Energy Metabolism in Cardiac Remodeling and Heart Failure

Fatty acids are the main substrates used by mitochondria to provide myocardial energy under normal conditions. During heart remodeling, however, the fuel preference switches to glucose. In the earlier stages of cardiac remodeling, changes in energy metabolism are considered crucial to protect the heart from irreversible damage. Furthermore, low fatty acid oxidation and the stimulus for glycolytic pathway lead to lipotoxicity, acidosis, and low adenosine triphosphate production. While myocardial function is directly associated with energy metabolism, the metabolic pathways could be potential targets for therapy in heart failure. © 2013 by Lippincott Williams & Wilkins.

Efeito do tabagismo passivo e do exercício físico associado sobre a expressão de transportador de glicose GLUT4 em músculos de ratos

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The role of proestrus on fertility and postovulatory uterine function in the cow

Modern protocols to synchronize ovulations for timed artificial insemination and timed embryo transfer that include manipulations in the proestrus period (i.e., between luteolysis and estrus) affect fertility in cattle. Specifically, stimulating pre-ovulatory follicle growth and exposure to estrogens after CL regression increase the proportion of cows pregnant and decrease late embryo mortality. Such effects may be due to both preovulatory actions of estrogens and post-ovulatory actions of progesterone, as concentrations of the later hormone may be changed in response to manipulations conducted during proestrus. In the first portion of this paper we describe strategies used recently to manipulate the proestrus period in protocols for synchronization of ovulation, and to present evidence of their effects on fertility. Manipulations of timing and prominence of sex steroids during the proestrus and early diestrus that affect fertility may act on targets such as the endometrium. This tissue expresses receptors for both estrogens and progesterone and these hormones change endometrial function to support conceptus growth and pregnancy maintenance. However, specific cellular and molecular mechanisms through which fertility is affected via manipulations of the proestrus are poorly understood. In the second portion of this paper we describe a well-defined animal model to study changes in endometrial function induced by manipulations conducted during the proestrus. Such manipulations induced endometrial changes on sex steroid receptors expression, cell proliferation, oxidative metabolism and eicosanoid synthesis in the uterus, but not on glucose transport to uterine lumen. In summary, evidence is accumulating to support a positive role of increasing duration and estrogen availability during the proestrus on fertility to synchronization protocols. Such positive effects may be through changes in endometrial function to stimulate conceptus growth and survival.

On the effect of aromatherapy with citrus fragrance in the therapy of major depressive disorder

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Previous Exercise Training Has a Beneficial Effect on Renal and Cardiovascular Function in a Model of Diabetes

Exercise training (ET) is an important intervention for chronic diseases such as diabetes mellitus (DM). However, it is not known whether previous exercise training intervention alters the physiological and medical complications of these diseases. We investigated the effects of previous ET on the progression of renal disease and cardiovascular autonomic control in rats with streptozotocin (STZ)-induced DM. Male Wistar rats were divided into five groups. All groups were followed for 15 weeks. Trained control and trained diabetic rats underwent 10 weeks of exercise training, whereas previously trained diabetic rats underwent 14 weeks of exercise training. Renal function, proteinuria, renal sympathetic nerve activity (RSNA) and the echocardiographic parameters autonomic modulation and baroreflex sensitivity (BRS) were evaluated. In the previously trained group, the urinary albumin/creatinine ratio was reduced compared with the sedentary diabetic and trained diabetic groups (p < 0.05). Additionally, RSNA was normalized in the trained diabetic and previously trained diabetic animals (p < 0.05). The ejection fraction was increased in the previously trained diabetic animals compared with the diabetic and trained diabetic groups (p < 0.05), and the myocardial performance index was improved in the previously trained diabetic group compared with the diabetic and trained diabetic groups (p < 0.05). In addition, the previously trained rats had improved heart rate variability and BRS in the tachycardic response and bradycardic response in relation to the diabetic group (p < 0.05). This study demonstrates that previous ET improves the functional damage that affects DM. Additionally, our findings suggest that the development of renal and cardiac dysfunction can be minimized by 4 weeks of ET before the induction of DM by STZ.

