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The nonessential amino acid asparagine can only be synthesized de novo by the enzymatic activity of asparagine synthetase (ASNS). While ASNS and asparagine have been implicated in the response to numerous metabolic stressors in cultured cells, the in vivo relevance of this enzyme in stress-related pathways remains unexplored. Here, we found ASNS to be expressed in pericentral hepatocytes, a population of hepatic cells specialized in xenobiotic detoxification. ASNS expression was strongly enhanced in 2 models of acute liver injury: carbon tetrachloride (C[Cl.sub.4]) and acetaminophen. We found that mice with hepatocyte-specific Asns deletion were more prone to pericentral liver damage than their control littermates after toxin exposure. This phenotype could be reverted by i.v. administration of asparagine. Unexpectedly, the stress- induced upregulation of ASNS involved an ATF4-independent, noncanonical pathway mediated by the nuclear receptor, liver receptor homolog 1 (LRH-1; NR5A2). Altogether, our data indicate that the induction of the asparagine-producing enzyme ASNS acts as an adaptive mechanism to constrain the necrotic wave that follows toxin administration and provide proof of concept that i.v. delivery of asparagine can dampen hepatotoxin-induced pericentral hepatocellular death., Introduction The liver is a highly structured tissue where oxygen-rich blood enters the hepatic lobule at peripheral portal triads and drains out through the central vein (reviewed in ref. 1). [...]

The systemic expression of the bile acid (BA) sensor farnesoid X receptor (FXR) has led to promising new therapies targeting cholesterol metabolism, triglyceride production, hepatic steatosis and biliary cholestasis. In contrast to systemic therapy, bile acid release during a meal selectively activates intestinal FXR. By mimicking this tissue-selective effect, the gut-restricted FXR agonist fexaramine (Fex) robustly induces enteric fibroblast growth factor 15 (FGF15), leading to alterations in BA composition, but does so without activating FXR target genes in the liver. However, unlike systemic agonism, we find that Fex reduces diet-induced weight gain, body-wide inflammation and hepatic glucose production, while enhancing thermogenesis and browning of white adipose tissue (WAT). These pronounced metabolic improvements suggest tissue-restricted FXR activation as a new approach in the treatment of obesity and metabolic syndrome.

BackgroundPhosphoinositide 3-kinase γ (PI3Kγ) signaling engaged by β-adrenergic receptors is pivotal in the regulation of myocardial contractility and remodeling. However, the role of PI3Kγ in catecholamine-induced arrhythmia is currently unknown.Methods and resultsMice lacking PI3Kγ (PI3Kγ(-/-)) showed runs of premature ventricular contractions on adrenergic stimulation that could be rescued by a selective β(2)-adrenergic receptor blocker and developed sustained ventricular tachycardia after transverse aortic constriction. Consistently, fluorescence resonance energy transfer probes revealed abnormal cAMP accumulation after β(2)-adrenergic receptor activation in PI3Kγ(-/-) cardiomyocytes that depended on the loss of the scaffold but not of the catalytic activity of PI3Kγ. Downstream from β-adrenergic receptors, PI3Kγ was found to participate in multiprotein complexes linking protein kinase A to the activation of phosphodiesterase (PDE) 3A, PDE4A, and PDE4B but not of PDE4D. These PI3Kγ-regulated PDEs lowered cAMP and limited protein kinase A-mediated phosphorylation of L-type calcium channel (Ca(v)1.2) and phospholamban. In PI3Kγ(-/-) cardiomyocytes, Ca(v)1.2 and phospholamban were hyperphosphorylated, leading to increased Ca(2+) spark occurrence and amplitude on adrenergic stimulation. Furthermore, PI3Kγ(-/-) cardiomyocytes showed spontaneous Ca(2+) release events and developed arrhythmic calcium transients.ConclusionsPI3Kγ coordinates the coincident signaling of the major cardiac PDE3 and PDE4 isoforms, thus orchestrating a feedback loop that prevents calcium-dependent ventricular arrhythmia.

