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1.
Neurochem Res ; 2021 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-33394221

RESUMO

Alzheimer's disease (AD) is a common neurodegenerative disease associated with deposition of ß-amyloid peptide (Aß). Platycodin D (PLD), a triterpenesaponin, may possess neuro-protective effect. In the current study, we aimed to explore the effects of PLD on Aß-induced inflammation and oxidative stress in microglial BV-2 cells. Our study showed that PLD treatment improved cell viability in Aß-induced BV-2 cells. PLD attenuated Aß-induced inflammation with deceased production of TNF-α, IL-1ß and IL-6 in Aß-induced BV-2 cells. PLD also mitigated the oxidative stress in Aß-induced BV-2 cells, as evidenced by deceased production of ROS and MDA, and increased SOD activity. Furthermore, the increased expression levels of TLR4 and p-p65 and decreased IκBα expression in the Aß-stimulated BV-2 cells were attenuated by PLD treatment. Overexpression of TLR4 reversed the anti-inflammatory effect of PLD in Aß-stimulated BV-2 cells. In addition, PLD treatment enhanced the Aß-stimulated increase in the expression levels of Nrf2, HO-1, and NQO1 in BV-2 cells. Knockdown of Nrf2 abrogated the anti-oxidative effect of PLD in Aß-stimulated BV-2 cells. In conclusion, these findings indicated that PLD protected BV-2 cells from Aß-induced oxidative stress and inflammation via regulating the TLR4/NF-κB and Nrf2/HO-1 signaling pathways. Thus, PLD may be a potential candidate for the treatment of AD.

2.
BMC Psychiatry ; 20(1): 392, 2020 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-32736652

RESUMO

BACKGROUND: This research aims to explore the life experiences of relocated earthquake survivors with PTSD and develop a conceptual framework for understanding their life experiences. METHOD: Interviews were conducted with twenty-three participants. The participant selection, data collection and analysis were based on grounded theory methodology. A theoretical model called "loss of homeland" was developed. RESULTS: Loss of homeland was the most important condition that influenced the relocated participants' self-identity, social connections, and meaning system. These aspects were categorized into existential changes, lost connections, and changes in identity. Post-disaster relocation threatens individuals' sense of meaning, integrity of self, and sense of belonging, affects every aspect of everyday life and shatters their inner and outer harmony. CONCLUSIONS: Further research guided by this theoretical model is needed to inform post-disaster mental health services and relocation policy. Mental health professionals and policy makers can make more informed decisions in terms of disaster relocation policy and manage post-disaster psychological disturbances by focusing on both places and people.

3.
Waste Manag ; 106: 145-154, 2020 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-32217443

RESUMO

Crushing is the key part for the recycling technology of waste printed circuit boards (WPCBs). In this study, the breakage and liberation effects of WPCBs was improved by heat pretreatment technology before crushing (HPBC). Based on the test results, 200 °C was found as the safe temperature for the HPBC of WPCBs. The fracture mode, particle size distribution, and enrichment characteristics of WPCBs were systematically studied. The experimental results show that the HPBC changed the breakage mode from longitudinal fracture to horizontal fracture and improved the liberation of metal from non-metal components. During the crushing process, the increase in the heat pretreatment time (30-120 min) and temperature (100-200 °C) can improve the crushing effect of WPCBs and increase the content of crushing products of -0.3 mm by 3.16% and 5.64%, respectively. Compared to the non-metallic components, the metal components have ductility and are difficult to break into -0.3 mm during the crushing process. HPBC can promote copper enrichment to narrow grain size. In the 0.3-1 mm range, the content of copper increased from 47.87% to 57.61%, an increase by 9.74%. The initial enrichment of copper was achieved by adjusting the crushing time. The recovery rate of copper can reach 85.66%, and the enrichment rate is 1.74 when 0.3-2 mm breaking product is used as enrichment. Therefore, HPBC can effectively improve the crushing and liberation effect of WPCBs and improve the enrichment rate, and thus is an effective pretreatment method.


