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1.
Nat Commun ; 14(1): 4812, 2023 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-37558654

RESUMO

Branched chain amino acid (BCAA) catabolic impairments have been implicated in several diseases. Branched chain ketoacid dehydrogenase (BCKDH) controls the rate limiting step in BCAA degradation, the activity of which is inhibited by BCKDH kinase (BDK)-mediated phosphorylation. Screening efforts to discover BDK inhibitors led to identification of thiophene PF-07208254, which improved cardiometabolic endpoints in mice. Structure-activity relationship studies led to identification of a thiazole series of BDK inhibitors; however, these inhibitors did not improve metabolism in mice upon chronic administration. While the thiophenes demonstrated sustained branched chain ketoacid (BCKA) lowering and reduced BDK protein levels, the thiazoles increased BCKAs and BDK protein levels. Thiazoles increased BDK proximity to BCKDH-E2, whereas thiophenes reduced BDK proximity to BCKDH-E2, which may promote BDK degradation. Thus, we describe two BDK inhibitor series that possess differing attributes regarding BDK degradation or stabilization and provide a mechanistic understanding of the desirable features of an effective BDK inhibitor.


Assuntos
Aminoácidos de Cadeia Ramificada , Tiofenos , Camundongos , Animais , Aminoácidos de Cadeia Ramificada/metabolismo , Fosforilação , Tiofenos/farmacologia , Oxirredutases/metabolismo
2.
Mol Metab ; 66: 101611, 2022 12.
Artigo em Inglês | MEDLINE | ID: mdl-36220546

RESUMO

OBJECTIVE: Branched chain amino acid (BCAA) catabolic defects are implicated to be causal determinates of multiple diseases. This work aimed to better understand how enhancing BCAA catabolism affected metabolic homeostasis as well as the mechanisms underlying these improvements. METHODS: The rate limiting step of BCAA catabolism is the irreversible decarboxylation by the branched chain ketoacid dehydrogenase (BCKDH) enzyme complex, which is post-translationally controlled through phosphorylation by BCKDH kinase (BDK). This study utilized BT2, a small molecule allosteric inhibitor of BDK, in multiple mouse models of metabolic dysfunction and NAFLD including the high fat diet (HFD) model with acute and chronic treatment paradigms, the choline deficient and methionine minimal high fat diet (CDAHFD) model, and the low-density lipoprotein receptor null mouse model (Ldlr-/-). shRNA was additionally used to knock down BDK in liver to elucidate liver-specific effects of BDK inhibition in HFD-fed mice. RESULTS: A rapid improvement in insulin sensitivity was observed in HFD-fed and lean mice after BT2 treatment. Resistance to steatosis was assessed in HFD-fed mice, CDAHFD-fed mice, and Ldlr-/- mice. In all cases, BT2 treatment reduced steatosis and/or inflammation. Fasting and refeeding demonstrated a lack of response to feeding-induced changes in plasma metabolites including insulin and beta-hydroxybutyrate and hepatic gene changes in BT2-treated mice. Mechanistically, BT2 treatment acutely altered the expression of genes involved in fatty acid oxidation and lipogenesis in liver, and upstream regulator analysis suggested that BT2 treatment activated PPARα. However, BT2 did not directly activate PPARα in vitro. Conversely, shRNA-AAV-mediated knockdown of BDK specifically in liver in vivo did not demonstrate any effects on glycemia, steatosis, or PPARα-mediated gene expression in mice. CONCLUSIONS: These data suggest that BT2 treatment acutely improves metabolism and liver steatosis in multiple mouse models. While many molecular changes occur in liver in BT2-treated mice, these changes were not observed in mice with AAV-mediated shRNA knockdown of BDK. All together, these data suggest that systemic BDK inhibition is required to improve metabolism and steatosis by prolonging a fasting signature in a paracrine manner. Therefore, BCAA may act as a "fed signal" to promote nutrient storage and reduced systemic BCAA levels as shown in this study via BDK inhibition may act as a "fasting signal" to prolong the catabolic state.


