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1.
Gene Ther ; 2020 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-32341485

RESUMO

Cardiovascular disease has become a major disease affecting health in the whole world. Gene therapy, delivering foreign normal genes into target cells to repair damages caused by defects and abnormal genes, shows broad prospects in treating different kinds of cardiovascular diseases. China has achieved great progress of basic gene therapy researches and pathogenesis of cardiovascular diseases in recent years. This review will summarize the latest research about gene therapy of proteins, epigenetics, including noncoding RNAs and genome-editing technology in myocardial infarction, cardiac ischemia-reperfusion injury, atherosclerosis, muscle atrophy, and so on in China. We wish to highlight some important findings about the essential roles of basic gene therapy in this field, which might be helpful for searching potential therapeutic targets for cardiovascular disease.

2.
Autophagy ; 16(3): 451-465, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31184563

RESUMO

The mechanisms underlying glucocorticoid (GC)-increased adiposity are poorly understood. Brown adipose tissue (BAT) acquires white adipose tissue (WAT) cell features defined as BAT whitening under certain circumstances. The aim of our current study was to investigate the possibility and mechanisms of GC-induced BAT whitening. Here, we showed that one-week dexamethasone (Dex) treatment induced BAT whitening, characterized by lipid droplet accumulation, in vitro and in vivo. Furthermore, autophagy and ATG7 (autophagy related 7) expression was induced in BAT by Dex, and treatment with the autophagy inhibitor chloroquine or adenovirus-mediated ATG7 knockdown prevented Dex-induced BAT whitening and fat mass gain. Moreover, Dex-increased ATG7 expression and autophagy was mediated by enhanced expression of BTG1 (B cell translocation gene 1, anti-proliferative) that stimulated activity of CREB1 (cAMP response element binding protein 1). The importance of BTG1 in this regulation was further demonstrated by the observed BAT whitening in adipocyte-specific BTG1-overexpressing mice and the attenuated Dex-induced BAT whitening and fat mass gain in mice with BTG1 knockdown in BAT. Taken together, we showed that Dex induces a significant whitening of BAT via BTG1- and ATG7-dependent autophagy, which might contribute to Dex-increased adiposity. These results provide new insights into the mechanisms underlying GC-increased adiposity and possible strategy for preventing GC-induced side effects via the combined use of an autophagy inhibitor.Abbreviations: ACADL: acyl-Coenzyme A dehydrogenase, long-chain; ACADM: acyl-Coenzyme A dehydrogenase, medium-chain; ACADS: acyl-Coenzyme A dehydrogenase, short-chain; ADIPOQ: adiponectin; AGT: angiotensinogen; Atg: autophagy-related; BAT: brown adipose tissue; BTG1: B cell translocation gene 1, anti-proliferative; CEBPA: CCAAT/enhancer binding protein (C/EBP), alpha; CIDEA: cell death-inducing DNA fragmentation factor, alpha subunit-like effector A; CPT1B: carnitine palmitoyltransferase 1b, muscle; CPT2: carnitine palmitoyltransferase 2; CQ: chloroquine; Dex: dexamethasone; eWAT: epididymal white adipose tissue; FABP4: fatty acid binding protein 4, adipocyte; FFAs: free fatty acids; GCs: glucocorticoids; NRIP1: nuclear receptor interacting protein 1; OCR: oxygen consumption rate; PBS: phosphate-buffered saline; PPARA: peroxisome proliferator activated receptor alpha; PPARG: peroxisome proliferator activated receptor gamma; PPARGC1A: peroxisome proliferator activated receptor, gamma, coactivator 1 alpha; PRDM16: PR domain containing 16; PSAT1: phosphoserine aminotransferase 1; RB1: RB transcriptional corepressor 1; RBL1/p107: RB transcriptional corepressor like 1; SQSTM1: sequestosome 1; sWAT: subcutaneous white adipose tissue; TG: triglycerides; UCP1: uncoupling protein 1 (mitochondrial, proton carrier); WT: wild-type.