Assuntos
Resíduo Eletrônico , Cobre , Calefação , Metais , Reciclagem
4.
Cell Signal ; 28(8): 1058-65, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27178152

RESUMO

Nucleobindin-1 (NUCB1), also known as Calnuc, is a highly conserved, multifunctional protein widely expressed in tissues and cells. It contains two EF-hand motifs which have been shown to play a crucial role in binding Ca(2+) ions. In this study, we applied comparative two-dimensional gel electrophoresis to characterize differentially expressed proteins in HA-CHIP over-expressed and endogenous CHIP depleted MC3T3-E1 stable cell lines, identifying NUCB1 as a novel CHIP/Stub1 targeted protein. NUCB1 interacts with and is down-regulated by CHIP by both proteasomal dependent and independent pathways, suggesting that CHIP-mediated down-regulation of nucleobindin-1 might play a role in osteoblast differentiation. The chaperone protein Hsp70 was found to be important for CHIP and NUCB1 interaction as well as CHIP-mediated NUCB1 down-regulation. Our findings provide new insights into understanding the stability regulation of NUCB1.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação para Baixo , Modelos Biológicos , Proteínas do Tecido Nervoso/metabolismo , Osteoblastos/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Diferenciação Celular , Linhagem Celular , Proteínas de Choque Térmico HSP70/metabolismo , Humanos , Camundongos , Nucleobindinas , Osteoblastos/citologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Ligação Proteica , Proteômica , Transdução de Sinais , Regulação para Cima
5.
J Chromatogr A ; 1452: 1-9, 2016 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-27207578

RESUMO

Peak detection is a critical step in chromatographic data analysis. In the present work, we developed a multi-scale Gaussian smoothing-based strategy for accurate peak extraction. The strategy consisted of three stages: background drift correction, peak detection, and peak filtration. Background drift correction was implemented using a moving window strategy. The new peak detection method is a variant of the system used by the well-known MassSpecWavelet, i.e., chromatographic peaks are found at local maximum values under various smoothing window scales. Therefore, peaks can be detected through the ridge lines of maximum values under these window scales, and signals that are monotonously increased/decreased around the peak position could be treated as part of the peak. Instrumental noise was estimated after peak elimination, and a peak filtration strategy was performed to remove peaks with signal-to-noise ratios smaller than 3. The performance of our method was evaluated using two complex datasets. These datasets include essential oil samples for quality control obtained from gas chromatography and tobacco plant samples for metabolic profiling analysis obtained from gas chromatography coupled with mass spectrometry. Results confirmed the reasonability of the developed method.


Assuntos
Cromatografia Gasosa/métodos , Cromatografia Gasosa-Espectrometria de Massas , Metabolômica/métodos , Distribuição Normal , Óleos Voláteis/química , Óleos Voláteis/isolamento & purificação , Controle de Qualidade , Razão Sinal-Ruído , Tabaco/química
6.
J Sep Sci ; 39(11): 2123-8, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-27059265

RESUMO

A method was developed for the determination of nine volatile N-nitrosamines in tobacco and smokeless tobacco products. The targets are N-nitrosodimethylamine, N-nitrosopyrrolidine, N-nitrosopiperidine, N-nitrosomorpholine, N-nitrosoethylmethylamine, N-nitrosodiethylamine, N-nitrosodipropylamine, N-nitrosobuylmethylmine, and N-nitrosodibutylamine. The samples were treated by dispersive solid-phase extraction using 1 g of primary secondary amine and 0.5 g of carbon and then analyzed by gas chromatography with tandem mass spectrometry with an electron impact ion source. The recoveries for the targets ranged from 84 to 118%, with <16% relative standard deviations at three spiking levels of 0.5, 1.25, and 2.5 ng/g. The limits of detection ranged from 0.03 to 0.15 ng/g. With the use of the proposed method, we detected the presence of six nitrosamines in the range of 0.4-30.7 ng/g. The study demonstrated that the method could be used as a rapid, convenient, and high-throughput method for N-nitrosamines analysis in tobacco matrix.


