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1.
Nat Commun ; 11(1): 4029, 2020 08 12.
Artigo em Inglês | MEDLINE | ID: mdl-32788597

RESUMO

In autosomal dominant optic atrophy (ADOA), caused by mutations in the mitochondrial cristae biogenesis and fusion protein optic atrophy 1 (Opa1), retinal ganglion cell (RGC) dysfunction and visual loss occur by unknown mechanisms. Here, we show a role for autophagy in ADOA pathogenesis. In RGCs expressing mutated Opa1, active 5' AMP-activated protein kinase (AMPK) and its autophagy effector ULK1 accumulate at axonal hillocks. This AMPK activation triggers localized hillock autophagosome accumulation and mitophagy, ultimately resulting in reduced axonal mitochondrial content that is restored by genetic inhibition of AMPK and autophagy. In C. elegans, deletion of AMPK or of key autophagy and mitophagy genes normalizes the axonal mitochondrial content that is reduced upon mitochondrial dysfunction. In conditional, RGC specific Opa1-deficient mice, depletion of the essential autophagy gene Atg7 normalizes the excess autophagy and corrects the visual defects caused by Opa1 ablation. Thus, our data identify AMPK and autophagy as targetable components of ADOA pathogenesis.


Assuntos
Autofagia , Atrofia Óptica Autossômica Dominante/complicações , Transtornos da Visão/complicações , Adenilato Quinase/metabolismo , Animais , Autofagia/genética , Axônios/patologia , Caenorhabditis elegans/metabolismo , Modelos Animais de Doenças , Ativação Enzimática , GTP Fosfo-Hidrolases/genética , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/metabolismo , Mitofagia , Mutação/genética , Fosforilação , Células Ganglionares da Retina/patologia
2.
Nat Commun ; 11(1): 3639, 2020 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-32686665

RESUMO

Integrated analysis of genomes, transcriptomes, proteomes and drug responses of cancer cell lines (CCLs) is an emerging approach to uncover molecular mechanisms of drug action. We extend this paradigm to measuring proteome activity landscapes by acquiring and integrating quantitative data for 10,000 proteins and 55,000 phosphorylation sites (p-sites) from 125 CCLs. These data are used to contextualize proteins and p-sites and predict drug sensitivity. For example, we find that Progesterone Receptor (PGR) phosphorylation is associated with sensitivity to drugs modulating estrogen signaling such as Raloxifene. We also demonstrate that Adenylate kinase isoenzyme 1 (AK1) inactivates antimetabolites like Cytarabine. Consequently, high AK1 levels correlate with poor survival of Cytarabine-treated acute myeloid leukemia patients, qualifying AK1 as a patient stratification marker and possibly as a drug target. We provide an interactive web application termed ATLANTiC (http://atlantic.proteomics.wzw.tum.de), which enables the community to explore the thousands of novel functional associations generated by this work.


Assuntos
Antineoplásicos/farmacologia , Neoplasias/tratamento farmacológico , Proteoma/metabolismo , Adenilato Quinase/metabolismo , Animais , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Biologia Computacional , Simulação por Computador , Citarabina/metabolismo , Citarabina/farmacologia , Desenvolvimento de Medicamentos , Genômica , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Neoplasias/metabolismo , Proteoma/genética , Proteômica , Cloridrato de Raloxifeno/metabolismo , Cloridrato de Raloxifeno/farmacologia , Receptores de Progesterona/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/fisiologia
3.
Life Sci ; 256: 117965, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32544463

RESUMO

BACKGROUND: Several studies have proved that physical activity (PA) regulates energetic metabolism associated with mitochondrial dynamics through AMPK activation in healthy subjects. Obesity, a condition that induces oxidative stress, mitochondrial dysfunction, and low AMPK activity leads to mitochondrial fragmentation. However, few studies describe the effect of PA on mitochondrial dynamics regulation in obesity. AIM: The present study aimed to evaluate the effect of a single session of PA on mitochondrial dynamics regulation as well as its effect on mitochondrial function and organization in skeletal muscles of obese rats (Zucker fa/fa). MAIN METHODS: Male Zucker lean and Zucker fa/fa rats aged 12 to 13 weeks were divided into sedentary and subjected-to-PA (single session swimming) groups. Gastrocnemius muscle was dissected into isolated fibers, mitochondria, mRNA, and total proteins for their evaluation. KEY FINDINGS: The results showed that PA increased the Mfn-2 protein level in the lean and obese groups, whereas Drp1 levels decreased in the obese group. OMA1 protease levels increased in the lean group and decreased in the obese group. Additionally, AMPK analysis parameters (expression, protein level, and activity) did not increase in the obese group. These findings correlated with the partial restoration of mitochondrial function in the obese group, increasing the capacity to maintain the membrane potential after adding calcium as a stressor, and increasing the transversal organization level of the mitochondria analyzed in isolated fibers. SIGNIFICANCE: These results support the notion that obese rats subjected to PA maintain mitochondrial function through mitochondrial fusion activation by an AMPK-independent mechanism.