The bile acid-responsive G protein-coupled receptor TGR5 is involved in several metabolic processes, and recent studies suggest that TGR5 activation may promote pathways that are protective against diet-induced diabetes. Here, we investigated the role of macrophage-specific TGR5 signaling in protecting adipose tissue from inflammation and associated insulin resistance. Examination of adipose tissue from obese mice lacking macrophage Tgr5 revealed enhanced inflammation, increased chemokine expression, and higher macrophage numbers compared with control obese animals. Moreover, macrophage-specific deletion of Tgr5 exacerbated insulin resistance in obese animals. Conversely, pharmacological activation of TGR5 markedly decreased LPS-induced chemokine expression in primary macrophages. This reduction was mediated by AKT-dependent activation of mTOR complex 1, which in turn induced the differential translation of the dominant-negative C/EBPβ isoform, liver inhibitory protein (LIP). Overall, these studies reveal a signaling pathway downstream of TGR5 that modulates chemokine expression in response to high-fat diet and suggest that targeting this pathway has the potential to be therapeutically exploited for prevention of chronic inflammatory diseases and type 2 diabetes mellitus., Introduction Type 2 diabetes mellitus (T2DM) is rising in incidence worldwide and constitutes a major public health burden (1). T2DM is characterized by hyperglycemia and is caused by progressive peripheral [...]

Background--Phosphoinositide 3-kinase y (PI3Ky) signaling engaged by /3-adrenergic receptors is pivotal in the regulation of myocardial contractility and remodeling. However, the role of PI3Ky in catecholamine-induced arrhythmia is currently unknown. Methods and Results--Mice lacking PI3Ky (PI3Ky-/-) showed runs of premature ventricular contractions on adrenergic stimulation that could be rescued by a selective β²-adrenergic receptor blocker and developed sustained ventricular tachycardia after transverse aortic constriction. Consistently, fluorescence resonance energy transfer probes revealed abnormal cAMP accumulation after β²-adrenergic receptor activation in PI3Kɣ-/- cardiomyocytes that depended on the loss of the scaffold but not of the catalytic activity of PI3Ky. Downstream from β-adrenergic receptors, PI3Kɣ was found to participate in multiprotein complexes linking protein kinase A to the activation of phosphodiesterase (PDE) 3A, PDE4A, and PDE4B but not of PDE4D. These PI3Ky-regulated PDEs lowered cAMP and limited protein kinase A-mediated phosphorylation of L-type calcium channel (Cav1.2) and phospholamban. In PI3K-ɣ-/- cardiomyocytes, Cav1.2 and phospholamban were hyperphosphorylated, leading to increased Ca2+ spark occurrence and amplitude on adrenergic stimulation. Furthermore, PI3K-ɣ-/- cardiomyocytes showed spontaneous Ca2+ release events and developed arrhythmic calcium transients. Conclusions--PI3Ky coordinates the coincident signaling of the major cardiac PDE3 and PDE4 isoforms, thus orchestrating a feedback loop that prevents calcium-dependent ventricular arrhythmia. [ABSTRACT FROM AUTHOR]

Summary: Adrenergic stimulation of the heart engages cAMP and phosphoinositide second messenger signaling cascades. Cardiac phosphoinositide 3-kinase p110γ participates in these processes by sustaining β-adrenergic receptor internalization through its catalytic function and by controlling phosphodiesterase 3B (PDE3B) activity via an unknown kinase-independent mechanism. We have discovered that p110γ anchors protein kinase A (PKA) through a site in its N-terminal region. Anchored PKA activates PDE3B to enhance cAMP degradation and phosphorylates p110γ to inhibit PIP3 production. This provides local feedback control of PIP3 and cAMP signaling events. In congestive heart failure, p110γ is upregulated and escapes PKA-mediated inhibition, contributing to a reduction in β-adrenergic receptor density. Pharmacological inhibition of p110γ normalizes β-adrenergic receptor density and improves contractility in failing hearts. [Copyright &y& Elsevier]

