STAT3α interacts with nuclear GSK3beta and cytoplasmic RISK pathway and stabilizes rhythm in the anoxic-reoxygenated embryonic heart

Activation of the Janus Kinase 2/Signal Transducer and Activator of Transcription 3 (JAK2/STAT3) pathway is known to play a key role in cardiogenesis and to afford cardioprotection against ischemia-reperfusion in adult. However, involvement of JAK2/STAT3 pathway and its interaction with other signaling pathways in developing heart transiently submitted to anoxia remains to be explored. Hearts isolated from 4-day-old chick embryos were submitted to anoxia (30min) and reoxygenation (80min) with or without the antioxidant MPG, the JAK2/STAT3 inhibitor AG490 or the PhosphoInositide-3-Kinase (PI3K)/Akt inhibitor LY-294002. Time course of phosphorylation of STAT3αtyrosine705 and Reperfusion Injury Salvage Kinase (RISK) proteins [PI3K, Akt, Glycogen Synthase Kinase 3beta (GSK3beta), Extracellular signal-Regulated Kinase 2 (ERK2)] was determined in homogenate and in enriched nuclear and cytoplasmic fractions of the ventricle. STAT3 DNA-binding was determined. The chrono-, dromo- and inotropic disturbances were also investigated by electrocardiogram and mechanical recordings. Phosphorylation of STAT3αtyr705 was increased by reoxygenation, reduced (~50%) by MPG or AG490 but not affected by LY-294002. STAT3 and GSK3beta were detected both in nuclear and cytoplasmic fractions while PI3K, Akt and ERK2 were restricted to cytoplasm. Reoxygenation led to nuclear accumulation of STAT3 but unexpectedly without DNA-binding. AG490 decreased the reoxygenation-induced phosphorylation of Akt and ERK2 and phosphorylation/inhibition of GSK3beta in the nucleus, exclusively. Inhibition of JAK2/STAT3 delayed recovery of atrial rate, worsened variability of cardiac cycle length and prolonged arrhythmias as compared to control hearts. Thus, besides its nuclear translocation without transcriptional activity, oxyradicals-activated STAT3α can rapidly interact with RISK proteins present in nucleus and cytoplasm, without dual interaction, and reduce the anoxia-reoxygenation-induced arrhythmias in the embryonic hear

Artificially elevated oxytocin concentrations in pet dogs are associated with higher proximity-maintenance and gazing towards the owners.

Abstract The relationship between dogs and their owners is characterized by an affective and enduring bond. It has been suggested that oxytocin might be the underlying mechanism driving this relationship, however evidence is mixed. In this study we tested whether intranasally administered oxytocin (compared to saline) would influence dogs' behavioural synchrony and shared attention towards their owners. Each individuals' pre and post administration oxytocin concentrations (measured in urine) were included in the analyses. Urinary oxytocin concentrations after administrations were positively associated with dogs' duration of social proximity and looking behaviours towards their owners supporting the role of oxytocin in modulating dogs' human-directed social behaviours

Transient anoxia and oxyradicals induce a region-specific activation of MAPKs in the embryonic heart

We have previously reported in the early septating embryonic heart that electromechanical disturbances induced by anoxia-reoxygenation are distinct in atria, ventricle, and outflow tract, and are attenuated in ventricle by opening of mitochondrial KATP (mitoKATP) channels. Here, we assessed the regional activation of mitogen-activated protein kinases (MAPKs) ERK, p38, and JNK in response to anoxia-reoxygenation and H2O2. Hearts isolated from 4-day-old chick embryos were subjected to 30-min anoxia and 60-min reoxygenation or exposed to H2O2 (50μM-1mM). The temporal pattern of activation of ERK, p38, and JNK in atria, ventricle, and outflow tract was determined using immunoblotting and/or kinase assay. The effect of the mitoKATP channel opener diazoxide (50μM) on JNK phosphorylation was also analyzed. Under basal conditions, total ERK and JNK were homogeneously distributed within the heart, whereas total p38 was the lowest in outflow tract. The phosphorylated/total form ratio of each MAPK was similar in all regions. Phosphorylation of ERK increased in atria and ventricle at the end of reoxygenation without change in outflow tract. Phosphorylation of p38 was augmented by anoxia in the three regions, and returned to basal level at the end of reoxygenation except in the outflow tract. JNK activity was not altered by anoxia-reoxygenation in atria and outflow tract. In ventricle, however, the diazoxide-inhibitable peak of JNK activity known to occur during reoxygenation was not accompanied by a change in phosphorylation level. H2O2 over 500μM impaired cardiac function, phosphorylated ERK in all the regions and p38 in atria and outflow tract, but did not affect JNK phosphorylation. At a critical stage of early cardiogenesis, anoxia, reoxygenation, exogenous H2O2 and opening of mitoKATP channels can subtly modulate ERK, p38, and JNK pathways in a region-specific manne