3.
Gastroenterology ; 156(4): 1098-1111, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30452920

RESUMO

BACKGROUND & AIMS: Activating transcription factor 4 (ATF4) regulates genes involved in the inflammatory response, amino acid metabolism, autophagy, and endoplasmic reticulum stress. We investigated whether its activity is altered in patients with inflammatory bowel diseases (IBDs) and mice with enterocolitis. METHODS: We obtained biopsy samples during endoscopy from inflamed and/or uninflamed regions of the colon from 21 patients with active Crohn's disease (CD), 22 patients with active ulcerative colitis (UC), and 38 control individuals without IBD and of the ileum from 19 patients with active CD and 8 individuals without IBD in China. Mice with disruption of Atf4 specifically in intestinal epithelial cells (Atf4ΔIEC mice) and Atf4-floxed mice (controls) were given dextran sodium sulfate (DSS) to induce colitis. Some mice were given injections of recombinant defensin α1 (DEFA1) and supplementation of l-alanyl-glutamine or glutamine in drinking water. Human and mouse ileal and colon tissues were analyzed by quantitative real-time polymerase chain reaction, immunoblots, and immunohistochemistry. Serum and intestinal epithelial cell (IEC) amino acids were measured by high-performance liquid chromatography-tandem mass spectrometry. Levels of ATF4 were knocked down in IEC-18 cells with small interfering RNAs. Microbiomes were analyzed in ileal feces from mice by using 16S ribosomal DNA sequencing. RESULTS: Levels of ATF4 were significantly decreased in inflamed intestinal mucosa from patients with active CD or active UC compared with those from uninflamed regions or intestinal mucosa from control individuals. ATF4 was also decreased in colonic epithelia from mice with colitis vs mice without colitis. Atf4ΔIEC mice developed spontaneous enterocolitis and colitis of greater severity than control mice after administration of DSS. Atf4ΔIEC mice had decreased serum levels of glutamine and reduced levels of antimicrobial peptides, such as Defa1, Defa4, Defa5, Camp, and Lyz1, in ileal Paneth cells. Atf4ΔIEC mice had alterations in ileal microbiomes compared with control mice; these changes were reversed by administration of glutamine. Injections of DEFA1 reduced the severity of spontaneous enteritis and DSS-induced colitis in Atf4ΔIEC mice. We found that expression of solute carrier family 1 member 5 (SLC1A5), a glutamine transporter, was directly regulated by ATF4 in cell lines. Overexpression of SLC1A5 in IEC-18 or primary IEC cells increased glutamine uptake and expression of antimicrobial peptides. Knockdown of ATF4 in IEC-18 cells increased expression of inflammatory cytokines, whereas overexpression of SLC1A5 in the knockdown cells reduced cytokine expression. Levels of SLC1A5 were decreased in inflamed intestinal mucosa of patients with CD and UC and correlated with levels of ATF4. CONCLUSIONS: Levels of ATF4 are decreased in inflamed intestinal mucosa from patients with active CD or UC. In mice, ATF4 deficiency reduces glutamine uptake by intestinal epithelial cells and expression of antimicrobial peptides by decreasing transcription of Slc1a5. ATF4 might therefore be a target for the treatment of IBD.


Assuntos
Fator 4 Ativador da Transcrição/deficiência , Peptídeos Catiônicos Antimicrobianos/metabolismo , Colite Ulcerativa/metabolismo , Doença de Crohn/metabolismo , Glutamina/metabolismo , Fator 4 Ativador da Transcrição/genética , Fator 4 Ativador da Transcrição/metabolismo , Adolescente , Adulto , Sistema ASC de Transporte de Aminoácidos/genética , Sistema ASC de Transporte de Aminoácidos/metabolismo , Animais , Estudos de Casos e Controles , Linhagem Celular , Colite/induzido quimicamente , Colite/metabolismo , Colite Ulcerativa/sangue , Colite Ulcerativa/patologia , Colo/citologia , Colo/metabolismo , Doença de Crohn/sangue , Doença de Crohn/patologia , Células Epiteliais , Feminino , Técnicas de Silenciamento de Genes , Glutamina/sangue , Glutamina/farmacologia , Humanos , Íleo/citologia , Íleo/metabolismo , Íleo/microbiologia , Mucosa Intestinal/metabolismo , Masculino , Camundongos , Microbiota/efeitos dos fármacos , Pessoa de Meia-Idade , Antígenos de Histocompatibilidade Menor/genética , Antígenos de Histocompatibilidade Menor/metabolismo , Celulas de Paneth/metabolismo , Adulto Jovem
4.
Mol Metab ; 20: 138-148, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30579932

