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1.
Adv Exp Med Biol ; 1155: 163-172, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31468395

RESUMO

Taurine (2-aminoethanesulfonic acid), a sulfur-containing ß-amino acid, is a free amino acid present in high concentrations in mammalian tissues. Taurine has pivotal roles in anti-oxidation, membrane stabilization, osmoregulation, anti-inflammation, and other process. In a DNA microarray analysis, we previously found that taurine markedly increases the mRNA expression of thioredoxin interacting protein (TXNIP) in Caco-2 cells. In this study, we investigated the effect of these taurine-induced changes in TXNIP on the function of Caco-2 cells. We found that taurine decreases glucose uptake in a dose-dependent manner. The taurine-induced decrease in glucose uptake was completely abolished by transfection with siRNA against TXNIP, suggesting that TXNIP is involved in the taurine-induced down-regulation of glucose uptake. We also revealed that taurine induces AMPK activation and further increases the intracellular ATP content in Caco-2 cells. These results suggest that taurine could regulate the function of Caco-2 cells via TXNIP induction, leading to extend our understanding of the functions of taurine.


Assuntos
Proteínas de Transporte/metabolismo , Glucose/metabolismo , Taurina/farmacologia , Trifosfato de Adenosina/metabolismo , Adenilato Quinase/metabolismo , Transporte Biológico , Células CACO-2 , Regulação para Baixo , Humanos , RNA Interferente Pequeno
2.
Nat Commun ; 10(1): 2943, 2019 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-31270333

RESUMO

Mutations exclusively in equilibrative nucleoside transporter 3 (ENT3), the only intracellular nucleoside transporter within the solute carrier 29 (SLC29) gene family, cause an expanding spectrum of human genetic disorders (e.g., H syndrome, PHID syndrome, and SHML/RDD syndrome). Here, we identify adult stem cell deficits that drive ENT3-related abnormalities in mice. ENT3 deficiency alters hematopoietic and mesenchymal stem cell fates; the former leads to stem cell exhaustion, and the latter leads to breaches of mesodermal tissue integrity. The molecular pathogenesis stems from the loss of lysosomal adenosine transport, which impedes autophagy-regulated stem cell differentiation programs via misregulation of the AMPK-mTOR-ULK axis. Furthermore, mass spectrometry-based metabolomics and bioenergetics studies identify defects in fatty acid utilization, and alterations in mitochondrial bioenergetics can additionally propel stem cell deficits. Genetic, pharmacologic and stem cell interventions ameliorate ENT3-disease pathologies and extend the lifespan of ENT3-deficient mice. These findings delineate a primary pathogenic basis for the development of ENT3 spectrum disorders and offer critical mechanistic insights into treating human ENT3-related disorders.


Assuntos
Células-Tronco Adultas/metabolismo , Proteínas de Transporte de Nucleosídeos/metabolismo , Adenosina/metabolismo , Adenilato Quinase/metabolismo , Células-Tronco Adultas/ultraestrutura , Aminoimidazol Carboxamida/análogos & derivados , Aminoimidazol Carboxamida/farmacologia , Animais , Autofagia , Transporte Biológico , Diferenciação Celular , Autorrenovação Celular , Metabolismo Energético , Ácidos Graxos/metabolismo , Células HEK293 , Humanos , Metabolismo dos Lipídeos , Lisossomos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Modelos Biológicos , Fenótipo , Ribonucleotídeos/farmacologia , Transdução de Sinais , Análise de Sobrevida , Serina-Treonina Quinases TOR/metabolismo
3.
Exp Parasitol ; 204: 107726, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31299264

RESUMO

The aims of this study were to evaluate if the use of copper oxide wire particles, isolated or in association with closantel, in lambs infected with Haemonchus contortus enhances the anthelmintic efficacy of closantel, as well as to evaluate the effects of treatment in hepatic energy metabolism, inflammatory markers and hematological and biochemical tests. The lambs were randomly divided into five groups (6 animals each), as follows: uninfected animals (Control); animals infected with H. contortus (HC); infected and treated with closantel (HC + CL); infected and treated with copper oxide wire particles (HC + Cu); and infected and treated with closantel plus copper oxide wire particles (HC + CL + Cu). The animals of infected groups were infected orally with H. contortus (5,000 L3 -larvae) and on day 14 post infection (p.i) the treatments were initiated. The egg per gram of feces (EPG), butyrylcholinesterase (BuChE), myeloperoxidase (MPO), adenylate kinase (AK) and pyruvate kinase (PK) activities and hematological and biochemical tests were evaluated. Treatments with copper oxide (isolated and associated) were able to reduce the EPG count on days 28, 35, 42 and 49 p.i when compared to HC group, while closantel was able to reduce EPG only from day 35 p.i. Moreover, treatment with closantel (isolated or associated) was able to prevent the inhibition of hepatic AK and PK activities caused by H. contortus infection, which may contribute to efficient intracellular energetic communication in order to maintain the balance between cellular ATP consumption and production. Butyrylcholinesterase and MPO activities were higher in infected lambs compared to uninfected, while treated groups showed lower enzymatic activity compared to the group HC. The use of all therapeutic protocols was able to reduce the EPG count. Based on these evidences, the use of copper oxide plus closantel may be considered an alternative to treat lambs infected by H. contortus.


