P99Nrg1beta enhances glucose uptake in cardiomyocytes via mTOR, Src and Akt

Background: Neuregulin (Nrg)1β is a growth factor that activates PI3K/Akt and Src/FAK via the ErbB2/ErbB4receptors. Although it is currently in clinical trial to treat haert failure, itremains unclear which cellular mechanisms are responsible for its cardioprotective actions. Here we tested if Nrg1β regulates glucose uptake in cardiomyocytes and analyzed the underlying signaling mechanisms. Methods: Neonatal rat ventricular myocytes were treated with Nrg1β (10ng/ml) in combination with the mTOR inhibitors PP242 (2mM) and rapamycin (20ng/ml), the ErbB2 inhibitor lapatinib (1mM), the Src inhibitor PP2 (5mM), the Akt inhibitor VIII (20mM), or vehicle. Cells were pre-incubated for 30 min with the inhibitors and proteins extracted 30 min after the addition of Nrg1β for analysis by Western blot. Glucose uptake was assessed by measuring the incorporation of 3H-D-glucose for 30 min. ErbB2 or ErbB4 receptors were knocked down withsiRNAfor 48h before Nrg1β treatment. Results: Similar to IGF-I and Insulin, Nrg1β caused a 1.9 fold increase in 3H-D-glucose incorporation (P< 0.01).Nrg1β induced phosphorylation of mTOR (S2448), Akt (S308) and FAK (Y861), as well as of the mTORC1 targets 4E-BP1, p70-S6K1 and ULK and the mTORC2 target Akt (S473). Lapatinib, PP242 and Akt inhibitor VIII blocked the Nrg1β-induced Akt-, mTOR-, p70-S6K1-, ULK-, and 4E-BP1-phosphorylation, indicating that these effects require ErbB2 and are mediated by Akt and mTOR. However, only lapatinib and Akt inhibitor VIII fully blocked the Nrg1β-induced glucose uptake; PP242 partially blocked it and rapamycin did not block it at all. These results suggest that Akt is required for Nrg1β-induced glucose uptake, and that mTORC2-dependent Akt phosphorylation mediates, at least in part, this response. PP2 blocked phosphorylation of FAK as expected, and it also partially blocked phosphorylation of Akt (S473) and p70-S6K1. PP2 also decreased general glucose uptake (0.6-fold of Ctl, p<0.05) and Nrg1β-induced glucose uptake (1.06-fold of Ctl, p=ns). Knock-down of ErbB4 receptor alone was sufficient to decrease both mTORC1 and mTORC2 signaling, whereas knock-down of ErbB2 affected only the mTORC2 targets. Conclusions: Our results show that Nrg1β increases glucose uptake in cardiomyocytes via Akt. We also show that Nrg1β activates mTORC1 via ErbB4 and mTORC2 via the ErbB2/ErbB4 heterodimer. Our data also support the hypothesis that Src/FAK is upstream of mTORC2 and mediates the Nrg1β-induced phosphorylation of Akt and glucose uptak

Look Who’s Talking:Using creative, playful arts-based methods in research with young children

Young children are often ignored or marginalised in the drive to address children’s participation and their wider set of rights. This is the case generally in social research, as well as within the field of Arts-Based Education Research. This article contributes to the growing literature on young children’s involvement in arts-based research, by providing a reflective account of our learning and playful engagement with children using creative methods. This small pilot project forms part of a larger international project titled Look Who’s Talking: Eliciting the Voices of Children from Birth to Seven, led by Professor Kate Wall at the University of Strathclyde. Visiting one nursery in Scotland, we worked with approximately 30 children from 3 to 5 years old. Seeking to connect with their play-based nursery experiences, we invited children to participate in a range of arts-based activities including drawing, craft-making, sculpting, a themed ‘play basket’ with various props, puppetry and videography. In this article, we develop reflective, analytical stories of our successes and dilemmas in the project. We were keen to establish ways of working with children that centred their own creativity and play, shaped by the materials we provided but not directed by us. However, we struggled to balance our own agenda with the more open-ended methods we had used. We argue that an intergenerational approach to eliciting voice with young children – in which adults are not afraid to shape the agenda, but do so in responsive, gradual and sensitive ways – creates the potential for a more inclusive experience for children that also meets researcher needs

Disruption of a GATA4/Ankrd1 Signaling Axis in Cardiomyocytes Leads to Sarcomere Disarray: Implications for Anthracycline Cardiomyopathy