RESUMO

OBJECTIVE: c-Jun, a prominent member of the activator protein 1 (AP-1) family, is involved in various physiology processes such as cell death and survival. However, a role of hepatic c-Jun in the whole-body metabolism is poorly understood. METHODS: We generated liver-specific c-Jun knock-out (c-jun△li) mice to investigate the effect of hepatic c-Jun on the whole-body physiology, particularly in blood glucose and body temperature. Primary hepatocytes were also used to explore a direct regulation of c-Jun in gluconeogenesis. RESULTS: c-jun△li mice showed higher hepatic gluconeogenic capacity compared with control mice, and similar results were obtained in vitro. In addition, fibroblast growth factor 21 (FGF21) expression was directly inhibited by c-Jun knockdown and adenovirus-mediated hepatic FGF21 over-expression blocked the effect of c-Jun on gluconeogenesis in c-jun△li mice. Interestingly, c-jun△li mice also exhibited higher body temperature, with induced thermogenesis and uncoupling protein 1 (UCP1) expression in brown adipose tissue (BAT). Furthermore, the body temperature became comparable between c-jun△li and control mice at thermoneutral temperature (30 °C). Moreover, the activity of sympathetic nervous system (SNS) was increased in c-jun△li mice and the higher body temperature was inhibited by beta-adrenergic receptor blocker injection. Finally, the activated SNS and increased body temperature in c-jun△li mice was most likely caused by the signals from the brain and hepatic vagus nerve, as the expression of c-Fos (the molecular marker of neuronal activation) was changed in several brain areas controlling body temperature and body temperature was decreased by selective hepatic vagotomy. CONCLUSIONS: These data demonstrate a novel function of hepatic c-Jun in the regulation of gluconeogenesis and body temperature via FGF21 and neural signals. Our results also provide novel insights into the organ crosstalk in the regulation of the whole-body physiology.


Assuntos
Temperatura Corporal , Fatores de Crescimento de Fibroblastos/metabolismo , Gluconeogênese , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Fígado/metabolismo , Nervo Vago/efeitos dos fármacos , Tecido Adiposo/metabolismo , Antagonistas Adrenérgicos beta/farmacologia , Animais , Células Cultivadas , Células Hep G2 , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Masculino , Camundongos , Propranolol/farmacologia , Transdução de Sinais
5.
J Chromatogr A ; 1588: 17-24, 2019 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-30587344

RESUMO

A novel solid phase microextraction fiber named graphene oxide incorporated poly acrylamide-ethylene glycol dimethacrylate (GO-poly AM-EDGMA) monolithic fiber has been successfully prepared in a fused silica capillary tube (250 µm, i.d.) via thermally initiated polymerization using acrylamide (AM) as the monomer, ethylene glycol dimethacrylate (EGDMA) as the crosslinker, dimethylformamide dispersed by GO as porogens, and then obtained by removing 1 cm wall from one end of the fused silica capillary tube. The fiber has large surface area (536 m2 g-1), pore capacity (0.694 cm3 g-1) good thermal stability (up to 320 ℃), long service life and good reproducibility (RSD <5% throughout 110 times) which facilitated for high throughput headspace solid phase microextraction (HS-SPME) coupled to gas chromatograph (GC) analysis. The HS-SPME/GC method using the new fiber was evaluated by the determination of five organophosphate esters (OPEs) in soil samples coupled to flame photometric detector (FPD). The proposed HS-SPME-GC/FPD method yielded satisfactory limits of quantification (0.03 ng g-1∼0.24 ng g-1); linearity (≥0.99), good intra- and inter-day precision expressed as relative standard deviations for a single fiber were in the range of 5.2-9.0% and 4.8-9.0%, respectively, and fiber-to-fiber reproducibility was in the range of 5.9-9.7%. The method was applied for the analysis of OPEs in environmental soil samples and the relative recoveries were found to be in the range from 80.1 to 105.6%. Based on these features, the new fiber has great potential for widespread use as a high throughput trace analysis tool.


Assuntos
Monitoramento Ambiental/métodos , Grafite/química , Metacrilatos/química , Organofosfatos/análise , Polietilenoglicóis/química , Poluentes do Solo/análise , Solo/química , Microextração em Fase Sólida/métodos , Cromatografia Gasosa , Polímeros/química , Reprodutibilidade dos Testes , Dióxido de Silício/química , Microextração em Fase Sólida/instrumentação
6.
Am J Cancer Res ; 8(11): 2284-2295, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30555744

RESUMO

The ubiquitin-specific protease 5 (USP5), a deubiquitinating enzyme, has been identified as a tumor promoter in several types of human cancer. However, the role of USP5 in non-small lung cancer (NSCLC) has not yet been elucidated. In this study, we found that USP5 was upregulated in NSCLC tissues compared with normal tissues. High expression of USP5 was correlated with large primary tumor size, poor differentiation and advanced TNM stage, and led to a significantly shorter overall survival (OS). USP5 overexpression enhanced, whereas USP5 silencing impaired the cell proliferation and colony formation of NSCLC cells in vitro. Moreover, knockdown of USP5 in H1299 cells inhibited tumor growth in vivo. Mechanistically, we found that USP5 deubiquitinated ß-catenin, prevented ubiquitination mediated ß-catenin degradation and promoted ß-catenin nuclear accumulation, leading to the activation of Wnt/ß-catenin signal pathway in NSCLC cells. Taken together, these findings suggest that USP5 functions as an oncogene in NSCLC and its oncogenic activity involves in part through Wnt/ß-catenin signal pathway.

