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1.
PLoS One ; 15(8): e0237108, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32750076

RESUMO

Anatomically terminal parts of the urinary, reproductive, and digestive systems of birds all connect to the cloaca. As the feces drain through the cloaca in chickens, the cloacal bacteria were previously believed to represent those of the digestive system. To investigate similarities between the cloacal microbiota and the microbiota of the digestive and reproductive systems, microbiota inhabiting the colon, cloaca, and magnum, which is a portion of the chicken oviduct of 34-week-old, specific-pathogen-free hens were analyzed using a 16S rRNA metagenomic approach using the Ion torrent sequencer and the Qiime2 bioinformatics platform. Beta diversity via unweighted and weighted unifrac analyses revealed that the cloacal microbiota was significantly different from those in the colon and the magnum. Unweighted unifrac revealed that the cloacal microbiota was distal from the microbiota in the colon than from the microbiota in the magnum, whereas weighted unifrac revealed that the cloacal microbiota was located further away from the microbiota in the magnum than from the microbiota inhabiting the colon. Pseudomonas spp. were the most abundant in the cloaca, whereas Lactobacillus spp. and Flavobacterium spp. were the most abundant species in the colon and the magnum. The present results indicate that the cloaca contains a mixed population of bacteria, derived from the reproductive, urinary, and digestive systems, particularly in egg-laying hens. Therefore, sampling cloaca to study bacterial populations that inhabit the digestive system of chickens requires caution especially when applied to egg-laying hens. To further understand the physiological role of the microbiota in chicken cloaca, exploratory studies of the chicken's cloacal microbiota should be performed using chickens of different ages and types.


Assuntos
Galinhas/microbiologia , Cloaca/microbiologia , Colo/microbiologia , Microbioma Gastrointestinal , Oviductos/microbiologia , Animais , Feminino , Flavobacterium/genética , Flavobacterium/patogenicidade , Lactobacillus/genética , Lactobacillus/patogenicidade , Metagenoma , Pseudomonas/genética , Pseudomonas/patogenicidade
2.
Virology ; 543: 13-19, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32056842

RESUMO

Orthohantaviruses are negative-sense, single-stranded RNA viruses harbored by rodents, shrews, moles, and bats. Of the shrew-borne orthohantaviruses in the Republic of Korea (ROK), Jeju orthohantavirus (Jeju virus, JJUV) was found on Jeju Island. This small-scale epidemiologic survey investigated the geographic distribution and molecular phylogeny of JJUV in the ROK. In 32 trapping sites, tissues of 84 Crocidura shantungensis were analyzed for JJUV RNA. JJUV RNA was detected in seven (8.3%) shrews captured on the Korean peninsula. The molecular epidemiologic survey demonstrated the prevalence of JJUV by geographic distribution. The RNA loads of JJUV were evaluated in various tissues. Entire coding sequences of tripartite genomes were recovered from two JJUV strains on the mainland. Phylogenetic relationships of the JJUV revealed a distinct geographic lineage of mainland strains from the strains on Jeju Island. This study sheds light on the molecular epidemiology, phylogeographic diversity, and virus-host co-divergence of JJUV, ROK.


Assuntos
Infecções por Hantavirus/veterinária , Hantavirus/genética , Musaranhos/virologia , Animais , Variação Genética , Hantavirus/classificação , Infecções por Hantavirus/epidemiologia , Filogenia , Filogeografia , República da Coreia/epidemiologia
3.
J Forensic Sci ; 64(4): 1092-1095, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30586152

RESUMO

Body transportation is defined as moving the body of the victim from the scene of a homicide to another location. We analyzed data on 513 South Korean homicide cases with convictions to compare 104 homicides involving the transportation of victims' bodies with 409 homicides that did not. Offenders who transported their victims' bodies were significantly younger and more likely to be related to the victims and were less likely to use sharp instruments and more likely to use blunt instruments or strangle the victims than offenders who did not. Victims of homicide involving body transportation were more likely to suffer injuries to the neck and head and less likely to suffer injuries to the torso. Finally, offenders who transported victims' bodies were more likely to have planned the offense and destroyed evidence. The present findings have practical implications for police investigations.