Audience effect on domestic dogs’ behavioural displays and facial expressions

In the present study we investigated the infuence of positive and negative arousal situations and the presence of an audience on dogs’ behavioural displays and facial expressions. We exposed dogs to positive anticipation, non-social frustration and social frustration evoking test sessions and measured pre and post-test salivary cortisol concentrations. Cortisol concentration did not increase during the tests and there was no diference in pre or post-test concentrations in the diferent test conditions, excluding a diferent level of arousal. Displacement behaviours of “looking away” and “snifng the environment” occurred more in the frustration-evoking situations compared to the positive anticipation and were correlated with cortisol concentrations. “Ears forward” occurred more in the positive anticipation condition compared to the frustration-evoking conditions, was positively infuenced by the presence of an audience, and negatively correlated to the pre-test cortisol concentrations, suggesting it may be a good indicator of dogs’ level of attention. “Ears fattener”, “blink”, “nose lick”, “tail wagging” and “whining” were associated with the presence of an audience but were not correlated to cortisol concentrations, suggesting a communicative component of these visual displays. These fndings are a frst step to systematically test which subtle cues could be considered communicative signals in domestic dogs

Improving Reconstituted HDL Composition for Efficient Post-Ischemic Reduction of Ischemia Reperfusion Injury

BACKGROUND: New evidence shows that high density lipoproteins (HDL) have protective effects beyond their role in reverse cholesterol transport. Reconstituted HDL (rHDL) offer an attractive means of clinically exploiting these novel effects including cardioprotection against ischemia reperfusion injury (IRI). However, basic rHDL composition is limited to apolipoprotein AI (apoAI) and phospholipids; addition of bioactive compound may enhance its beneficial effects. Objective The aim of this study was to investigate the role of rHDL in post-ischemic model, and to analyze the potential impact of sphingosine-1-phosphate (S1P) in rHDL formulations. Methods and RESULTS: The impact of HDL on IRI was investigated using complementary in vivo , ex vivo and in vitro IRI models. Acute post-ischemic treatment with native HDL significantly reduced infarct size and cell death in the ex vivo , isolated heart (Langendorff) model and the in vivo model (-48%, p<0.01). Treatment with rHDL of basic formulation (apoAI + phospholipids) had a non-significant impact on cell death in vitro and on the infarct size ex vivo and in vivo . In contrast, rHDL containing S1P had a highly significant, protective influence ex vivo , and in vivo (-50%, p<0.01). This impact was comparable with the effects observed with native HDL. Pro-survival signaling proteins, Akt, STAT3 and ERK1/2 were similarly activated by HDL and rHDL containing S1P both in vitro (isolated cardiomyocytes) and in vivo . CONCLUSION: HDL afford protection against IRI in a clinically relevant model (post-ischemia). rHDL is significantly protective if supplemented with S1P. The protective impact of HDL appears to target directly the cardiomyocyte

High density lipoprotein/sphingosine-1-phosphate-induced cardioprotection: Role of STAT3 as part of the SAFE pathway

High density lipoprotein (HDL) cholesterol has beneficial effects beyond its atheroprotective function in reverse cholesterol transport, including cardioprotection against ischemia reperfusion (IR) injuries. Two major constituents of HDL, namely the structural protein apolipoprotein AI (apoAI) and the sphingolipid sphingosine-1-phosphate (S1P) appear to contribute to this cardioprotective effect via the activation of intrinsic prosurvival signaling pathways that still remain to be clarified.   Recently, a powerful prosurvival signaling pathway, termed the survivor activating factor enhancement (SAFE) pathway, which involves the activation of signal transducer and activator of transcription 3 (STAT3) and tumor necrosis factor α (TNF), has been shown to protect against ischemia-reperfusion injuries. The present review summarizes the evidence for the roles of HDL and S1P in cardioprotection and discusses the signaling pathways that have been implicated. It thus provides support for our contention that S1P should be considered in potential formulations of reconstituted HDL (reHDL) that may be tested for cardioprotection against coronary artery disease via the activation of the SAFE pathway

Pharmacological Intervention to Modulate HDL: What Do We Target?

The cholesterol concentrations of low-density lipoprotein (LDL) and high-density lipoprotein (HDL) have traditionally served as risk factors for cardiovascular disease. As such, novel therapeutic interventions aiming to raise HDL cholesterol have been tested in the clinical setting. However, most trials led to a significant increase in HDL cholesterol with no improvement in cardiovascular events. The complexity of the HDL particle, which exerts multiple physiological functions and is comprised of a number of subclasses, has raised the question as to whether there should be more focus on HDL subclass and function rather than cholesterol quantity. We review current data regarding HDL subclasses and subclass-specific functionality and highlight how current lipid modifying drugs such as statins, cholesteryl ester transfer protein inhibitors, fibrates and niacin often increase cholesterol concentrations of specific HDL subclasses. In addition this review sets out arguments suggesting that the HDL3 subclass may provide better protective effects than HDL2