Assuntos
Mitocôndrias/patologia , Fibras Musculares Esqueléticas/patologia , Obesidade/patologia , Condicionamento Físico Animal , Adenilato Quinase/metabolismo , Animais , Biomarcadores/metabolismo , Citrato (si)-Sintase/metabolismo , DNA Mitocondrial/metabolismo , Regulação da Expressão Gênica , Masculino , Potencial da Membrana Mitocondrial , Dinâmica Mitocondrial , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Tamanho do Órgão , Estresse Oxidativo , Fosforilação , Ratos Zucker
4.
Clin Sci (Lond) ; 134(12): 1537-1553, 2020 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-32556103

RESUMO

Hyperuricaemia (HUA) significantly increases the risk of metabolic syndrome and is strongly associated with the increased prevalence of high serum free fatty acids (FFAs) and insulin resistance. However, the underlying mechanisms are not well established, especially the effect of uric acid (UA) on adipose tissue, a vital organ in regulating whole-body energy and FFA homeostasis. In the present study, we noticed that adipocytes from the white adipose tissue of patients with HUA were hypertrophied and had decreased UCP1 expression. To test the effects of UA on adipose tissue, we built both in vitro and in vivo HUA models and elucidated that a high level of UA could induce hypertrophy of adipocytes, inhibit their hyperplasia and reduce their beige-like characteristics. According to mRNA-sequencing analysis, UA significantly decreased the expression of leptin in adipocytes, which was closely related to fatty acid metabolism and the AMPK signalling pathway, as indicated by KEGG pathway analysis. Moreover, lowering UA using benzbromarone (a uricosuric agent) or metformin-induced activation of AMPK expression significantly attenuated UA-induced FFA metabolism impairment and adipose beiging suppression, which subsequently alleviated serum FFA elevation and insulin resistance in HUA mice. Taken together, these observations confirm that UA is involved in the aetiology of metabolic abnormalities in adipose tissue by regulating leptin-AMPK pathway, and metformin could lessen HUA-induced serum FFA elevation and insulin resistance by improving adipose tissue function via AMPK activation. Therefore, metformin could represent a novel treatment strategy for HUA-related metabolic disorders.


Assuntos
Adipócitos/patologia , Tecido Adiposo Bege/patologia , Tecido Adiposo Branco/patologia , Ácidos Graxos não Esterificados/sangue , Hiperuricemia/sangue , Hiperuricemia/tratamento farmacológico , Resistência à Insulina , Metformina/uso terapêutico , Células 3T3-L1 , Adenilato Quinase/metabolismo , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Tecido Adiposo Bege/efeitos dos fármacos , Tecido Adiposo Branco/efeitos dos fármacos , Adulto , Animais , Ativação Enzimática , Feminino , Humanos , Hipertrofia , Leptina/metabolismo , Lipogênese , Lipólise , Masculino , Metformina/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Transdução de Sinais , Triglicerídeos/metabolismo , Ácido Úrico/sangue
5.
Life Sci ; 249: 117498, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-32142765

RESUMO

AIMS: Doxorubicin (DOX) is an effective anthracycline anticancer drug. However, the clinical usage of it is limited due to its severe cardiotoxicity side effects. Metformin (Met) is a kind of first-line antihyperglycemic drug which has a potential protective effect on the heart,it is often used for oral treatment of type 2 diabetes. In this study, we explored whether Met could attenuate cardiotoxicity induced by DOX. MATERIALS AND METHODS: For the sake of exploring the Met protective effect and mechanism, we established the DOX-induced cardiotoxicity models both in H9C2 cells incubated with 5 µM DOX in vitro and Sprague-Dawley rats treated with 20 mg/kg cumulative dose of DOX. KEY FINDINGS: Met is able to inhibit growth inhibition and apoptosis of H9C2 cells induced by DOX. The heart indexes of rats were examined to evaluate the Met cardiotoxicity protection. Met improved the abnormal indexes, serum markers of cardiac heart injury, echocardiography, electrocardiogram, cardiac pathology, cardiomyocyte apoptosis, and oxidative stress markers induced by DOX. Furthermore, in vivo and in vitro studies demonstrated that Met protected against DOX-induced increasing cleaved caspase-3 and Bax. Met also prevented the downregulation of Bcl-2, activated the AMPK pathway, and inhibited the MAPK pathway. SIGNIFICANCE: Met showed protective effects on DOX-induced cardiotoxicity by reducing oxidative stress and apoptosis, as well as regulating AMPK and MAPK signaling pathways.