Doxorubicin (Adriamycin) is an effective anti-cancer drug, but its clinical usage is limited by a dose-dependent cardiotoxicity characterized by widespread sarcomere disarray and loss of myofilaments. Cardiac ankyrin repeat protein (CARP, ANKRD1) is a transcriptional regulatory protein that is extremely susceptible to doxorubicin; however, the mechanism(s) of doxorubicin-induced CARP depletion and its specific role in cardiomyocytes have not been completely defined. We report that doxorubicin treatment in cardiomyocytes resulted in inhibition of CARP transcription, depletion of CARP protein levels, inhibition of myofilament gene transcription, and marked sarcomere disarray. Knockdown of CARP with small interfering RNA (siRNA) similarly inhibited myofilament gene transcription and disrupted cardiomyocyte sarcomere structure. Adenoviral overexpression of CARP, however, was unable to rescue the doxorubicin-induced sarcomere disarray phenotype. Doxorubicin also induced depletion of the cardiac transcription factor GATA4 in cardiomyocytes. CARP expression is regulated in part by GATA4, prompting us to examine the relationship between GATA4 and CARP in cardiomyocytes. We show in co-transfection experiments that GATA4 operates upstream of CARP by activating the proximal CARP promoter. GATA4-siRNA knockdown in cardiomyocytes inhibited CARP expression and myofilament gene transcription, and induced extensive sarcomere disarray. Adenoviral overexpression of GATA4 (AdV-GATA4) in cardiomyocytes prior to doxorubicin exposure maintained GATA4 levels, modestly restored CARP levels, and attenuated sarcomere disarray. Interestingly, siRNA-mediated depletion of CARP completely abolished the Adv-GATA4 rescue of the doxorubicin-induced sarcomere phenotype. These data demonstrate co-dependent roles for GATA4 and CARP in regulating sarcomere gene expression and maintaining sarcomeric organization in cardiomyocytes in culture. The data further suggests that concurrent depletion of GATA4 and CARP in cardiomyocytes by doxorubicin contributes in large part to myofibrillar disarray and the overall pathophysiology of anthracycline cardiomyopathy

Neuregulin-1β promotes glucose uptake via PI3K/Akt in neonatal rat cardiomyocytes

Nrg1beta is critically involved in cardiac development and also maintains function of the adult heart. Studies conducted in animal models showed that it improves cardiac performance under a range of pathological conditions, which led to its introduction in clinical trials to treat heart failure. Recent work also implicated Nrg1beta in the regenerative potential of neonatal and adult hearts. The molecular mechanisms whereby Nrg1beta acts in cardiac cells are still poorly understood. In the present study, we analyzed the effects of Nrg1beta on glucose uptake in neonatal rat ventricular myocytes and investigated to what extent mTOR/Akt signaling pathways are implicated. We show that Nrg1beta enhances glucose uptake in cardiomyocytes as efficiently as IGF-I and insulin. Nrg1beta causes phosphorylation of ErbB2 and ErbB4 and rapidly induces the phosphorylation of FAK (Tyr(861)), Akt (Thr(308) and Ser(473)), and its effector AS160 (Thr(642)). Knockdown of ErbB2 or ErbB4 reduces Akt phosphorylation and blocks the glucose uptake. The Akt inhibitor VIII and the PI3K inhibitors LY-294002 and Byl-719 abolish Nrg1beta-induced phosphorylation and glucose uptake. Finally, specific mTORC2 inactivation after knockdown of rictor blocks the Nrg1beta-induced increases in Akt-p-Ser(473) but does not modify AS160-p-Thr(642) or the glucose uptake responses to Nrg1beta. In conclusion, our study demonstrates that Nrg1beta enhances glucose uptake in cardiomyocytes via ErbB2/ErbB4 heterodimers, PI3Kalpha, and Akt. Furthermore, although Nrg1beta activates mTORC2, the resulting Akt-Ser(473) phosphorylation is not essential for glucose uptake induction. These new insights into pathways whereby Nrg1beta regulates glucose uptake in cardiomyocytes may contribute to the understanding of its regenerative capacity and protective function in heart failure

Chromosome numbers for the Italian flora: 14

In this contribution, new chromosome data obtained on material collected in Italy are presented. It includes the first counts for four subspecies of the Italian endemic Centaurea aplolepa Moretti, i.e. C. aplolepa subsp. aplolepa, C. aplolepa subsp. bertolonii, C. aplolepa subsp. levantina, and C. aplolepa subsp. parvula. In addition, the first chromosome count for an Italian population of Silene canescens (Caryophyllaceae) is provided

Canonical correlation analysis for gene-based pleiotropy discovery.

Genome-wide association studies have identified a wealth of genetic variants involved in complex traits and multifactorial diseases. There is now considerable interest in testing variants for association with multiple phenotypes (pleiotropy) and for testing multiple variants for association with a single phenotype (gene-based association tests). Such approaches can increase statistical power by combining evidence for association over multiple phenotypes or genetic variants respectively. Canonical Correlation Analysis (CCA) measures the correlation between two sets of multidimensional variables, and thus offers the potential to combine these two approaches. To apply CCA, we must restrict the number of attributes relative to the number of samples. Hence we consider modules of genetic variation that can comprise a gene, a pathway or another biologically relevant grouping, and/or a set of phenotypes. In order to do this, we use an attribute selection strategy based on a binary genetic algorithm. Applied to a UK-based prospective cohort study of 4286 women (the British Women's Heart and Health Study), we find improved statistical power in the detection of previously reported genetic associations, and identify a number of novel pleiotropic associations between genetic variants and phenotypes. New discoveries include gene-based association of NSF with triglyceride levels and several genes (ACSM3, ERI2, IL18RAP, IL23RAP and NRG1) with left ventricular hypertrophy phenotypes. In multiple-phenotype analyses we find association of NRG1 with left ventricular hypertrophy phenotypes, fibrinogen and urea and pleiotropic relationships of F7 and F10 with Factor VII, Factor IX and cholesterol levels