AIMS: Insulin/insulin-like growth factor-1 (IGF-1) signaling plays an important role in many biological processes. The class IA isoform of phosphoinositide 3-kinase (PI3K) is an important downstream effector of the insulin/IGF-1 signaling pathway. The aim of this study is to examine the effect of persistent activation of PI3K on gene expression and markers of cellular senescence in murine hearts. MAIN METHODS: Transgenic mice expressing a constitutively active PI3K in a heart-specific manner were analyzed at the ages of 3 and 20months. Effects of persistent activation of PI3K on gene expression were comprehensively analyzed using microarrays. KEY FINDINGS: Upon comprehensive gene expression profiling, the genes whose expression was increased included those for several heat shock chaperons. The amount and nuclear localization of a forkhead box O (FOXO) protein was increased. In addition, the gene expression of insulin receptor substrate-2 decreased, and that of phosphatase and tensin homolog deleted on chromosome ten (PTEN) increased, suggesting that the persistent activation of PI3K modified the expression of molecules of insulin/IGF-1 signaling. The expression of markers of cellular senescence, such as senescence-associated beta-galactosidase activity, cell cycle inhibitors, proinflammatory cytokines, and lipofuscin, did not differ between old wild-type and caPI3K mice. SIGNIFICANCE: The persistent activation of PI3K modified the expression of molecules of insulin/IGF-1 signaling pathway in a transgenic mouse line. Markers of cellular senescence were not changed in the aged mutant mice.

Since donor T-cells' allorecognition of host antigens is a prerequisite prerequisite for the onset of graft versus host disease (GVHD), blocking their cellular signaling pathways can decrease the severity of GVHD. We hypothesized that epigallocatechin-3-gallate (EGCG), due to its strong affinity to macromolecules, would adhere to surface molecules of donor T-cells, inhibit their allorecognition and attenuate GVHD in the recipient. We tested the hypothesis by treating donor splenocytes with EGCG in both in vitro and in vivo murine GVHD models. EGCG treatment decreased the proliferation of donor cells in MLR cultures and secretion of IL-2 and INF-γ. It also reduced the epitope detection of CD3ε, CD4 and CD28 but did not down-regulate the protein expression of these molecules, suggesting blockage of cell surface stimulatory signals. Similarly, EGCG treatment did not decrease mRNA expression for some of these molecules but decreased mitogen-induced cell proliferation, indicating that EGCG did not interfere the transcription of these genes but affected cell proliferation pathways. Furthermore, EGCG-treated donor splenocytes when transplanted into immunocompromized recipient mice decreased of proliferation, and the treatment extended the recipients' survival at least during the early stage of GVHD. These results strongly suggest that EGCG attenuates by both blocking specific cell surface molecules the donor T-cells' proliferation pathways.

OBJECTIVE To assess the feasibility of quantitative myocardial perfusion imaging (MPI) in acute myocardial infarction (AMI), using multi-row detector CT (MDCT) with a model-based deconvolution method. DESIGN, SETTING, PATIENTS AND INTERVENTIONS: Fifteen normal subjects with normal coronary arteries and 26 patients with AMI after reperfusion therapy underwent MPI with MDCT. Perfusion parameters: tissue blood flow (TBF), tissue blood volume (TBV) and mean transit time (MTT) were obtained and compared with clinical parameters, angiography and single-photon emission CT (SPECT) data. Furthermore, the MPI data were compared with data from myocardial magnetic resonance (MR) in 10 subjects. RESULTS The TBF and TBV of infarcted myocardium were significantly lower than those of non-infarcted areas (TBF, 51.96±19.42 vs 108.84±13.29 ml/100 g/min, p<0.01; TBV, 4.47±2.23 vs 9.79±2.58 ml/100 g, p<0.01). The MTT of infarcted areas did not differ from that of non-infarcted areas. The defect areas on TBV colour maps were significantly associated with peak creatine kinase level, QRS score and SPECT defect score. The ratio of TBF or TBV in the epicardial to endocardial side was significantly higher in infarct myocardium with good collateral circulation than in myocardium with poor/no collateral circulation (p<0.01 for both). The TBF measurements with CT- and MR-MPI were in good agreement by linear regression analysis (R=0.55, p<0.01). CONCLUSIONS This study demonstrated that MDCT perfusion imaging with deconvolution analysis could quantitatively detect myocardial perfusion abnormalities in patients with AMI and may provide the basis for the non-invasive and quantitative assessment of myocardial infarction.

