RESUMO
Triacylglycerol (TAG) and glycogen are the two major metabolites for carbon storage in most eukaryotic organisms. We investigated the glycogen metabolism of the oleaginous Yarrowia lipolytica and found that this yeast accumulates up to 16% glycogen in its biomass. Assuming that elimination of glycogen synthesis would result in an improvement of lipid accumulation, we characterized and deleted the single gene coding for glycogen synthase, YlGSY1. The mutant was grown under lipogenic conditions with glucose and glycerol as substrates and we obtained up to 60% improvement in TAG accumulation compared to the wild-type strain. Additionally, YlGSY1 was deleted in a background that was already engineered for high lipid accumulation. In this obese background, TAG accumulation was also further increased. The highest lipid content of 52% was found after 3 days of cultivation in nitrogen-limited glycerol medium. Furthermore, we constructed mutants of Y. lipolytica and Saccharomyces cerevisiae that are deleted for both glycogen and TAG synthesis, demonstrating that the ability to store carbon is not essential. Overall, this work showed that glycogen synthesis is a competing pathway for TAG accumulation in oleaginous yeasts and that deletion of the glycogen synthase has beneficial effects on neutral lipid storage.
Assuntos
Proteínas Fúngicas/genética , Glicogênio Sintase/genética , Glicogênio/biossíntese , Engenharia Metabólica/métodos , Triglicerídeos/biossíntese , Yarrowia/metabolismo , Biomassa , Carbono/metabolismo , Fermentação , Proteínas Fúngicas/metabolismo , Deleção de Genes , Expressão Gênica , Glucose/metabolismo , Glicerol/metabolismo , Glicogênio/antagonistas & inibidores , Glicogênio Sintase/deficiência , Cinética , Metabolismo dos Lipídeos , Nitrogênio/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Yarrowia/genéticaRESUMO
Lafora disease (LD) is a fatal progressive myoclonus epilepsy characterized neuropathologically by aggregates of abnormally structured glycogen and proteins (Lafora bodies [LBs]), and neurodegeneration. Whether LBs could be prevented by inhibiting glycogen synthesis and whether they are pathogenic remain uncertain. We genetically eliminated brain glycogen synthesis in LD mice. This resulted in long-term prevention of LB formation, neurodegeneration, and seizure susceptibility. This study establishes that glycogen synthesis is requisite for LB formation and that LBs are pathogenic. It opens a therapeutic window for potential treatments in LD with known and future small molecule inhibitors of glycogen synthesis.
Assuntos
Glicogênio/antagonistas & inibidores , Glicogênio/biossíntese , Doença de Lafora/prevenção & controle , Animais , Modelos Animais de Doenças , Fosfatases de Especificidade Dupla/genética , Técnicas de Inativação de Genes , Glicogênio Sintase/genética , Glicogênio Sintase/metabolismo , Doença de Lafora/patologia , Doença de Lafora/fisiopatologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Tirosina Fosfatases não ReceptorasRESUMO
Brain glycogen could be a critical energy source for brain activity when the glucose supply from the blood is inadequate (hypoglycaemia). Although untested, it is hypothesized that during prolonged exhaustive exercise that induces hypoglycaemia and muscular glycogen depletion, the resultant hypoglycaemia may cause a decrease in brain glycogen. Here,we tested this hypothesis and also investigated the possible involvement of brain monoamines with the reduced levels of brain glycogen. For this purpose,we exercised male Wistar rats on a treadmill for different durations (30-120 min) at moderate intensity (20 m min⻹) and measured their brain glycogen levels using high-power microwave irradiation (10 kW). At the end of 30 and 60 min of running, the brain glycogen levels remained unchanged from resting levels, but liver and muscle glycogen decreased. After 120 min of running, the glycogen levels decreased significantly by â¼37-60% in five discrete brain loci (the cerebellum 60%, cortex 48%, hippocampus 43%, brainstem 37% and hypothalamus 34%) compared to those of the sedentary control. The brain glycogen levels in all five regions after running were positively correlated with the respective blood and brain glucose levels. Further, in the cortex, the levels of methoxyhydroxyphenylglycol (MHPG) and 5-hydroxyindoleacetic acid (5-HIAA), potential involved in degradation of the brain glycogen, increased during prolonged exercise and negatively correlated with the glycogen levels. These results support the hypothesis that brain glycogen could decrease with prolonged exhaustive exercise. Increased monoamines together with hypoglycaemia should be associated with the development of decreased brain glycogen, suggesting a new clue towards the understanding of central fatigue during prolonged exercise.