Assuntos
Adenilato Quinase/metabolismo , Antibióticos Antineoplásicos/toxicidade , Doxorrubicina/toxicidade , Coração/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases , Metformina/farmacologia , Animais , Linhagem Celular , Ratos , Ratos Sprague-Dawley
6.
Metabolism ; 106: 154191, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32112822

RESUMO

BACKGROUND: Low-density lipoprotein receptor-related protein 1 (LRP1) plays a key role in fatty acid metabolism and glucose homeostasis. In the context of dyslipemia, LRP1 is upregulated in the heart. Our aim was to evaluate the impact of cardiomyocyte LRP1 deficiency on high fat diet (HFD)-induced cardiac and metabolic alterations, and to explore the potential mechanisms involved. METHODS: We used TnT-iCre transgenic mice with thoroughly tested suitability to delete genes exclusively in cardiomyocytes to generate an experimental mouse model with conditional Lrp1 deficiency in cardiomyocytes (TNT-iCre+-LRP1flox/flox). FINDINGS: Mice with Lrp1-deficient cardiomyocytes (cm-Lrp1-/-) have a normal cardiac function combined with a favorable metabolic phenotype against HFD-induced glucose intolerance and obesity. Glucose intolerance protection was linked to higher hepatic fatty acid oxidation (FAO), lower liver steatosis and increased whole-body energy expenditure. Proteomic studies of the heart revealed decreased levels of cardiac pro-atrial natriuretic peptide (pro-ANP), which was parallel to higher ANP circulating levels. cm-Lrp1-/- mice showed ANP signaling activation that was linked to increased fatty acid (FA) uptake and increased AMPK/ ACC phosphorylation in the liver. Natriuretic peptide receptor A (NPR-A) antagonist completely abolished ANP signaling and metabolic protection in cm-Lrp1-/- mice. CONCLUSIONS: These results indicate that an ANP-dependent axis controlled by cardiac LRP1 levels modulates AMPK activity in the liver, energy homeostasis and whole-body metabolism.


Assuntos
Resistência à Insulina/genética , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Miócitos Cardíacos/metabolismo , Obesidade/genética , Adenilato Quinase/metabolismo , Animais , Fator Natriurético Atrial/metabolismo , Células Cultivadas , Dieta Hiperlipídica , Intolerância à Glucose/genética , Intolerância à Glucose/metabolismo , Intolerância à Glucose/patologia , Metabolismo dos Lipídeos/genética , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/deficiência , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Camundongos Transgênicos , Miócitos Cardíacos/patologia , Obesidade/metabolismo , Obesidade/patologia
7.
Anticancer Res ; 40(3): 1405-1417, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32132037

RESUMO

BACKGROUND/AIM: Niclosamide is an antihe-minthic drug that has shown cytotoxic effects on non-small cell lung carcinoma (NSCLC) cells. However, the exact mechanisms underlying the anti-tumour activity of niclosamide in NSCLC cancer cells remains to be defined. The aim of this study was to evaluate the antitumor activity of niclosamide in human A549 and CL1-5 non-small cell lung cancer cells using in vitro and in vivo. MATERIALS AND METHODS: We investigated the effects of niclosamide on cell viability, apoptosis, the mitochondrial membrane potential (MMP; Δϕm), and autophagy and apoptosis-related protein expression in human A549 and CL1-5 non-small cell lung cancer cells. RESULTS: Niclosamide induced mainly caspase-independent apoptosis through apoptosis-inducible factor (AIF) translocation to the nucleus upon mitochondria damage. Moreover, niclosamide-induced autophagy may act as adaptive response against apoptosis. AMPK/AKT/mTOR pathway were involved in niclosamide-induced cell death and autophagy in response to ATP depletion. Furthermore, niclosamide efficiently suppressed tumor growth and induce autophagy in vivo. CONCLUSION: Niclosamide induced apoptosis by activating the intrinsic and caspase-independent pathway in human A549 and CL1-5 non-small cell lung cancer cells. Therefore, niclosamide is a potential candidate for anti-NSCLC therapy.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Niclosamida/farmacologia , Células A549 , Trifosfato de Adenosina/metabolismo , Adenilato Quinase/metabolismo , Animais , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/patologia , Processos de Crescimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Ativação Enzimática/efeitos dos fármacos , Humanos , Neoplasias Pulmonares/patologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Distribuição Aleatória , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Am J Physiol Endocrinol Metab ; 318(5): E613-E624, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32154742