Assuntos
Fígado Gorduroso , PPAR alfa , Animais , Camundongos , 3-Metil-2-Oxobutanoato Desidrogenase (Lipoamida)/metabolismo , Aminoácidos de Cadeia Ramificada/metabolismo , Jejum , Camundongos Knockout , RNA Interferente Pequeno
3.
Am J Physiol Endocrinol Metab ; 301(5): E825-35, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21771968

RESUMO

Adipose tissue controls energy homeostasis and systemic insulin sensitivity through the elaboration of a series of cytokines and hormones, collectively termed "adipokines." We and others have identified Lcn2 as a novel adipokine, but its exact role in obesity-induced insulin resistance remains controversial. The aim of this study was to examine the metabolic phenotype of Lcn2(-/-) mice to clarify the role of Lcn2 in metabolism. Male and female Lcn2(-/-) and wild-type (WT) littermates were placed on either chow or high-fat diet (HFD) to characterize their metabolic phenotype. Studies included body weight and body composition, glucose and insulin tolerance tests, and adipokine expression studies in serum and in white adipose tissue (WAT). Neither chow nor HFD cohorts showed any differences in body weight or body composition. Chow-fed Lcn2(-/-) mice did not exhibit any difference in glucose homeostasis compared with WT mice. Fasting serum glucose levels were lower in the chow-fed Lcn2(-/-) mice, but this finding was not seen in the HFD cohort. Serum adiponectin, leptin, resistin, and RBP4 levels were not different between WT and Lcn2(-/-) on chow diet. HFD-fed male Lcn2(-/-) mice did display a small improvement in glucose tolerance, but no difference in insulin sensitivity was seen in either male or female Lcn2(-/-) mice on HFD. We conclude that the global ablation of Lcn2 has a minimal effect on obesity-associated glucose intolerance but does not appear to affect either age- or obesity-mediated insulin resistance in vivo.


Assuntos
Proteínas de Fase Aguda/fisiologia , Dieta Hiperlipídica , Intolerância à Glucose/etiologia , Lipocalinas/fisiologia , Proteínas Oncogênicas/fisiologia , Proteínas de Fase Aguda/genética , Proteínas de Fase Aguda/metabolismo , Animais , Composição Corporal/genética , Composição Corporal/fisiologia , Peso Corporal/genética , Peso Corporal/fisiologia , Feminino , Predisposição Genética para Doença , Glucose/metabolismo , Intolerância à Glucose/complicações , Intolerância à Glucose/genética , Intolerância à Glucose/metabolismo , Teste de Tolerância a Glucose , Resistência à Insulina/genética , Lipocalina-2 , Lipocalinas/genética , Lipocalinas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Obesos , Obesidade/complicações , Obesidade/genética , Obesidade/metabolismo , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/metabolismo , Fenótipo
4.
J Pharmacol Exp Ther ; 326(1): 135-43, 2008 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-18388257

RESUMO

Patients on a statin regimen have a decreased risk of death due to bacterial sepsis. We have found that protection by simvastatin includes the inhibition of host cell invasion by Staphylococcus aureus, the most common etiologic agent of sepsis. Inhibition was due in part to depletion of isoprenoid intermediates within the cholesterol biosynthesis pathway and led to the cytosolic accumulation of the small GTPases CDC42, Rac, and RhoB. Actin stress fiber disassembly required for host invasion was attenuated by simvastatin and by the inhibition of phosphoinositide 3-kinase (PI3K) activity. PI3K relies on coupling to prenylated proteins, such as this subset of small GTPases, for access to membrane-bound phosphoinositide to mediate stress fiber disassembly. Therefore, we examined whether simvastatin restricts PI3K cellular localization. In response to simvastatin, the PI3K isoform p85, coupled to these small-GTPases, was sequestered within the cytosol. From these findings, we propose a mechanism whereby simvastatin restricts p85 localization, inhibiting the actin dynamics required for bacterial endocytosis. This approach may provide the basis for protection at the level of the host in invasive infections by S. aureus.


Assuntos
Sinvastatina/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Terpenos/metabolismo , Linhagem Celular , Células Cultivadas , Humanos , Fatores Hospedeiros de Integração/antagonistas & inibidores , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Staphylococcus aureus/citologia
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