Low efficiencies of gene targeting via homologous recombination (HR) have limited basic research and applications using human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs). Here, we show highly and equally efficient gene knockout and knock-in at both transcriptionally active (HPRT1, KU80, LIG1, LIG3) and inactive (HB9) loci in these cells using high-capacity helper-dependent adenoviral vectors (HDAdVs). Without the necessity of introducing artificial DNA double-strand breaks, 7-81% of drug-resistant colonies were gene-targeted by accurate HR, which were not accompanied with additional ectopic integrations. Even at the motor neuron-specific HB9 locus, the enhanced green fluorescent protein (EGFP) gene was accurately knocked in in 23-57% of drug-resistant colonies. In these clones, induced differentiation into the HB9-positive motor neuron correlated with EGFP expression. Furthermore, HDAdV infection had no detectable adverse effects on the undifferentiated state and pluripotency of hESCs and hiPSCs. These results suggest that HDAdV is one of the best methods for efficient and accurate gene targeting in hESCs and hiPSCs and might be especially useful for therapeutic applications.

Back-to-back hadron pair yields in d+Au and p+p collisions at √s(NN)=200 GeV were measured with the PHENIX detector at the Relativistic Heavy Ion Collider. Rapidity separated hadron pairs were detected with the trigger hadron at pseudorapidity |η|<0.35 and the associated hadron at forward rapidity (deuteron direction, 3.0<η<3.8). Pairs were also detected with both hadrons measured at forward rapidity; in this case, the yield of back-to-back hadron pairs in d+Au collisions with small impact parameters is observed to be suppressed by a factor of 10 relative to p+p collisions. The kinematics of these pairs is expected to probe partons in the Au nucleus with a low fraction x of the nucleon momenta, where the gluon densities rise sharply. The observed suppression as a function of nuclear thickness, p(T), and η points to cold nuclear matter effects arising at high parton densities.

We present measurements of J/ψ yields in d+Au collisions at sqrt[s(NN)]=200  GeV recorded by the PHENIX experiment and compare them with yields in p+p collisions at the same energy per nucleon-nucleon collision. The measurements cover a large kinematic range in J/ψ rapidity (-2.2<y<2.4) with high statistical precision and are compared with two theoretical models: one with nuclear shadowing combined with final state breakup and one with coherent gluon saturation effects. In order to remove model dependent systematic uncertainties we also compare the data to a simple geometric model. The forward rapidity data are inconsistent with nuclear modifications that are linear or exponential in the density weighted longitudinal thickness, such as those from the final state breakup of the bound state.

OBJECTIVES Diabetes is the clinical consequence of the loss of the majority of the β-cell population and failure to regenerate new pancreatic β cells. The current therapies based on β-cell replacement have failed to achieve β-cell renewal and thus, long-term insulin freedom. We have hypothesized that early rejection of endothelial elements within the islet grafts may seriously hamper islet regeneration in both native and islet grafts. METHODS In the present study, we analyzed the role of endothelial cells to activate pancreatic stem cells during islet regeneration. Mice were pretreated with or without endothelial pharmacological ablation of endothelial cells, followed by an acute β-cell injury using a single intraperitoneal injection of streptozotocin. We performed comparative morphometric analyses of recovered pancreata on days 3, 7, 10, and 30 after streptozotocin injury, staining with bromodeoxyuridine (BrdU) for representative cell types, β cells, endothelial elements, and stem cells. Blood glucose levels were measured continuously after the injury to monitor the capacity for metabolic control. RESULTS Morphometric analyses revealed an increasing number of cells over time to be stained with a stem cell and BrdU markers among animals only injured with streptozotocin but not with endothelial ablation. Notably, on day 10, stem cell markers were dramatically decrease nearly to basal levels, with appearance of numerous insulin-positive cells. Intact vessels with cobblestone-shaped endothelial elements were observed in direct proportion to the better outcomes, both by morphometric and by metabolic parameters. In contrast, fewer insulin-positive cells were observed in pancreata that had been ablated of endothelial cells showing extensive collapse of endocrine functions. CONCLUSIONS We observed that endothelial elements promoted stem cell proliferation and islet regeneration after a β-cell insult. We believe that preservation of endothelial cells positively affects the process of pancreatic regeneration.