Assuntos
Encéfalo/metabolismo , Glicogênio/antagonistas & inibidores , Glicogênio/metabolismo , Condicionamento Físico Animal/métodos , Resistência Física/fisiologia , Animais , Glicemia/metabolismo , Teste de Esforço/métodos , Glicogênio/biossíntese , Masculino , Projetos Piloto , Ratos , Ratos Wistar , Fatores de TempoRESUMO
The pharmacological properties of 1,4-dideoxy-1,4-imino-d-arabinitol (DAB), a potent inhibitor of glycogen phosphorylase and synthase activity in liver preparations, were characterized in different brain tissue preparations as a prerequisite for using it as a tool to investigate brain glycogen metabolism. Its inhibitory effect on glycogen phosphorylase was studied in homogenates of brain tissue and astrocytes and IC50-values close to 400 nM were found. However, the concentration of DAB needed for inhibition of glycogen shunt activity, i.e. glucose metabolism via glycogen, in intact astrocytes was almost three orders of magnitude higher. Additionally, such complete inhibition required a pre-incubation period, a finding possibly reflecting a limited permeability of the astrocytic membrane. DAB did not affect the accumulation of 2-deoxyglucose-6-phosphate indicating that the transport of DAB is not mediated by the glucose transporter. DAB had no effect on enzymes involving glucose-6-phosphate, i.e. glucose-6-phosphate dehydrogenase, phosphoglucoisomerase and hexokinase. Furthermore, DAB was evaluated in a functional preparation of the isolated mouse optic nerve, in which its presence severely reduced the ability to sustain evoked compound action potentials in the absence of glucose, a condition in which glycogen serves as an important energy substrate. Based on the experimental findings, DAB can be used to evaluate glycogen shunt activity and its functional importance in intact brain tissue and cells at a concentration of 300-1000 muM and a pre-incubation period of 1 h.
Assuntos
Arabinose/farmacologia , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Glicogênio/antagonistas & inibidores , Glicogênio/metabolismo , Álcoois Açúcares/farmacologia , Animais , Animais Recém-Nascidos , Células Cultivadas , Relação Dose-Resposta a Droga , Glucose/antagonistas & inibidores , Glucose/metabolismo , Imino Furanoses/farmacologia , CamundongosRESUMO
This study aimed to investigate how Akkermansia muciniphila can implicate type 2 diabetes mellitus and the mechanisms underlying the effects A. muciniphila on type 2 diabetes mellitus. Normal and streptozotocin-induced diabetic Sprague-Dawley rats were orally administered with A. muciniphila and solvent. After 4 weeks of treatment, diabetic rats orally administered with live or pasteurized A. muciniphila exhibited significant increase in the blood concentration of high-density lipoprotein, and decrease in the hepatic glycogen, serum plasminogen activator inhibitor-1, tumor necrosis factor-α, lipopolysaccharide, malondialdehyde and total glucagon-like peptide-1. Moreover, diabetic rats orally administered with A. muciniphila showed significantly increased species alpha diversity and gene function in gut microbes. These results indicated that A. muciniphila can improve liver function, reduce gluco/lipotoxicity, alleviate oxidative stress, suppress inflammation and normalize intestine microbiota of the host animal, thereby ameliorating type 2 diabetes mellitus. Akkermansia muciniphila might be considered as one of the ideal new probiotics used in the management of type 2 diabetes mellitus in future.
