. (<b>B</b>) Immunostaining of tyrosine-phosphorylated STAT3 (pY-STAT3) in hearts pre-ischemia or 3 h after reperfusion (n = 5 for each group). Representative photographs of the hearts from each group are shown. Values are expressed as the percentage of pY-STAT3–positive cells in the hearts pre-ischemia or 6 h after reperfusion. *<i>P</i> < 0.05 vs. pre-ischemia. Scale bar = 100 μm. AU = arbitrary units.</p

Confirmation of myocardial-specific SOCS3 deletion in SOCS3-CKO mice.

<p>mRNA was prepared from the hearts and livers of wild-type (WT) mice 3 h after lipopolysaccharide (LPS) injection, and real-time PCR analyses for SOCS3 were performed. Values normalized to GAPDH are expressed (n = 5 per group). *<i>P</i> < 0.05 vs. WT pre-injection, §<i>P</i> < 0.05 vs. WT LPS injection.</p

Reduction of apoptosis and mitochondrial cytochrome c release in SOCS3-CKO mice during IRI.

<p>(<b>A</b>) Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assays of hearts from wild-type (WT) or SOCS3-CKO mice 6 h after reperfusion (n = 5 per group). Representative images of the ischemic areas of hearts from each group. The graph shows the percentage of TUNEL-positive cells. *<i>P</i> < 0.05 vs. WT sham, §<i>P</i> < 0.05 vs. WT IRI. (<b>B</b>) The cytosolic (Cyto) and mitochondrial (Mito) fractions were prepared from the left ventricles of WT or SOCS3-CKO mice 6 h after reperfusion and subjected to western blotting using an antibody against cytochrome c. The cytosolic marker α-tubulin and the mitochondrial marker VDAC served as internal controls for WT mice (n = 6 per group). The graph represents quantitative differences in cytochrome c expression. *<i>P</i> < 0.05 vs. WT sham, §<i>P</i> < 0.05 vs. WT IRI. AU, arbitrary units; IRI, ischemia reperfusion injury. (<b>C</b>) Total cell lysates were prepared from the left ventricles of wild-type (WT) or SOCS3-CKO mice pre-ischemia or 6 h after reperfusion, and western blots were probed with antibodies raised against cleaved caspase 8, cleaved caspase 3, and serine-phosphorylated STAT3 (pS-STAT3). The graphs represent quantitative differences in expression among cleaved caspase 8, cleaved caspase 3, and PS-STAT3 (n = 5 per group). §<i>P</i> < 0.05 vs. WT pre-ischemia, *<i>P</i> < 0.05 vs. WT 6 h after IRI. AU, arbitrary units; IRI, ischemia reperfusion injury.</p