Navegando por Autor "Kettelhut, Isis do Carmo"
Agora exibindo 1 - 8 de 8
Resultados por página
Opções de Ordenação
Item Calcitonin gene-related peptide exerts inhibitory effects on autophagy in the heart of mice.(2021) Schavinski, Aline Zanatta; Machado, Juliano; Morgan, Henrique Jorge Novaes; Silva, Natalia Lautherbach Ennes da; Gomes, Sílvia de Paula; Kettelhut, Isis do Carmo; Navegantes, Luiz Carlos CarvalhoCalcitonin Gene-Related Peptide (CGRP) is a potent vasodilator peptide widely distributed in the central nervous system and various peripheral tissues, including cardiac muscle. However, its role in heart protein metabolism remains unknown. We examined the acute effects of CGRP on autophagy and the related signaling pathways in the heart mice and cultured neonatal cardiomyocytes. CGRP (100 μg kg− 1 ; s.c.) or 0.9 % saline was injected in awake male C57B16 mice, and the metabolic profile was determined up to 60 min. In fed mice, CGRP drastically increased glycemia and reduced insulinemia, an effect that was accompanied by reduced cardiac phosphoryla- tion levels of Akt at Ser473 without affecting FoxO. Despite these catabolic effects, CGRP acutely inhibited autophagy as estimated by the decrease in LC3II:LC3I and autophagic flux. In addition, the fasting-induced autophagic flux in mice hearts was entirely abrogated by one single injection of CGRP. In parallel, CGRP stimulated PKA/CREB and mTORC1 signaling and increased the phosphorylation of Unc51-like kinase-1 (ULK1), an essential protein in autophagy initiation. Similar effects were observed in cardiomyocytes, in which CGRP also inhibited autophagic flux and stimulated Akt and FoxO phosphorylation. These findings suggest that CGRP in vivo acutely suppresses autophagy in the heart of fed and fasted mice, most likely through the activation of PKA/ mTORC1 signaling but independent of Akt.Item CAMP-dependent protein kinase inhibits FoxO activity and regulates skeletal muscle plasticity in mice.(2020) Silveira, Wilian de Assis; Gonçalves, Dawit Alberto Pinheiro; Machado, Juliano; Silva, Natalia Lautherbach Ennes da; Borges, Danilo Lustrino; Gomes, Sílvia de Paula; Pereira, Marcelo G.; Miyabara, Elen Haruka; Sandri, Marco; Kettelhut, Isis do Carmo; Navegantes, Luiz Carlos CarvalhoAlthough we have shown that catecholamines suppress the activity of the Ubiquitin- Proteasome System (UPS) and atrophy-related genes expression through a cAMP-de- pendent manner in skeletal muscle from rodents, the underlying mechanisms remain unclear. Here, we report that a single injection of norepinephrine (NE; 1 mg kg−1; s.c) attenuated the fasting-induced up-regulation of FoxO-target genes in tibialis anterior (TA) muscles by the stimulation of PKA/CREB and Akt/FoxO1 signaling pathways. In addition, muscle-specific activation of PKA by the overexpression of PKA catalytic subunit (PKAcat) suppressed FoxO reporter activity induced by (1) a wild-type; (2) a non-phosphorylatable; (3) a non-phosphorylatable and non-acet- ylatable forms of FoxO1 and FoxO3; (4) downregulation of FoxO protein content, and probably by (5) PGC-1α up-regulation. Consistently, the overexpression of the PKAcat inhibitor (PKI) up-regulated FoxO activity and the content of Atrogin-1 and MuRF1, as well as induced muscle fiber atrophy, the latter effect being prevented by the overexpression of a dominant negative (d. n.) form of FoxO (d.n.FoxO). The sustained overexpression of PKAcat induced fiber-type transition toward a smaller, slower, and more oxidative phenotype and improved muscle resistance to fatigue. Taken together, our data provide the first evidence that endogenous PKA activity is required to restrain the basal activity of FoxO and physiologically important to maintain skeletal muscle mass.Item Expression of glycerokinase in brown adipose tissue is stimulated by the sympathetic nervose system.(2003) Festuccia, William Tadeu Lara; Cota, Renata Guerra de Sá; Kawashita, Nair Honda; Garófalo, Maria Antonieta Rissato; Evangelista, Elísio Alberto; Rodrigues, Vanderlei; Kettelhut, Isis do Carmo; Migliorini, Renato HéliosExpression of glycerokinase in brown adipose tissue is stimulated by the sympathetic nervous system. Am J Physiol Regul Integr Comp Physiol 284: R1536–R1541, 2003; 10.1152/ajpregu.00764.2002.