RESUMO

The angiopoietin-like protein (ANGPTL) family represents a promising therapeutic target for dyslipidemia, which is a feature of obesity and type 2 diabetes (T2DM). The aim of the present study was to determine the metabolic role of ANGPTL8 and to investigate its nutritional, hormonal, and molecular regulation in key metabolic tissues. The regulation of Angptl8 gene expression by insulin and glucose was quantified using a combination of in vivo insulin clamp experiments in mice and in vitro experiments in primary and cultured hepatocytes and adipocytes. The role of AMPK signaling was examined, and the transcriptional control of Angptl8 was determined using bioinformatic and luciferase reporter approaches. The metabolism of Angptl8 knockout mice (ANGPTL8-/-) was examined following chow and high-fat diets (HFD). Insulin acutely increased Angptl8 expression in liver and adipose tissue, which involved the CCAAT/enhancer-binding protein (C/EBPß) transcription factor. In insulin clamp experiments, glucose further enhanced Angptl8 expression in the presence of insulin in adipose tissue. The activation of AMPK signaling antagonized the effect of insulin on Angptl8 expression in hepatocytes and adipocytes. The ANGPTL8-/- mice had improved glucose tolerance and displayed reduced fed and fasted plasma triglycerides. However, there was no change in body weight or steatosis in ANGPTL8-/- mice after the HFD. These data show that ANGPTL8 plays important metabolic roles in mice that extend beyond triglyceride metabolism. The finding that insulin, glucose, and AMPK signaling regulate Angptl8 expression may provide important clues about the distinct function of ANGPTL8 in these tissues.


Assuntos
Tecido Adiposo/metabolismo , Proteínas Semelhantes a Angiopoietina/metabolismo , Glucose/metabolismo , Homeostase/fisiologia , Fígado/metabolismo , Células 3T3-L1 , Adenilato Quinase/metabolismo , Proteínas Semelhantes a Angiopoietina/genética , Animais , Dieta Hiperlipídica , Regulação da Expressão Gênica , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Insulina/farmacologia , Camundongos , Camundongos Knockout , Transdução de Sinais/efeitos dos fármacos
9.
Am J Physiol Endocrinol Metab ; 318(5): E791-E805, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32182124

RESUMO

Irisin, a newly identified myokine, is critical to modulating body metabolism and biological homeostasis. However, whether irisin protects the skeletal muscles against metabolic stresses remains unknown. In this study, we determine the effect of irisin on high glucose and fatty acid-induced damages using irisin-overexpressed mouse C2C12 (irisin-C2C12) myoblasts and skeletal muscle from irisin-injected mice. Compared with empty vector-transfected control C2C12 cells, irisin overexpression resulted in a marked increase in cell viability and decrease in apoptosis under high-glucose stress. Progression of the cell cycle into the G2/M phase in the proliferative condition was also observed with irisin overexpression. Furthermore, glucose uptake, glycogen accumulation, and phosphorylation of AMPKα/insulin receptor (IR) ß-subunit/Erk1/2 in response to insulin stimulation were enhanced by irisin overexpression. In irisin-C2C12 myoblasts, these responses of phosphorylation were preserved under palmitate treatment, which induced insulin resistance in the control cells. These effects of irisin were reversed by inhibiting AMPK with compound C. In addition, high glucose-induced suppression of the mitochondrial membrane potential was also prevented by irisin. Moreover, suppression of IR in irisin-C2C12 myoblasts by cotransfection of shRNA against IR also mitigated the effects of irisin while not affecting AMPKα phosphorylation. As an in vivo study, soleus muscles from irisin-injected mice showed elevated phosphorylation of AMPKα and Erk1/2 and glycogen contents. Our results indicate that irisin counteracts the stresses generated by high glucose and fatty acid levels and irisin overexpression serves as a novel approach to elicit cellular protection. Furthermore, AMPK activation is a crucial factor that regulates insulin action as a downstream target.


Assuntos
Adenilato Quinase/metabolismo , Fibronectinas/farmacologia , Glucose/farmacologia , Mioblastos/efeitos dos fármacos , Ácido Palmítico/farmacologia , Receptor de Insulina/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Fibronectinas/genética , Fibronectinas/metabolismo , Resistência à Insulina/fisiologia , Camundongos , Mioblastos/metabolismo , Fosforilação/efeitos dos fármacos , Transdução de Sinais/fisiologia
10.
Chem Biol Interact ; 317: 108943, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31926917

RESUMO

Epidemiological studies have shown that cigarette smoking is beneficial in ulcerative colitis and that nicotine may be responsible for this effect. However, the mechanism remains unclear. In a previous study, nicotine was found to induce autophagy in intestinal cells. Here, we evaluated the effect of nicotine-induced autophagy in a dextran sodium sulfate (DSS)-induced colitis mouse model. C57BL/6 adult male mice drank DSS water solution freely for seven consecutive days, and then tap water was administered. The effect of nicotine treatment was examined in the DSS model, including colon length, disease severity, histology of the colon tissue, and inflammation levels. Moreover, autophagy levels were detected by Western blot analysis (LC3II/LC3I, p62, and beclin-1). The levels of DSS-induced colitis were significantly decreased following nicotine treatment. The disease activity score, body weight, histologic damage scores, and the level of colonic inflammatory factors of nicotine-treated mice all decreased compared to those of the control mice. Additionally, nicotine enhanced the expression of LC3II/LC3I and beclin-1 but decreased the p62 protein level. Inhibiting autophagy by 3-MA attenuated the protective effects of nicotine on colitis. Additionally, both in vitro and in vivo experiments showed changes in AMPK-mTOR-P70S6K during this process. These results suggest that nicotine improved colitis by regulating autophagy and provided a protective effect against DSS-induced colitis.