Assuntos
Nitrosaminas/análise , Extração em Fase Sólida , Produtos do Tabaco/análise , Tabaco sem Fumaça/análise , Tabaco/química , Compostos Orgânicos Voláteis/análise , Cromatografia Gasosa , Espectrometria de Massas em Tandem
7.
Inhal Toxicol ; 28(2): 89-94, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26865272

RESUMO

To accurately estimate the risk of inhaling cigarette smoke containing toxic chemicals, it is important that the distribution of these chemicals is accurately measured in cigarette smoke aerosol particles of various sizes. In this study, a single-channel smoking machine was directly coupled to an electrical low-pressure impactor. The particles of mainstream cigarette smoke were collected using 12 polyester films, and the particulate matter (PM) was characterized. Nicotine, tobacco-specific N-nitrosamines (TSNAs, including NNN, NAT, NAB, and NNK), polycyclic aromatic hydrocarbons (PAHs, including benzo(a)pyrene (BaP), benzo(a)anthracene, and chrysene), and heavy metals (including Cr, As, Cd, and Pb) present in the particles of different sizes were analyzed by GC, HPLC-MS/MS, GC/MS, or ICP-MS, respectively. The results demonstrated that the nicotine, TSNAs, PAHs, and heavy metals in mainstream cigarette smoke were dispersed over a particle size ranging from 0.1 µm to 2.0 µm, and the concentration of these toxic chemicals initially increased and then decreased the particle size grew. The distribution of nicotine was uniform for the PM in the size ranges of less than 0.1 µm, 0.1-1.0 µm, and 1.0-2.0 µm, TSNAs and heavy metals in particles of less 0.1 µm were more abundant, and PAHs in fine particles were also more abundant.


Assuntos
Tamanho da Partícula , Material Particulado/química , Fumaça/análise , Produtos do Tabaco/análise , Metais Pesados/química , Nicotina/química
8.
Cell Biochem Biophys ; 70(3): 1799-802, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25062968

RESUMO

The objective of the study was to explore the effects of olanzapine-fluoxetine combination (OFC) treatment of major depressive disorders on the quality of life in the acute treatment period. Methods were prospective and observational design. One hundred and three patients of major depressive disorders were observed. One group of 53 patients received OFC treatment (OFC group); the other group of 50 patients received the treatment of duloxetine (duloxetine group). Two groups were needed to be observed 8 weeks. Observed indicators were Hamilton Depression Rating Scale for Depression (HAMD-24) and four factor scores: the slow, sleep disorders, anxiety/somatization, and hopelessness, Clinical Global Impression-Severity of Illness (CGI-S), WHO quality of life scale (WHOQOL-BREF), and sub-rate measurements. HAMD-24 and four factor scores observation time were assessed before and after treatment; 1, 2, 4, 8 weeks, WHOQOL-BREF score, and sub-time measurements were assessed before treatment and 8 weeks after treatment. HAMD-24 scores of OFC patients in the first week were significantly lower than those of the duloxetine group. The sleep factor scores of OFC patients were significantly lower than those of the duloxetine group in 4 and 8 weeks. By the end of 8 weeks, OFC group was rated significantly lower than the duloxetine group in the physical area. In the acute treatment period, OFC treatment effected faster than the single duloxetine in patients with major depressive disorders. OFC effected within 1 week and was better than the single duloxetine in improving the sleep and physical conditions.


Assuntos
Benzodiazepinas/uso terapêutico , Transtorno Depressivo Maior/tratamento farmacológico , Fluoxetina/uso terapêutico , Qualidade de Vida , Adulto , Benzodiazepinas/efeitos adversos , Transtorno Depressivo Maior/psicologia , Esquema de Medicação , Combinação de Medicamentos , Cloridrato de Duloxetina , Feminino , Fluoxetina/efeitos adversos , Humanos , Masculino , Pessoa de Meia-Idade , Escalas de Graduação Psiquiátrica , Transtornos do Sono-Vigília/etiologia , Transtornos Somatoformes/etiologia , Tiofenos/efeitos adversos , Tiofenos/uso terapêutico
9.
J Psychiatry Neurosci ; 39(5): 304-11, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24866415

