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
For the first time, the study of the antioxidant activity, the characterization of the phytoconstituants, and the evaluation of in vitro and in vivo toxicity of A. djiboutiensis leave and latex are performed. The antioxidant activity of both latex (ADL) and the methanolic extract of leaves (ADM) is determined using 1-diphenyl-2-picrylhydrazyl (DPPH), 2,2'-azino-bis 3-ethylbenzothiazoline-6-sulphonic acid (ABTS) scavenging radical methods and ferric reducing/antioxidant power (FRAP) assay. The phytochemical study of latex is done using Liquid Chromatography-Mass Spectrometry (LC-MS/MS) and a molecular networking-based approach. The evaluation of in vivo toxicity is performed on mice by oral gavage with a suspension of ADL. Our results show that weak antioxidant activity of ADL and ADM in opposition to their high polyphenol, 83.01 mg and 46.4 mg expressed in gallic acid equivalent (GAE)/g of dry weight (DW), respectively, and flavonoid contents 13.12 mg and 4.25 mg expressed in quercetin equivalent (QE)/g dry weight (DW), respectively. Using the Global Natural Products Social Molecular Networking (GNPS) website, nine (9) anthraquinones derivatives, ten (10) chromones derivatives, two (2) flavonols/ chromones isomers are annotated in the molecular network. The treated mice do not display abnormalities in their general physical appearance and biochemistry parameters, compared to the controls. Only glucose and calcium levels are slightly higher in male treated mice compared to the vehicles.
Assuntos
Aloe/química , Antioxidantes/farmacologia , Extratos Vegetais/farmacologia , Animais , Temperatura Corporal/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Cromatografia Líquida/métodos , Djibuti , Feminino , Masculino , Camundongos , Extratos Vegetais/administração & dosagem , Espectrometria de Massas em Tandem/métodosRESUMO
Young mammals possess a limited regenerative capacity in some tissues, which is lost upon maturation. We investigated whether cellular senescence might play a role in such loss during liver regeneration. We found that following partial hepatectomy, the senescence-associated genes p21, p16Ink4a, and p19Arf become dynamically expressed in different cell types when regenerative capacity decreases, but without a full senescent response. However, we show that treatment with a senescence-inhibiting drug improves regeneration, by disrupting aberrantly prolonged p21 expression. This work suggests that senescence may initially develop from heterogeneous cellular responses, and that senotherapeutic drugs might be useful in promoting organ regeneration.
Assuntos
Compostos de Bifenilo/farmacologia , Inibidor de Quinase Dependente de Ciclina p21/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Fígado/fisiologia , Nitrofenóis/farmacologia , Regeneração/efeitos dos fármacos , Sulfonamidas/farmacologia , Animais , Células Cultivadas , Senescência Celular/efeitos dos fármacos , Inibidor p16 de Quinase Dependente de Ciclina/genética , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Animais , Piperazinas/farmacologiaRESUMO
Hepatocellular carcinoma (HCC) is the second leading cause of cancer death worldwide. While curative approaches for early stage HCC exist, effective treatment options for advanced HCC are lacking. Furthermore, there are no efficient chemopreventive strategies to limit HCC development once cirrhosis is established. One challenge for drug development is unsatisfactory animal models. In this article, we describe an orthotopic xenograft mouse model of human liver cancer cell lines through image-guided injection into the liver. This technique provides a less invasive yet highly efficient approach to engraft human HCC into mouse liver. Similarly, image-guided injections are used to deliver chemotherapeutics locally, enabling reduction in potential systemic adverse effects, while reducing the required dose for a therapeutic effect. In summary, this image-guided strategy provides a novel and convenient approach to improve current HCC mouse models. © 2019 The Authors. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.