Assuntos
Adenilato Quinase/metabolismo , Autofagia/efeitos dos fármacos , Colite/prevenção & controle , Nicotina/farmacologia , Serina-Treonina Quinases TOR/metabolismo , Adenina/análogos & derivados , Adenina/farmacologia , Adenilato Quinase/genética , Animais , Colite/induzido quimicamente , Sulfato de Dextrana/toxicidade , Regulação da Expressão Gênica/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Quinases S6 Ribossômicas 70-kDa/genética , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Serina-Treonina Quinases TOR/genética
11.
Anticancer Res ; 40(1): 323-333, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31892583

RESUMO

BACKGROUND/AIM: Despite the Warburg effect, mitochondria play an essential role in the survival and maintenance of cancer cells. Thus, mitochondria have been considered a target for anticancer agents. Here, we identified a mitochondria-targeting anticancer agent from natural products. MATERIALS AND METHODS: Morphological and functional changes in mitochondria were determined by a fluorescence-based High Content Imaging System. Using human non-small cell lung cancer (NSCLC) cell lines (H1299, H226B, and A549), cell viability and colony formation assays, cell cycle analysis, and immunoblotting were performed to determine cytotoxic and proapoptotic effects of papuamine. RESULTS: Using a natural product chemical library, we identified papuamine as an active compound to inhibit viability and ATP production of NSCLC cells. Papuamine depleted intracellular ATP by causing mitochondrial dysfunction, as indicated by the loss of the mitochondrial membrane potential and increased mitochondrial superoxide generation. Papuamine significantly inhibited viability and colony formation of NSCLC cells by inducing apoptosis. CONCLUSION: Papuamine has a potential as a novel mitochondria-targeting anticancer agent.


Assuntos
Alcaloides/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Mitocôndrias/patologia , Células A549 , Trifosfato de Adenosina/metabolismo , Adenilato Quinase/metabolismo , Alcaloides/química , Alcaloides/farmacologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Sobrevivência Celular/efeitos dos fármacos , Humanos , Neoplasias Pulmonares/patologia , Mitocôndrias/efeitos dos fármacos , Ensaio Tumoral de Célula-Tronco , Regulação para Cima/efeitos dos fármacos
12.
Exp Mol Pathol ; 113: 104371, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31917290

RESUMO

As the number of elderly patients increases, some patients with heart problems may also need surgery. The purpose of this study was to investigate whether dexmedetomidine (DEX), a common used anesthetic, was beneficial to the patients with heart problems. Myocardial cells induced by doxorubicin (DOX) was to simulate the myocardium injury in vitro. H9c2 cells were treated with DOX, DEX/DOX, Compound C and Compound C/DEX/DOX, respectively. The expression of p-AMPK, AMPK, p-GSK3ß, GSK3ß, Bcl2, Bax, Cleaved caspase3, Caspase3, TXNIP, NLRP3, ASC, Cleaved caspase-1 and Caspase-1 were analyzed by Western blot. CCK-8 assay and flow cytometry analysis were used to detect the cell viability and cell apoptosis. The levels of TNF-α, IL-1ß and IL-18 were detected by ELISA assay and the levels of NO, ROS, LDH, SOD, MDA and taurine were detected by corresponding assay kits. As a result, DEX promoted the cell viability and inhibited the inflammation, oxidative stress and apoptosis. In addition, DEX suppressed the expression of taurine, TXNIP, NLRP3, ASC and cleaved caspase-1 and activated the expression of p-AMPK and p-GSK3ß. However, those above changes could be reversed by Compound C. In conclusion, this study indicated that DEX could reduce the inflammation and apoptosis of DOX-induced myocardial cells through activating the AMPK-GSK3ß signaling pathway. Because of the above effects of DEX, it may be beneficial for surgical patients with heart problems.