RESUMO

BACKGROUND: Obsessive-compulsive disorder (OCD) is a common, heritable neuropsychiatric disorder, hypothetically underpinned by dysfunction of brain cortical-striatal-thalamic-cortical (CSTC) circuits; however, the extent of brain functional abnormalities in individuals with OCD is unclear, and the genetic basis of this disorder is poorly understood. We determined the whole brain functional connectivity patterns in patients with OCD and their healthy first-degree relatives. METHODS: We used resting-state fMRI to measure functional connectivity strength in patients with OCD, their healthy first-degree relatives and healthy controls. Whole brain functional networks were constructed by measuring the temporal correlations of all brain voxel pairs and further analyzed using a graph theory approach. RESULTS: We enrolled 39 patients with OCD, 20 healthy first-degree relatives and 39 healthy controls in our study. Compared with healthy controls, patients with OCD showed increased functional connectivity primarily within the CSTC circuits and decreased functional connectivity in the occipital cortex, temporal cortex and cerebellum. Moreover, patients with OCD and their first-degree relatives exhibited overlapping increased functional connectivity strength in the bilateral caudate nucleus, left orbitofrontal cortex (OFC) and left middle temporal gyrus. LIMITATIONS: Potential confounding factors, such as medication use, heterogeneity in symptom clusters and comorbid disorders, may have impacted our findings. CONCLUSION: Our preliminary results suggest that patients with OCD have abnormal resting-state functional connectivity that is not limited to CSTC circuits and involves abnormalities in additional large-scale brain systems, especially the limbic system. Moreover, resting-state functional connectivity strength abnormalities in the left OFC, bilateral caudate nucleus and left middle temporal gyrus may be neuroimaging endophenotypes for OCD.


Assuntos
Encéfalo/fisiopatologia , Família , Transtorno Obsessivo-Compulsivo/fisiopatologia , Adulto , Mapeamento Encefálico , Comorbidade , Endofenótipos , Feminino , Humanos , Imagem por Ressonância Magnética , Masculino , Vias Neurais/fisiopatologia , Transtorno Obsessivo-Compulsivo/tratamento farmacológico , Transtorno Obsessivo-Compulsivo/epidemiologia , Processamento de Sinais Assistido por Computador
10.
Arthritis Rheumatol ; 66(7): 1854-63, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24578159

RESUMO

OBJECTIVE: Carboxyl terminus of Hsp70-interacting protein (CHIP or STUB1) is an E3 ligase that regulates the stability of several proteins involved in tumor growth and metastasis. However, the role of CHIP in bone growth and bone remodeling in vivo has not been reported. This study was undertaken to investigate the role and mechanism of CHIP in regulation of bone mass and bone remodeling. METHODS: The bone phenotype of Chip(-/-) mice was assessed by histologic, histomorphometric, and micro-computed tomographic analyses. The mechanism by which CHIP regulates the degradation of tumor necrosis factor receptor-associated factor 6 (TRAF6) and the inhibition of NF-κB signaling was examined by immunoprecipitation, Western blot, and luciferase reporter assays. RESULTS: Deletion of the Chip gene led to an osteopenic phenotype and increased osteoclast formation. TRAF6, an adaptor protein that is a key regulator of NF-κB signaling and is critical for RANKL-induced osteoclastogenesis, was up-regulated in osteoclasts from Chip(-/-) mice. CHIP interacted with TRAF6 to promote TRAF6 ubiquitination and proteasome degradation. Further, CHIP inhibited p65 nuclear translocation, leading to the repression of TRAF6-mediated NF-κB transcription. CONCLUSION: CHIP inhibits NF-κB signaling by promoting TRAF6 degradation and plays an important role in osteoclastogenesis and bone remodeling. These findings suggest that CHIP may be a novel therapeutic target in bone loss-associated disorders.


Assuntos
Desenvolvimento Ósseo/fisiologia , Remodelação Óssea/fisiologia , Osteoclastos/metabolismo , Fator 6 Associado a Receptor de TNF/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Doenças Ósseas Metabólicas/genética , Doenças Ósseas Metabólicas/metabolismo , Doenças Ósseas Metabólicas/patologia , Reabsorção Óssea/genética , Reabsorção Óssea/metabolismo , Reabsorção Óssea/patologia , Linhagem Celular , Células HEK293 , Humanos , Macrófagos/citologia , Camundongos , Camundongos Knockout , NF-kappa B/metabolismo , Osteoclastos/patologia , Fenótipo , Transdução de Sinais/fisiologia , Ubiquitina-Proteína Ligases/genética , Ubiquitinação/fisiologia
11.
Biochem Biophys Res Commun ; 446(1): 387-92, 2014 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-24613385