Assuntos
Anti-Helmínticos/administração & dosagem , Cobre/administração & dosagem , Hemoncose/veterinária , Inflamação/veterinária , Salicilanilidas/administração & dosagem , Doenças dos Ovinos/tratamento farmacológico , Abomaso/metabolismo , Adenilato Quinase/metabolismo , Administração Oral , Animais , Anti-Helmínticos/farmacologia , Anti-Helmínticos/uso terapêutico , Análise Química do Sangue/veterinária , Butirilcolinesterase/sangue , Cápsulas , Metabolismo Energético/efeitos dos fármacos , Contagem de Eritrócitos/veterinária , Fezes/parasitologia , Hemoncose/complicações , Hemoncose/tratamento farmacológico , Hemoncose/metabolismo , Hematócrito/veterinária , Hemoglobinas/análise , Inflamação/tratamento farmacológico , Inflamação/prevenção & controle , Fígado/efeitos dos fármacos , Fígado/enzimologia , Fígado/metabolismo , Masculino , Contagem de Ovos de Parasitas/veterinária , Peroxidase/sangue , Piruvato Quinase/metabolismo , Distribuição Aleatória , Salicilanilidas/farmacologia , Salicilanilidas/uso terapêutico , Ovinos , Doenças dos Ovinos/metabolismo , Doenças dos Ovinos/prevenção & controle
4.
Phys Rev Lett ; 122(23): 238102, 2019 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-31298900

RESUMO

The enormous catalytic power of natural enzymes relies on the ability to overcome the bottleneck event in the enzymatic cycle, yet the underlying physical mechanisms are not fully understood. Here, by performing molecular simulations of the whole enzymatic cycle for a model multisubstrate enzyme with a dynamic energy landscape model, we show that multisubstrate enzymes can utilize steric frustration to facilitate the rate-limiting product-release step. During the enzymatic cycles, the bottleneck product is actively squeezed out by the binding of a new substrate at the neighboring site through the population of a substrate-product cobound complex, in which the binding pockets are frustrated due to steric incompatibility. Such steric frustration thereby enables an active mechanism of product release driven by substrate-binding energy, facilitating the enzymatic cycle.


Assuntos
Enzimas/química , Enzimas/metabolismo , Modelos Químicos , Adenilato Quinase/química , Adenilato Quinase/metabolismo , Modelos Moleculares , Conformação Proteica , Relação Estrutura-Atividade , Termodinâmica
5.
Parasitol Int ; 72: 101929, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31108219

RESUMO

In Leishmania spp. ATP utilizing enzymes serves as a key role in preserving integrity of host cells for survival of parasite. Earlier reports suggested that Adenylate kinase (AK) a phosphotransferase enzyme released by Leishmania donovani secretome, involved in modulating levels of NTPs. In the present study, we cloned, expressed and characterized recombinant putative AK. Based on a sequence and phylogeny analysis, we identified the prominent features of the seven AK isoforms of Leishmania donovani and assigned our putative AK as LdAK2a. The Km value of LdAK2a for ATP and AMP substrate were 204 µM and 184 µM, respectively and Vmax was calculated as 1.6 µmol min-1 mg-1 protein. Ap5A, a known inhibitor of AK inhibited LdAK2a with estimated Ki values of 280 nM and 230 nM for ATP and AMP respectively. CD spectral studies were carried out to estimate its structural stability. Recombinant LdAK2a was found to prevent ATP mediated cell cytolysis of Raw 264.7 macrophages in vitro, which was determined by LDH assay and MMP assay. This is the first report which validates that Leishmanial AK2a can prevent ATP mediated cytolysis of macrophage cells and thereby probably play a role in preserving integrity of host cells for survival of parasite.


Assuntos
Trifosfato de Adenosina/metabolismo , Adenilato Quinase/metabolismo , Interações Hospedeiro-Parasita , Leishmania donovani/enzimologia , Macrófagos/patologia , Macrófagos/parasitologia , Adenilato Quinase/genética , Animais , Morte Celular , Cinética , Leishmaniose Visceral , Camundongos , Células RAW 264.7
6.
Mol Med Rep ; 19(6): 4935-4945, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31059086