Heart failure is associated with a change in cardiac energy metabolism. SIRT1 is a NAD(+)-dependent protein deacetylase, and important in the regulation of cellular energy metabolism. To examine the role of SIRT1 in cardiac energy metabolism, we created transgenic mice overexpressing SIRT1 in a cardiac-specific manner, and investigated cardiac functional reserve, energy reserve, substrate uptake, and markers of mitochondrial function. High overexpression of SIRT1 caused dilated cardiomyopathy. Moderate overexpression of SIRT1 impaired cardiac diastolic function, but did not cause heart failure. Fatty acid uptake was decreased and the number of degenerated mitochondria was increased dependent on SIRT1 gene dosage. Markers of reactive oxygen species were decreased. Changes in morphology and reactive oxygen species were associated with the reduced expression of genes related to mitochondrial function and autophagy. In addition, the respiration of isolated mitochondria was decreased. Cardiac function was normal in transgenic mice expressing a low level of SIRT1 at baseline, but the mice developed cardiac dysfunction upon pressure overload. In summary, the constitutive overexpression of SIRT1 reduced cardiac function associated with impaired mitochondria in mice.

BACKGROUND: Cachexia, namely body wasting, is a common complication in cases of congestive heart failure (CHF). Although, neurohumoral and immune abnormalities are associated with the condition, precisely how the imbalance of catabolism and anabolism is responsible for the wasting process is not known. METHODS: We analyzed markers of cachexia in Dahl salt-sensitive rats which show marked hypertension with preserved systolic function at 11weeks and CHF at 17-19weeks of age. We also analyzed the change in hepatic metabolism associated with CHF since liver plays a central role in the systemic regulation of catabolism and anabolism. RESULTS: In CHF rats, a failure to grow was observed and blood hepatic protein levels were decreased associated with increased blood proinflammatory cytokine levels, indicating that Dahl rats serve as a model of cardiac cachexia. Food intake was reduced, and blood sugar and insulin levels were decreased. Despite the apparent fasting condition, blood fatty acid levels were decreased and triglycerides levels were increased. In CHF rats, liver incorporated more glucose, the gene expression related to gluconeogenesis was decreased, the gene expression related to lipogenesis was increased, and the triglyceride content of the liver was increased. The paradoxical production of triglycerides synthesis in fasting rats was associated with a proinflammatory response in liver. CONCLUSIONS: The Dahl salt-sensitive rat can be used as a model of cardiac cachexia. The cachexia was associated with abnormal hepatic metabolism that might work as a maladaptive response during the progression of CHF.

BACKGROUND It remains unclear whether patients with chronic heart failure (CHF) and advanced left ventricular (LV) dysfunction on β-blocker therapy benefit from exercise training (ET). METHODS AND RESULTS We studied 45 CHF patients with advanced LV dysfunction [ejection fraction (LVEF)< 25%] and impaired exercise tolerance [normalized peak oxygen uptake (PVO₂)< 70%] receiving a β-blocker: 33 patients participated in a cardiac rehabilitation program with ET (ET group) and 12 did not (inactive control group). Exercise capacity, LV dimension and plasma B-type natriuretic peptide (BNP) were assessed before and after a 3-month study period. At baseline, both groups had markedly reduced LVEF (ET group 18 ± 4% vs. Control group 18 ± 5%, NS) and impaired exercise capacity (normalized PVO₂ 51 ± 10% vs. 55 ± 9%, NS). Although one patient in the ET group withdrew from the program due to worsening CHF, no serious cardiac events occurred during the ET sessions. After 3 months, the ET group (n = 24) had significantly improved PVO₂ by 16 ± 15%(1,005 ± 295 to 1,167 ± 397ml/min, P < 0.001), while the PVO₂ of the control group was unchanged. LV end-diastolic dimension decreased in both groups to a similar extent, but plasma BNP was significantly decreased only in the ET group (432 to 214 pg/ml, P < 0.05). CONCLUSIONS The data indicate that in CHF patients with advanced LV dysfunction on β-blocker therapy, ET successfully improves exercise capacity and BNP without adversely affecting LV remodeling or causing serious cardiac complications.