Assuntos
Diabetes Mellitus Experimental/dietoterapia , Microbioma Gastrointestinal/efeitos dos fármacos , Hipoglicemiantes/farmacologia , Probióticos/farmacologia , Verrucomicrobia/fisiologia , Animais , HDL-Colesterol/agonistas , HDL-Colesterol/imunologia , HDL-Colesterol/metabolismo , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/imunologia , Diabetes Mellitus Experimental/microbiologia , Fezes/microbiologia , Microbioma Gastrointestinal/imunologia , Regulação da Expressão Gênica/efeitos dos fármacos , Peptídeo 1 Semelhante ao Glucagon/genética , Peptídeo 1 Semelhante ao Glucagon/imunologia , Glicogênio/antagonistas & inibidores , Glicogênio/imunologia , Glicogênio/metabolismo , Inflamação , Mucosa Intestinal/metabolismo , Intestinos/efeitos dos fármacos , Intestinos/imunologia , Lipopolissacarídeos/antagonistas & inibidores , Lipopolissacarídeos/imunologia , Lipopolissacarídeos/metabolismo , Fígado/efeitos dos fármacos , Fígado/imunologia , Fígado/metabolismo , Masculino , Malondialdeído/antagonistas & inibidores , Malondialdeído/imunologia , Malondialdeído/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Inibidor 1 de Ativador de Plasminogênio/genética , Inibidor 1 de Ativador de Plasminogênio/imunologia , Ratos , Ratos Sprague-Dawley , Estreptozocina , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
The effects of vitamin C, vitamin E and vitamin B12 on the noise-induced acute change in hepatic glycogen content in rats were investigated. The exposure of rats to 95 dB and 110 dB of noise acutely reduced their hepatic glycogens. Vitamin C (ascorbic acid) and vitamin E (alpha-tocopherol) attenuated the noise-induced acute reduction in the hepatic glycogen contents. This result suggests that antioxidants could reduce the change via reactive oxygen species. Vitamin B12 (cobalamin) delayed the noise-induced change, a finding that suggests that vitamin B12 could postpone the acute change via compensating for vitamin B12 deficiency.
Assuntos
Ácido Ascórbico/farmacologia , Glicogênio/metabolismo , Fígado/metabolismo , Ruído , Vitamina B 12/farmacologia , Vitamina E/farmacologia , Animais , Antioxidantes/farmacologia , Feminino , Glicogênio/antagonistas & inibidores , Fígado/efeitos dos fármacos , Ruído/efeitos adversos , Ratos , Ratos Wistar , Estresse Fisiológico/metabolismoRESUMO
MicroRNAs (miRNAs) play important roles in epithelial-to-mesenchymal transition (EMT). Moreover, hyperglycaemia induces damage to renal tubular epithelial cells, which may lead to EMT in diabetic nephropathy. However, the effects of miRNAs on EMT in diabetic nephropathy are poorly understood. In the present study, we found that the level of microRNA-23b (miR-23b) was significantly decreased in high glucose (HG)-induced human kidney proximal tubular epithelial cells (HK2) and in kidney tissues of db/db mice. Overexpression of miR-23b attenuated HG-induced EMT, whereas knockdown of miR-23b induced normal glucose (NG)-mediated EMT in HK2 cells. Mechanistically, miR-23b suppressed EMT in diabetic nephropathy by targeting high mobility group A2 (HMGA2), thereby repressing PI3K-AKT signalling pathway activation. Additionally, HMGA2 knockdown or inhibition of the PI3K-AKT signalling pathway with LY294002 mimicked the effects of miR-23b overexpression on HG-mediated EMT, whereas HMGA2 overexpression or activation of the PI3K-AKT signalling pathway with BpV prevented the effects of miR-23b on HG-mediated EMT. We also confirmed that overexpression of miR-23b alleviated EMT, decreased the expression levels of EMT-related genes, ameliorated renal morphology, glycogen accumulation, fibrotic responses and improved renal functions in db/db mice. Taken together, we showed for the first time that miR-23b acts as a suppressor of EMT in diabetic nephropathy through repressing PI3K-AKT signalling pathway activation by targeting HMGA2, which maybe a potential therapeutic target for diabetes-induced renal dysfunction.