Assuntos
Restos Mortais , Homicídio/estatística & dados numéricos , Transportes/estatística & dados numéricos , Adolescente , Adulto , Distribuição por Idade , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Vítimas de Crime/estatística & dados numéricos , Criminosos/estatística & dados numéricos , Feminino , Medicina Legal , Humanos , Lactente , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , República da Coreia , Ferimentos e Lesões/patologia , Adulto Jovem
4.
FASEB J ; 33(3): 4341-4354, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30592630

RESUMO

Fibrosis is characterized by the increased accumulation of extracellular matrix (ECM), which drives abnormal cell proliferation and progressive organ dysfunction in many inflammatory and metabolic diseases. Studies have shown that halofuginone, a racemic halogenated derivative, inhibits glutamyl-prolyl-transfer RNA-synthetase (EPRS)-mediated fibrosis. However, the mechanism by which this occurs is unclear. We explored the mechanistic aspects of how EPRS could develop liver fibrotic phenotypes in cells and animal models. Treatment with TGF-ß1 up-regulated fibronectin and collagen I levels in LX2 hepatic stellate cells. This effect was inhibited in prolyl-transfer RNA synthetase (PRS)-suppressed LX2 cells. Using the promoter luciferase assay, TGF-ß1-mediated collagen I, α1 chain transcription and γ2 basal laminin transcription in LX2 cells were down-regulated by EPRS suppression, suggesting that EPRS may play roles in ECM production at transcriptional levels. Furthermore, signal transducer and activator of transcription (STAT) signaling activation was involved in the effects of TGF-ß1 on ECM expression in a PRS-dependent manner. This was mediated via a protein-protein complex formation consisting of TGF-ß1 receptor, EPRS, Janus kinases, and STAT6. Additionally, ECM expression in fibrotic livers overlapped with EPRS expression along fibrotic septa regions and was positively correlated with STAT6 activation in carbon tetrachloride-treated mice. This was less obvious in livers of Eprs-/+ mice. These findings suggest that, during fibrosis development, EPRS plays roles in nontranslational processes of ECM expression via intracellular signaling regulation upon TGF-ß1 stimulation.-Song, D.-G., Kim, D., Jung, J. W., Nam, S. H., Kim, J. E., Kim, H.-J., Kim, J. H., Lee, S.-J., Pan, C.-H., Kim, S., Lee, J. W. Glutamyl-prolyl-tRNA synthetase induces fibrotic extracellular matrix via both transcriptional and translational mechanisms.


Assuntos
Aminoacil-tRNA Sintetases/metabolismo , Matriz Extracelular/metabolismo , Biossíntese de Proteínas/genética , Transcrição Genética/genética , Aminoacil-tRNA Sintetases/genética , Animais , Linhagem Celular , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Regulação para Baixo/genética , Matriz Extracelular/genética , Fibrose/genética , Fibrose/metabolismo , Células Estreladas do Fígado/metabolismo , Humanos , Fígado/metabolismo , Cirrose Hepática/genética , Cirrose Hepática/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais/genética , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo , Regulação para Cima/genética
5.
Genetica ; 146(2): 211-226, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29441472

RESUMO

Many peninsulas in the temperate zone played an important role as refugia of various flora and fauna, and the southern Korean Peninsula also served as a refugium for many small mammals in East Asia during the Pleistocene. The Asian lesser white-toothed shrew, Crocidura shantungensis, is a widely distributed species in East Asia, and is an appropriate model organism for exploring the role of the Korean Peninsula as a refugium of small mammals. Here, we investigated phylogenetic relationships and genetic diversity based on the entire sequence of the mitochondrial cytochrome b gene (1140 bp). A Bayesian tree for 98 haplotypes detected in 228 C. shantungensis specimens from East Asia revealed the presence of three major groups with at least 5 subgroups. Most haplotypes were distributed according to their geographic proximity. Pairwise FST's and analysis of molecular variance (AMOVA) revealed a high degree of genetic differentiation and variance among regions as well as among populations within region, implying little gene flow among local populations. Genetic evidence from South Korean islands, Jeju-do Island of South Korea, and Taiwan leads us to reject the hypothesis of recent population expansion. We observed unique island-type genetic characteristics consistent with geographic isolation and resultant genetic drift. Phylogeographic inference, together with estimates of genetic differentiation and diversity, suggest that the southern most part the Korean Peninsula, including offshore islands, played an important role as a refugium for C. shantungensis during the Pleistocene. However, the presence of several refugia on the mainland of northeast Asia is also proposed.