beta(1) Adrenergic receptor is key to cold- and diet-induced thermogenesis in mice

Brown adipose tissue (BAT) is predominantly regulated by the sympathetic nervous system (SNS) and the adrenergic receptor signaling pathway. Knowing that a mouse with triple beta-receptor knockout (KO) is cold intolerant and obese, we evaluated the independent role played by the beta(1) isoform in energy homeostasis. First, the 30 min i.v. infusion of norepinephrine (NE) or the beta(1) selective agonist dobutamine (DB) resulted in similar interscapular BAT (iBAT) thermal response in WT mice. Secondly, mice with targeted disruption of the beta(1) gene (KO of beta(1) adrenergic receptor (beta 1KO)) developed hypothermia during cold exposure and exhibited decreased iBAT thermal response to NE or DB infusion. Thirdly, when placed on a high-fat diet (HFD; 40% fat) for 5 weeks, beta 1KO mice were more susceptible to obesity than WT controls and failed to develop diet-induced thermogenesis as assessed by BAT Ucp1 mRNA levels and oxygen consumption. Furthermore, beta 1KO mice exhibited fasting hyperglycemia and more intense glucose intolerance, hypercholesterolemia, and hypertriglyceridemia when placed on the HFD, developing marked non-alcoholic steatohepatitis. In conclusion, the beta(1) signaling pathway mediates most of the SNS stimulation of adaptive thermogenesis. Journal of Endocrinology (2012) 214, 359-365

Ruolo delle specie reattive dell'ossigeno e di caveole/raft lipidici nella biosegnalazione in linee cellulari umane

Membrane lipid rafts are detergent-resistant microdomains containing glycosphingolipids, cholesterol and glycosylphosphatidylinositol-linked proteins; they seem to be actively involved in many cellular processes including signal transduction, apoptosis, cell adhesion and migration. Lipid rafts may represent important functional platforms where redox signals are produced and transmitted in response to various agonists or stimuli. In addition, a new concept is emerging that could be used to define the interactions or amplification of both redox signalling and lipid raft-associated signalling. This concept is characterized by redox-mediated feed forward amplification in lipid platforms. It is proposed that lipid rafts are formed in response to various stimuli; for instance, NAD(P)H oxidase (Nox) subunits are aggregated or recruited in these platforms, increasing Nox activity. Superoxide and hydrogen peroxide generation could induce various regulatory activities, such as the induction of glucose transport activity and proliferation in leukaemia cells. The aim of our study is to probe: i) the involvement of lipid rafts in the modulation of the glucose transporter Glut1 in human acute leukemia cells; ii) the involvement of plasma membrane caveolae/lipid rafts in VEGF-mediated redox signaling via Nox activation in human leukemic cells; iii) the role of p66shc, an adaptor protein, in VEGF signaling and ROS production in endothelial cells (ECs); iv) the role of Sindecan-2, a transmembrane heparan sulphate proteoglycan, in VEGF signaling and physiological response in ECs and v) the antioxidant and pro-apoptotic activities of simple dietary phenolic acids, i. e. caffeic, syringic and protocatechuic acids in leukemia cells, characterized by a very high ROS content. Our results suggest that the role played by NAD(P)H oxidase-derived ROS in the regulation of glucose uptake, proliferation and migration of leukaemia and endothelial cells could likely occur through the control of lipid raft-associated signalling.

ROS generate dalle NAD(P)H ossidasi nella segnalazione redox in linee cellulari leucemiche