Assuntos
Nefropatias Diabéticas/genética , Células Epiteliais/metabolismo , Proteína HMGA2/genética , MicroRNAs/genética , Animais , Linhagem Celular , Cromonas/farmacologia , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/patologia , Modelos Animais de Doenças , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Transição Epitelial-Mesenquimal/genética , Regulação da Expressão Gênica , Glucose/farmacologia , Glicogênio/antagonistas & inibidores , Glicogênio/biossíntese , Proteína HMGA2/metabolismo , Humanos , Túbulos Renais Proximais/efeitos dos fármacos , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/patologia , Camundongos , Camundongos Transgênicos , MicroRNAs/agonistas , MicroRNAs/antagonistas & inibidores , MicroRNAs/metabolismo , Morfolinas/farmacologia , Oligorribonucleotídeos/genética , Oligorribonucleotídeos/metabolismo , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de SinaisRESUMO
We examined the possibility that protein kinase C (PKC) is chronically activated and may contribute to impaired glycogen synthesis and insulin resistance in soleus muscles of hyperinsulinemic type II diabetic Goto-Kakizaki (GK) rats. Relative to nondiabetic controls, PKC enzyme activity and levels of immunoreactive PKC-alpha, beta, epsilon, and delta were increased in membrane fractions and decreased cytosolic fractions of GK soleus muscles. In addition, PKC-theta levels were decreased in both membrane and cytosol fractios, whereas PKC-zeta levels were not changed in either fraction in GK soleus muscles. These increases in membrane PKC (alpha, beta, epsilon, and delta) could not be accounted for by alterations in PKC mRNA or total PKC levels but were associated with increases in membrane diacylglycerol (DAG) and therefore appeared to reflect translocative activation of PKC. In evaluation of potential causes for persistent PKC activation, membrane PKC levels were decreased in soleus muscles of hyperglycemic streptozotocin (STZ)-induced diabetic rats; thus, a role for simple hyperglycemia as a cause of PKC activation in GK rats was not evident in the STZ model. In support of the possibility that hyperinsulinemia contributed to PKC activation in GK soleus muscles, we found that DAG levels were increased, and PKC was translocated, in soleus muscles of both (1) normoglycemic hyperinsulinemic obese/aged rats and (2) mildly hyperglycemic hyperinsulinemic obese/Zucker rats. In keeping with the possibility that PKC activation may contribute to impaired glycogen synthase activation in GK muscles, phorbol esters inhibited, and a PKC inhibitor, RO 31-8220, increased insulin effects on glycogen synthesis in soleus muscles incubated in vitro. Our findings suggested that: (1) hyperinsulinemia, as observed in type II diabetic GK rats and certain genetic and nongenetic forms of obesity in rats, is associated with persistent translocation and activation of PKC in soleus muscles, and (2) this persistent PKC activation may contribute to impaired glycogen synthesis and insulin resistance.
Assuntos
Envelhecimento , Diabetes Mellitus Tipo 2/enzimologia , Glicogênio/biossíntese , Resistência à Insulina , Músculo Esquelético/enzimologia , Obesidade/enzimologia , Proteína Quinase C/biossíntese , Adipócitos/enzimologia , Animais , Membrana Celular/enzimologia , Citosol/enzimologia , Ativação Enzimática , Glicogênio/antagonistas & inibidores , Isoenzimas/biossíntese , Fígado/enzimologia , Masculino , Proteína Quinase C beta , Proteína Quinase C-alfa , Proteína Quinase C-delta , Proteína Quinase C-épsilon , RNA Mensageiro/biossíntese , Ratos , Ratos Mutantes , Ratos Wistar , Ratos Zucker , Valores de Referência , Transcrição GênicaRESUMO
In healthy individuals, glycogen recovery after a strong depletion is known to be rapid and insulin independent during the initial phase, and subsequently, slow and insulin dependent. Free fatty acids (FFAs) as a putative source of insulin resistance (IR) could thus impair glycogen recovery during the second period. Using in vivo 13C nuclear magnetic resonance (NMR), we studied the effect of long-chain triglyceride emulsion on gastrocnemius glycogen resynthesis during a 3-h recovery period after 90 min of moderate exercise consisting of plantar flexion on overnight-fasted healthy men (n = 8). In separate experiments, each subject was infused with 10% Ivelip (0.015 ml x kg(-1) x min(-1)) or 10% glycerol (0.13 mg x kg(-1) x min(-1)). NMR spectra were acquired before and at the end of the exercise and during the recovery period. Whole-body glucose and lipid oxidation rates (indirect calorimetry), plasma insulin, C-peptide, glucose, lactate, beta-hydroxybutyrate, triglycerides, and FFAs were determined. Glycogen consumption was 47.6 +/- 4.5% (glycerol) and 49.7 +/- 4.8% (Ivelip) of the initial glycogen. An acquired IR in the Ivelip group was significant at the onset of the recovery period by homeostasis model assessment (P = 0.002). Glycogen resynthesis in the glycerol group appeared faster during the 1st h than during the subsequent 2nd h of the postexercise period. The glycogen resynthesis level was significantly lower in the Ivelip group than in the glycerol group during the recovery period (P = 0.04 during the 1st h and P = 0.001 during the next 2 h). During the recovery, plasma lactate and whole-body oxidation rates were similar in the two groups, whereas glycemia was significantly higher in the Ivelip group. A decreased cellular uptake of glucose as a substrate for glycogenosynthesis, rather than a competition between oxidation of carbohydrate and FFA, is discussed.