The nonessential amino acid asparagine can only be synthesized de novo by the enzymatic activity of asparagine synthetase (ASNS). While ASNS and asparagine have been implicated in the response to numerous metabolic stressors in cultured cells, the in vivo relevance of this enzyme in stress-related pathways remains unexplored. Here, we found ASNS to be expressed in pericentral hepatocytes, a population of hepatic cells specialized in xenobiotic detoxification. ASNS expression was strongly enhanced in 2 models of acute liver injury: carbon tetrachloride (CCl4) and acetaminophen. We found that mice with hepatocyte-specific Asns deletion were more prone to pericentral liver damage than their control littermates after toxin exposure. This phenotype could be reverted by i.v. administration of asparagine. Unexpectedly, the stress-induced upregulation of ASNS involved an ATF4-independent, noncanonical pathway mediated by the nuclear receptor, liver receptor homolog 1 (LRH-1; NR5A2). Altogether, our data indicate that the induction of the asparagine-producing enzyme ASNS acts as an adaptive mechanism to constrain the necrotic wave that follows toxin administration and provide proof of concept that i.v. delivery of asparagine can dampen hepatotoxin-induced pericentral hepatocellular death. [ABSTRACT FROM AUTHOR]

During severe or chronic hepatic injury, biliary epithelial cells (BECs) undergo rapid activation into proliferating progenitors, a crucial step required to establish a regenerative process known as ductular reaction (DR). While DR is a hallmark of chronic liver diseases, including advanced stages of non-alcoholic fatty liver disease (NAFLD), the early events underlying BEC activation are largely unknown. Here, we demonstrate that BECs readily accumulate lipids during high-fat diet feeding in mice and upon fatty acid treatment in BEC-derived organoids. Lipid overload induces metabolic rewiring to support the conversion of adult cholangiocytes into reactive BECs. Mechanistically, we found that lipid overload activates the E2F transcription factors in BECs, which drive cell cycle progression while promoting glycolytic metabolism. These findings demonstrate that fat overload is sufficient to reprogram BECs into progenitor cells in the early stages of NAFLD and provide new insights into the mechanistic basis of this process, revealing unexpected connections between lipid metabolism, stemness, and regeneration., Competing Interests: EY, GE, AP, AJ, PD, KH, LF, JA, GS, KS No competing interests declared, (© 2023, Yildiz et al.)

Objective: We evaluated whether phosphatidylinositol 3-kinase gamma (PI3Kgamma) plays a role in reparative neovascularization and endothelial progenitor cell (EPC) function., Methods and Results: Unilateral limb ischemia was induced in mice lacking the PI3Kgamma gene (PI3Kgamma-/-) or expressing a catalytically inactive mutant (PI3Kgamma(KD/KD)) and wild-type controls (WT). Capillarization and arteriogenesis were reduced in PI3Kgamma-/- ischemic muscles resulting in delayed reperfusion compared with WT, whereas reparative neovascularization was preserved in PI3Kgamma(KD/KD). In PI3Kgamma-/- muscles, endothelial cell proliferation was reduced, apoptosis was increased, and interstitial space was infiltrated with leukocytes but lacked cKit+ progenitor cells that in WT muscles typically surrounded arterioles. PI3Kgamma is constitutively expressed by WT EPCs, with expression levels being upregulated by hypoxia. PI3Kgamma-/- EPCs showed a defect in proliferation, survival, integration into endothelial networks, and migration toward SDF-1. The dysfunctional phenotype was associated with nuclear constraining of FOXO1, reduced Akt and eNOS phosphorylation, and decreased nitric oxide (NO) production. Pretreatment with an NO donor corrected the migratory defect of PI3Kgamma-/- EPCs. PI3Kgamma(KD/KD) EPCs showed reduced Akt phosphorylation, but constitutive activation of eNOS and preserved proliferation, survival, and migration., Conclusions: We newly demonstrated that PI3Kgamma modulates angiogenesis, arteriogenesis, and vasculogenesis by mechanisms independent from its kinase activity.