7.
Org Biomol Chem ; 16(40): 7488-7497, 2018 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-30272759

RESUMO

The heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) was reported to participate in the development of a variety of tumors. BC15 is a DNA aptamer targeting hnRNP A1. Firstly, through sequence truncation, we identified 31-mer sequence BC15-31 as the core sequence of BC15 with a strong binding affinity and high selectivity to the hnRNP A1 protein. Isothymidine (isoT) modification was then applied for the structural optimization of BC15-31, systematic modification and biological evaluation were carried out. Incorporation of isoT in the 1,3 sites at the 5'-end of BC15-31 can significantly enhance the protein affinity. Chemical modifications close to the 3'-end can greatly improve the stability of the aptamer. Furthermore, BC15-31 modified with isoT at both the 5'-end and 3'-end displayed an additive effect with enhanced bioactivity and stability at the same time. Our study strategy on BC15 provides a useful guideline for chemical modification and optimization of the aptamer for further clinical application.


Assuntos
Aptâmeros de Nucleotídeos/química , Aptâmeros de Nucleotídeos/metabolismo , Ribonucleoproteína Nuclear Heterogênea A1/metabolismo , Timidina/química , Células A549 , Aptâmeros de Nucleotídeos/genética , Aptâmeros de Nucleotídeos/farmacologia , Sequência de Bases , Proliferação de Células/efeitos dos fármacos , Células Hep G2 , Humanos , Ligação Proteica , Relação Estrutura-Atividade , Especificidade por Substrato
8.
Spectrochim Acta A Mol Biomol Spectrosc ; 195: 157-164, 2018 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-29414573

RESUMO

A new aluminum ion fluorescent probe (4-(diethylamino)-2-hydroxybenzylidene)isoquinoline-1-carbohydrazide (HL1) has been conveniently synthesized and characterized. HL1 exhibited a highly selective and pronounced enhancement for Al3+ in the fluorescence emission over other common cations by forming a 2:1 complex, with a recognition mechanism based on excited-state intramolecular proton transfer (ESIPT) and intramolecular charge transfer (ICT). The strong fluorescent emission can be observed even at ppm level concentration of the probe in the presence of Al3+ with 41 fold intensity enhancement at 545 nm. HL1 displays good linear relationship with Al3+ in the low concentration and the limit of detection is 8.08 × 10-8 mol/L. Similar molecules with different substituents on salicylaldehyde phenyl ring were synthesized for studying the structure-activity relationship. Density-functional theory (DFT) calculations are in agreement with the proposed mechanism. It is confirmed that HL1 could be used to detect Al3+ ions in real sample by fluorescence spectrometry and Al3+ ions in cells by bioimaging.

9.
Diabetes ; 67(4): 569-580, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29321171

RESUMO

Although the central nervous system has been implicated in glucocorticoid-induced gain of fat mass, the underlying mechanisms are poorly understood. The aim of this study was to investigate the possible involvement of hypothalamic serum- and glucocorticoid-regulated kinase 1 (SGK1) in glucocorticoid-increased adiposity. It is well known that SGK1 expression is induced by acute glucocorticoid treatment, but it is interesting that we found its expression to be decreased in the arcuate nucleus of the hypothalamus, including proopiomelanocortin (POMC) neurons, following chronic dexamethasone (Dex) treatment. To study the role of SGK1 in POMC neurons, we produced mice that developed or experienced adult-onset SGK1 deletion in POMC neurons (PSKO). As observed in Dex-treated mice, PSKO mice exhibited increased adiposity and decreased energy expenditure. Mice overexpressing constitutively active SGK1 in POMC neurons consistently had the opposite phenotype and did not experience Dex-increased adiposity. Finally, Dex decreased hypothalamic α-melanocyte-stimulating hormone (α-MSH) content and its precursor Pomc expression via SGK1/FOXO3 signaling, and intracerebroventricular injection of α-MSH or adenovirus-mediated FOXO3 knockdown in the arcuate nucleus largely reversed the metabolic alterations in PSKO mice. These results demonstrate that POMC SGK1/FOXO3 signaling mediates glucocorticoid-increased adiposity, providing new insights into the mechanistic link between glucocorticoids and fat accumulation and important hints for possible treatment targets for obesity.