Assuntos
Xenoenxertos/fisiologia , Neoplasias Hepáticas Experimentais/terapia , Camundongos , Transplante Heterólogo/métodos , Ultrassom/métodos , Animais , Carcinoma Hepatocelular/terapia , Neoplasias Hepáticas/terapia , Transplante Heterólogo/instrumentação , Ultrassom/instrumentaçãoRESUMO
Strict regulation of Ca2+ homeostasis is essential for normal cellular physiology. Store-operated Ca2+ entry (SOCE) is a major mechanism controlling basal Ca2+ levels and intracellular Ca2+ store refilling, and abnormal SOCE severely impacts on human health. Overactive SOCE results in excessive extracellular Ca2+ entry due to dominant STIM1 or ORAI1 mutations and has been associated with tubular aggregate myopathy (TAM) and Stormorken syndrome (STRMK). Both disorders are spectra of the same disease and involve muscle weakness, myalgia and cramps, and additional multi-systemic signs including miosis, bleeding diathesis, hyposplenism, dyslexia, short stature and ichthyosis. To elucidate the physiological consequences of STIM1 over-activation, we generated a murine model harboring the most common TAM/STRMK mutation and characterized the phenotype at the histological, ultrastructural, metabolic, physiological and functional level. In accordance with the clinical picture of TAM/STRMK, the Stim1R304W/+ mice manifested muscle weakness, thrombocytopenia, skin and eye anomalies and spleen dysfunction, as well as additional features not yet observed in patients such as abnormal bone architecture and immune system dysregulation. The murine muscles exhibited contraction and relaxation defects as well as dystrophic features, and functional investigations unraveled increased Ca2+ influx in myotubes. In conclusion, we provide insight into the pathophysiological effect of the STIM1 R304W mutation in different cells, tissues and organs and thereby significantly contribute to a deeper understanding of the pathomechanisms underlying TAM/STRMK and other human disorders involving aberrant Ca2+ homeostasis and affecting muscle, bones, platelets or the immune system.
Assuntos
Transtornos Plaquetários/genética , Dislexia/genética , Ictiose/genética , Transtornos de Enxaqueca/genética , Miose/genética , Miopatias Congênitas Estruturais/genética , Proteínas de Neoplasias/genética , Baço/anormalidades , Molécula 1 de Interação Estromal/genética , Animais , Transtornos Plaquetários/fisiopatologia , Osso e Ossos/metabolismo , Osso e Ossos/patologia , Sinalização do Cálcio/genética , Modelos Animais de Doenças , Dislexia/fisiopatologia , Eritrócitos Anormais , Olho/metabolismo , Olho/patologia , Técnicas de Introdução de Genes , Humanos , Ictiose/patologia , Ictiose/fisiopatologia , Sistema Imunitário/patologia , Proteínas Sensoras de Cálcio Intracelular/genética , Proteínas de Membrana/genética , Camundongos , Transtornos de Enxaqueca/fisiopatologia , Miose/fisiopatologia , Fadiga Muscular/genética , Debilidade Muscular/genética , Debilidade Muscular/patologia , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Mutação/genética , Miopatias Congênitas Estruturais/fisiopatologia , Proteína ORAI1/genética , Pele/metabolismo , Pele/patologia , Baço/fisiopatologiaRESUMO
BACKGROUND: Autosomal dominant gain-of-function mutations in human stimulator of interferon genes (STING) lead to a severe autoinflammatory disease called STING-associated vasculopathy with onset in infancy that is associated with enhanced expression of interferon-stimulated gene transcripts. OBJECTIVE: The goal of this study was to analyze the phenotype of a new mouse model of STING hyperactivation and the role of type I interferons in this system. METHODS: We generated a knock-in model carrying an amino acid substitution (V154M) in mouse STING, corresponding to a recurrent mutation seen in human patients with STING-associated vasculopathy with onset in infancy. Hematopoietic development and tissue histology were analyzed. Lymphocyte activation and proliferation were assessed in vitro. STING V154M/wild-type (WT) mice were crossed to IFN-α/ß receptor (IFNAR) knockout mice to evaluate the type I interferon dependence of the mutant Sting phenotype recorded. RESULTS: In STING V154M/WT mice we detected variable expression of inflammatory infiltrates in the lungs and kidneys. These mice showed a marked decrease in survival and developed a severe combined immunodeficiency disease (SCID) affecting B, T, and natural killer cells, with an almost complete lack of antibodies and a significant expansion of monocytes and granulocytes. The blockade in B- and T-cell development was present from early immature stages in bone marrow and thymus. In addition, in vitro experiments revealed an intrinsic proliferative defect of mature T cells. Although the V154M/WT mutant demonstrated increased expression of interferon-stimulated genes, the SCID phenotype was not reversed in STING V154M/WT IFNAR knockout mice. However, the antiproliferative defect in T cells was rescued partially by IFNAR deficiency. CONCLUSIONS: STING gain-of-function mice developed an interferon-independent SCID phenotype with a T-cell, B-cell, and natural killer cell developmental defect and hypogammaglobulinemia that is associated with signs of inflammation in lungs and kidneys. Only the intrinsic proliferative defect of T cells was partially interferon dependent.