Assuntos
Variação Genética , Musaranhos/genética , Animais , Citocromos b/genética , Haplótipos , Coreia (Geográfico) , Filogeografia , Musaranhos/classificação
6.
Cancer Res Treat ; 50(3): 646-657, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28724284

RESUMO

Purpose: Early prediction of treatment outcomes represents an essential step towards increased treatment efficacy and survival in patients with hepatocellular carcinoma (HCC). In this study, we performed two-dimensional electrophoresis (2-DE) followed by protein profiling to identify biomarkers predictive of therapeutic outcomes in patients with HCC who received liver-directed therapy (LDTx) involving local radiotherapy (RT), and studied the underlying mechanisms of the identified proteins. Materials and Methods: 2-DE analysis was conducted by pooling sera from patients with a good or poor prognosis; serum proteomic profiles of the two groups were compared and analyzed using matrixassisted laser desorption/ionization time-of-flight mass spectrometry. Identified proteins were confirmed via enzyme-linked immunosorbent assay. An invasion assay was performed after overexpression and knockdown of target protein in Huh7 cells. Results: Levels of inter-alpha inhibitor H4 (ITIH4), fibrinogen gamma chain, keratin 9/1 complex, carbonic anhydrase I, and carbonmonoxyhemoglobin S were changed by more than 4-fold in response to LDTx. In particular, pre-LDTx ITIH4 expression was more than 5-fold higher in patients with a good prognosis, compared to patients with a poor prognosis. The migration ability of Huh7 cells was significantly suppressed and enhanced by ITIH4 overexpression and knockdown, respectively. The tumors of patients with HCC and a good prognosis expressed high levels of ITIH4, compared to those of patients with a poor prognosis. Conclusion: Taken together, ITIH4 may be a potential therapeutic target that could inhibit cancer metastasis, as well as a prognostic marker for patients with HCC who are receiving LDTx.


Assuntos
Biomarcadores Tumorais/sangue , Carcinoma Hepatocelular/terapia , Fluoruracila/administração & dosagem , Glicoproteínas/sangue , Neoplasias Hepáticas/terapia , Proteínas Secretadas Inibidoras de Proteinases/sangue , Proteômica/métodos , Adulto , Idoso , Proteínas Sanguíneas , Carcinoma Hepatocelular/sangue , Linhagem Celular Tumoral , Movimento Celular , Quimiorradioterapia , Eletroforese em Gel Bidimensional , Feminino , Fluoruracila/uso terapêutico , Regulação Neoplásica da Expressão Gênica , Humanos , Infusões Intra-Arteriais , Neoplasias Hepáticas/sangue , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Análise de Sobrevida , Resultado do Tratamento , Regulação para Cima
7.
Mitochondrial DNA A DNA Mapp Seq Anal ; 29(4): 650-658, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-28657486

RESUMO

Onychodactylus koreanus, a hynobiid salamander species endemic to the Korean Peninsula, can be regarded as a strict ecological specialist, probably vulnerable to anthropogenic environmental modifications and climate change. We used mitochondrial cytochrome b gene to analyze the genetic diversity and phylogenetic structure of O. koreanus from 19 populations collected in an attempt to cover its major distribution within South Korea. A total of 76 haplotypes of O. koreanus obtained in our analyses could be subdivided into three phylogenetic clades, KR, NE and SE. Clade KR haplotypes occur in most of the regions throughout the Korean Peninsula with four distinct subclades (KR I-IV). Clade NE and SE haplotypes were only observed in two populations YY and YS, respectively. Haplotype sharing was scarce even among populations in geographical proximity, and most of the populations were represented by a single clade or subclade, indicating the low level of gene flow among populations. O. koreanus likely originated from the historical southward dispersal of its ancestral lineages following divergence from Chinese O. zhaoermii that was recovered as the sister of O. koreanus in our phylogenetic analysis. Our results have critical implications for the taxonomic status of O. koreanus and its long-term management plan.