Assuntos
Exercício Físico/fisiologia , Glicogênio/antagonistas & inibidores , Glicogênio/biossíntese , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Triglicerídeos/farmacologia , Adulto , Isótopos de Carbono , Emulsões , Ácidos Graxos não Esterificados/sangue , Glicerol/sangue , Glicerol/farmacologia , Humanos , Infusões Intravenosas , Resistência à Insulina/fisiologia , Metabolismo dos Lipídeos , Espectroscopia de Ressonância Magnética , Masculino , Oxirredução , Valores de Referência , Triglicerídeos/sangueRESUMO
Rat pancreatic islets cultured for 1 to 5 days in the presence of 20 to 80 mmol/L D-glucose accumulate glycogen in a time- and concentration-dependent manner. When the glycogen-rich islets are incubated for 6 to 10 minutes in the absence of D-glucose, the rate of glycogenolysis is grossly proportional to the glycogen content. Exogenous D-glucose (7 to 20 mmol/L) inhibits glycogenolysis. This inhibitory effect opposes the increase in glycolytic flux attributable to the utilization of exogenous glucose. Both the inhibitory effect of D-glucose on glycogenolysis and the utilization of exogenous hexose tend to be higher with alpha- than with beta-D-glucose. In light of these findings, it is proposed that the interference of D-glucose with glycogenolysis might play a role in the paradoxical changes in insulin output and its altered anomeric specificity in response to D-glucose administration, as is often encountered in non-insulin-dependent diabetic subjects and experimental models of B-cell glucotoxicity.
Assuntos
Glucose/efeitos adversos , Glicogênio/metabolismo , Ilhotas Pancreáticas/efeitos dos fármacos , Animais , Técnicas de Cultura , Feminino , Glicogênio/antagonistas & inibidores , Glicogênio/farmacologia , Ratos , Fatores de TempoRESUMO
Thirty strength-trained subjects were randomized to carbohydrate (CHO) or placebo (Pla) groups and lifted weights for 2 h (10 exercises, 4 sets each, 10 repetitions, with 2- to 3-min rest intervals). Subjects received 10 ml x kg(-1) x h(-1) CHO (6%) or Pla beverages during the weight training bout. Blood, saliva, and vastus lateralis muscle biopsy samples were collected before and after exercise. Blood cell counts were determined, and plasma was analyzed for IL-6, IL-10, IL-1 receptor antagonist (IL-1ra), IL-8, and cortisol. Muscle was analyzed for glycogen content and relative gene expression of 13 cytokines (IL-1alpha, IL-1beta, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p35, IL-12p40, IL-15, IFN-gamma, TNF-alpha) by use of real-time quantitative RT-PCR. Significant but modest increases were measured for plasma IL-6, IL-10, IL-1ra, and IL-8, but the pattern of increase did not differ between CHO and Pla groups. The rate of decrease in muscle glycogen content did not differ between CHO and Pla (P = 0.463). Muscle cytokine mRNA was detected preexercise for IL-1beta, IL-6, IL-15, IL-8, and TNF-alpha, and of these, IL-1beta, IL-6, IL-8, and TNF-alpha were significantly increased after the 2-h weight training bout. The increase in mRNA (fold difference from preexercise) did not differ between CHO and Pla groups. In summary, CHO vs. Pla ingestion did not alter modest increases measured for plasma IL-6, IL-10, IL-1ra, and IL-8, and muscle gene expression for IL-1beta, IL-6, IL-8, and TNF-alpha in strength-trained subjects lifting weights intensively for 2 h.