Assuntos
Linfócitos B/fisiologia , Inflamação/genética , Células Matadoras Naturais/imunologia , Proteínas de Membrana/genética , Mutação/genética , Imunodeficiência Combinada Severa/genética , Linfócitos T/fisiologia , Agamaglobulinemia , Animais , Diferenciação Celular/genética , Modelos Animais de Doenças , Humanos , Interferon Tipo I/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor de Interferon alfa e beta/genéticaRESUMO
Hepatocellular carcinoma (HCC) is the only cancer for which non-invasive diagnosis is recognized by international guidelines. Contrast agent free ultrasound imaging, computed tomography (CT) and/or magnetic resonance imaging are techniques used for early detection and confirmation. Clinical evidence depicts that CT is 30% less precise as compared to MRI for detection of small tumors. In our work, we have reported some novel tools that can enhance the sensitivity and precision of CT applied to preclinical research (micro-CT). Our system, containing non-toxic nano-droplets loaded with iodine has high contrasting properties, liver and hepatocyte specificity and strong liver persistence. Micro-CT was performed on HCC model implanted in nude mice by intrahepatic injection. Contrast agent was administrated intravenously. This method allows an unprecedented high precision of detection, quantitative measurement of tumor volume and quantitative follow-up of the tumor development.
Assuntos
Carcinoma Hepatocelular/diagnóstico por imagem , Carcinoma Hepatocelular/patologia , Halogenação , Neoplasias Hepáticas/diagnóstico por imagem , Neoplasias Hepáticas/patologia , Fígado/diagnóstico por imagem , Nanotecnologia , Microtomografia por Raio-X/métodos , Animais , Linhagem Celular Tumoral , Proliferação de Células , Transformação Celular Neoplásica , Emulsões , Feminino , Humanos , Fígado/patologia , Camundongos , UltrassonografiaRESUMO
ATP6AP2 codes for the (pro)renin receptor and is an essential component of vacuolar H+ ATPase. Activating (pro)renin for conversion of Angiotensinogen to Angiotensin makes ATP6AP2 attractive for drug intervention. Tissue-specific ATP6AP2 inactivation in mouse suggested a strong impact on various organs. Consistent with this, we found that embryonic ablation of Atp6ap2 resulted in both male hemizygous lethality and female haploinsufficiency. Next, we examined the phenotype of an induced inactivation in the adult animal, most akin to detect potential effect of functional interference of ATP6AP2 through drug therapy. Induced ablation of Atp6ap2, even without equal efficiency in all tissues (aorta, brain and kidney), resulted in rapid lethality marked by weight loss, changes in nutritional as well as blood parameters, leukocyte depletion, and bone marrow hypoplasia. Upon Atp6ap2 ablation, the colon demonstrated a rapid disruption of crypt morphology, aberrant proliferation, cell-death activation, as well as generation of microadenomas. Consequently, disruption of ATP6AP2 is extremely poorly tolerated in the adult, and severely affects various organ systems demonstrating that ATP6AP2 is an essential gene implicated in basic cellular mechanisms and necessary for multiple organ function. Accordingly, any potential drug targeting of this gene product must be strictly assessed for safety.