Discovery of the Nox family has led to the concept that ROS are “intentionally” generated and are biologically functional in various cell types. Over the last decades, ROS have been shown to be involved in several physiological and pathological processes and ROS producing enzymes have been suggested as a target for drug development. The mechanism involved in the prosurvival effect of cytokines on the human acute myeloid leukaemia cell lines M07e and B1647 is investigated. A decrease in intracellular reactive oxygen species (ROS) content, glucose transport activity and cell survival was observed in the presence of inhibitors of plasma membrane ROS sources, such as DPI and apocynin, and by small interference RNA for NOX2 in M07e cells. Furthermore, Nox generated ROS are required to sustain B1647 cell viability and proliferation; in fact, antioxidants such as EUK-134 or Nox inhibitors and siRNA direct cells to apoptotic cell death, suggesting that manipulation of cellular NOX2 and NOX4 could affect survival of leukemic cells. Moreover, hydrogen peroxide has been long thought to be freely diffusible but recent evidence suggest that specific mammalian aquaporin homologues (AQP8) possess the capacity to channel H2O2 across membrane. In this thesis is shown that inhibition of aquaporins diminishes intracellular ROS accumulation either when H2O2 is produced by Nox enzymes or when is added exogenously to the medium. These data suggest that specific inhibition of Nox enzymes and AQP8 could be an interesting novel anti-leukemic strategy.

Characteristics of a spina bifida population including North American Caucasian and Hispanic individuals.

BACKGROUND: Meningomyelocele (MM) is a common human birth defect. MM is a disorder of neural development caused by contributions from genes and environmental factors that result in the NTD and lead to a spectrum of physical and neurocognitive phenotypes. METHODS: A multidisciplinary approach has been taken to develop a comprehensive understanding of MM through collaborative efforts from investigators specializing in genetics, development, brain imaging, and neurocognitive outcome. Patients have been recruited from five different sites: Houston and the Texas-Mexico border area; Toronto, Canada; Los Angeles, California; and Lexington, Kentucky. Genetic risk factors for MM have been assessed by genotyping and association testing using the transmission disequilibrium test. RESULTS: A total of 509 affected child/parent trios and 309 affected child/parent duos have been enrolled to date for genetic association studies. Subsets of the patients have also been enrolled for studies assessing development, brain imaging, and neurocognitive outcomes. The study recruited two major ethnic groups, with 45.9% Hispanics of Mexican descent and 36.2% North American Caucasians of European descent. The remaining patients are African-American, South and Central American, Native American, and Asian. Studies of this group of patients have already discovered distinct corpus callosum morphology and neurocognitive deficits that associate with MM. We have identified maternal MTHFR 667T allele as a risk factor for MM. In addition, we also found that several genes for glucose transport and metabolism are potential risk factors for MM. CONCLUSIONS: The enrolled patient population provides a valuable resource for elucidating the disease characteristics and mechanisms for MM development.

The Effect of Heavy Metal Stress on Avian Proximal Tubule Urate Secretion

In both humans and birds, urate is an important antioxidant when maintained at normal plasma concentrations. Though human kidneys primarily reabsorb filtered urate, while those of birds perform mostly secretion, both maintain urate levels at ~300microM. The importance of maintaining urate levels within the homeostatic range was observed when the study of several prominent diseases revealed an association with hyperuricemia. This study examined the effect of elevated zinc concentration on avian urate secretion. Here, acute exposure of chicken proximal tubule epithelial cells (cPTCs) to zinc stress had no effect on urate secretion, but prolonged zinc-induced cellular stress inhibited active transepithelial urate secretion with no change in Mrp4 expression, glucose transport, or transepithelial resistance. Moreover, zinc had no effect on urate transport by isolated brush border membrane vesicles, suggesting involvement of a more complex cellular stress adaptation. Previous work has demonstrated that AMP-activated protein kinase (AMPK), a critical metabolic regulator, conserves energy during cellular stress by shutting down ATP-utilizing processes and activating ATP-generating processes. Pharmacological activation of AMPK by AICAR produced decreased urate secretion by cPTCs similar to the effect seen with prolonged exposure to zinc, while the AMPK inhibitor Compound C prevented both AICAR and zinc inhibition of urate secretion, suggesting a stress induced mechanism of regulation. Supported by NSF. IACUC #A08-046.