RESUMO

Previous reports have indicated a potential link between microRNA (miR)­214 and hypoxia. In the present study, the biological functions and potential mechanisms of miR­214 were determined, as well as its correlation with HIF­1α signaling in non­small cell lung cancer (NSCLC) cells. Quantitative polymerase chain reaction revealed that miR­214 expression was upregulated in lung cancer tissues compared with adjacent normal tissues. miR­214 mimics were transfected into A549 cells, and MTT, colony formation, invasion and wound healing assays were performed. It was demonstrated that miR­214 mimic transfection promoted the invasion, proliferation and migration of A549 cells. Furthermore, miR­214 inhibitor transfection decreased H1299 cell invasion, proliferation and migration. Next, the association between miR­214 expression and the HIF­1α signaling cascade was examined. It was demonstrated that miR­214 mimics upregulated the expression of hypoxia­inducible factor (HIF)­1α, vascular endothelial growth factor (VEGF), adenylate kinase 3 and matrix metalloproteinase (MMP)2, whereas miR­214 inhibitor downregulated the expression of these factors. Using prediction software, it was demonstrated that tumor suppressor ING4 was a target of miR­214. A luciferase reporter assay confirmed that ING4 was a direct target of miR­214. There was a negative correlation between ING4 and miR­214 expression in lung cancer tissues. In addition, ING4 siRNA and plasmid was transfected into cells in order to validate its effect on HIF­1α, MMP2 and VEGF expression. ING4 overexpression downregulated HIF­1α and its targets MMP2 and VEGF, while ING4 siRNA upregulated HIF­1α, MMP2 and VEGF. In conclusion, it was demonstrated that miR­214 targeted ING4 in lung cancer cells, and upregulated the HIF­1α cascade, leading to MMP2 and VEGF upregulation. This approach may help to clarify the role of miRNA in non­small lung cancer and may be a new therapeutic target for non­small lung cancer.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Proteínas de Homeodomínio/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias Pulmonares/patologia , MicroRNAs/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Células A549 , Adenilato Quinase/genética , Adenilato Quinase/metabolismo , Adulto , Idoso , Antagomirs/metabolismo , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Feminino , Proteínas de Homeodomínio/antagonistas & inibidores , Proteínas de Homeodomínio/genética , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Neoplasias Pulmonares/metabolismo , Masculino , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Pessoa de Meia-Idade , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Proteínas Supressoras de Tumor/antagonistas & inibidores , Proteínas Supressoras de Tumor/genética , Fator A de Crescimento do Endotélio Vascular/genética
7.
Eur J Med Chem ; 175: 234-246, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31082766

RESUMO

Selenocyanates and diselenides are potential antitumor agents. Here we report two series of selenium derivatives related to selenocyanates and diselenides containing carboxylic, amide and imide moieties. These compounds were screened for their potency and selectivity against seven tumor cell lines and two non-malignant cell lines. Results showed that MCF-7 cells were especially sensitive to the treatment, with seven compounds presenting GI50 values below 10 µM. Notably, the carboxylic selenocyanate 8b and the cyclic imide 10a also displayed high selectivity for tumor cells. Treatment of MCF-7 cells with these compounds resulted in cell cycle arrest at S phase, increased levels of pJNK and pAMPK and caspase independent cell death. Autophagy inhibitors wortmannin and chloroquine partially prevented 8b and 10a induced cell death. Consistent with autophagy, increased Beclin1 and LC3-IIB and reduced SQSTM1/p62 levels were detected. Our results point to 8b and 10a as autophagic cell death inducers.


Assuntos
Adenilato Quinase/metabolismo , Autofagia/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Citostáticos/farmacologia , MAP Quinase Quinase 4/metabolismo , Compostos Organosselênicos/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Citostáticos/administração & dosagem , Citostáticos/química , Relação Dose-Resposta a Droga , Ativação Enzimática , Humanos , Compostos Organosselênicos/administração & dosagem , Compostos Organosselênicos/química , Fase S/efeitos dos fármacos
8.
Int J Nanomedicine ; 14: 2861-2877, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31118612

RESUMO

Background: Combination therapy remains a promising strategy for treating neurodegenerative diseases, although green synthesis of gold nanoparticles for treating chronic neuroinflammation and studying their efficacy in treating neuroinflammation-mediated neurodegenerative diseases is not well assessed. Results: Here, Ephedra sinica Stapf (ES) extract was used as the reducing, capping, and stabilizing agent for gold nanoparticle synthesis. We developed ES extract-capped gold nanoparticles (ES-GNs) and investigated their anti-neuroinflammatory properties in microglia. ES-GNs displayed maximum absorption at 538 nm in ultraviolet-visible spectroscopy. Dynamic light scattering assessment revealed that ES-GN diameter was 57.6±3.07 nm, with zeta potential value of -24.6±0.84 mV. High resolution-transmission electron microscopy confirmed the spherical shape and average diameter (35.04±4.02 nm) of ES-GNs. Crystalline structure of ES-GNs in optimal conditions was determined by X-ray powder diffraction, and elemental gold presence was confirmed by energy-dispersive X-ray spectroscopy. Fourier transform-infrared spectroscopy confirmed gold nanoparticle synthesis using ES. Anti-neuroinflammatory properties of ES-GNs on production of pro-inflammatory mediators (nitric oxide, prostaglandin E2, and reactive oxygen species) and cytokines (tumor necrosis factor-α, IL-1ß, and IL-6) in lipopolysaccharide (LPS)-stimulated microglia were investigated by ELISA and flow cytometry. ES-GNs significantly attenuated LPS-induced production of pro-inflammatory mediators and cytokines, which was related to suppressed transcription and translation of inducible nitric oxide synthase and cyclooxygenase-2, determined by RT-PCR and western blotting. ES-GNs downregulated upstream signaling pathways (IκB kinase-α/ß, nuclear factor-κB, Janus-activated kinase /signal transducers and activators of transcription, mitogen-activated protein kinase , and phospholipase D) of pro-inflammatory mediators and cytokines in LPS-stimulated microglia. Anti-neuroinflammatory properties of ES-GNs were mediated by ES-GNs-induced AMP-activated protein kinase)-mediated nuclear erythroid 2-related factor 2 /antioxidant response element signaling. Conclusion: Collectively, these findings provide a new insight on the role of ES-GNs in treating chronic neuroinflammation-induced neurodegenerative diseases.