Assuntos
Insuficiência de Múltiplos Órgãos/mortalidade , Insuficiência de Múltiplos Órgãos/patologia , ATPases Translocadoras de Prótons/deficiência , Receptores de Superfície Celular/deficiência , Animais , Técnicas de Inativação de Genes , Camundongos , Análise de SobrevidaRESUMO
Fragile X-associated tremor/ataxia syndrome (FXTAS) is a neurodegenerative disorder caused by a limited expansion of CGG repeats in the 5' UTR of FMR1. Two mechanisms are proposed to cause FXTAS: RNA gain-of-function, where CGG RNA sequesters specific proteins, and translation of CGG repeats into a polyglycine-containing protein, FMRpolyG. Here we developed transgenic mice expressing CGG repeat RNA with or without FMRpolyG. Expression of FMRpolyG is pathogenic, while the sole expression of CGG RNA is not. FMRpolyG interacts with the nuclear lamina protein LAP2ß and disorganizes the nuclear lamina architecture in neurons differentiated from FXTAS iPS cells. Finally, expression of LAP2ß rescues neuronal death induced by FMRpolyG. Overall, these results suggest that translation of expanded CGG repeats into FMRpolyG alters nuclear lamina architecture and drives pathogenesis in FXTAS.
Assuntos
Ataxia/genética , Proteínas de Ligação a DNA/metabolismo , Proteína do X Frágil da Deficiência Intelectual/genética , Síndrome do Cromossomo X Frágil/genética , Proteínas de Membrana/metabolismo , Lâmina Nuclear/metabolismo , Peptídeos/genética , Biossíntese de Proteínas , RNA Mensageiro/metabolismo , Tremor/genética , Expansão das Repetições de Trinucleotídeos/genética , Animais , Ataxia/metabolismo , Encéfalo/metabolismo , Encéfalo/patologia , Proteína do X Frágil da Deficiência Intelectual/metabolismo , Síndrome do Cromossomo X Frágil/metabolismo , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Lâmina Nuclear/patologia , Peptídeos/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Tremor/metabolismoRESUMO
Mutations of the AMP-activated kinase gamma 2 subunit (AMPKγ2), N488I (AMPKγ2NI) and R531G (AMPKγ2RG), are associated with Wolff-Parkinson-White (WPW) syndrome, a cardiac disorder characterized by ventricular pre-excitation in humans. Cardiac-specific transgenic overexpression of human AMPKγ2NI or AMPKγ2RG leads to constitutive AMPK activation and the WPW phenotype in mice. However, overexpression of these mutant proteins also caused profound, non-physiological increase in cardiac glycogen, which might abnormally alter the true phenotype. To investigate whether physiological levels of AMPKγ2NI or AMPKγ2RG mutation cause WPW syndrome and metabolic changes in other organs, we generated two knock-in mouse lines on the C57BL/6N background harboring mutations of human AMPKγ2NI and AMPKγ2RG, respectively. Similar to the reported phenotypes of mice overexpressing AMPKγ2NI or AMPKγ2RG in the heart, both lines developed WPW syndrome and cardiac hypertrophy; however, these effects were independent of cardiac glycogen accumulation. Compared with AMPKγ2WT mice, AMPKγ2NI and AMPKγ2RG mice exhibited reduced body weight, fat mass, and liver steatosis when fed with a high fat diet (HFD). Surprisingly, AMPKγ2RG but not AMPKγ2NI mice fed with an HFD exhibited severe kidney injury characterized by glycogen accumulation, inflammation, apoptosis, cyst formation, and impaired renal function. These results demonstrate that expression of AMPKγ2NI and AMPKγ2RG mutations at physiological levels can induce beneficial metabolic effects but that this is accompanied by WPW syndrome. Our data also reveal an unexpected effect of AMPKγ2RG in the kidney, linking lifelong constitutive activation of AMPK to a potential risk for kidney dysfunction in the context of an HFD.