Behavioural Analysis of Dogs’ Response to Threatening and Neutral Conspecific Video Stimuli

Dogs’ displacement behaviours and some facial expressions have been suggested to function as appeasement signals, reducing the occurrences of aggressive interactions. The present study had the objectives of using naturalistic videos, including their auditory stimuli, to expose a population of dogs to a standardised conflict (threatening dog) and non-conflict (neutral dog) situation and to measure the occurrence of displacement behaviours and facial expressions under the two conditions. Video stimuli were recorded in an ecologically valid situation: two different female pet dogs barking at a stranger dog passing by (threatening behaviour) or panting for thermoregulation (neutral behaviour). Video stimuli were then paired either with their natural sound or an artificial one (pink noise) matching the auditory characteristics. Fifty-six dogs were exposed repeatedly to the threatening and neutral stimuli paired with the natural or artificial sound. Regardless of the paired auditory stimuli, dogs looked significantly more at the threatening than the neutral videos (χ2(56, 1) = 138.867, p < 0.001). They kept their ears forward more in the threatening condition whereas ears were rotated more in the neutral condition. Contrary to the hypotheses, displacement behaviours of sniffing, yawning, blinking, lip-wiping (the tongue wipes the lips from the mouth midpoint to the mouth corner), and nose-licking were expressed more in the neutral than the threatening condition. The dogs tested showed socially relevant cues, suggesting that the experimental paradigm is a promising method to study dogs’ intraspecific communication. Results suggest that displacement behaviours are not used as appeasement signals to interrupt an aggressive encounter but rather in potentially ambiguous contexts where the behaviour of the social partner is difficult to predict

Appeasement function of displacement behaviours? Dogs' behavioural displays exhibited towards threatening and neutral humans

Appeasement signals are behavioural patterns displaying an animal's non-aggressive attitude and are hypothesized to reduce the aggressive behaviours in the receiver. In domestic dogs, specific displacement behaviours (i.e., behavioural patterns exhibited without an apparent function related to the ongoing situation), have been suggested to function as appeasement signals. To test this possibility, we assessed whether the occurrence of these behaviours was dependent on a social conflict context, predicting that, if displacement behaviours also function as appeasement signals, they should be more prevalent in a conflict vs. non-conflict context. Fifty-three dogs were exposed to two unfamiliar humans approaching them in either a mildly threatening or neutral way. We categorized the attitude of the dogs towards the strangers as "reactive", i.e., barking and lunging towards the stimulus, and "non-reactive", i.e., remaining passive in front of the stimuli. We coded dogs' displacement activities and modelled their duration or frequency as a function of the interaction between the test condition and the attitude of the dog. Displacement behaviours of "blinking", "nose licking" and "lip wiping" were associated with a "non-reactive" attitude, independently from the test condition, confirming an association with a non-aggressive intention. "Head turning" was associated with a "non-reactive" attitude in the threatening condition. In conclusion, dogs with a non-aggressive attitude exhibited more putative appeasement signals; however, these were not strictly associated with a conflict-ridden situation, calling for further investigation of their function

HDL protects against myocardial ischemia reperfusion injury via miR-34b and miR-337 expression which requires STAT3

Purpose: High density lipoprotein (HDL) protects against myocardial infarction via mechanisms that remain unclear. STAT3 (signal transducer and activator of transcription 3) plays a key role in HDL-induced cardioprotection. In the heart, microRNAs (miRNAs) are involved in ischemia reperfusion injury. We therefore investigated whether the cardioprotective effect of HDL modulates miRNAs as a downstream target of STAT3 activation. Methods: STAT3 cardiomyocyte deficient mice (STAT3-KO) and wildtype littermates (STAT3-WT) were submitted to left coronary ligature and reperfused (IR) with or without injection of HDL. Infarct size (IS) was determined and cardiac miRNA expression was evaluated after reperfusion in sham, IR and IR+HDL hearts by microarray analysis. In vitro, neonatal rat ventricular cardiomyocytes were submitted to hypoxia with or without HDL incubation. Cell viability and miRNA expression were analysed. Results: In vivo, HDL reduced IS from 40.5±4.3% to 24.4±2.1% (p<0.05) in STAT3-WT mice. HDL failed to protect in STAT3-KO mice. In STAT3-WT mice, both miR-34b and miR-337 were increased in IR compared to sham and IR+HDL groups (p<0.05). These miRNAs were not modulated in STAT3-KO mice. In vitro, incubation with HDL improved cell viability against hypoxia (p<0.05). The expression of miR-34b and miR-337 was increased by hypoxia and reduced by HDL treatment (p<0.05). In cardiomyocytes transfected with miRNA mimics, HDL failed to improve cell viability against hypoxia. Conclusions: Our study, performed both in vivo and in vitro, delineates a novel cardioprotective signalling pathway activated by HDL, involving STAT3-mediated decrease of miR-34b and miR-337 expression