Assuntos
Anti-Inflamatórios/farmacologia , Ephedra sinica/química , Ouro/farmacologia , Nanopartículas Metálicas/química , Microglia/patologia , Extratos Vegetais/farmacologia , Adenilato Quinase/metabolismo , Animais , Elementos de Resposta Antioxidante/genética , Antioxidantes/farmacologia , Morte Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Citocinas/metabolismo , Difusão Dinâmica da Luz , Mediadores da Inflamação/metabolismo , Nanopartículas Metálicas/ultraestrutura , Camundongos , Microglia/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Espectroscopia de Infravermelho com Transformada de Fourier , Difração de Raios X
10.
J Biol Regul Homeost Agents ; 33(2): 517-523, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30972993

RESUMO

This study aimed to investigate the effect of different exercise loads on sex hormones and expressions of relevant genes in hypothalamus in obese mice. Sixty weaning male C57BL/6 mice were used as subjects. Among them, 15 mice were randomly classified into the normal diet group (CON group), and the remaining 45 mice into high-fat diet group (MOB group). The obesity was successfully achieved by high-fat diet 10 weeks later. Then the rats were randomly divided into three groups based on weight, namely, obesity control group (OBC group), obesity with moderate-intensity exercise group (MOBC group), and obesity with high-intensity exercise group (HOBC group), with 15 mice in each group. Mice in the MOBC group and HOBC group were offered 8 weeks of swimming training, and the exercise time increased incrementally until 2 h and 4 h per day. After the training was over, ELISA method was used to determine the serum levels of Adiponectin (Adipo), luteotropic hormone (LH), follicle-stimulating hormone (FSH) and testosterone (T). Real-time PCR was implemented to detect the transcriptional levels of genes of Adipo and other relevant proteins in the hypothalamus. The result showed that compared with the CON group, there was a significant reduction in the serum levels of Adipo, LH, FSH and T in the OBC group (P<0.05). As compared with the OBC group, the serum levels of Adipo, LH, FSH and T increased significantly in the OBC group (P<0.05). There was a significant increase in the transcriptional levels of Adipo, Adipo receptor 1 (Adipo R1), and AMP-activated protein kinase (AMPK) in the OBC group (P<0.05) compared to in the CON group; meanwhile, the transcriptional levels of kisspeptin (Kiss) and gonadotropin-releasing hormone (GnRH) decreased significantly (P<0.05). In conclusion, long-term moderate-intensity exercise could improve the negative effect of obesity on sex development. Long-term high-intensity exercises could not improve the inhibitory effect of obesity on sex development.


Assuntos
Hormônios Esteroides Gonadais/sangue , Hipotálamo/metabolismo , Obesidade/genética , Condicionamento Físico Animal , Adenilato Quinase/metabolismo , Adiponectina/sangue , Animais , Hormônio Foliculoestimulante/sangue , Hormônio Liberador de Gonadotropina/metabolismo , Kisspeptinas/metabolismo , Hormônio Luteinizante/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Distribuição Aleatória , Testosterona/sangue
11.
Colloids Surf B Biointerfaces ; 179: 421-428, 2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-31003168

RESUMO

Combination therapy remains a promising approach to ameliorate cerebral ischemia injury. Nevertheless, the primary mechanism of the neuroprotective properties of Dictyopteris divaricata extract-capped gold nanoparticles (DD-GNPs) is not completely understood. DD-GNPs displayed maximum absorption at 525 nm and a diameter of 62.6 ± 1.2 nm, with a zeta potential value of -26.1 ± 0.6 mV. High resolution-transmission electron microscopy confirmed the spherical shape and average diameter (28.01 ± 2.03 nm). Crystalline structure and gold nanoparticle synthesis of DD-GNPs were determined by X-ray powder diffraction, and the presence of elemental gold was confirmed by energy-dispersive X-ray spectroscopy and Fourier transform-infrared spectroscopy. We examined the neuroprotective properties of DD-GNPs and explored their potential mechanisms in human SH-SY5Y neuroblastoma cells treated with oxygen and glucose deprivation/reoxygenation (OGD/R). We found that DD-GNPs inhibited OGD/R-induced release of lactate dehydrogenase (LDH), loss of cell viability, and production of reactive oxygen species. This neuroprotection was accompanied by regulation of apoptosis-related proteins, as indicated by decreased levels of cleaved-caspase-3, cleaved-PARP, cleaved-caspase-9, p53, p21, and Bax, as well as an increased level of Bcl-2. Notably, the neuroprotective effects of DD-GNPs were partially abolished by HO-1, NQO1, Nrf2, and AMPK knockdown. Our results established that DD-GNPs effectively attenuated OGD/R-stimulated neuronal injury, as evidenced by reduced neuronal injury. Even though the accumulating evidence has indicated the low toxicity and minimal side effects of GNPs, experimental clinical trials of DD-GNPs are still limited because of the lack of knowledge regarding the effects of DD-GNPs as neuroprotective agents against neurodegenerative diseases.


