RESUMO
Insulin resistance is a major public health burden that often results in other comorbidities including type 2 diabetes, nonalcoholic fatty liver disease (NAFLD), and cardiovascular disease. An insulin sensitizer has the potential to become a disease-modifying therapy. It remains an unmet medical need to identify therapeutics that target the insulin signaling pathway to treat insulin resistance. Low-molecular-weight protein tyrosine phosphatase (LMPTP) negatively regulates insulin signaling and has emerged as a potential therapeutic target for insulin sensitization. Genetic studies have demonstrated that LMPTP is positively associated with obesity in humans and promotes insulin resistance in rodents. A recent study showed that pharmacological inhibition or genetic deletion of LMPTP protects mice from high-fat diet-induced insulin resistance and diabetes. Here, we show that loss of LMPTP by genetic deletion has no significant effects on improving glucose tolerance in lean or diet-induced obese mice. Furthermore, our data demonstrate that LMPTP deficiency potentiates cardiac hypertrophy that leads to mild cardiac dysfunction. Our findings suggest that the development of LMPTP inhibitors for the treatment of insulin resistance and type 2 diabetes should be reevaluated, and further studies are needed to characterize the molecular and pathophysiological role of LMPTP.NEW & NOTEWORTHY Inhibition of LMPTP with a small-molecule inhibitor, Cmpd23, improves glucose tolerance in mice as reported earlier. However, genetic deficiency of the LMPTP-encoding gene, Acp1, has limited effects on glucose metabolism but leads to mild cardiac hypertrophy in mice. The findings suggest the potential off-target effects of Cmpd23 and call for reevaluation of LMPTP as a therapeutic target for the treatment of insulin resistance and type 2 diabetes.
Assuntos
Diabetes Mellitus Tipo 2 , Resistência à Insulina , Animais , Cardiomegalia/genética , Cardiomegalia/metabolismo , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/genética , Dieta Hiperlipídica , Glucose/metabolismo , Insulina/metabolismo , Resistência à Insulina/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Proteínas Tirosina Fosfatases/metabolismo , Proteínas Tirosina Fosfatases/uso terapêutico , MagrezaRESUMO
Obesity is a major contributing factor to the pathogenesis of Type 2 diabetes. Multiple human genetics studies suggest that high activity of the low molecular weight protein tyrosine phosphatase (LMPTP) promotes metabolic syndrome in obesity. We reported that LMPTP is a critical promoter of insulin resistance in obesity by regulating liver insulin receptor signaling and that inhibition of LMPTP reverses obesity-associated diabetes in mice. Since LMPTP is expressed in adipose tissue but little is known about its function, here we examined the role of LMPTP in adipocyte biology. Using conditional knockout mice, we found that selective deletion of LMPTP in adipocytes impaired obesity-induced subcutaneous adipocyte hypertrophy. We assessed the role of LMPTP in adipogenesis in vitro, and found that LMPTP deletion or knockdown substantially impaired differentiation of primary preadipocytes and 3T3-L1 cells into adipocytes, respectively. Inhibition of LMPTP in 3T3-L1 preadipocytes also reduced adipogenesis and expression of proadipogenic transcription factors peroxisome proliferator activated receptor gamma (PPARγ) and CCAAT/enhancer-binding protein alpha. Inhibition of LMPTP increased basal phosphorylation of platelet-derived growth factor receptor alpha (PDGFRα) on activation motif residue Y849 in 3T3-L1, resulting in increased activation of the mitogen-associated protein kinases p38 and c-Jun N-terminal kinase and increased PPARγ phosphorylation on inhibitory residue S82. Analysis of the metabolome of differentiating 3T3-L1 cells suggested that LMPTP inhibition decreased cell glucose utilization while enhancing mitochondrial respiration and nucleotide synthesis. In summary, we report a novel role for LMPTP as a key driver of adipocyte differentiation via control of PDGFRα signaling.