Assuntos
Proteínas Quinases Ativadas por AMP/genética , Mutação , Insuficiência Renal/genética , Síndrome de Wolff-Parkinson-White/genética , Animais , Apoptose , Modelos Animais de Doenças , Técnicas de Introdução de Genes , Inflamação/genética , Inflamação/patologia , Rim/metabolismo , Rim/patologia , Masculino , Camundongos Endogâmicos C57BL , Insuficiência Renal/patologia , Síndrome de Wolff-Parkinson-White/patologiaRESUMO
Partial monosomy 21 (PM21) is a rare chromosomal abnormality that is characterized by the loss of a variable segment along human chromosome 21 (Hsa21). The clinical phenotypes of this loss are heterogeneous and range from mild alterations to lethal consequences, depending on the affected region of Hsa21. The most common features include intellectual disabilities, craniofacial dysmorphology, short stature, and muscular and cardiac defects. As a complement to human genetic approaches, our team has developed new monosomic mouse models that carry deletions on Hsa21 syntenic regions in order to identify the dosage-sensitive genes that are responsible for the symptoms. We focus here on the Ms5Yah mouse model, in which a 7.7-Mb region has been deleted from the App to Runx1 genes. Ms5Yah mice display high postnatal lethality, with a few surviving individuals showing growth retardation, motor coordination deficits, and spatial learning and memory impairments. Further studies confirmed a gene dosage effect in the Ms5Yah hippocampus, and pinpointed disruptions of pathways related to cell adhesion (involving App, Cntnap5b, Lgals3bp, Mag, Mcam, Npnt, Pcdhb2, Pcdhb3, Pcdhb4, Pcdhb6, Pcdhb7, Pcdhb8, Pcdhb16 and Vwf). Our PM21 mouse model is the first to display morphological abnormalities and behavioural phenotypes similar to those found in affected humans, and it therefore demonstrates the major contribution that the App-Runx1 region has in the pathophysiology of PM21.
Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/genética , Monossomia/genética , Deleção de Sequência/genética , Animais , Animais Recém-Nascidos , Comportamento Animal , Peso Corporal , Cromossomos Humanos Par 21/genética , Análise por Conglomerados , Subunidade alfa 2 de Fator de Ligação ao Core/deficiência , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Modelos Animais de Doenças , Comportamento Exploratório , Feto/anormalidades , Feto/patologia , Dosagem de Genes , Regulação da Expressão Gênica no Desenvolvimento , Hipocampo/metabolismo , Hipocampo/patologia , Humanos , Aprendizagem em Labirinto , Memória , Camundongos , Anotação de Sequência Molecular , Atividade Motora , Análise de Sequência com Séries de Oligonucleotídeos , Software , Aprendizagem Espacial , Transcriptoma/genéticaRESUMO
According to the hitherto accepted view, neutrophils kill ingested microorganisms by subjecting them to high concentrations of highly toxic reactive oxygen species (ROS) and bringing about myeloperoxidase-catalysed halogenation. We show here that this simple scheme, which for many years has served as a satisfactory working hypothesis, is inadequate. We find that mice made deficient in neutrophil-granule proteases but normal in respect of superoxide production and iodinating capacity, are unable to resist staphylococcal and candidal infections. We also show that activation provokes the influx of an enormous concentration of ROS into the endocytic vacuole. The resulting accumulation of anionic charge is compensated for by a surge of K+ ions that cross the membrane in a pH-dependent manner. The consequent rise in ionic strength engenders the release of cationic granule proteins, including elastase and cathepsin G, from the anionic sulphated proteoglycan matrix. We show that it is the proteases, thus activated, that are primarily responsible for the destruction of the bacteria.