Molecular mechanisms for the insulin sensitizing activity of rexinoids in skeletal muscle of diabetic (Db/Db) mice

Rexinoids are synthetic agonists for the retinoid X receptors (RXRs), a member of the nuclear receptor family of ligand-activated transcription factors. Rexinoids have been shown to lower serum glucose and insulin levels in animal models of type 2 diabetes. However the mechanisms that are responsible for the insulin-sensitizing action of rexinoids are largely unknown. Skeletal muscle accounts for the majority of insulin-regulated whole-body glucose disposal and impaired insulin action in muscle is an important contributor to the pathophysiology of type 2 diabetes. Glucose transport is a rate-limiting step in glucose utilization. The goal of these studies is to examine the mechanisms of the anti-diabetic activity of rexinoids in skeletal muscle of diabetic db/db mice. The results we have obtained showed that treatment of db/db mice with rexinoids for two weeks resulted in a significant increase in insulin-stimulated glucose transport activity in skeletal muscle. Insulin stimulates glucose transport in muscle via the regulation of both the insulin receptor substrate-1 (IRS-1)/Akt pathway and the Cbl-associated protein (CAP)/Cbl pathway. Rexinoids increased the insulin-stimulated IRS-1 tyrosine phosphorylation and Akt phosphorylation without effects on the activity of the CAP/Cbl pathway. The effects of rexinoids on the IRS-1/Akt pathway were associated with a decrease in the level of IRS-1 Serine 307 phosphorylation as well as qualitative and quantitative alterations in the fatty acyl-CoAs present within the muscle cells. In addition, rexinoids increased the expression of uncoupling protein 3 (UCP3) and activation of AMPK in diabetic muscle. This effect may also enhance the IRS-1/Akt signaling. We believe that it is the concerted activation of the IRS-1/Akt and AMPK signaling systems, a pharmacological mechanism that as far as we know, is unique to rexinoids, that results in the anti-diabetic effects of these drugs. Our results also suggest that the glucose-lowering mechanism of rexinoids is distinct from that of the thiazolidinediones (TZDs), peroxisome proliferator-activated receptor γ (PPARγ) agonists with well-characterized anti-diabetic activity. Rexinoids appear to represent a novel class of insulin sensitizers, with potential applications for the treatment of type 2 diabetes. ^

Functional growth hormone (GH) receptors and GH are expressed by preimplantation mouse embryos: A role for GH in early embryogenesis?

The results of this study challenge the widely held view that growth hormone (GH) acts only during the postnatal period. RNA phenotyping shows transcripts for the GH receptor and GH-binding protein in mouse preimplantation embryos of all stages from fertilized eggs (day 1) to blastocysts (day 4). An antibody specific to the cytoplasmic region of the GH receptor revealed receptor protein expression, first in two-cell embryos, the stage of activation of the embryonic genome (day 2), and in all subsequent stages. In cleavage-stage embryos this immunoreactivity was localized mainly to the nucleus, but clear evidence of membrane labeling was apparent in blastocysts. GH receptor immunoreactivity was also observed in cumulus cells associated with unfertilized oocytes but not in the unfertilized oocytes. The blastocyst receptor was demonstrated to be functional, exhibiting the classic bell-shaped dose–response curves for GH stimulation of both 3-O-methyl glucose transport and protein synthesis. Maximal stimulation of 40–50% was seen for both responses at less than 1 ng/ml recombinant GH, suggesting a role for maternal GH. However mRNA transcripts for GH were also detected from the morula stage (day 3) by using reverse transcription–PCR, and GH immunoreactivity was seen in blastocysts. These observations raise the possibility of a paracrine/autocrine GH loop regulating embryonic development in its earliest stages.