Assuntos
Adiposidade/efeitos dos fármacos , Dexametasona/farmacologia , Proteína Forkhead Box O3/genética , Glucocorticoides/farmacologia , Proteínas Imediatamente Precoces/genética , Neurônios/efeitos dos fármacos , Pró-Opiomelanocortina/metabolismo , Proteínas Serina-Treonina Quinases/genética , Adiposidade/genética , Animais , Núcleo Arqueado do Hipotálamo/efeitos dos fármacos , Núcleo Arqueado do Hipotálamo/metabolismo , Metabolismo Energético/efeitos dos fármacos , Proteína Forkhead Box O3/metabolismo , Hipotálamo/citologia , Hipotálamo/efeitos dos fármacos , Hipotálamo/metabolismo , Proteínas Imediatamente Precoces/metabolismo , Masculino , Camundongos , Neurônios/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , alfa-MSH/efeitos dos fármacos , alfa-MSH/metabolismo
10.
J Sep Sci ; 41(5): 1104-1111, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29239108

RESUMO

A sensitive, accurate, and cost effective method for the quantification of trimethyl phosphate, which is highly polar and volatile, in environmental water is presented. Trimethyl phosphate was headspace solid-phase microextracted on a molecularly imprinted polymeric fiber, and then the fiber was thermally desorbed in the gas chromatograph injector, and the compound was determined. The trimethyl phosphate imprinted polymeric fiber was prepared by copolymerization in a fused silica capillary tube and obtained by removal of the wall of fused silica capillary tube. The monolithic fiber displayed good selectivity toward trimethyl phosphate among its structural analogues. It was thermally stable up to 320°C so that it can withstand the high temperature of the gas chromatograph injector for desorption. The factors influencing the performance of its headspace solid-phase microextraction were studied. Under the optimal conditions, the method for quantification of trimethyl phosphate in environmental water was well developed. It exhibited significant linearity, the lowest limit of quantification to date, and good recoveries. Using this method, trimethyl phosphate was detected in five out of seven environmental water samples at concentration levels from 0.28 to 1.22 µg/L, illustrating the heavy pollution of trimethyl phosphate in environmental water.

12.
Oncotarget ; 8(36): 60390-60400, 2017 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-28947979

RESUMO

This study evaluated the short-term effects of intensity-modulated radiotherapy (IMRT) and cisplatin concurrent chemo-radiotherapy (CCRT) on attention in patients with nasopharyngeal cancer (NPC). Timely detection and early prevention of cognitive decline are important in cancer patients, because long-term cognitive effects may be permanent and irreversible. Thirty-eight NPC patients treated with IMRT (17/38) or CCRT (21/38) and 38 healthy controls were recruited for the study. Neuropsychological tests were administered to each patient before treatment initiation and within a week after treatment completion. Changes in attention performance over time were evaluated using difference values (D-values). Decreased attention was already observable in patients with NPC prior to treatment. Baseline quotient scores for auditory attention, auditory and visual vigilance, and auditory speed were lower in patients treated with CCRT than in healthy controls (P=0.037, P=0.001, P=0.007, P=0.032, respectively). Auditory stamina D-values were higher in patients treated with IMRT alone (P=0.042), while full-scale response control quotient D-values were lower in patients treated with CCRT (P=0.030) than in healthy controls. Gender, depression, education, and sleep quality were each related to decreased attention and response control. Our results showed that IMRT had no negative acute effects on attention in NPC patients, while CCRT decreased response control.

13.
PLoS One ; 12(7): e0180504, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28678882

RESUMO

This study aimed to determine whether insertion of an intrathecal catheter following accidental dural puncture (ADP) in obstetric patients can reduce the incidence of post-dural puncture headache (PDPH) and the requirement of a therapeutic epidural blood patch (TEBP). This was also compared with relocating the epidural catheter at a different vertebral interspace. A retrospective study was performed, as well as a meta-analysis of the literature to further validate our findings. We reviewed the records of 86 obstetric patients who suffered from ADP during epidural anesthesia or combined spinal-epidural anesthesia from October 2015 to November 2016 at our institution. Although, there was no significant decrease in the incidence of PDPH (P = 0.08), the requirement for a TEBP (P = 0.025) was significantly reduced in the intrathecal catheter group compared with the relocated group. In the meta-analysis, 13 eligible studies including 1044 obstetric patients were finally identified. To estimate the pooled risk ratios (RRs), fixed or random effect models were used depending on the heterogeneity. We initially found that an intrathecal catheter significantly reduced the incidence of PDPH (pooled RR = 0.823; 95% CI = 0.700-0.967; P = 0.018) and the requirement of a TEBP (pooled RR = 0.616; 95% CI = 0.443-0.855; P = 0.004). Our study shows that insertion of an intrathecal catheter following ADP might be an effective and dependable method for reducing the risk of a PDPH and requirement for a TEBP in obstetric patients.