RESUMO

Transforming growth factor-ß (TGF-ß) signaling plays an important role in regulation of a wide variety of cellular processes. Canonical TGF-ß signaling is mediated by Smads which were further regulated by several factors. We previously reported that E3 ubiquitin ligase CHIP (carboxyl terminus of Hsc70-interacting protein, also named Stub1) controlled the sensitivity of TGF-ß signaling by modulating the basal level of Smad3 through ubiquitin-mediated degradation. Here, we present evidence that Hsp70 and Hsp90 regulate the complex formation of Smad3/CHIP. Furthermore, we observed that over-expressed Hsp70 or inhibition of Hsp90 by geldanamycin (GA) leads to facilitated CHIP-induced ubiquitination and degradation of Smad3, which finally enhances TGF-ß signaling. In contrast, over-expressed Hsp90 antagonizes CHIP mediated Smad3 ubiquitination and degradation and desensitizes cells in response to TGF-ß signaling. Taken together, our data reveal an opposite role of Hsp70 and Hsp90 in regulating TGF-ß signaling by implicating CHIP-mediated Smad3 ubiquitination and degradation. This study provides a new insight into understanding the regulation of the TGF-ß signaling by chaperones.


Assuntos
Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Células COS , Linhagem Celular , Chlorocebus aethiops , Células HEK293 , Proteínas de Choque Térmico HSP70/química , Proteínas de Choque Térmico HSP90/química , Humanos , Vison , Complexos Multiproteicos/química , Complexos Multiproteicos/metabolismo , Proteólise , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Transdução de Sinais , Proteína Smad3/química , Proteína Smad3/metabolismo , Ubiquitina-Proteína Ligases/química , Ubiquitinação
12.
PLoS One ; 8(12): e83931, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24358320

RESUMO

BACKGROUND: Previous studies have demonstrated that structural deficits and functional connectivity imbalances might underlie the pathophysiology of obsessive-compulsive disorder (OCD). The purpose of the present study was to investigate gray matter deficits and abnormal resting-state networks in patients with OCD and further investigate the association between the anatomic and functional alterations and clinical symptoms. METHODS: Participants were 33 treatment-naïve OCD patients and 33 matched healthy controls. Voxel-based morphometry was used to investigate the regions with gray matter abnormalities and resting-state functional connectivity analysis was further conducted between each gray matter abnormal region and the remaining voxels in the brain. RESULTS: Compared with healthy controls, patients with OCD showed significantly increased gray matter volume in the left caudate, left thalamus, and posterior cingulate cortex, as well as decreased gray matter volume in the bilateral medial orbitofrontal cortex, left anterior cingulate cortex, and left inferior frontal gyrus. By using the above morphologic deficits areas as seed regions, functional connectivity analysis found abnormal functional integration in the cortical-striatum-thalamic-cortical (CSTC) circuits and default mode network. Subsequent correlation analyses revealed that morphologic deficits in the left thalamus and increased functional connectivity within the CSTC circuits positively correlated with the total Y-BOCS score. CONCLUSION: This study provides evidence that morphologic and functional alterations are seen in CSTC circuits and default mode network in treatment-naïve OCD patients. The association between symptom severity and the CSTC circuits suggests that anatomic and functional alterations in CSTC circuits are especially important in the pathophysiology of OCD.


Assuntos
Córtex Cerebral/fisiopatologia , Corpo Estriado/fisiopatologia , Vias Neurais , Transtorno Obsessivo-Compulsivo/patologia , Transtorno Obsessivo-Compulsivo/fisiopatologia , Adolescente , Adulto , Comportamento , Mapeamento Encefálico , Estudos de Casos e Controles , Feminino , Humanos , Imagem por Ressonância Magnética , Masculino , Reprodutibilidade dos Testes , Tálamo/fisiopatologia , Adulto Jovem
13.
Protein Expr Purif ; 90(1): 27-33, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23648869

RESUMO

Protein transduction domains (PTDs), such as the TAT peptide derived from HIV Tat protein, may transduce macromolecules into cells. In the present study, the TAT peptide-fused artificial transcription factors (ATFs) were generated by fusion of the N-terminal TAT peptide with SV40 promoter-targeted three-fingered C2H2 zinc finger proteins and the KRAB transcriptional repression domain. The fusion proteins were then expressed in an E .coli system and purified by Ni-NTA affinity chromatography. The purified fusion proteins were tested on mammalian cell lines CHO DG44 and L929. TAT-ATF-S, which contains the zinc fingers that bind to the SV40 promoter with high specificity, exhibited the desired transcriptional repression activity to the reported genes, indicating the successful cellular delivery and desired conformation of TAT-ATF-S. Our study has provided a new strategy for intracellular ATF delivery.