Assuntos
Apoptose/efeitos dos fármacos , Dexmedetomidina/farmacologia , Doxorrubicina/efeitos adversos , Inflamação/patologia , Miócitos Cardíacos/patologia , Adenilato Quinase/metabolismo , Animais , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Glicogênio Sintase Quinase 3 beta/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Óxido Nítrico/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Taurina/metabolismo
13.
Food Chem Toxicol ; 135: 111043, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31836553

RESUMO

Tylorrhynchus heterochaetus (Hechong in Chinese) has been used in Chinese traditional medicine for treating various diseases. This study was aimed to assess the anti-fatigue effect of T. heterochaetus on Kunming mice and its primary mechanism of action using forced running, rotating rod and weight-loaded swimming tests. Low (2.70 mg/0.5 mL/20 g), medium (5.41 mg/0.5 mL/20 g) and high (6.58 mg/0.5 mL/20 g) doses of T. heterochaetus aqueous extract were treated to mice for 28 days. Among the doses, the low and medium doses showed significant (p ≤ 0.05) anti-fatigue effect on the weight-loaded swimming test. Also, T. heterochaetus extract showed significant (p ≤ 0.05) effects on fatigue-related blood parameters by increasing the GLU, TG and LDH levels and decreasing the LA, CK and BUN levels. The levels of liver and skeletal muscle glycogen were also significantly (p ≤ 0.05) increased after treatment. Further, on Western blot analysis, it has been found that T. heterochaetus enhanced the expressions of AMPK and PGC-1α in the liver and skeletal muscles of mice. From the study, our outcomes suggest that T. heterochaetus possess an anti-fatigue effect through the AMPK-linked pathway and thereby it can regularize the energy metabolism.


Assuntos
Adenilato Quinase/metabolismo , Produtos Biológicos/farmacologia , Fadiga/prevenção & controle , Poliquetos/química , Animais , Nitrogênio da Ureia Sanguínea , Peso Corporal/efeitos dos fármacos , Creatina Quinase/metabolismo , Glucose/metabolismo , Glicogênio/metabolismo , L-Lactato Desidrogenase/metabolismo , Ácido Láctico/metabolismo , Fígado/metabolismo , Camundongos , Músculo Esquelético/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Teste de Desempenho do Rota-Rod , Corrida , Natação , Triglicerídeos/metabolismo
14.
J Biotechnol ; 307: 202-207, 2020 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-31672531

RESUMO

Addressing the challenges associated with the development of in vitro biocatalytic carboxylate reductions for potential applications, important aspects of the co-factor regeneration systems and strategies for minimizing over-reduction were investigated. The ATP recycling can be performed with similarly high efficiency exploiting the polyphosphate source by combining Meiothermus ruber polyphosphate kinase and adenylate kinase or with Sinorhizobium meliloti polyphosphate kinase instead of the latter. Carboxylate reductions with the enzyme candidates used in this work allow operating at co-factor concentrations of adenosine 5'-triphosphate and ß-nicotinamide adenine dinucleotide 2'-phosphate of 100 µM and, thereby, reducing the amounts of alcohols formed by side activities in the enzyme preparations. This study confirmed the expected benefits of carboxylic acid reductases in chemoselectively reducing the carboxylates to the corresponding aldehydes while leaving reductively-sensitive nitro, ester and cyano groups intact.


Assuntos
Aldeídos/metabolismo , Bactérias/enzimologia , Ácidos Carboxílicos/metabolismo , Fosfotransferases (Aceptor do Grupo Fosfato)/metabolismo , Sinorhizobium meliloti/enzimologia , Trifosfato de Adenosina/metabolismo , Adenilato Quinase/genética , Adenilato Quinase/metabolismo , Álcoois , Bactérias/genética , Sistema Livre de Células , NADP/metabolismo , Fosfotransferases (Aceptor do Grupo Fosfato)/genética , Sinorhizobium meliloti/genética
15.
J Nutr ; 150(4): 672-684, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-31858105

RESUMO

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) is the most common liver disease in the world. Hepatic de novo lipogenesis (DNL) has been suggested to contribute to the pathogenesis of NAFLD. Recent studies have demonstrated that niacin (NA) modulates hepatic DNL through GPR109A. However, the underlying mechanism remains largely unknown. OBJECTIVES: This study aims to elucidate the potential molecular mechanism by which GPR109A inhibits hepatic DNL. METHODS: C57BL/6 wild-type (WT) and Gpr109a knockout (KO) mice (male, 5 wk old) were fed a high-fat diet (60% energy from fat) firstly for 6 wk to generate a diet-induced obese model. Subsequently, they were randomly divided into 4 groups for the next 8-9 wk: WT mice with oral water [WT + vehile (VE)], WT mice with oral NA (50 mM, dissolved in water) (WT + NA), KO mice with oral water (KO + VE), and KO mice with oral NA (50 mM) (KO + NA). Mechanisms were examined in HepG2 cells. Body composition, liver histology, biomarkers of hepatic function, lipid accumulation, and lipid synthesis signals in HepG2 cells were measured. RESULTS: Upon activation, GPR109A apparently protected against obesity and hepatic steatosis (P < 0.05). The concentrations of hepatic Tnf-α in the WT + NA group were about 50% of those in the WT + VE group (P < 0.05). The activities of serum alanine transaminase and aspartate transaminase were 26.7% and 53.5% lower in the WT + NA group than in the WT + VE group, respectively (P < 0.05). In HepG2 cells, activation of GPR109A resulted in remarkable inhibition of oleic acid-induced lipid accumulation via a protein kinase C-extracellular signal-regulated kinase-1/2-AMP-activated protein kinase signaling pathway. CONCLUSIONS: NA inhibits hepatic lipogenesis in C57BL/6 mice through a GPR109A-mediated signaling pathway, consistent with the mechanistic studies in HepG2 cells, suggesting its potential for treatment of NAFLD and other fatty liver diseases.