Assuntos
Anestesia Epidural/efeitos adversos , Cefaleia Pós-Punção Dural/prevenção & controle , Adulto , Feminino , Humanos , Cefaleia Pós-Punção Dural/etiologia , Gravidez , Estudos Retrospectivos , Comportamento de Redução do Risco
14.
Org Biomol Chem ; 15(5): 1174-1182, 2017 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-28084479

RESUMO

Aptamers are useful tools in molecular imaging due to their numerous attractive properties, such as excellent affinity and selectivity to diverse types of target molecules and biocompatibility. We carried out structure-activity relationship studies with the tenascin-C (TN-C) binding aptamer GBI-10, which is a promising candidate in tumor imaging. To increase the tumor targeting ability and nuclease resistance under physiological conditions, systematic modifications of GBI-10 with single and multiple 2'-deoxyinosine (2'-dI) or d-/l-isonucleoside (d-/l-isoNA) were performed. Results indicated that sector 3 of the proposed secondary structure is the most important region for specific binding with TN-C. By correlating the affinity of eighty-four GBI-10 derivatives with their predicted secondary structure by Zuker Mfold, we first validated the preferred secondary structure at 37 °C. We found that d-/l-isoNA modified GBI-10 derivatives exhibited improved affinity to the target as well as plasma stability. Affinity measurement and confocal imaging analysis highlighted one potent compound: 4AL/26TL/32TL, which possessed a significantly increased targeting ability to tumor cells. These results revealed the types of modified nucleotides, and the position and number of substituents in GBI-10 that were critical to the TN-C binding ability. Stabilized TN-C-binding DNA aptamers were prepared and they could be further developed for tumor imaging. Our strategy to introduce 2'-dI and d-/l-isoNA modifications after the selection process is likely to be generally applicable to improve the in vivo stability of aptamers without compromising their binding ability.


Assuntos
Glioma/diagnóstico por imagem , Imagem Molecular , Tenascina/química , Células 3T3 , Animais , Aptâmeros de Nucleotídeos/química , Células Cultivadas , Fluorescência , Humanos , Camundongos , Simulação de Dinâmica Molecular
15.
Diabetes ; 66(3): 640-650, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-27993927

RESUMO

Although many functions of activating transcription factor 4 (ATF4) are identified, a role of ATF4 in the hypothalamus in regulating energy homeostasis is unknown. Here, we generated adult-onset agouti-related peptide neuron-specific ATF4 knockout (AgRP-ATF4 KO) mice and found that these mice were lean, with improved insulin and leptin sensitivity and decreased hepatic lipid accumulation. Furthermore, AgRP-ATF4 KO mice showed reduced food intake and increased energy expenditure, mainly because of enhanced thermogenesis in brown adipose tissue. Moreover, AgRP-ATF4 KO mice were resistant to high-fat diet-induced obesity, insulin resistance, and liver steatosis and maintained at a higher body temperature under cold stress. Interestingly, the expression of FOXO1 was directly regulated by ATF4 via binding to the cAMP-responsive element site on its promoter in hypothalamic GT1-7 cells. Finally, Foxo1 expression was reduced in the arcuate nucleus (ARC) of the hypothalamus of AgRP-ATF4 KO mice, and adenovirus-mediated overexpression of FOXO1 in ARC increased the fat mass in AgRP-ATF4 KO mice. Collectively, our data demonstrate a novel function of ATF4 in AgRP neurons of the hypothalamus in energy balance and lipid metabolism and suggest hypothalamic ATF4 as a potential drug target for treating obesity and its related metabolic disorders.


Assuntos
Fator 4 Ativador da Transcrição/genética , Núcleo Arqueado do Hipotálamo/metabolismo , Metabolismo Energético/genética , Resistência à Insulina/genética , Fígado/metabolismo , Neurônios/metabolismo , Proteína Relacionada com Agouti/metabolismo , Animais , Dieta Hiperlipídica , Ingestão de Alimentos/genética , Proteína Forkhead Box O1/metabolismo , Homeostase , Hipotálamo/citologia , Hipotálamo/metabolismo , Insulina/metabolismo , Leptina/metabolismo , Metabolismo dos Lipídeos/genética , Masculino , Camundongos , Camundongos Knockout , Obesidade/metabolismo
16.
Anal Chim Acta ; 935: 113-20, 2016 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-27543020