Assuntos
Adipócitos/metabolismo , Adipócitos/patologia , Adipogenia , Proteínas Tirosina Fosfatases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Gordura Subcutânea/patologia , Células 3T3-L1 , Adipogenia/genética , Animais , Diferenciação Celular/genética , Respiração Celular , Tamanho Celular , Transporte de Elétrons , Deleção de Genes , Regulação da Expressão Gênica , Glucose/metabolismo , Glicólise , Hipertrofia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Metaboloma , Camundongos , Camundongos Knockout , Mitocôndrias/metabolismo , Modelos Biológicos , PPAR gama/metabolismo , Fosforilação , Fosfosserina/metabolismo , Proteínas Tirosina Fosfatases/antagonistas & inibidores , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Transdução de Sinais , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
BACKGROUND: Protein tyrosine phosphatase 1B (PTP1B) and low molecular weight protein tyrosine phosphatase (LMPTP) are implicated in the development of metabolic disorders. Yet, their role in progenitor stem cell adipogenic differentiation and modulation of mitochondrial dynamics remains elusive. METHODS: In this study, we decided to investigate whether inhibition of PTP1B and LMPTP enhance adipogenic differentiation of metabolically impaired progenitor stem cells via modulation of mitochondrial bioenergetics and dynamics. Cells were cultured under adipogenic conditions in the presence of PTP1B and LMPTP inhibitors, and were subjected to the analysis of the main adipogenic-related and mitochondrial-related genes using RT-qPCR. Protein levels were established with western blot while mitochondrial morphology with MicroP software. RESULTS: Selective inhibitors of both PTP1B and MPTP enhanced adipogenic differentiation of metabolically impaired progenitor stem cells. We have observed enhanced expression of PPARy and adiponectin in treated cells. What is more, increased antioxidative defence and alternations in mitochondrial bioenergetics were observed. We have found that inhibition of PTP1B as well as C23 activates oxidative phosphorylation and enhances mitochondrial fusion contributing to enhanced adipogenesis. CONCLUSIONS: The presented data provides evidence that the application of PTP1B and LMPTP inhibitors enhances adipogenesis through the modulation of mitochondrial dynamics. Video abstract.
Assuntos
Adipogenia/genética , Mitocôndrias/genética , Fosfoproteínas/genética , Proteína Tirosina Fosfatase não Receptora Tipo 1/genética , Proteínas Tirosina Fosfatases/genética , Proteínas de Ligação a RNA/genética , Adipócitos/citologia , Adipócitos/metabolismo , Adiponectina/genética , Animais , Apoptose/genética , Diferenciação Celular/genética , Cavalos , Humanos , Mitocôndrias/metabolismo , Dinâmica Mitocondrial/genética , Fosforilação Oxidativa , Estresse Oxidativo/genética , PPAR gama/genética , Células-Tronco/citologia , Células-Tronco/metabolismo , NucleolinaRESUMO
Equine metabolic syndrome (EMS) is a cluster of metabolic disorders, such as obesity, hyperinsulinemia, and hyperleptinemia, as well as insulin resistance (IR). In accordance with the theory linking obesity and IR, excessive accumulation of lipids in insulin-sensitive tissues (lipotoxicity), like liver, alters several cellular functions, including insulin signaling. Therefore, the purpose of the study was to isolate equine hepatic progenitor-like cells (HPCs) and assess whether inhibition of low molecular weight protein tyrosine phosphatase (LMPTP) affects the expression of genes involved in macroautophagy, chaperone-mediated autophagy (CMA), endoplasmic reticulum stress, and mitochondrial dynamics in a palmitate-induced IR model. We demonstrated that LMPTP inhibition significantly enhanced expression of heat shock cognate 70 kDa protein (HSC70), lysosome-associated membrane protein 2 (LAMP2), and parkin (PRKN), all master regulators of selective autophagy. We also observed downregulation of C/EBP homologous protein (CHOP), activating transcription factor 6 (ATF6) and binding immunoglobulin protein encoded by the HSPA gene. Moreover, LMPTP inhibition increased alternative splicing of X-box binding protein 1 (XBP1), suggesting high endonuclease activity of inositol-requiring enzyme 1 alpha (IRE1α). Taken together, our data provide convincing evidence that LMPTP inhibition reverses palmitate-induced insulin resistance and lipotoxicity. In conclusion, this study highlights the role of LMPTP in the regulation of CMA, mitophagy, and ER stress, and provides a new in vitro model for studying HPC lipotoxicity in pre-clinical research.
