GHRH agonist MR-409 protects ß-cells from streptozotocin-induced diabetes.

Patients with type 1 diabetes (T1D) suffer from insufficient functional ß-cell mass, which results from infiltration of inflammatory cells and cytokine-mediated ß-cell death. Previous studies demonstrated the beneficial effects of agonists of growth hormone-releasing hormone receptor (GHRH-R), such as MR-409 on preconditioning of islets in a transplantation model. However, the therapeutic potential and protective mechanisms of GHRH-R agonists on models of T1D diabetes have not been explored. Using in vitro and in vivo models of T1D, we assessed the protective propertie of the GHRH agonist, MR409 on ß-cells. The treatment of insulinoma cell lines and rodent and human islets with MR-409 induces Akt signaling by induction of insulin receptor substrate 2 (IRS2), a master regulator of survival and growth in ß-cells, in a PKA-dependent manner. The increase in cAMP/PKA/CREB/IRS2 axis by MR409 was associated with decrease in ß-cell death and improved insulin secretory function in mouse and human islets exposed to proinflammatory cytokines. The assessment of the effects of GHRH agonist MR-409 in a model of T1D induced by low-dose streptozotocin showed that mice treated with MR-409 exhibited better glucose homeostasis, higher insulin levels, and preservation of ß-cell mass. Increased IRS2 expression in ß-cells in the group treated with MR-409 corroborated the in vitro data and provided evidence for the underlying mechanism responsible for beneficial effects of MR-409 in vivo. Collectively, our data show that MR-409 is a novel therapeutic agent for the prevention and treatment of ß-cells death in T1D.

Novel roles of mTORC2 in regulation of insulin secretion by actin filament remodeling.

Mammalian target of rapamycin (mTOR) kinase is an essential hub where nutrients and growth factors converge to control cellular metabolism. mTOR interacts with different accessory proteins to form complexes 1 and 2 (mTORC), and each complex has different intracellular targets. Although mTORC1's role in ß-cells has been extensively studied, less is known about mTORC2's function in ß-cells. Here, we show that mice with constitutive and inducible ß-cell-specific deletion of RICTOR (ßRicKO and ißRicKO mice, respectively) are glucose intolerant due to impaired insulin secretion when glucose is injected intraperitoneally. Decreased insulin secretion in ßRicKO islets was caused by abnormal actin polymerization. Interestingly, when glucose was administered orally, no difference in glucose homeostasis and insulin secretion were observed, suggesting that incretins are counteracting the mTORC2 deficiency. Mechanistically, glucagon-like peptide-1 (GLP-1), but not gastric inhibitory polypeptide (GIP), rescued insulin secretion in vivo and in vitro by improving actin polymerization in ßRicKO islets. In conclusion, mTORC2 regulates glucose-stimulated insulin secretion by promoting actin filament remodeling.NEW & NOTEWORTHY The current studies uncover a novel mechanism linking mTORC2 signaling to glucose-stimulated insulin secretion by modulation of the actin filaments. This work also underscores the important role of GLP-1 in rescuing defects in insulin secretion by modulating actin polymerization and suggests that this effect is independent of mTORC2 signaling.

Exercise-Stimulated ROS Sensitive Signaling Pathways in Skeletal Muscle.

Physical exercise represents a major challenge to whole-body homeostasis, provoking acute and adaptative responses at the cellular and systemic levels. Different sources of reactive oxygen species (ROS) have been described in skeletal muscle (e.g., NADPH oxidases, xanthine oxidase, and mitochondria) and are closely related to the physiological changes induced by physical exercise through the modulation of several signaling pathways. Many signaling pathways that are regulated by exercise-induced ROS generation, such as adenosine monophosphate-activated protein kinase (AMPK), mitogen activated protein kinase (MAPK), nuclear respiratory factor2 (NRF2), and PGC-1α are involved in skeletal muscle responses to physical exercise, such as increased glucose uptake, mitochondriogenesis, and hypertrophy, among others. Most of these adaptations are blunted by antioxidants, revealing the crucial role played by ROS during and after physical exercise. When ROS generation is either insufficient or exacerbated, ROS-mediated signaling is disrupted, as well as physical exercise adaptations. Thus, an understanding the limit between "ROS that can promote beneficial effects" and "ROS that can promote harmful effects" is a challenging question in exercise biology. The identification of new mediators that cause reductive stress and thereby disrupt exercise-stimulated ROS signaling is a trending on this topic and are covered in this current review.