Assuntos
Glucose/deficiência , Ouro/química , Nanopartículas Metálicas/química , Fármacos Neuroprotetores/farmacologia , Oxigênio/farmacologia , Feófitas/química , Extratos Vegetais/farmacologia , Adenilato Quinase/metabolismo , Elementos de Resposta Antioxidante/genética , Antioxidantes/farmacologia , Linhagem Celular Tumoral , Difusão Dinâmica da Luz , Ativação Enzimática/efeitos dos fármacos , Heme Oxigenase-1/metabolismo , Humanos , Nanopartículas Metálicas/ultraestrutura , NAD(P)H Desidrogenase (Quinona)/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Neurotoxinas/toxicidade , Transdução de Sinais/efeitos dos fármacos , Espectroscopia de Infravermelho com Transformada de Fourier , Difração de Raios X
12.
Life Sci ; 227: 30-38, 2019 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-31002918

RESUMO

The gradual energy dissipation of all organisms allows adapting to energy demands. Pathological situations of uncured diseases such as cancer, diabetes, and other obesity-related diseases are caused by an abrupt energy imbalance. As an energy sensor, AMP-activated kinase (AMPK) can regulate the cellular energy status. In case of increased energy demands or insufficient nutrient supply, cells digest their own interior, which is called autophagy. AMPK-mediated autophagy regulates various metabolic and physiological processes and is dysregulated in different chronic conditions. Because of AMPK's critical role in physiology and pathology, it is an emerging target for both prevention and treatment of these uncured diseases. This review discusses the multifaceted role of AMPK on cancer cell survival and inhibition mechanism. First, we discuss the dual role of AMPK on cancer progression and suppression, and we discuss how different AMPK subunit combinations influence the tumor progression and suppression. Next, we discuss what could be the centering point of AMPK that supports promotion or inhibition of the cancer cell growth. Furthermore, we review the role of connecting mechanism of AMPK-mediated molecular intermediates on cancer cell survival and inhibition pathways. Finally, we discuss how AMPK can affect DNA damage and repairing mechanisms, and immune response of host cell and cancer cells.


Assuntos
Adenilato Quinase/metabolismo , Adenilato Quinase/fisiologia , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Autofagia , Sobrevivência Celular , Dano ao DNA , Metabolismo Energético/fisiologia , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo
13.
Cell Mol Biol (Noisy-le-grand) ; 65(2): 1-6, 2019 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-30860465

RESUMO

Presently, curcumin derivatives had been paid more attention in view of their high bioavailability or water solubility, which herein possibly replaced the curcumin for their functional applications in future. Here, one novel chemically synthesized curcumin derivative, ZYX01, was used to identify anti-proliferation activity of human non-small lung cancer cells A549 and its anti-proliferative mechanism. Our study showed that ZYX01 could induce autophagic death of A549 cells by morphological observation, MTT assay, acridine orange staining and MDC assay, which possess a dose-and time-dependent manner. ZYX01-treated A549 cells possessed an increase in LC3-II/LC3-I ratio, upregulation of beclin-1 and downregulation of p62 expression. We further confirmed the cellular AMPK/ULK1/Beclin-1 signaling pathway in A549 cells after ZYX01 treatment. The anti-migration effect of ZYX01 in A549 cells was also explored by wound healing assay and transwell experiment. Current results had confirmed that ZYX01 induced A549 cells autophagy through AMPK/ULK1/Beclin-1 pathway and shed light on the future study on the anti-cancer molecular mechanism.


Assuntos
Adenilato Quinase/metabolismo , Proteína Homóloga à Proteína-1 Relacionada à Autofagia/metabolismo , Autofagia/efeitos dos fármacos , Proteína Beclina-1/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Curcumina/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Pulmonares/patologia , Transdução de Sinais , Células A549 , Carcinoma Pulmonar de Células não Pequenas/enzimologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Curcumina/química , Vesículas Citoplasmáticas/efeitos dos fármacos , Vesículas Citoplasmáticas/metabolismo , Humanos , Neoplasias Pulmonares/enzimologia , Proteínas Associadas aos Microtúbulos/metabolismo
14.
Life Sci ; 226: 33-46, 2019 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-30898646