Assuntos
DNA Mitocondrial/genética , Especiação Genética , Variação Genética , Genoma Mitocondrial , Urodelos/genética , Animais , China , Citocromos b/genética , Genes Mitocondriais , Mitocôndrias/genética , Filogenia , República da Coreia , Especificidade da Espécie
8.
Oncotarget ; 8(48): 83480-83494, 2017 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-29137358

RESUMO

Transmembrane 4 L six family member 5 (TM4SF5) is highly expressed in hepatocellular carcinoma tissues and enhances migration in two-dimensional environments. Here, we investigated how TM4SF5 is involved in diverse pro-metastatic phenotypes in in vivo-like three-dimensional (3D) extracellular matrix gels. TM4SF5-positive cells aggressively formed invasive foci in 3D Matrigel, depending on TM4SF5-mediated signaling activity, cytoskeletal organization, and matrix metallopeptidase (MMP) 2-mediated extracellular remodeling, whereas TM4SF5-null cells did not. The TM4SF5-null cells did, however, form invasive foci in 3D Matrigel following inhibition of Rho-associated protein kinase or addition of collagen I, suggesting that collagen I compensated for TM4SF5 expression. Similarly, TM4SF5-positive cells expressing vascular endothelial-cadherin formed network-like vasculogenic mimicry in 3D Matrigel and collagen I mixture gels, whereas TM4SF5-negative cells in the mixture gels displayed the network structures only upon further treatment with epidermal growth factor. The foci formation also required MMP2-mediated remodeling of the extracellular matrix. Co-cultures exhibited TM4SF5-positive or cancer-associated fibroblasts at the outward edges of TM4SF5-null cell clusters. Compared with TM4SF5-null cells, TM4SF5-positive cells in 3D collagen gels showed a more invasive outgrowth with dramatic invadopodia. These observations suggest that TM4SF5 plays roles in the promotion of diverse metastatic properties with fewer environmental requirements than TM4SF5-negative cells.

9.
Oncotarget ; 8(25): 41242-41255, 2017 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-28465485

RESUMO

BACKGROUND & AIMS: Although immunotherapy has emerged as an attractive therapy for refractory cancers, its limited efficacy in hepatocellular carcinoma (HCC) suggests the need for a combination strategy that can either enhance or complement therapeutic effect. We investigated whether combination of immune checkpoint blockade (ICB) and radiation could enhance antitumor effect in a murine HCC model. METHODS: Using murine HCC, HCa-1, the effect of radiation on programmed death-ligand1 (PD-L1) expression was determined by real-time PCR, flow cytometry, and western blotting. Signaling pathways involved in altered PD-L1 expression were examined. Tumor growth and survival rate were evaluated for a combination of anti-PD-L1 and radiation. Immunological parameters in the tumor were assessed using flow cytometry and histological study. RESULTS: Radiation upregulated PD-L1 expression in tumor cells through IFN-γ/STAT3 signaling, which could facilitate therapeutic action of anti-PD-L1. Combination of anti-PD-L1 and radiation significantly suppressed tumor growth compared to treatment with anti-PD-L1 alone or radiation alone group (P<0.01). Survival was significantly improved in the combination group compared to anti-PD-L1 alone or radiation alone group (7-week survival rate; 90% vs. 0% or 30%, respectively, P<0.001). The underlying mechanism involved increasing apoptosis, decreasing tumor cell proliferation, as well as restoration of CD8+ T cell functions. CONCLUSIONS: The combination of anti-PD-L1 and radiation significantly improved the antitumor effect shown in tumor growth delay as well as in survival, supporting a novel combination strategy of immunoradiotherapy in HCC.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Carcinoma Hepatocelular/terapia , Neoplasias Hepáticas Experimentais/terapia , Animais , Anticorpos Monoclonais/imunologia , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/imunologia , Antígeno B7-H1/metabolismo , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Terapia Combinada , Células Hep G2 , Humanos , Imunoterapia/métodos , Neoplasias Hepáticas Experimentais/imunologia , Neoplasias Hepáticas Experimentais/metabolismo , Masculino , Camundongos Endogâmicos C3H , Radioterapia/métodos , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/efeitos da radiação , Análise de Sobrevida , Carga Tumoral/efeitos dos fármacos , Carga Tumoral/imunologia , Carga Tumoral/efeitos da radiação
10.
PLoS One ; 11(1): e0146463, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26745884