Assuntos
Adenilato Quinase/metabolismo , Lipogênese/fisiologia , Sistema de Sinalização das MAP Quinases/fisiologia , Niacina/administração & dosagem , Hepatopatia Gordurosa não Alcoólica/prevenção & controle , Receptores Acoplados a Proteínas-G/fisiologia , Animais , Dieta Hiperlipídica , Técnicas de Silenciamento de Genes , Células Hep G2 , Humanos , Lipogênese/efeitos dos fármacos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Hepatopatia Gordurosa não Alcoólica/etiologia , Obesidade/etiologia , Obesidade/prevenção & controle , Receptores Acoplados a Proteínas-G/deficiência , Transdução de Sinais
16.
Mol Cell Endocrinol ; 499: 110591, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31546019

RESUMO

Metformin (MET) is the most widely prescribed hypoglycemic drug in type 2 diabetes and Polycystic Ovary Syndrome. Besides its effects on glucose metabolism, MET exerts beneficial effects on these patients' fertility. However, the exact mechanisms of action of MET on female fertility are still unclear. In this work, we analyzed a possible direct effect of MET on ovarian cells. We found expression of the organic cation transporters OCT1, OCT2 and OCT3, responsible for MET uptake into the cells, in rat granulosa cells and human cumulus cells. Furthermore, MET increased pAMPK and decreased VEGF levels both in vivo and in rat granulosa cells in culture. These last effects were reversed when OCTs were inhibited. Our results suggest that MET acts directly on ovarian cells regulating cell metabolism and VEGF expression. Our findings are relevant to optimize PCOS fertility treatment and to explore ovarian MET actions in other female pathologies.


Assuntos
Adenilato Quinase/metabolismo , Células do Cúmulo/citologia , Metformina/administração & dosagem , Fatores de Transcrição de Octâmero/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Adulto , Animais , Proliferação de Células/efeitos dos fármacos , Células do Cúmulo/efeitos dos fármacos , Células do Cúmulo/metabolismo , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Metformina/farmacologia , Modelos Animais , Fosforilação/efeitos dos fármacos , Ratos
17.
Dis Markers ; 2019: 8186091, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31827645

RESUMO

Breast cancer (BC) is a type of malignant tumor originating from the epithelial tissue of the mammary gland, and about 20% of breast cancers are human epidermal growth factor receptor 2 positive (HER2+), which is a subtype with more aggression. Recently, HER2-positive breast cancer is often accompanied by poor prognosis of patients, and targeted therapy showed a promising prospect. To combat this disease, novel therapeutic targets are still needed. Adenylate kinase 4 (AK4) is a member of the adenylate kinase family and is expressed in the mitochondrial matrix. AK4 is involved in multiple cellular functions such as energy metabolism homeostasis. Interestingly, AK4 was observed highly expressed in several tumor tissues, and the involvement of AK4 in cancer development was generally revealed. However, the possible role of AK4 on the growth and development of breast cancer is still unclear. Here, we investigated the possible functions of AK4 on the progression of HER2-positive breast cancer. We found the high expression of AK4 in HER2-positive breast cancer tissues from patients who received surgical treatment. Additionally, AK4 expression levels were obviously correlated with clinical-pathological features, including pTNM stage (P = 0.017) and lymph node metastasis (P = 0.046). We mechanically confirmed that AK4 depletion showed the obvious impairment of cell proliferation and invasion in MCF7 and MDA-MB-231 cells. AK4 also facilitates tumor growth and metastasis of HER2-positive breast cancer in vivo. In conclusion, we identified and mechanically confirmed that AK4 is a novel therapeutic target of HER2-positive breast cancer.