Dual oxidase 1 limits the IFNγ-associated antitumor effect of macrophages.

BACKGROUND: Macrophages play pivotal roles in tumor progression and the response to anticancer therapies, including radiotherapy (RT). Dual oxidase (DUOX) 1 is a transmembrane enzyme that plays a critical role in oxidant generation. METHODS: Since we found DUOX1 expression in macrophages from human lung samples exposed to ionizing radiation, we aimed to assess the involvement of DUOX1 in macrophage activation and the role of these macrophages in tumor development. RESULTS: Using Duox1-/- mice, we demonstrated that the lack of DUOX1 in proinflammatory macrophages improved the antitumor effect of these cells. Furthermore, intratumoral injection of Duox1-/- proinflammatory macrophages significantly enhanced the antitumor effect of RT. Mechanistically, DUOX1 deficiency increased the production of proinflammatory cytokines (IFNγ, CXCL9, CCL3 and TNFα) by activated macrophages in vitro and the expression of major histocompatibility complex class II in the membranes of macrophages. We also demonstrated that DUOX1 was involved in the phagocytotic function of macrophages in vitro and in vivo. The antitumor effect of Duox1-/- macrophages was associated with a significant increase in IFNγ production by both lymphoid and myeloid immune cells. CONCLUSIONS: Our data indicate that DUOX1 is a new target for macrophage reprogramming and suggest that DUOX1 inhibition in macrophages combined with RT is a new therapeutic strategy for the management of cancers.

Muscle Redox Signaling: Engaged in Sickness and in Health.

Even though physical activity is known to perturb the redox homeostasis and create a pro-oxidative muscular environment, robust evidence has confirmed precise, powerful, and beneficial effects of regular physical activity on health. Physical exercise can activate redox-sensitive intracellular signaling pathways via reactive oxygen species (ROS)-related pathways leading to modification of muscle function through genomic and nongenomic mechanisms. However, ROS-mediated signaling also has deleterious effects on skeletal muscle function, which has been observed in several pathological conditions, such as cancer, obesity, and diabetes, among others. One of the most challenging issues debated on this topic is that of the levels of redox signaling that promote either beneficial or harmful effects to our bodies. This Forum discusses the latest progress in muscle redox signaling with emphasis on muscle physiology and physiopathology. Antioxid. Redox Signal. 33, 539-541.

HIV-1 Tat protein induces DNA damage in human peripheral blood B-lymphocytes via mitochondrial ROS production.

Human immunodeficiency virus (HIV) infection is associated with B-cell malignancies in patients though HIV-1 is not able to infect B-cells. The rate of B-cell lymphomas in HIV-infected individuals remains high even under the combined antiretroviral therapy (cART) that reconstitutes the immune function. Thus, the contribution of HIV-1 to B-cell oncogenesis remains enigmatic. HIV-1 induces oxidative stress and DNA damage in infected cells via multiple mechanisms, including viral Tat protein. We have detected elevated levels of reactive oxygen species (ROS) and DNA damage in B-cells of HIV-infected individuals. As Tat is present in blood of infected individuals and is able to transduce cells, we hypothesized that it could induce oxidative DNA damage in B-cells promoting genetic instability and malignant transformation. Indeed, incubation of B-cells isolated from healthy donors with purified Tat protein led to oxidative stress, a decrease in the glutathione (GSH) levels, DNA damage and appearance of chromosomal aberrations. The effects of Tat relied on its transcriptional activity and were mediated by NF-κB activation. Tat stimulated oxidative stress in B-cells mostly via mitochondrial ROS production which depended on the reverse electron flow in Complex I of respiratory chain. We propose that Tat-induced oxidative stress, DNA damage and chromosomal aberrations are novel oncogenic factors favoring B-cell lymphomas in HIV-1 infected individuals.