Assuntos
Carboidratos/administração & dosagem , Sistema Imunitário/efeitos dos fármacos , Sistema Imunitário/fisiologia , Resistência Física , Levantamento de Peso/fisiologia , Administração Oral , Adulto , Contagem de Células Sanguíneas , Citocinas/sangue , Citocinas/genética , Citocinas/metabolismo , Expressão Gênica , Glicogênio/antagonistas & inibidores , Humanos , Hidrocortisona/sangue , Masculino , Músculo Esquelético/metabolismo , RNA Mensageiro/metabolismo , Fatores de TempoRESUMO
S mutans strain MT703 from an active carious lesion in the tooth of a child had type e specificity and showed a cross-reaction with the Lancefield group E cell wall streptococcal polysaccharide antigen. Heat-killed cells MT703 adhered to a glass surface in the presence of CGT MT703 and sucrose. Pretreatment of the cells with anti-MT703 whole cell serums inhibited adherecne. The removal of glycerol teichoic acid antibody and group E antibody from the MT703 serum did not result in a loss of inhibitory activity. Antiserum with or without adsorption significantly inhibited glucan synthesis by CGT from sucrose. Antibodies specific for the polyglycerol phosphate of teichoic acid did not inhibit adherence. Anti-group E serum and serums specific for other types of S mutans, did not show adherence inhibitory activity except for an occasional type c specific antiserum. Antibody specific for the type e antigen produced significant inhibition of the binding of CGT to the MT703 cell wall, and adherence of these cells did not occur. Antibody to CGT inhibited glucan synthesis. Treatment of the cells with dextranase, dextran antibody, or trypsin caused a significant reduction in adherence. The results suggest that the type antigen and dextran on the surface of the S mutans type e cell are functional in adherence, and that these polymers are associated with cell wall protein.
Assuntos
Soros Imunes/farmacologia , Streptococcus mutans/patogenicidade , Streptococcus/patogenicidade , Glucose/análogos & derivados , Glucosiltransferases/metabolismo , Glucosiltransferases/farmacologia , Glicogênio/antagonistas & inibidores , Concentração de Íons de Hidrogênio , Polissacarídeos Bacterianos/biossíntese , Sorotipagem , Especificidade da Espécie , Streptococcus mutans/classificação , Streptococcus mutans/enzimologia , Streptococcus mutans/imunologia , Streptococcus mutans/metabolismoRESUMO
Loss of muscle mass related to anti-cancer therapy is a major concern in cancer patients, being associated with important clinical endpoints including survival, treatment toxicity and patient-related outcomes. We investigated effects of voluntary exercise during cisplatin treatment on body weight, food intake as well as muscle mass, strength and signalling. Mice were treated weekly with 4 mg/kg cisplatin or saline for 6 weeks, and randomized to voluntary wheel running or not. Cisplatin treatment induced loss of body weight (29.8%, P < 0.001), lean body mass (20.6%, P = 0.001), as well as anorexia, impaired muscle strength (22.5% decrease, P < 0.001) and decreased glucose tolerance. In addition, cisplatin impaired Akt-signalling, induced genes related to protein degradation and inflammation, and reduced muscle glycogen content. Voluntary wheel running during treatment attenuated body weight loss by 50% (P < 0.001), maintained lean body mass (P < 0.001) and muscle strength (P < 0.001), reversed anorexia and impairments in Akt and protein degradation signalling. Cisplatin-induced muscular inflammation was not prevented by voluntary wheel running, nor was glucose tolerance improved. Exercise training may preserve muscle mass in cancer patients receiving cisplatin treatment, potentially improving physical capacity, quality of life and overall survival.