Assuntos
Adenilato Quinase/metabolismo , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/patologia , Proliferação de Células , Neoplasias Pulmonares/secundário , Receptor ErbB-2/metabolismo , Animais , Apoptose , Neoplasias da Mama/metabolismo , Neoplasias da Mama/terapia , Terapia Combinada , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/terapia , Metástase Linfática , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Invasividade Neoplásica , Prognóstico , Taxa de Sobrevida , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Phys Rev E ; 100(5-1): 052409, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31869999

RESUMO

Natural enzymes often have enormous catalytic power developed by evolution. Revealing the underlying physical strategy used by enzymes to achieve high catalysis efficiency is one of the central focuses in the field of biological physics. Our recent work demonstrated that multisubstrate enzymes can utilize steric frustration encountered in the substrate-product cobound complex to overcome the bottleneck of the enzymatic cycle [W. Li et al., Phys. Rev. Lett. 122, 238102 (2019)10.1103/PhysRevLett.122.238102]. However, the key atomic-level interactions by which the steric frustration contributes to the enzymatic cycle remain elusive. In this work we study the microscopic mechanism for the role of the substrate-product frustration on the key physical steps in the enzymatic cycle of adenylate kinase (AdK), a multisubstrate enzyme catalyzing the reversible phosphoryl transfer reaction ATP+AMP⇋ADP+ADP. By using atomistic molecular dynamics simulations with enhanced sampling, we showed that the competitive interactions from the phosphate groups of the substrate ATP and product ADP in the ATP-ADP cobound complex of the AdK lead to local frustration in the binding pockets. Such local frustration disrupts the hydrogen bond network around the binding pockets, which causes lowered barrier height for the opening of the enzyme conformations and expedited release of the bottleneck product ADP. Our results directly demonstrated from the atomistic level that the local frustration in the active sites of the enzyme can be utilized to facilitate the key physical steps of the enzymatic cycle, providing numerical evidence to the predictions of the previous theoretical work.


Assuntos
Adenilato Quinase/metabolismo , Simulação de Dinâmica Molecular , Difosfato de Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Adenilato Quinase/química , Sítios de Ligação , Cinética , Ligação Proteica , Conformação Proteica , Termodinâmica
19.
Cell Rep ; 29(13): 4540-4552.e8, 2019 12 24.
Artigo em Inglês | MEDLINE | ID: mdl-31875559

RESUMO

A growing body of evidence suggests that changes in fat metabolism may have a significant effect on lifespan. Accumulation of lipid deposits in non-adipose tissue appears to be critical for age-related pathologies and may also contribute to the aging process itself. We established a model of lipid storage in muscle cells of C. elegans to reveal a mechanism that promotes longevity non-cell-autonomously. Here, we describe how muscle-specific activation of adipose triglyceride lipase (ATGL) and the phospholipase A2 (PLA2) ortholog IPLA-7 collectively affect inter-tissular communication and systemic adaptation that requires the activity of AMP-dependent protein kinase (AMPK) and a highly conserved nuclear receptor outside of the muscle. Our data suggest that muscle-specific bioactive lipid signals, or "lipokines," are generated following triglyceride breakdown and that these signals impinge on a complex network of genes that modify the global lipidome, consequently extending the lifespan.


Assuntos
Caenorhabditis elegans/metabolismo , Lipidômica , Lipídeos/química , Longevidade/fisiologia , Músculos/metabolismo , Adenilato Quinase/metabolismo , Animais , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Dieta , Ativação Enzimática , Hidrólise , Gotículas Lipídicas/metabolismo , Lipólise , Células Musculares/metabolismo , Especificidade de Órgãos , Fatores de Transcrição/metabolismo
20.
Food Funct ; 10(11): 7509-7522, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31670355

RESUMO

Enterotoxigenic Escherichia coli (ETEC) triggers diarrhea in humans and livestock. We have previously showed that ETEC promotes intestinal epithelial cell apoptosis and increases gamma-aminobutyric acid (GABA) concentration in the jejunum, suggesting that GABA might mediate ETEC-induced apoptosis. Here, we found that GABA alleviates ETEC-induced intestinal barrier dysfunctions, including ETEC-induced apoptosis both in vivo and in vitro. Interestingly, the alleviation of GABA on ETEC-induced apoptosis largely depends on autophagy. Mechanistically, GABA attenuates ETEC-induced apoptosis via activating GABAAR signaling and the AMPK-autophagy pathway. These findings highlight that maintaining intestinal GABA concentration could alleviate intestinal ETEC infection.


Assuntos
Adenilato Quinase/metabolismo , Apoptose/efeitos dos fármacos , Escherichia coli Enterotoxigênica , Células Epiteliais/microbiologia , Infecções por Escherichia coli/metabolismo , Receptores de GABA-A/metabolismo , Ácido gama-Aminobutírico/farmacologia , Adenilato Quinase/genética , Animais , Autofagia/efeitos dos fármacos , Autofagia/fisiologia , Linhagem Celular , Células Epiteliais/efeitos dos fármacos , Infecções por Escherichia coli/patologia , Mucosa Intestinal/citologia , Receptores de GABA-A/genética , Suínos
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