Assuntos
HIV-1/metabolismo , Proteínas Recombinantes de Fusão/genética , Produtos do Gene tat do Vírus da Imunodeficiência Humana/genética , Animais , Sítios de Ligação , Células CHO , Cricetinae , Cricetulus , Produtos do Gene tat/genética , HIV-1/genética , Estrutura Terciária de Proteína , Proteínas Recombinantes de Fusão/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Dedos de Zinco , Produtos do Gene tat do Vírus da Imunodeficiência Humana/metabolismo
14.
J Biomol Screen ; 18(4): 378-87, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23042077

RESUMO

Chronic hepatitis B virus (HBV) infection is an independent risk factor for the development of hepatocellular carcinoma (HCC). The HBV HBx gene is frequently identified as an integrant in the chromosomal DNA of patients with HCC. HBx encodes the X protein (HBx), a putative viral oncoprotein that affects transcriptional regulation of several cellular genes. Therefore, HBx may be an ideal target to impede the progression of HBV infection-related HCC. In this study, integrated HBx was transcriptionally downregulated using an artificial transcription factor (ATF). Two three-fingered Cys2-His2 zinc finger (ZF) motifs that specifically recognized two 9-bp DNA sequences regulating HBx expression were identified from a phage-display library. The ZF domains were linked into a six-fingered protein that specified an 18-bp DNA target in the Enhancer I region upstream of HBx. This DNA-binding domain was fused with a Krüppel-associated box (KRAB) transcriptional repression domain to produce an ATF designed to downregulate HBx integrated into the Hep3B HCC cell line. The ATF significantly repressed HBx in a luciferase reporter assay. Stably expressing the ATF in Hep3B cells resulted in significant growth arrest, whereas stably expressing the ATF in an HCC cell line lacking integrated HBx (HepG2) had virtually no effect. The targeted downregulation of integrated HBx is a promising novel approach to inhibiting the progression of HBV infection-related HCC.


Assuntos
Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , Transativadores/genética , Fatores de Transcrição/metabolismo , Dedos de Zinco , Sequência de Aminoácidos , Sequência de Bases , Carcinoma Hepatocelular/patologia , Proliferação de Células , Regulação para Baixo/genética , Elementos Facilitadores Genéticos/genética , Genes Reporter , Genoma Humano/genética , Células Hep G2 , Humanos , Neoplasias Hepáticas/patologia , Dados de Sequência Molecular , Regiões Promotoras Genéticas/genética , Ligação Proteica/genética , Proteínas Repressoras , Fatores de Transcrição/química , Proteínas Virais Reguladoras e Acessórias
15.
Biosci Biotechnol Biochem ; 76(10): 1909-12, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23047106

RESUMO

An optimized method based on tetracycline-inducible gene expression system T-REx was developed to screen and evaluate Tet repressor (TetR)-expressing cell lines using enhanced green fluorescence protein (EGFP) as reporter gene. To verify the effectiveness of the method, two TetR-expressing Chinese hamster ovary (CHO) cell lines, CHO-TR B2 (stringent) and B5 (less stringent), in which the EGFP genes were variantly controlled by tetracycline, were used to construct cell lines expressing the anti-apoptosis gene survivin upon induction with tetracycline. The resulting stable clones were analyzed for survivin expression. The analysis showed that all four B5-derived clones exhibited leaky survivin expression in the absence of tetracycline, while the B2-derived clones did not. DNA laddering and annexin V/PI staining assays further indicated that although tetracycline-inducible expression of survivin conferred resistance to NH4Cl- and staurosporine-induced apoptosis in both the B2- and the B5-derived stable cell lines, the B2-derived cell lines showed more stringent regulation in the absence of tetracycline. This represents successful utilization of the present screening method.