RESUMO

In order to highly enrich two liver cancer biomarkers (hexanal and 2-butanone) in human blood, in this study, natural nontoxic polydopamine/dialdehyde starch/chitosan (PD/DAS/CHI) coating material was synthesized and immobilized on the inner wall of polytetrafluoro-ethlyene (PTFE) tube. It was used to develop the method based on in-tube solid-phase microextraction (IT-SPME) with in-situ derivatization (ISD) coupled to high performance liquid chromatography for the determination of the above mentioned two liver cancer biomarkers in human blood. The simple, rapid and sensitive IT-SPME-ISD method can be finished within 11 min. Under optimum conditions, the limits of detection (LODs) were 1.4 and 1.6 nmol L(-1) for hexanal and 2-butanone, respectively. The relative recoveries from real human blood samples were in the range from 70% to 91% with the intra- and inter-day precisions less than 7.2%. Furthermore, this method was successfully applied for the analysis of hexanal and 2-butanone in blood samples from healthy people with 0.42 ± 0.05 and 0.34 ± 0.04 µmol L(-1), while liver cancer patients with 1.90 ± 0.07  µmol L(-1) and 0.91 ± 0.07 µmol L(-1), respectively. The t-test's results showed there is a statistically significant difference between the data from healthy persons and liver cancer patients. Hence, the developed method might be applied in the screening of suspected liver cancer patients.


Assuntos
Biomarcadores Tumorais/sangue , Neoplasias/sangue , Microextração em Fase Sólida , Idoso , Quitosana , Feminino , Humanos , Indóis , Masculino , Pessoa de Meia-Idade , Tamanho da Partícula , Polímeros , Amido/análogos & derivados
17.
J Biol Chem ; 291(35): 18536-46, 2016 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-27405764

RESUMO

Although numerous biological functions of the activating transcription factor 4 (ATF4) have been identified, a direct effect of ATF4 on alcoholic liver steatosis has not been described previously. The aim of our current study is to investigate the role of ATF4 in alcoholic liver steatosis and elucidate the underlying mechanisms. Here, we showed that the expression of ATF4 is induced by ethanol in hepatocytes in vitro and in vivo, and liver-specific ATF4 knock-out mice are resistant to ethanol-induced liver steatosis, associated with stimulated hepatic AMP-activated protein kinase (AMPK) activity. Furthermore, adenovirus-mediated AMPK knockdown significantly reversed the suppressive effects of ATF4 deficiency on ethanol-induced liver steatosis in mice. In addition, ethanol-fed ATF4 knock-out mice exhibit AMPK-dependent inhibition of fatty acid synthase and stimulation of carnitine palmitoyltransferase 1 (CPT1) in the liver. Moreover, hepatic Tribbles homolog 3 (TRB3) expression was stimulated by ethanol in an ATF4-dependent manner, and adenovirus-mediated TRB3 knockdown blocked ATF4-dependent ethanol-induced AMPK inhibition and triglyceride accumulation in AML-12 cells. Finally, TRB3 directly interacted with AMPK to suppress its phosphorylation. Taken together, these results identify the ATF4-TRB3-AMPK axis as a novel pathway responsible for ethanol-induced liver steatosis.


Assuntos
Fator 4 Ativador da Transcrição/metabolismo , Fígado Gorduroso Alcoólico/metabolismo , Hepatócitos/metabolismo , Fígado/metabolismo , Transdução de Sinais , Triglicerídeos/biossíntese , Proteínas Quinases Ativadas por AMP/biossíntese , Proteínas Quinases Ativadas por AMP/genética , Fator 4 Ativador da Transcrição/genética , Animais , Proteínas de Ciclo Celular/biossíntese , Proteínas de Ciclo Celular/genética , Linhagem Celular , Etanol/efeitos adversos , Etanol/farmacologia , Fígado Gorduroso Alcoólico/genética , Fígado Gorduroso Alcoólico/patologia , Hepatócitos/patologia , Fígado/patologia , Masculino , Camundongos , Camundongos Knockout , Especificidade de Órgãos/genética , Triglicerídeos/genética
18.
Sci Signal ; 9(428): ra50, 2016 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-27188441

RESUMO

Liver steatosis, a condition in which lipid accumulates in liver cells, is a leading cause of many liver diseases. The livers of patients with hepatocellular carcinoma, a cancer characterized by liver steatosis, have decreased abundance of the transcription cofactor BTG1 (B cell translocation gene 1). We showed that the livers of db/db mice, which are a genetic model of obesity, had decreased BTG1 mRNA and protein abundance. BTG1 overexpression ameliorated liver steatosis in db/db mice, whereas knockdown of BTG1 induced liver steatosis in wild-type mice. Consistent with these changes, we found that BTG1 decreased triglyceride accumulation in cultured hepatocytes. BTG1 overexpression inhibited the expression of the gene encoding stearoyl-CoA desaturase 1 (SCD1), an enzyme involved in the synthesis of fatty acids, by suppressing the activity of activating transcription factor 4 (ATF4). Knockdown of SCD1 prevented liver steatosis in wild-type mice induced by knockdown of BTG1. Conversely, the ability of BTG1 overexpression to ameliorate liver steatosis in db/db mice was negated by ATF4 overexpression. Moreover, BTG1 transgenic mice were resistant to liver steatosis induced by a high-carbohydrate diet. BTG1 abundance was decreased by this diet through a pathway that involved mammalian target of rapamycin (mTOR), ribosomal protein S6 kinase 1 (S6K1), and cAMP response element-binding protein (CREB). Together, our study identifies a role of BTG1 in regulating hepatic lipid metabolism and specifically in preventing ATF4 and SCD1 from inducing liver steatosis.