Assuntos
Anorexia/prevenção & controle , Cisplatino/farmacologia , Músculo Esquelético/efeitos dos fármacos , Atrofia Muscular/prevenção & controle , Condicionamento Físico Animal , Animais , Anorexia/induzido quimicamente , Anorexia/metabolismo , Anorexia/fisiopatologia , Peso Corporal/efeitos dos fármacos , Feminino , Expressão Gênica , Intolerância à Glucose/induzido quimicamente , Intolerância à Glucose/metabolismo , Intolerância à Glucose/fisiopatologia , Glicogênio/antagonistas & inibidores , Glicogênio/biossíntese , Camundongos , Força Muscular/efeitos dos fármacos , Força Muscular/fisiologia , Músculo Esquelético/fisiologia , Atrofia Muscular/induzido quimicamente , Atrofia Muscular/metabolismo , Atrofia Muscular/fisiopatologia , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Corrida/fisiologia , Transdução de SinaisRESUMO
Apelin, a novel adipokine, is the specific endogenous ligand of G protein-coupled receptor APJ. Consistent with its putative role as an adipokine, apelin has been linked to states of insulin resistance. However, the function of apelin in hepatic insulin resistance, a vital part of insulin resistance, and its underlying mechanisms still remains unclear. Here we define the impacts of apelin on TNF-α-induced reduction of glycogen synthesis in the hepatocytes. Our studies indicate that apelin reversed TNF-α-induced reduction of glycogen synthesis in HepG2 cells, mouse primary hepatocytes and liver tissues of C57BL/6J mice by improving JNK-IRS1-AKT-GSK pathway. Moreover, Western blot revealed that APJ, but not apelin, expressed in the hepatocytes and liver tissues of mice. We found that F13A, a competitive antagonist for G protein-coupled receptor APJ, suppressed the effects of apelin on TNF-α-induced reduction of glycogen synthesis in the hepatocytes, suggesting APJ is involved in the function of apelin. In conclusion, we show novel evidence suggesting that apelin ameliorates TNF-α-induced reduction of glycogen synthesis in the hepatocytes through G protein-coupled receptor APJ. Apelin appears as a beneficial adipokine with anti-insulin resistance properties, and thus as a promising therapeutic target in metabolic disorders.
Assuntos
Glicogênio/biossíntese , Hepatócitos/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Fígado/efeitos dos fármacos , Receptores Acoplados a Proteínas G/genética , Adipocinas , Animais , Apelina , Receptores de Apelina , Regulação da Expressão Gênica/efeitos dos fármacos , Glicogênio/agonistas , Glicogênio/antagonistas & inibidores , Células Hep G2 , Hepatócitos/citologia , Hepatócitos/metabolismo , Humanos , Resistência à Insulina/genética , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Fígado/citologia , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Cultura Primária de Células , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Receptores Acoplados a Proteínas G/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/farmacologiaAssuntos
Glucose/metabolismo , Insulina/farmacologia , Lítio/farmacologia , Músculos/metabolismo , Potássio/farmacologia , Rubídio/farmacologia , Sódio/farmacologia , Animais , Diafragma/metabolismo , Interações Medicamentosas , Glucose/análise , Glicogênio/antagonistas & inibidores , Glicogênio/metabolismo , Técnicas In Vitro , Cinética , Lactatos/biossíntese , Masculino , Concentração Osmolar , Ratos , Compostos de Sulfidrila/farmacologia , Fatores de TempoAssuntos
Fígado/citologia , Animais , Nucléolo Celular/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Doença Crônica , Citoplasma/efeitos dos fármacos , Retículo Endoplasmático/efeitos dos fármacos , Fluorenos/toxicidade , Glicogênio/antagonistas & inibidores , Complexo de Golgi/efeitos dos fármacos , Fígado/efeitos dos fármacos , Masculino , Microscopia Eletrônica , Mitocôndrias/efeitos dos fármacos , Mitocôndrias Hepáticas/efeitos dos fármacos , Ratos , Ratos EndogâmicosRESUMO
Sry is transiently activated in pre-Sertoli cells of the gonadal ridge to initiate testis differentiation in mice. In pre-Sertoli cells, however, the cellular events induced immediately after the onset of Sry expression remain largely unknown. Here we show that testis-specific glycogen accumulation in pre-Sertoli cells is one of the earliest cellular events downstream of Sry action. In developing XY gonads, glycogen accumulation starts to occur in pre-Sertoli cells from around 11.15 dpc (tail somite 14 stage) in a center-to-pole pattern similar to the initial Sry expression profile. Glycogen accumulation was also found in XX male gonads of Sry-transgenic embryos, but not in XX female gonads of wildtype embryos at any developmental stage. In vitro analyses using various culture conditions suggest that testis-specific glycogen deposition is a tissue-autonomous event that can be induced even in serum-free conditions and in a culture of gonadal explants without adjacent mesonephros. Moreover, glycogen accumulation in pre-Sertoli cells was significantly inhibited in vitro by the PI3K inhibitor LY294002, but not by the MEK inhibitor PD98059. Active phospho-AKT (PI3K effector) showed a high degree of accumulation in gonadal somatic cells of genital ridges in a testis-specific manner, both in vitro and in vivo. Therefore, these findings suggest that immediately after the onset of Sry expression, activation of the PI3K-AKT pathway promotes testis-specific glycogen storage in pre-Sertoli cells. To the best of our knowledge, this is a novel Sry-downstream cellular event which preserves this readily available energy source in Sertoli cells for testis-specific morphogenesis and hormone production.