RESUMO

BACKGROUND: Human urothelial carcinoma associated 1 (UCA1) has been recognized as an oncogenic lncRNA in various cancers, except infantile hemangioma (IH). This study attempts to explore the functional role of lncRNA UCA1 in IH. METHODS: qRT-PCR was carried out to detect the expression of lncRNA UCA1 in human IH tissues. Two hemangioma cell lines (EOMA and HemECs) were transfected with shRNAs specific for lncRNA UCA1, or a plasmid for expression lncRNA UCA1. The expression of miR-200c in cell was suppressed or overexpressed by miRNA-mediated transfection. CCK-8 assay, flow cytometry, Transwell assay, and Western blot were performed to detect cell survival, migration and invasion. RESULTS: LncRNA UCA1 was up-regulated in proliferating-phase hemangioma samples, as compared to involuting-phase. Silence of lncRNA UCA1 significantly reduced EOMA cells viability, migration and invasion, and induced apoptosis. These observations were coupled with the down-regulations of CyclinD1, CDK6 and CDK4, the cleavage of caspase-3 and caspase-9, as well as the decreased expression levels of MMP-9 and Vimentin. miR-200c was highly expressed in lncRNA UCA1 silenced-cells. Besides, the anti-tumor effects of lncRNA UCA1 silence towards EOMA cells were reversed by miR-200c suppression. Same effects of lncRNA UCA1 and miR-200c on HemECs cells were observed. Furthermore, silence of lncRNA UCA1 repressed mTOR, AMPK and Wnt/ß-catenin signaling via a miR-200c-dependent fashion. CONCLUSION: This study evidences that silence of lncRNA UCA1 inhibits hemangioma cells growth, migration and invasion possibly via its regulation on miR-200c expression and the activation of mTOR, AMPK and Wnt/ß-catenin signaling pathways.


Assuntos
Hemangioma/genética , Hemangioma/patologia , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Adenilato Quinase/genética , Adenilato Quinase/metabolismo , Animais , Apoptose/fisiologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Feminino , Hemangioma/metabolismo , Humanos , Lactente , Recém-Nascido , Masculino , Camundongos , MicroRNAs/genética , Invasividade Neoplásica , RNA Longo não Codificante/genética , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Ativação Transcricional , Regulação para Cima , Via de Sinalização Wnt
15.
Curr Med Sci ; 39(1): 37-43, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30868489

RESUMO

This study aimed to verify the effects of berberine (BBR) on the fat metabolism proteins involved in the sirtuin 3 (SIRT3)/adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK)/acetyl-CoA carboxylase (ACC) pathway in the liver tissues of rats with high-fat diet (HFD)-induced non-alcoholic fatty liver disease (NAFLD). Forty-eight rats were randomly divided into the normal control (NC) group, HFD group or BBR group, with 16 rats in each group. After 8 and 16 weeks of treatment, serum and liver samples were collected. Subsequently, body parameters, biochemical parameters and liver pathology were examined. The expression levels of proteins involved in the SIRT3/AMPK/ACC pathway in the liver were detected by Western blotting. After 8 and 16 weeks of a HFD, the successful establishment of rat models with different degrees of NAFLD was confirmed by hematoxylin and eosin (H&E) and Oil Red O staining. NAFLD rat models exhibited obesity and hyperlipidemia, and the protein expression levels of SIRT3, p-AMPK, p-ACC, and CPT-1A in the liver were significantly decreased compared to those in the NC group. The concurrent administration of BBR with the HFD effectively improved serum and liver lipid profiles and ameliorated liver injury. Furthermore, the protein expression levels of SIRT3, p-AMPK, p-ACC, and CPT-1A in the liver were significantly increased in the BBR group as compared with those in the HFD group. In conclusion, our data suggest that the mechanism by which BBR ameliorates HFD-induced hepatic steatosis may be related to the activation of the SIRT3/AMPK/ACC pathway in the liver.


Assuntos
Berberina/administração & dosagem , Dieta Hiperlipídica/efeitos adversos , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Obesidade/induzido quimicamente , Transdução de Sinais/efeitos dos fármacos , Acetil-CoA Carboxilase/metabolismo , Adenilato Quinase/metabolismo , Animais , Berberina/farmacologia , Carnitina O-Palmitoiltransferase/metabolismo , Modelos Animais de Doenças , Metabolismo dos Lipídeos/efeitos dos fármacos , Masculino , Hepatopatia Gordurosa não Alcoólica/induzido quimicamente , Hepatopatia Gordurosa não Alcoólica/metabolismo , Obesidade/complicações , Obesidade/metabolismo , Fosforilação/efeitos dos fármacos , Distribuição Aleatória , Ratos , Sirtuínas/metabolismo
16.
Parasitol Res ; 118(5): 1573-1579, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30815727

RESUMO

Parasitic infections caused by protozoan belonging to genus Eimeria are considered important for the poultry industry, due to their severe intestinal lesions and high mortality rates, causing significant economic losses. Although several mechanisms of coccidiosis pathogenesis are known, the effects of this infection on intestinal enzymes linked to adenosine triphosphate (ATP) metabolism, as creatine kinase (CK), adenylate kinase (AK), and pyruvate kinase (PK), remain unknown. Thus, the aim of this study was to evaluate whether coccidiosis impairs enzymes linked ATP metabolism in the intestine of chicken chicks. For this, 42 animals that were 2 days old were divided into two groups: uninfected (the negative control group) and experimentally infected on second day of life (the positive control group). On days 5, 10, and 15 post-infection (PI), fecal samples were collected for oocyst counts; intestinal tissue was collected in order to evaluate CK, AK, and PK activities, as well as parameters of the oxidative stress and histopathology. On days 10 and 15 PI, infected animals showed high counts of oocysts in fecal samples and intestinal lesions compared to the control group. Cytosolic CK activity was higher in infected animals on days 10 and 15 PI compared to the control group, while mitochondrial CK activity was lower on days 5, 10, and 15 PI. Also, AK activity was lower in infected animals on days 10 and 15 PI compared to control group, while no differences were observed between groups regarding PK activity. In relation to parameters of oxidative stress, intestinal lipid peroxidation and reactive oxygen species levels were higher in infected animals on days 10 and 15 PI compared to the control group, while non-protein thiol levels were lower on day 10 PI. On the 15th day, infected animals had lower body weight (P < 0.05). Based on this evidence, inhibition of mitochondrial CK activity causes an impairment of intestinal energetic homeostasis possibly through depletion on ATP levels, although the cytosolic CK activity acted as an attempt to restore the mitochondrial ATP levels through a feedback mechanism. Moreover, the impairment on energy metabolism appears to be mediated by excessive production of intestinal ROS, as well as oxidation of lipids and thiol groups.


