RESUMO
BACKGROUND: Sepsis induces loss of skeletal muscle mass by activating the ubiquitin proteasome (UPS) and autophagy systems. Although muscle protein synthesis in healthy neonatal piglets is responsive to amino acids (AA) stimulation, it is not known if AA can prevent the activation of muscle protein degradation induced by sepsis. We hypothesize that AA attenuate the sepsis-induced activation of UPS and autophagy in neonates. METHODS: Newborn pigs were infused for 8 h with liposaccharide (LPS) (0 and 10 µg·kg(-1)·h(-1)), while circulating glucose and insulin were maintained at fasting levels; circulating AA were clamped at fasting or fed levels. Markers of protein degradation and AA transporters in longissimus dorsi (LD) were examined. RESULTS: Fasting AA increased muscle microtubule-associated protein light 1 chain 3 II (LC3-II) abundance in LPS compared to control, while fed AA levels decreased LC3-II abundance in both LPS and controls. There was no effect of AA supplementation on activated protein kinase (AMP), forkhead box O1 and O4 phosphorylation, nor on sodium-coupled neutral AA transporter 2 and light chain AA transporter 1, muscle RING-finger protein-1 and muscle Atrophy F-Box/Atrogin-1 abundance. CONCLUSION: These findings suggest that supplementation of AA antagonize autophagy signal activation in skeletal muscle of neonates during endotoxemia.
Assuntos
Aminoácidos/sangue , Autofagia/efeitos dos fármacos , Endotoxemia/fisiopatologia , Insulina/sangue , Músculo Esquelético/patologia , Aminoácidos de Cadeia Ramificada/sangue , Animais , Animais Recém-Nascidos , Glicemia/análise , Nitrogênio da Ureia Sanguínea , Endotoxemia/sangue , Modelos Biológicos , Complexo de Endopeptidases do Proteassoma/metabolismo , Sepse/fisiopatologia , Sus scrofa , Suínos , TemperaturaRESUMO
Neonatal pigs are used as a model to study and optimize the clinical treatment of infants who are unable to maintain oral feeding. Using this model, we have shown previously that pulsatile administration of leucine during continuous feeding over 24 h via orogastric tube enhanced protein synthesis in skeletal muscle compared with continuous feeding alone. To determine the long-term effects of leucine pulses, neonatal piglets (n = 11-12/group) were continuously fed formula via orogastric tube for 21 days, with an additional parenteral infusion of either leucine (CON + LEU; 800 µmol·kg-1·h-1) or alanine (CON + ALA) for 1 h every 4 h. The results show that body and muscle weights and lean gain were â¼25% greater, and fat gain was 48% lower in CON + LEU than CON + ALA; weights of other tissues were unaffected by treatment. Fractional protein synthesis rates in longissimus dorsi, gastrocnemius, and soleus muscles were â¼30% higher in CON + LEU compared with CON + ALA and were associated with decreased Deptor abundance and increased mTORC1, mTORC2, 4E-BP1, and S6K1 phosphorylation, SNAT2 abundance, and association of eIF4E with eIF4G and RagC with mTOR. There were no treatment effects on PKB, eIF2α, eEF2, or PRAS40 phosphorylation, Rheb, SLC38A9, v-ATPase, LAMTOR1, LAMTOR2, RagA, RagC, and LAT1 abundance, the proportion of polysomes to nonpolysomes, or the proportion of mRNAs encoding rpS4 or rpS8 associated with polysomes. Our results demonstrate that pulsatile delivery of a leucine supplement during 21 days of continuous enteral feeding enhances lean growth by stimulating the mTORC1-dependent translation initiation pathway, leading to protein synthesis in skeletal muscle of neonates.