—The effect of cold exposure (4°C) or prolonged norepinephrine infusion on the activity and mRNA levels of glycerokinase (GyK) was investigated in rat interscapular brown adipose tissue (BAT). Cold exposure for 12 and 24 h induced increases of 30% and 100%, respectively, in the activity of BAT GyK, which was paralleled by twofold and fourfold increase in enzyme mRNA levels. BAT hemidenervation resulted in reductions of 50% and 30% in GyK activity and in mRNA levels, respectively, in denervated pads from rats kept at 25°C, and suppressed in these pads the cold-induced increases in both GyK activity and mRNA levels. The increase in GyK activity induced by cold exposure was not affected by phenoxybenzamine, but was markedly inhibited by previous administration of propranolol or actinomycin D. BAT GyK activity did not change significantly after 6 h of continuous subcutaneous infusion of norepinephrine (20 g/h), but increased twofold and fourfold after 12 and 24 h, with no further increase after 72 h of infusion. Norepinephrine infusion also activated mRNA production, but the effect was comparatively smaller than that on enzyme activity. -Adrenergic agonists also stimulated GyK activity with the following relative magnitude of response: CL316243 ( 3) isoproterenol (non-selective) dobutamine ( 1). In vitro rates of incorporation of glycerol into glyceride-glycerol were increased in BAT from rats exposed to cold. The data suggest that in conditions of a sustained increase in BAT sympathetic flow there is a stimulation of GyK gene expression at the pretranslational level, with increased enzyme activity, mediated by -adrenoreceptors, mainly 3.Item Phosphodiesterase 4 inhibition restrains muscle proteolysis in diabetic rats by activating PKA and EPAC/Akt effectors and inhibiting FoxO factors.(2021) Arcaro, Carlos Alberto; Assis, Renata Pires; Oliveira, Juliana Oriel; Zanon, Neusa Maria; Gomes, Sílvia de Paula; Navegantes, Luiz Carlos Carvalho; Kettelhut, Isis do Carmo; Brunett, Iguatemy Lourenço; Baviera, Amanda MartinsAim: There is growing evidence about the ability of cyclic adenosine monophosphate (cAMP) signaling and nonselective phosphodiesterase (PDE) inhibitors on mitigate muscle atrophy. PDE4 accounts for the major cAMP hydrolyzing activity in skeletal muscles, therefore advances are necessary about the consequences of treatment with PDE4 inhibitors on protein breakdown in atrophied muscles. We postulated that rolipram (selective PDE4 inhibitor) may activate cAMP downstream effectors, inhibiting proteolytic systems in skeletal muscles of diabetic rats. Main methods: Streptozotocin-induced diabetic rats were treated with 2 mg/kg rolipram for 3 days. Changes in the levels of components belonging to the proteolytic machineries in soleus and extensor digitorum longus (EDL) muscles were investigated, as well as cAMP effectors. Key findings: Treatment of diabetic rats with rolipram decreased the levels of atrogin-1 and MuRF-1 in soleus and EDL, and reduced the activities of calpains and caspase-3; these findings partially explains the low ubiquitin conjugates levels and the decreased proteasome activity. The inhibition of muscle proteolysis may be occurring due to phosphorylation and inhibition of forkhead box O (FoxO) factors, probably as a consequence of the increased cAMP levels, followed by the activation of PKA and Akt effectors. Akt activation may be associated with the increased levels of exchange protein directly activated by cAMP (EPAC). As a result, rolipram treatment spared muscle mass in diabetic rats. Significance: The antiproteolytic responses associated with PDE4 inhibition may be helpful to motivate future investigations about the repositioning of PDE4 inhibitors for the treatment of muscle wasting conditions.Item Role of ubiquitin-proteasome-dependent proteolytic process in degradation of muscle proteic from diabetic rabbits.(2001) Galban, Victor Diaz; Evangelista, Elísio Alberto; Migliorini, Renato Hélios; Kettelhut, Isis do CarmoThe activity of ATP, ubiquitin (Ub)-dependent proteases partially purified from skeletal muscle (psoas) from alloxan diabetic rabbits was determined at different periods of insulin deficiency. Two days after alloxan injection, no change was observed in the activity of ATP, Ub-dependent proteases, but this activity increased 3 and 5 days after diabetes induction, attaining 181% of control values on the 5th day. However, after this early rise, the activity of muscle ATP, Ub-dependent proteases decreased, returning to values that did not differ significantly from controls 7 and 10 days after alloxan injection. After 15 days, the activity of these proteases was 57% lower than in muscle from control rabbits. Both the initial increase and the subsequent fall in the activity of the enzymes were prevented by insulin treatment of alloxan diabetic rabbits. The data suggest that Ub-proteasomedependent proteolysis have an important role in the control of muscle protein degradation and may be regulated by insulin.Item Schistosoma mansoni : functional proteasomes are required for development in the vertebrate host.