Assuntos
Morte Celular Autofágica , Estresse do Retículo Endoplasmático , Doenças dos Cavalos/enzimologia , Fígado/enzimologia , Síndrome Metabólica/enzimologia , Proteínas Tirosina Fosfatases/metabolismo , Células-Tronco/enzimologia , Animais , Doenças dos Cavalos/patologia , Cavalos , Resistência à Insulina , Fígado/patologia , Síndrome Metabólica/patologia , Ácido Palmítico/toxicidade , Células-Tronco/patologiaRESUMO
Purpose: Protein tyrosine phosphatases (PTPs) play an essential way in diseases including cancer, obesity, diabetes and autoimmune disorders. As a member of PTPs, low molecular weight PTP (LMPTP) has been a well-recognized anti-insulin resistance target in obesity. However, the number of reported LMPTP inhibitors is limited. Our research aims to discover a novel LMPTP inhibitor and evaluate its biological activity against insulin resistance. Methods: A virtual screening pipeline based on the X-ray co-crystal complex of LMPTP was constructed. Enzyme inhibition assay and cellular bioassay were used to evaluate the activity of screened compounds. Results: The screening pipeline rendered 15 potential hits from Specs chemical library. Enzyme inhibition assay identified compound F9 (AN-465/41163730) as a potential LMPTP inhibitor with a K i value of 21.5 ± 7.3 µM. Cellular bioassay showed F9 could effectively increase the glucose consumption of HepG2 cells as a result of releasing insulin resistance by regulating PI3K-Akt pathway. Conclusion: In summary, this study presents a versatile virtual screening pipeline for potential LMPTP inhibitor discovery and provides a novel-scaffold lead compound that is worthy of further modification to get more potent LMPTP inhibitors.
Assuntos
Resistência à Insulina , Tirosina , Humanos , Peso Molecular , Fosfatidilinositol 3-Quinases , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/química , Proteínas Tirosina Fosfatases/metabolismo , Obesidade , Proteína Tirosina Fosfatase não Receptora Tipo 1RESUMO
OBJECTIVES: Non-alcoholic fatty liver disease (NAFLD) is considered a well-known pathology that is determined without using alcohol and has emerged as a growing public health problem. Lipotoxicity is known to promote hepatocyte death, which, in the context of NAFLD, is termed lipoapoptosis. The severity of NAFLD correlates with the degree of hepatocyte lipoapoptosis. Protein-tyrosine phosphatases (PTP) including PTP1B and Low molecular weight PTP (LMPTP), are negative regulators of the insulin signaling pathway and are considered a promising therapeutic target in the treatment of diabetes. In this study, we hypothesized that the inhibition of PTP1B and LMPTP may potentially prevent hepatocyte apoptosis, mitochondrial dysfunction and endoplasmic reticulum (ER) stress onset, following lipotoxicity induced using a free fatty acid (FFA) mixture. METHODS: HepG2 cells were cultured in the presence or absence of two PTP inhibitors, namely MSI-1436 and Compound 23, prior to palmitate/oleate overloading. Apoptosis, ER stress, oxidative stress, and mitochondrial dynamics were then evaluated by either MUSE or RT-qPCR analysis. RESULTS: The obtained data demonstrate that the inhibition of PTP1B and LMPTP prevents apoptosis induced by palmitate and oleate in the HepG2 cell line. Moreover, mitochondrial dynamics were positively improved following inhibition of the enzyme, with concomitant oxidative stress reduction and ER stress abrogation. CONCLUSION: In conclusion, PTP's inhibitory properties may be a promising therapeutic strategy for the treatment of FFA-induced lipotoxicity in the liver and ultimately in the management of the NAFLD condition.