Assuntos
Técnicas Genéticas , Proteínas Inibidoras de Apoptose/genética , Tetraciclina/farmacologia , Animais , Células CHO , Cricetinae , Cricetulus , Expressão Gênica/efeitos dos fármacos , Genes Reporter/genética , Proteínas de Fluorescência Verde/genética , Proteínas Repressoras/genética
16.
J Bone Miner Res ; 27(7): 1607-18, 2012 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-22407479

RESUMO

Differentiation of committed precursor cells into the osteoblast lineage is tightly regulated by several factors, including Runx2 and BMP2. We previously reported that C terminus of Hsc70-interacting protein/STIP1 homology and U-Box containing protein 1 (CHIP/STUB1) negatively regulated osteoblast differentiation through promoting Runx2 protein degradation. However, how CHIP is regulated during osteoblast differentiation remains unknown. In this study, we found that miR-764-5p is up-expressed during the osteoblast differentiation in calvarial and osteoblast progenitor cells, coupled with down-expression of CHIP protein. We observed that forced expression or inhibition of miR-764-5p decreased or increased the CHIP protein level through affecting its translation by targeting the 3'-UTR region. Perturbation of miR-764-5p resulted in altered differentiation fate of osteoblast progenitor cells and the role of miR-764-5p was reversed by overexpression of CHIP, whereas depletion of CHIP impaired the effect of miR-764-5p. Our data showed that miR-764-5p positively regulates osteoblast differentiation from osteoblast progenitor cells by repressing the translation of CHIP protein.


Assuntos
Regulação da Expressão Gênica , MicroRNAs/metabolismo , Osteoblastos/citologia , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/fisiologia , Regiões 3' não Traduzidas , Células 3T3 , Animais , Diferenciação Celular , Proteínas de Choque Térmico HSC70/metabolismo , Imunoglobulina G/química , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/fisiologia , Biossíntese de Proteínas , Estrutura Terciária de Proteína , Ubiquitina-Proteína Ligases/antagonistas & inibidores
17.
J Affect Disord ; 138(3): 313-21, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22331021

RESUMO

BACKGROUND: Abnormality of orbitofronto-striatal circuits was postulated in obsessive-compulsive disorder (OCD). The aim of this study is to test the abnormality hypothesis of orbitofronto-striatal circuits and explore whether there are any other dysfunctional brain regions in OCD using a resting-state functional magnetic resonance imaging (fMRI), and further investigate the relationship between the whole-brain voxel-based spontaneous neuronal activity of patients with OCD and clinical characteristics. METHODS: 23 patients with OCD and 23 age- and gender-matched normal controls were examined using resting-state fMRI, and amplitude of low-frequency fluctuation (ALFF) approach was used to analyze fMRI data. RESULTS: Compared with normal controls, patients with OCD presented increased ALFF in the bilateral orbitofrontal cortex (OFC), anterior cingulate cortex (ACC) as well as decreased ALFF in the bilateral cerebellum and parietal cortex (P<0.01, corrected). Additionally, the ALFF values in bilateral OFC were positively correlated with total Y-BOCS scores (P<0.005, uncorrected). CONCLUSION: Our findings added an expanding literature to the abnormality hypothesis of orbitofronto-striatal circuits and showed the changed spontaneous neuronal activity of the parietal cortex and cerebellum may also play an important role in the pathophysiology in patients with OCD.


Assuntos
Encéfalo/fisiopatologia , Imagem por Ressonância Magnética , Transtorno Obsessivo-Compulsivo/fisiopatologia , Adolescente , Adulto , Feminino , Humanos , Masculino , Transtorno Obsessivo-Compulsivo/diagnóstico , Transtorno Obsessivo-Compulsivo/patologia , Adulto Jovem
18.
Int J Hyperthermia ; 27(3): 275-85, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21501029