(2005) Cota, Renata Guerra de Sá; Borges, William de Castro; Evangelista, Elísio Alberto; Kettelhut, Isis do Carmo; Rodrigues, VanderleiProteasomes are multi-subunit proteases involved in several mechanisms and thought to contribute to the regulation of cellular homeostasis. Here, we report for the Wrst time biochemical evidence for the existence of a ubiquitin–proteasome proteolytic pathway in this parasite. Proteasomes from both cercariae and adult worms exhibited a high preference for hydrolysis of the substrate Suc- LLVY-AMC, although in the cercariae extract the rate of hydrolysis was 50% lower when compared to adult worms extracts. The same diVerence in proteasome activities was observed when endogenous proteins were broken down in the presence of ATP and ubiquitin. Additionally, accumulation of high molecular weight conjugates was observed when cercariae were pre-incubated with proteasome inhibitors. Finally, we present evidence that during experimental schistosomiasis, proteasome inhibitors were able to reduce the number of lung stage schistosomula, reduce the worm burden and consequently decrease the egg output in infected mice.Item Sympathetic innervation suppresses the autophagic-lysosomal system in brown adipose tissue under basal and cold-stimulated conditions.(2020) Mioranza, Franciele Przygodda; Silva, Natalia Lautherbach Ennes da; Buzelle, Samyra Lopes; Gonçalves, Dawit Alberto Pinheiro; Assis, Ana Paula; Gomes, Sílvia de Paula; Garófalo, Maria Antonieta Rissato; Heck, Lilian Carmo; Matsuo, Flávia Sayuri; Mota, Ryerson Fonseca; Osako, Mariana Kiomy; Kettelhut, Isis do Carmo; Navegantes, Luiz Carlos CarvalhoThe sympathetic nervous system (SNS) activates cAMP signaling and promotes trophic effects on brown adipose tissue (BAT) through poorly understood mechanisms. Because norepinephrine has been found to induce antiproteolytic effects on muscle and heart, we hypothesized that the SNS could inhibit autophagy in interscapular BAT (IBAT). Here, we describe that selective sympathetic denervation of rat IBAT kept at 25°C induced atrophy, and in parallel dephosphorylated forkhead box class O (FoxO), and increased cathepsin activity, autophagic flux, autophagosome formation, and expression of autophagy-related genes. Conversely, cold stimulus (4°C) for up to 72 h induced thermogenesis and IBAT hypertrophy, an anabolic effect that was associated with inhibition of cathepsin activity, autophagic flux, and autophagosome formation. These effects were abrogated by sympathetic denervation, which also upregulated Gabarapl1 mRNA. In addition, the cold-driven sympathetic activation stimulated the mechanistic target of rapamycin (mTOR) pathway, leading to the enhancement of protein synthesis, evaluated in vivo by puromycin incorporation, and to the inhibitory phosphorylation of Unc51-like kinase-1, a key protein in the initiation of autophagy. This coincided with a higher content of exchange protein-1 directly activated by cAMP (Epac1), a cAMP effector, and phosphorylation of Akt at Thr308, all these effects being abolished by denervation. Systemic treatment with norepinephrine for 72 h mimicked most of the cold effects on IBAT. These data suggest that the noradrenergic sympathetic inputs to IBAT restrain basal autophagy via suppression of FoxO and, in the setting of cold, stimulate protein synthesis via the Epac/Akt/mTOR-dependent pathway and suppress the autophagosome formation, probably through posttranscriptional mechanisms.Item Urocortin 2 promotes hypertrophy and enhances skeletal muscle function through cAMP and insulin/IGF-1 signaling pathways.(2022) Silva, Natalia Lautherbach Ennes da; Gonçalves, Dawit Alberto Pinheiro; Silveira, Wilian de Assis; Gomes, Sílvia de Paula; Valentim, Rafael Rossi; Zanon, Neusa Maria; Pereira, Marcelo G.; Miyabara, Elen Haruka; Navegantes, Luiz Carlos Carvalho; Kettelhut, Isis do CarmoObjective: Although it is well established that urocortin 2 (Ucn2), a peptide member of the corticotrophin releasing factor (CRF) family, and its specific corticotrophin-releasing factor 2 receptor (CRF2R) are highly expressed in skeletal muscle, the role of this peptide in the regulation of skeletal muscle mass and protein metabolism remains elusive. Methods: To elucidate the mechanisms how Ucn2 directly controls protein metabolism in skeletal muscles of normal mice, we carried out genetic tools, physiological and molecular analyses of muscles in vivo and in vitro. Results: Here, we demonstrated that Ucn2 overexpression activated cAMP signaling and promoted an expressive muscle hypertrophy associated with higher rates of protein synthesis and activation of Akt/mTOR and ERK1/2 signaling pathways. Furthermore, Ucn2 induced a decrease in mRNA levels of atrogin-1 and in autophagic flux inferred by an increase in the protein content of LC3-I, LC3-II and p62. Accordingly, Ucn2 reduced both the transcriptional activity of FoxO in vivo and the overall protein degradation in vitro through an inhibition of lysosomal proteolytic activity. In addition, we demonstrated that Ucn2 induced a fast-to-slow fiber type shift and improved fatigue muscle resistance, an effect that was completely blocked in muscles co-transfected with mitogen-activated protein kinase phosphatase 1 (MKP-1), but not with dominant-negative Akt mutant (Aktmt). Conclusions: These data suggest that Ucn2 triggers an anabolic and anti-catabolic response in skeletal muscle of normal mice probably through the activation of cAMP cascade and participation of Akt and ERK1/2 signaling. These findings open new perspectives in the development of therapeutic strategies to cope with the loss of muscle mass.