Assuntos
Fenômenos Fisiológicos Celulares/efeitos dos fármacos , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Metabolismo Energético/fisiologia , Glicogênio/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Diferenciação Sexual , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Animais , Bussulfano/farmacologia , Cromonas/farmacologia , Proteínas de Ligação a DNA/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Feminino , Glicogênio/antagonistas & inibidores , Injeções Intraperitoneais , Masculino , Camundongos , Camundongos Endogâmicos ICR , Camundongos Transgênicos , Morfolinas/farmacologia , Proteínas Nucleares/efeitos dos fármacos , Técnicas de Cultura de Órgãos/métodos , Especificidade de Órgãos/fisiologia , Células de Sertoli/metabolismo , Caracteres Sexuais , Análise para Determinação do Sexo , Diferenciação Sexual/genética , Proteína da Região Y Determinante do Sexo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Testículo/embriologia , Testículo/metabolismo , Testículo/ultraestrutura , Fatores de Transcrição/efeitos dos fármacosRESUMO
Improved methodology was used to establish that the phosphorylation of a serine located 10 residues from the N-terminus of glycogen synthase (N10) increases from 0.12 mol.mol-1 to 0.54 mol.mol-1 in vivo in response to adrenalin. The only 'N10 kinase' detected in muscle extracts was casein kinase-1 (CK1), although its activity was unaffected by injection of adrenalin in vivo or by incubation with cyclic-AMP-dependent protein kinase and MgATP in vitro. Prior phosphorylation of the serine residue N7 by phosphorylase kinase increased sixfold the rate of phosphorylation of glycogen synthase by CK1, and altered the specificity of CK1 so that it phosphorylated the serine residue N10 specifically. Stoichiometric phosphorylation of N7 decreased the activity ratio (+/- glucose 6-phosphate) of glycogen synthase from 0.80 to 0.45, and subsequent phosphorylation of N10 to 0.8 mol.mol-1 produced a further decrease to 0.17, demonstrating that N10 phosphorylation inhibits glycogen synthase. The major 'N10 phosphatase' in skeletal muscle extracts was identified as the glycogen-associated form of protein phosphatase-1 (PP1G), accounting for approximately 75% of the N10 phosphatase activity in the extracts and about 90% of the activity in isolated glycogen particles. Phosphorylation of N10, after prior phosphorylation of N7, decreased the rate of dephosphorylation of N7. These results, in conjunction with previous findings, establish that adrenalin inhibits glycogen synthase by increasing the phosphorylation of N7, N10 and three further serines located 30, 34 and 38 residues from the start of the C-terminal CNBr peptide (termed the region C30-C38). They also indicate that increased phosphorylation of N10, the region C30-C38, and perhaps N7, is initiated through the inhibition of PP1G by adrenalin, which results from phosphorylation of its glycogen-targetting subunit by cyclic-AMP-dependent protein kinase [Hubbard, M.J. & Cohen, P. (1989) Eur. J. Biochem. 186, 711-716]. The conclusion that direct phosphorylation of glycogen synthase by cyclic-AMP-dependent protein kinase makes little contribution to inhibition by adrenalin, is at variance with the teachings of the major textbooks of biochemistry.