Dissecting thyroid hormone transport and metabolism in dendritic cells.

We reported thyroid hormone (TH) receptor expression in murine dendritic cells (DCs) and 3,5,3'-triiodothyronine (T3)-dependent stimulation of DC maturation and ability to develop a Th1-type adaptive response. Moreover, an increased DC capacity to promote antigen-specific cytotoxic T-cell activity, exploited in a DC-based antitumor vaccination protocol, was revealed. However, putative effects of the main circulating TH, l-thyroxine (T4) and the mechanisms of TH transport and metabolism at DC level, crucial events for TH action at target cell level, were not known. Herein, we show that T4 did not reproduce those registered T3-dependent effects, finding that may reflect a homoeostatic control to prevent unspecific systemic activation of DCs. Besides, DCs express MCT10 and LAT2 TH transporters, and these cells mainly transport T3 with a favored involvement of MCT10 as its inhibition almost prevented T3 saturable uptake mechanism and reduced T3-induced IL-12 production. In turn, DCs express iodothyronine deiodonases type 2 and 3 (D2, D3) and exhibit both enzymatic activities with a prevalence towards TH inactivation. Moreover, T3 increased MCT10 and LAT2 expression and T3 efflux from DCs but not T3 uptake, whereas it induced a robust induction of D3 with a parallel slight reduction in D2. These findings disclose pivotal events involved in the mechanism of action of THs on DCs, providing valuable tools for manipulating the immunogenic potential of these cells. Furthermore, they broaden the knowledge of the TH mechanism of action at the immune system network.

Sustained IGF-1 Secretion by Adipose-Derived Stem Cells Improves Infarcted Heart Function.

The mechanism by which stem cell-based therapy improves heart function is still unknown, but paracrine mechanisms seem to be involved. Adipose-derived stem cells (ADSCs) secrete several factors, including insulin-like growth factor-1 (IGF-1), which may contribute to myocardial regeneration. Our aim was to investigate whether the overexpression of IGF-1 in ADSCs (IGF-1-ADSCs) improves treatment of chronically infarcted rat hearts. ADSCs were transduced with a lentiviral vector to induce IGF-1 overexpression. IGF-1-ADSCs transcribe100- to 200-fold more IGF-1 mRNA levels compared to nontransduced ADSCs. IGF-1 transduction did not alter ADSC immunophenotypic characteristics even under hypoxic conditions. However, IGF-1-ADSCs proliferate at higher rates and release greater amounts of growth factors such as IGF-1, vascular endothelial growth factor (VEGF), and hepatocyte growth factor (HGF) under normoxic and hypoxic conditions. Importantly, IGF-1 secreted by IGF-1-ADSCs is functional given that Akt-1 phosphorylation was remarkably induced in neonatal cardiomyocytes cocultured with IGF-1-ADSCs, and this increase was prevented with phosphatidylinositol 3-kinase (PI3K) inhibitor treatment. Next, we tested IGF-1-ADSCs in a rat myocardial infarction (MI) model. MI was performed by coronary ligation, and 4 weeks after MI, animals received intramyocardial injections of either ADSCs (n = 7), IGF-1-ADSCs (n = 7), or vehicle (n = 7) into the infarcted border zone. Left ventricular function was evaluated by echocardiography before and after 6 weeks of treatment, and left ventricular hemodynamics were assessed 7 weeks after cell injection. Notably, IGF-1-ADSCs improved left ventricular ejection fraction and cardiac contractility index, but did not reduce scar size when compared to the ADSC-treated group. In summary, transplantation of ADSCs transduced with IGF-1 is a superior therapeutic approach to treat MI compared to nontransduced ADSCs, suggesting that gene and cell therapy may bring additional benefits to the treatment of MI.

When an Intramolecular Disulfide Bridge Governs the Interaction of DUOX2 with Its Partner DUOXA2.