Assuntos
Fator 4 Ativador da Transcrição/metabolismo , Fígado Gorduroso/metabolismo , Metabolismo dos Lipídeos , Fígado/metabolismo , Proteínas de Neoplasias/metabolismo , Estearoil-CoA Dessaturase/metabolismo , Animais , Carcinoma Hepatocelular/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Ácidos Graxos/metabolismo , Células HEK293 , Hepatócitos/metabolismo , Humanos , Neoplasias Hepáticas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Obesidade/metabolismo , RNA Mensageiro/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Triglicerídeos/metabolismo
19.
Diabetes ; 65(8): 2151-63, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27207553

RESUMO

Friend of GATA 2 (FOG2) is a transcriptional cofactor involved mostly in cardiac function. The aim of this study was to investigate the role of hepatic FOG2 in insulin sensitivity and lipid accumulation. FOG2 overexpression by adenovirus-expressing FOG2 (Ad-FOG2) significantly attenuates insulin signaling in hepatocytes in vitro. Opposite effects were observed when FOG2 was knocked down through adenovirus-expressing small hairpin RNA for FOG2 (Ad-shFOG2). Furthermore, FOG2 knockdown by Ad-shFOG2 ameliorated insulin resistance in leptin receptor-mutated (db/db) mice, and FOG2 overexpression by Ad-FOG2 attenuated insulin sensitivity in C57BL/6J wild-type (WT) mice. In addition, Ad-FOG2 reduced, whereas Ad-shFOG2 promoted, hepatic triglyceride (TG) accumulation in WT mice under fed or fasted conditions, which was associated with increased or decreased hepatic peroxisome proliferator-activated receptor α (PPARα) expression, respectively. Moreover, the improved insulin sensitivity and increased hepatic TG accumulation by Ad-shFOG2 were largely reversed by adenovirus-expressing PPARα (Ad-PPARα) in WT mice. Finally, we generated FOG2 liver-specific knockout mice and found that they exhibit enhanced insulin sensitivity and elevated hepatic TG accumulation, which were also reversed by Ad-PPARα. Taken together, the results demonstrate a novel function of hepatic FOG2 in insulin sensitivity and lipid metabolism through PPARα.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Resistência à Insulina/genética , Metabolismo dos Lipídeos/genética , Fígado/metabolismo , PPAR alfa/metabolismo , Fatores de Transcrição/metabolismo , Animais , Glicemia/metabolismo , Colesterol/metabolismo , Proteínas de Ligação a DNA/genética , Ácidos Graxos não Esterificados/metabolismo , Feminino , Teste de Tolerância a Glucose , Glicogênio/metabolismo , Insulina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , PPAR alfa/genética , Receptores para Leptina/genética , Receptores para Leptina/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Fatores de Transcrição/genética , Triglicerídeos/metabolismo
20.
Autophagy ; 12(3): 592-3, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26760678

RESUMO

Although many biological functions of MAPK1/ERK2-MAPK3/ERK1 (mitogen-activated protein kinase 1/3) have been reported, a direct effect of MAPK1/3 on hepatic lipid metabolism remains largely unknown. We recently showed that activation of MAPK1/3 ameliorates liver steatosis in LEPR (leptin receptor)-deficient (db/db) mice, a classic animal model for liver steatosis. Consistent with these results, knockdown of MAPK1/3 promotes liver steatosis in C57/B6J wild-type (WT) mice. Autophagic flux and ATG7 (autophagy related 7) levels are increased by MAPK1/3 activation or decreased by MAPK1/3 knockdown in livers and primary hepatocytes. Blockade of autophagic flux by chloroquine (CQ) or ATG7 knockdown reverses the ameliorated liver steatosis in MAPK1/3-activated db/db mice. Together, these findings identify a beneficial role for MAPK1/3 in liver steatosis that is mediated by ATG7-dependent autophagy, which provides novel insights into the mechanisms underlying liver steatosis and create a rationale for targeting MAPK1/3 in the treatment of liver steatosis.


Assuntos
Proteína 7 Relacionada à Autofagia/metabolismo , Autofagia , Metabolismo dos Lipídeos , Fígado/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Animais , Fígado/enzimologia , Camundongos Endogâmicos C57BL
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