Assuntos
Trifosfato de Adenosina/metabolismo , Adenilato Quinase/metabolismo , Galinhas/parasitologia , Coccidiose/veterinária , Creatina Quinase/metabolismo , Eimeria/metabolismo , Mitocôndrias/metabolismo , Piruvato Quinase/metabolismo , Animais , Coccidiose/metabolismo , Metabolismo Energético/fisiologia , Glicólise/fisiologia , Homeostase , Enteropatias/parasitologia , Intestinos/parasitologia , Intestinos/patologia , Mitocôndrias/enzimologia , Estresse Oxidativo , Fosforilação , Doenças das Aves Domésticas/parasitologia , Espécies Reativas de Oxigênio/metabolismo , Ganho de Peso
17.
Int J Mol Sci ; 20(6)2019 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-30875837

RESUMO

The effects of RNA on in-cell NMR spectroscopy and ribosomes on the kinetic activity of several metabolic enzymes are reviewed. Quinary interactions between labelled target proteins and RNA broaden in-cell NMR spectra yielding apparent megadalton molecular weights in-cell. The in-cell spectra can be resolved by using cross relaxation-induced polarization transfer (CRINEPT), heteronuclear multiple quantum coherence (HMQC), transverse relaxation-optimized, NMR spectroscopy (TROSY). The effect is reproduced in vitro by using reconstituted total cellular RNA and purified ribosome preparations. Furthermore, ribosomal binding antibiotics alter protein quinary structure through protein-ribosome and protein-mRNA-ribosome interactions. The quinary interactions of Adenylate kinase, Thymidylate synthase and Dihydrofolate reductase alter kinetic properties of the enzymes. The results demonstrate that ribosomes may specifically contribute to the regulation of biological activity.


Assuntos
Antibacterianos/farmacologia , Ressonância Magnética Nuclear Biomolecular/métodos , RNA/metabolismo , Ribossomos/química , Ribossomos/metabolismo , Adenilato Quinase/química , Adenilato Quinase/metabolismo , Domínio Catalítico , Modelos Moleculares , Ligação Proteica/efeitos dos fármacos , Estrutura Quaternária de Proteína , Teoria Quântica , Ribossomos/efeitos dos fármacos , Ribossomos/genética , Tetra-Hidrofolato Desidrogenase/química , Tetra-Hidrofolato Desidrogenase/metabolismo , Timidilato Sintase/química , Timidilato Sintase/metabolismo
18.
Proc Natl Acad Sci U S A ; 116(11): 5154-5159, 2019 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-30804206

RESUMO

A high-fat diet (HFD) causes obesity-associated morbidities involved in macroautophagy and chaperone-mediated autophagy (CMA). AMPK, the mediator of macroautophage, has been reported to be inactivated in HFD-caused renal injury. However, PAX2, the mediator for CMA, has not been reported in HFD-caused renal injury. Here we report that HFD-caused renal injury involved the inactivation of Pax2 and Ampk, and the activation of soluble epoxide hydrolase (sEH), in a murine model. Specifically, mice fed on an HFD for 2, 4, and 8 wk showed time-dependent renal injury, the significant decrease in renal Pax2 and Ampk at both mRNA and protein levels, and a significant increase in renal sEH at mRNA, protein, and molecular levels. Also, administration of an sEH inhibitor, 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl)urea, significantly attenuated the HFD-caused renal injury, decreased renal sEH consistently at mRNA and protein levels, modified the renal levels of sEH-mediated epoxyeicosatrienoic acids (EETs) and dihydroxyeicosatrienoic acids (DHETs) as expected, and increased renal Pax2 and Ampk at mRNA and/or protein levels. Furthermore, palmitic acid (PA) treatment caused significant increase in Mcp-1, and decrease in both Pax2 and Ampk in murine renal mesangial cells (mRMCs) time- and dose-dependently. Also, 14(15)-EET (a major substrate of sEH), but not its sEH-mediated metabolite 14,15-DHET, significantly reversed PA-induced increase in Mcp-1, and PA-induced decrease in Pax2 and Ampk. In addition, plasmid construction revealed that Pax2 may positively regulate Ampk transcriptionally in mRMCs. This study provides insights into and therapeutic target for the HFD-mediated renal injury.