RESUMO

Radiotherapy (RT) is a potent anti-tumor modality. However, unwanted effects including increased recurrence and metastasis that involve factors such as cytokines, which induce complex molecular mechanisms, have also been reported. In a previous study, we showed that interleukin (IL)-12 and radiotherapy combination treatment suppressed tumor growth and metastasis in a hepatoma mouse model. In this study, we investigated the mechanism underlying the IL-12 anti-tumor effect during radiotherapy. In tumor-bearing mice, irradiation decreased IL-12 expression in the tumors and spleens. However, a number of dendritic cells infiltrated into the tumors in which IL-12 expression did not decrease. To further study the underlying detailed mechanism for this decrease in IL-12, LPS-stimulated bone marrow-derived dendritic cells (BMDCs) were irradiated, and then IL-12- and IL-6-associated molecules were examined in irradiated tumors and BMDCs. Irradiation resulted in IL-12 suppression and IL-6 increase. IL-6 and signal transducer and activator of transcription 3 (STAT3) inhibitors restored the irradiation-induced IL-12 decrease via suppression of C-Rel activation. Taken together, our study suggests that irradiation-induced IL-6 can decrease IL-12 production through the inhibition of C-Rel phosphorylation by the IL-6/STAT3 signaling pathway.


Assuntos
Células Dendríticas/metabolismo , Interleucina-12/biossíntese , Interleucina-6/fisiologia , Proteínas Proto-Oncogênicas c-rel/fisiologia , Fator de Transcrição STAT3/metabolismo , Animais , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/radioterapia , Linhagem Celular Tumoral , Células Dendríticas/imunologia , Células Dendríticas/efeitos da radiação , Lipopolissacarídeos/farmacologia , Neoplasias Hepáticas Experimentais/imunologia , Neoplasias Hepáticas Experimentais/metabolismo , Neoplasias Hepáticas Experimentais/radioterapia , Masculino , Camundongos Endogâmicos C3H , Transplante de Neoplasias , Transdução de Sinais
11.
Oncotarget ; 6(40): 42905-22, 2015 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-26556867

RESUMO

Tissue inhibitors of metalloproteinases (TIMPs) control extracellular matrix (ECM) homeostasis by inhibiting the activity of matrix metalloproteinases (MMPs), which are associated with ECM turnover. Recent studies have revealed that TIMPs are implicated in tumorigenesis in both MMP-dependent and MMP-independent manners. We examined a mechanism by which TIMP-2 stimulated lung adenocarcinoma cell proliferation, independent of MMP inhibition. The stimulation of growth by TIMP-2 in A549 cells required c-Src kinase activation. c-Src kinase activity, induced by TIMP-2, concomitantly increased FAK, phosphoinositide 3-kinase (PI3-kinase)/AKT, and ERK1/2 activation. Selective knockdown of integrin α3ß1, known as a TIMP-2 receptor, did not significantly change TIMP-2 growth promoting activity. Furthermore, we showed that high TIMP-2 expression in lung adenocarcinomas is associated with a worse prognosis from multiple cohorts, especially for stage I lung adenocarcinoma. Through integrated analysis of The Cancer Genome Atlas data, TIMP-2 expression was significantly associated with the alteration of driving genes, c-Src activation, and PI3-kinase/AKT pathway activation. Taken together, our results demonstrate that TIMP-2 stimulates lung adenocarcinoma cell proliferation through c-Src, FAK, PI3-kinase/AKT, and ERK1/2 pathway activation in an MMP-independent manner.