AIMS: The dual oxidase 2 (DUOX2) protein belongs to the NADPH oxidase (NOX) family. As H2O2 generator, it plays a key role in both thyroid hormone biosynthesis and innate immunity. DUOX2 forms with its maturation factor, DUOX activator 2 (DUOXA2), a stable complex at the cell surface that is crucial for the H2O2-generating activity, but the nature of their interaction is unknown. The contribution of some cysteine residues located in the N-terminal ectodomain of DUOX2 in a surface protein-protein interaction is suggested. We have investigated the involvement of different cysteine residues in the formation of covalent bonds that could be of critical importance for the function of the complex. RESULTS: We report the identification and the characterization of an intramolecular disulfide bond between cys-124 of the N-terminal ectodomain and cys-1162 of an extracellular loop of DUOX2, which has important functional implications in both export and activity of DUOX2. This intramolecular bridge provides structural support for the formation of interdisulfide bridges between the N-terminal domain of DUOX2 and the two extracellular loops of its partner, DUOXA2. INNOVATION: Both stability and function of the maturation factor, DUOXA2, are dependent on the oxidative folding of DUOX2, indicating that DUOX2 displays a chaperone-like function with respect to its partner. CONCLUSIONS: The oxidative folding of DUOX2 that takes place in the endoplasmic reticulum (ER) appears to be a key event in the trafficking of the DUOX2/DUOXA2 complex as it promotes an appropriate conformation of the N-terminal region, which is propitious to subsequent covalent interactions with the maturation factor, DUOXA2.

A change in liver metabolism but not in brown adipose tissue thermogenesis is an early event in ovariectomy-induced obesity in rats.

Menopause is associated with increased visceral adiposity and disrupted glucose homeostasis, but the underlying molecular mechanisms related to these metabolic changes are still elusive. Brown adipose tissue (BAT) plays a key role in energy expenditure that may be regulated by sexual steroids, and alterations in glucose homeostasis could precede increased weight gain after ovariectomy. Thus, the aim of this work was to evaluate the metabolic pathways in both the BAT and the liver that may be disrupted early after ovariectomy. Ovariectomized (OVX) rats had increased food efficiency as early as 12 days after ovariectomy, which could not be explained by differences in feces content. Analysis of isolated BAT mitochondria function revealed no differences in citrate synthase activity, uncoupling protein 1 expression, oxygen consumption, ATP synthesis, or heat production in OVX rats. The addition of GDP and BSA to inhibit uncoupling protein 1 decreased oxygen consumption in BAT mitochondria equally in both groups. Liver analysis revealed increased triglyceride content accompanied by decreased levels of phosphorylated AMP-activated protein kinase and phosphorylated acetyl-CoA carboxylase in OVX animals. The elevated expression of gluconeogenic enzymes in OVX and OVX + estradiol rats was not associated with alterations in glucose tolerance test or in serum insulin but was coincident with higher glucose disposal during the pyruvate tolerance test. Although estradiol treatment prevented the ovariectomy-induced increase in body weight and hepatic triglyceride and cholesterol accumulation, it was not able to prevent increased gluconeogenesis. In conclusion, the disrupted liver glucose homeostasis after ovariectomy is neither caused by estradiol deficiency nor is related to increased body mass.

Granulocyte-colony stimulating factor treatment of chronic myocardial infarction.

PURPOSE: The aim of this study was to investigate the impact of granulocyte-colony stimulating factor (G-CSF) administration on cardiac function of rats with chronic myocardial infarction through two different protocols: high dose short term and low dose long term protocols. METHODS: Wistar rats were submitted to MI surgery and after 4 weeks they received recombinant human G-CSF (Filgrastim) or vehicle subcutaneously. We tested the classical protocol (50 microg/kg/day during 7 days) and the long term low dose treatment (four cycles of 5 days of 10 microg/kg/day). Cardiac performance was evaluated before, 4 and 6 weeks after G-CSF injections by electro- and echocardiography, hemodynamic and treadmill exercise test. RESULTS: All infarcted groups exhibited impaired function compared to sham operated animals. Moreover, all cardiac functional parameter were not different between G-CSF and Vehicle group at resting conditions as well as after treadmill exercise stress test, despite intense white blood cell mobilization in both protocols at all time points. Hypertrophy was not different and infarct size was similar in histological analysis CONCLUSIONS: These data clearly show that G-CSF treatment was unable to restore cardiac function impaired by myocardial infarction either with classical approach or long term low dose administration.