Assuntos
Adenilato Quinase/metabolismo , Dieta Hiperlipídica , Epóxido Hidrolases/antagonistas & inibidores , Rim/lesões , Fator de Transcrição PAX2/metabolismo , Animais , Sistema Enzimático do Citocromo P-450/metabolismo , Modelos Animais de Doenças , Eicosanoides/metabolismo , Inibidores Enzimáticos/farmacologia , Epóxido Hidrolases/metabolismo , Hipertrofia , Rim/patologia , Células Mesangiais/efeitos dos fármacos , Células Mesangiais/metabolismo , Células Mesangiais/patologia , Camundongos , Ácido Palmítico , Compostos de Fenilureia/farmacologia , Piperidinas/farmacologia , Solubilidade , Fatores de Tempo , Ganho de Peso
19.
Int J Mol Med ; 43(4): 1778-1788, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30720055

RESUMO

Acid­sensing ion channel 1a (ASIC1a), member of the degenerin/epithelial sodium channel protein superfamily, serves a critical role in various physiological and pathological processes. The aim of the present study was to examine the role of ASIC1a in the autophagy of rat articular chondrocytes. Autophagy was induced by acidic stimulation in rat articular chondrocytes and the extent of autophagy was evaluated via the expression levels of microtubule­associated protein 1 light chain 3II, Beclin1 and uncoordinated­51 like kinase1. Suppression of ASIC1a was achieved using small interfering RNA technology and/or inhibitor psalmotoxin­1. The expression levels of autophagy markers were measured by western blot analysis and reverse transcription­quantitative polymerase chain reaction methods. Intracellular calcium ([Ca2+]i) was analyzed using a Ca2+­imaging method. Additionally, protein expression levels of the Ca2+/calmodulin­dependent protein kinase kinase ß (CaMKKß)/5'­monophosphate­activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) pathway were measured by western blot analysis. The results showed that autophagy was increased in a pH­ and time­dependent manner with exposure to an acidic environment. In addition, silencing ASIC1a significantly decreased the expression levels of autophagy makers, accompanied by abrogation of the acid­induced [Ca2+]i increase. Furthermore, silencing of ASIC1a downregulated the levels of CaMKKß/ß­actin and phosphorylated (p­) AMPK/AMPK, and upregulated the levels of p­mTOR/mTOR. These results indicated that ASIC1a is a potent regulator of autophagy in chondrocytes, which may be associated with decreased Ca2+ influx and the CaMKKß/AMPK/mTOR pathway.


Assuntos
Canais Iônicos Sensíveis a Ácido/metabolismo , Autofagia , Sinalização do Cálcio , Condrócitos/citologia , Condrócitos/metabolismo , Ácidos , Adenilato Quinase/metabolismo , Animais , Autofagia/efeitos dos fármacos , Biomarcadores/metabolismo , Quelantes de Cálcio/farmacologia , Sinalização do Cálcio/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Condrócitos/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Inativação Gênica , Concentração de Íons de Hidrogênio , Cirrose Hepática/genética , Cirrose Hepática/patologia , Masculino , Modelos Biológicos , Peptídeos/toxicidade , Ratos Sprague-Dawley , Venenos de Aranha/toxicidade , Serina-Treonina Quinases TOR/metabolismo
20.
Biochem Biophys Res Commun ; 510(2): 284-289, 2019 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-30700382

RESUMO

Although autophagy and phagocytosis are involved in the regulation of host inflammatory response to bacterial infection in macrophages, the underlying mechanisms have not been completely elucidated. In the present study, we found that infecting RAW264.7 macrophages with Staphylococcus aureus (S. aureus) activated multiple signaling pathways including phosphatidylinositide 3-kinase (PI3K)/protein kinase B (Akt), nuclear factor-κB (NF-κB), and Rac1, as well as triggered autophagy. LY294002, a specific PI3K activity inhibitor, significantly decreased autophagy and phagocytosis of macrophages upon S. aureus infection. Similarly, knockdown of Beclin1 by specific siRNA significantly inhibited autophagy and phagocytosis of S. aureus-infected macrophages. Additionally, we showed that although administration of Beclin1 siRNA had no effects on phosphorylation of Akt (p-Akt), inhibition of PI3K activity by LY294002 significantly decreased the expression of Beclin1, suggesting that Beclin1 is a downstream molecular of PI3K. Furthermore, inhibition of autophagy significantly increased the production of NF-κB-dependent TNFα/IL-1ß in S. aureus-infected macrophages. Collectively, these findings demonstrated, for the first time, that the PI3K/Akt-Beclin1 signaling pathway positively regulates phagocytosis and negatively mediates NF-κB-dependent inflammation in S. aureus-infected macrophages.


Assuntos
Adenilato Quinase/metabolismo , Proteína Beclina-1/metabolismo , Macrófagos/microbiologia , Subunidade p50 de NF-kappa B/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Infecções Estafilocócicas/metabolismo , Animais , Autofagia , Interleucina-1beta/metabolismo , Macrófagos/metabolismo , Camundongos , Fagocitose , Células RAW 264.7 , Transdução de Sinais , Staphylococcus aureus , Fator de Necrose Tumoral alfa/metabolismo
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