RESUMO

PURPOSE: Whole body hyperthermia (WBH) has been regarded as a promising alternative therapy to cure late stage cancer with metastasis. As the final biological and therapeutic effects are dependent on the specific protocol, the potential of using a microwave-based WBH approach for metastasis inhibition is established and its typical results are discussed. MATERIALS AND METHODS: The effectiveness of a 30-min whole body hyperthermia (WH) on animals, raised to a rectal temperature of 40.2° ± 0.3°C for 30 min followed by 84 h observation by 2450 MHz microwave irradiation, were evaluated. In an experimental lung metastasis model by injection of B16-F10 melanoma, lungs were removed from sacrificed mice 16 days after tumour implantation, and the expression of heat shock protein, inter-cellular adhesion molecule 1 (ICAM-1), proliferating cell nuclear antigen (PCNA) and cyclin D(1) was examined. CD4(+), CD8(+) and NK cell subpopulation in peripheral blood were measured by flow cytometry before and after the last treatment. RESULTS: The best therapeutic effect was obtained when the mice were treated with WBH in combination with the initial chemotherapy with cis-diaminodichloroplatinum (CDDP) and dacarbazine (DTIC) (p < 0.05). The WBH alone has an advantage of reduced toxicity and lower cost. Heat shock protein (HSP) expression increased in the hyperthermia groups. Reduction of PCNA and cyclin D(1) was observed in the mice treated with WH alone or in combination with chemotherapy. In the hyperthermia groups, CD4(+)/CD8(+) decreased while the NK increased slightly. CONCLUSIONS: The whole body hyperthermia protocol described in this work inhibits B16 tumour metastasis by inhibiting cell proliferation, neovascularisation and stimulating favourable immune responses. It demonstrated that WBH treatment benefits therapy of metastasis cancers.


Assuntos
Hipertermia Induzida/métodos , Imunidade/imunologia , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Melanoma Experimental/terapia , Metástase Neoplásica/patologia , Animais , Antineoplásicos/uso terapêutico , Cisplatino/uso terapêutico , Dacarbazina/uso terapêutico , Feminino , Febre/imunologia , Neoplasias Pulmonares/secundário , Camundongos , Camundongos Endogâmicos C57BL
19.
Biotechnol Lett ; 33(7): 1293-300, 2011 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-21380526

RESUMO

To optimize Chinese hamster ovary (CHO) cell culture to recombinant protein therapeutic production, we stably overexpressed survivin and cyclin D1 in three CHO DG44-derived cell lines. The modifications conferred increases of 56-94% in S-phase fractions and decreases of 33-43% in early-stage apoptosis fractions. Clone 6.3, which expressed the highest levels of survivin and cyclin D1, reached significantly greater cell densities in suspension (2.7 × 10(6) cells/ml) following serum deprivation. Nude mice inoculated with the modified cells showed no tumorigenesis suggesting that the CHO DG44-derived cell lines are viable candidates for biopharmaceutical production.


Assuntos
Apoptose , Células CHO/fisiologia , Meios de Cultura Livres de Soro , Ciclina D1/biossíntese , Expressão Gênica , Proteínas Inibidoras de Apoptose/biossíntese , Animais , Técnicas de Cultura de Células/métodos , Sobrevivência Celular , Cricetinae , Cricetulus , Camundongos , Camundongos Nus , Dados de Sequência Molecular , Proteínas Recombinantes/metabolismo , Análise de Sequência de DNA
20.
J Bone Miner Res ; 26(8): 1964-73, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21445983

RESUMO

The spermatogenesis associated 4 gene (Spata4, previously named TSARG2) was demonstrated to participate in spermatogenesis. Here we report that Spata4 is expressed in osteoblasts and that overexpression of Spata4 accelerates osteoblast differentiation and mineralization in MC3T3-E1 cells. We found that Spata4 interacts with p-Erk1/2 in the cytoplasm and that overexpression of Spata4 enhances the phosphorylation of Erk1/2. Intriguingly, we observed that Spata4 increases the transcriptional activity of Runx2, a critical transcription factor regulating osteoblast differentiation. We showed that Spata4-activated Runx2 is through the activation of Erk1/2. Consistent with this observation, we found that overexpression of Spata4 increases the expression of osteoblastic marker genes, including osteocalcin (Ocn), Bmp2, osteopontin (Opn), type 1 collagen, osterix (Osx), and Runx2. We concluded that Spata4 promotes osteoblast differentiation and mineralization through the Erk-activated Runx2 pathway. Our findings provided new evidence that Spata4 plays a role in regulation of osteoblast differentiation.


Assuntos
Diferenciação Celular , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Osteoblastos/citologia , Osteoblastos/enzimologia , Proteínas/metabolismo , Transdução de Sinais , Animais , Biomarcadores/metabolismo , Calcificação Fisiológica , Linhagem Celular , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Citoplasma/metabolismo , Fêmur/citologia , Fêmur/metabolismo , Regulação da Expressão Gênica , Camundongos , Fosforilação , Ligação Proteica , Transporte Proteico , Crânio/citologia , Crânio/metabolismo
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