The Formation of an Insulin-responsive Vesicular Cargo Compartment Is an Early Event in 3T3-L1 Adipocyte Differentiation

Differentiating 3T3-L1 cells exhibit a dramatic increase in the rate of insulin-stimulated glucose transport during their conversion from proliferating fibroblasts to nonproliferating adipocytes. On day 3 of 3T3-L1 cell differentiation, basal glucose transport and cell surface transferrin binding are markedly diminished. This occurs concomitant with the formation of a distinct insulin-responsive vesicular pool of intracellular glucose transporter 1 (GLUT1) and transferrin receptors as assessed by sucrose velocity gradients. The intracellular distribution of the insulin-responsive aminopeptidase is first readily detectable on day 3, and its gradient profile and response to insulin at this time are identical to that of GLUT1. With further time of differentiation, GLUT4 is expressed and targeted to the same insulin-responsive vesicles as the other three proteins. Our data are consistent with the notion that a distinct insulin-sensitive vesicular cargo compartment forms early during fat call differentiation and its formation precedes GLUT4 expression. The development of this compartment may result from the differentiation-dependent inhibition of constitutive GLUT1 and transferrin receptor trafficking such that there is a large increase in, or the new formation of, a population of postendosomal, insulin-responsive vesicles.

Exercise-induced changes in expression and activity of proteins involved in insulin signal transduction in skeletal muscle: Differential effects on insulin-receptor substrates 1 and 2

Level of physical activity is linked to improved glucose homeostasis. We determined whether exercise alters the expression and/or activity of proteins involved in insulin-signal transduction in skeletal muscle. Wistar rats swam 6 h per day for 1 or 5 days. Epitrochlearis muscles were excised 16 h after the last exercise bout, and were incubated with or without insulin (120 nM). Insulin-stimulated glucose transport increased 30% and 50% after 1 and 5 days of exercise, respectively. Glycogen content increased 2- and 4-fold after 1 and 5 days of exercise, with no change in glycogen synthase expression. Protein expression of the glucose transporter GLUT4 and the insulin receptor increased 2-fold after 1 day, with no further change after 5 days of exercise. Insulin-stimulated receptor tyrosine phosphorylation increased 2-fold after 5 days of exercise. Insulin-stimulated tyrosine phosphorylation of insulin-receptor substrate (IRS) 1 and associated phosphatidylinositol (PI) 3-kinase activity increased 2.5- and 3.5-fold after 1 and 5 days of exercise, despite reduced (50%) IRS-1 protein content after 5 days of exercise. After 1 day of exercise, IRS-2 protein expression increased 2.6-fold and basal and insulin-stimulated IRS-2 associated PI 3-kinase activity increased 2.8-fold and 9-fold, respectively. In contrast to IRS-1, IRS-2 expression and associated PI 3-kinase activity normalized to sedentary levels after 5 days of exercise. Insulin-stimulated Akt phosphorylation increased 5-fold after 5 days of exercise. In conclusion, increased insulin-stimulated glucose transport after exercise is not limited to increased GLUT4 expression. Exercise leads to increased expression and function of several proteins involved in insulin-signal transduction. Furthermore, the differential response of IRS-1 and IRS-2 to exercise suggests that these molecules have specialized, rather than redundant, roles in insulin signaling in skeletal muscle.

Hexose permeation pathways in Plasmodium falciparum-infected erythrocytes

Plasmodium falciparum requires glucose as its energy source to multiply within erythrocytes but is separated from plasma by multiple membrane systems. The mechanism of delivery of substrates such as glucose to intraerythrocytic parasites is unclear. We have developed a system for robust functional expression in Xenopus oocytes of the P. falciparum asexual stage hexose permease, PfHT1, and have analyzed substrate specificities of PfHT1. We show that PfHT1 (a high-affinity glucose transporter, Km ≈ 1.0 mM) also transports fructose (Km ≈ 11.5 mM). Fructose can replace glucose as an energy source for intraerythrocytic parasites. PfHT1 binds fructose in a furanose conformation and glucose in a pyranose form. Fructose transport by PfHT1 is ablated by mutation of a single glutamine residue, Q169, which is predicted to lie within helix 5 of the hexose permeation pathway. Glucose transport in the Q169N mutant is preserved. Comparison in oocytes of transport properties of PfHT1 and human facilitative glucose transporter (GLUT)1, an archetypal mammalian hexose transporter, combined with studies on cultured P. falciparum, has clarified hexose permeation pathways in infected erythrocytes. Glucose and fructose enter erythrocytes through separate permeation pathways. Our studies suggest that both substrates enter parasites via PfHT1.