Assuntos
Leucina/farmacologia , Proteínas Musculares/efeitos dos fármacos , Músculo Esquelético/efeitos dos fármacos , Biossíntese de Proteínas/efeitos dos fármacos , Alanina/farmacologia , Sistema A de Transporte de Aminoácidos/efeitos dos fármacos , Sistema A de Transporte de Aminoácidos/metabolismo , Animais , Animais Recém-Nascidos , Músculos do Dorso , Suplementos Nutricionais , Nutrição Enteral , Infusões Parenterais , Leucina/administração & dosagem , Alvo Mecanístico do Complexo 1 de Rapamicina , Alvo Mecanístico do Complexo 2 de Rapamicina , Complexos Multiproteicos/efeitos dos fármacos , Complexos Multiproteicos/metabolismo , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Fosforilação/efeitos dos fármacos , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Proteínas Quinases S6 Ribossômicas 90-kDa/efeitos dos fármacos , Proteínas Quinases S6 Ribossômicas 90-kDa/metabolismo , Proteínas Ribossômicas/efeitos dos fármacos , Proteínas Ribossômicas/genética , Sus scrofa , Suínos , Serina-Treonina Quinases TOR/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismoRESUMO
INTRODUCTION: Leucine (Leu) activates mammalian target of rapamycin (mTOR) to upregulate protein synthesis (PS). RESULTS: PS in skeletal muscles, heart, liver, pancreas, and jejunum, but not kidney, were greater in low protein supplemented with Leu (LP+L) than LP, but lower than high protein (HP). In longissimus dorsi muscle, protein kinase B phosphorylation was similar in LP and LP+L, but lower than HP. Although less than HP, p70 ribosomal S6 kinase 1 (S6K1) and eukaryotic initiation factor (eIF) 4E binding protein 1 (4EBP1) association with regulatory associated protein of mammalian target of rapamycin was greater in LP+L than LP, resulting in higher S6K1 and 4EBP1 phosphorylation. Feeding LP+L vs. LP decreased 4EBP1·eIF4E and increased eIF4E·eIF4G formation, but not to HP. Similar results were obtained for S6K1 and 4EBP1 phosphorylation in gastrocnemius, masseter, heart, liver, pancreas, and jejunum, but not kidney. eIF2α and elongation factor 2 phosphorylation was unaffected by treatment. DICUSSION: Our results suggest that enteral Leu supplementation of a low protein diet enhances PS in most tissues through mTOR complex 1 pathways. METHODS: To examine enteral Leu effects on PS and signaling activation, 5-d-old piglets were fed for 24 h diets containing: (i) LP, (ii) LP+L, or (iii) HP.
Assuntos
Leucina/uso terapêutico , Músculo Esquelético/metabolismo , Miocárdio/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Aminoácidos/metabolismo , Animais , Animais Recém-Nascidos , Glicemia/metabolismo , Suplementos Nutricionais , Nutrição Enteral/métodos , Fator de Iniciação 4E em Eucariotos/química , Fator de Iniciação Eucariótico 4G/química , Fatores de Iniciação em Eucariotos/química , Glicólise , Insulina/sangue , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Suínos , Fatores de Tempo , Distribuição TecidualRESUMO
Protein synthesis and eukaryotic initiation factor (eIF) activation are increased in skeletal muscle of neonatal pigs parenterally infused with amino acids. Leucine appears to be the most effective single amino acid to trigger these effects. To examine the response to enteral leucine supplementation, overnight food-deprived 5-d-old pigs were gavage fed at 0 and 60 min a: 1) low-protein diet (LP); 2) LP supplemented with leucine (LP+L) to equal leucine in the high-protein diet (HP); or 3) HP diet. Diets were isocaloric and equal in lactose. Fractional protein synthesis rates and translation initiation control mechanisms were examined in skeletal muscles and visceral tissues 90 min after feeding. Protein synthesis rates in longissimus dorsi, gastrocnemius, and masseter muscles, heart, jejunum, kidney, and pancreas, but not liver, were greater in the LP+L group compared with the LP group and did not differ from the HP group. Feeding LP+L and HP diets compared with the LP diet increased phosphorylation of mammalian target of rapamycin (mTOR), 4E-binding protein 1, ribosomal protein S6 kinase-1, and eIF4G and formation of the active eIF4E·eIF4G complex in longissimus dorsi muscle. In all tissues except liver, activation of mTOR effectors increased in pigs fed LP+L and HP vs. LP diets. Our results suggest that leucine supplementation of a low-protein meal stimulates protein synthesis in muscle and most visceral tissues to a rate similar to that achieved by feeding a high-protein meal and this stimulation involves activation of mTOR downstream effectors.