Assuntos
Adenocarcinoma/metabolismo , Neoplasias Pulmonares/metabolismo , Transdução de Sinais/fisiologia , Inibidor Tecidual de Metaloproteinase-2/metabolismo , Adenocarcinoma/mortalidade , Adenocarcinoma/patologia , Western Blotting , Proteína Tirosina Quinase CSK , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Ativação Enzimática/fisiologia , Quinase 1 de Adesão Focal/metabolismo , Regulação Neoplásica da Expressão Gênica/fisiologia , Técnicas de Silenciamento de Genes , Humanos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Sistema de Sinalização das MAP Quinases/fisiologia , Metaloproteinases da Matriz/metabolismo , Mutagênese Sítio-Dirigida , Fosfatidilinositol 3-Quinases/metabolismo , Prognóstico , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transcriptoma , Transfecção , Quinases da Família src/metabolismo
12.
Nanoscale ; 6(20): 11898-903, 2014 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-25175492

RESUMO

Hollow SnO2 spheres functionalized by Au catalysts were synthesized via the use of Au-decorated block copolymer (Au-BCP) sphere templates. Uniformly distributed Au nanoparticles on BCP spheres were prepared by the infiltration of Au precursors into polystyrene-b-poly(4-vinylpyridine) (PS-b-P4VP) spheres. A thin SnO2 layer was coated on the Au-BCP spheres using RF sputtering at room temperature without morphological deformation of the spheres. The Au nanoparticles were uniformly transferred from the Au-BCP spheres to the inner shells of the hollow SnO2 spheres followed by decomposition of BCP spheres. The Au-loaded hollow SnO2 spheres exhibited a superior H2S sensitivity (Rair/Rgas = 17.4 at 5 ppm) with remarkably selective characteristics with a minor response (Rair/Rgas < 2.5 at 5 ppm) toward other interfering gases. Our results pave the way for a new catalyst loading method using Au-BCP spheres for the uniformly distributed Au NPs on the SnO2 layers.


Assuntos
Testes Respiratórios/métodos , Ouro/química , Halitose/diagnóstico , Polímeros/química , Compostos de Estanho/química , Catálise , Gases , Humanos , Sulfeto de Hidrogênio/química , Nanopartículas Metálicas/química , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Nanopartículas , Nanotecnologia , Oxigênio/química , Poliestirenos/química , Polivinil/química , Piridinas/química , Propriedades de Superfície , Temperatura
13.
J Korean Med Sci ; 29(8): 1138-44, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25120326

RESUMO

Charcot-Marie-Tooth disease (CMT) is the most common inherited motor and sensory neuropathy. Previous studies have found that, according to CMT patients, neuropathic pain is an occasional symptom of CMT. However, neuropathic pain is not considered to be a significant symptom associated with CMT and, as a result, no studies have investigated the pathophysiology underlying neuropathic pain in this disorder. Thus, the first animal model of neuropathic pain was developed by our laboratory using an adenovirus vector system to study neuropathic pain in CMT. To this end, glycyl-tRNA synthetase (GARS) fusion proteins with a FLAG-tag (wild type [WT], L129P and G240R mutants) were expressed in spinal cord and dorsal root ganglion (DRG) neurons using adenovirus vectors. It is known that GARS mutants induce GARS axonopathies, including CMT type 2D (CMT2D) and distal spinal muscular atrophy type V (dSMA-V). Additionally, the morphological phenotypes of neuropathic pain in this animal model of GARS-induced pain were assessed using several possible markers of pain (Iba1, pERK1/2) or a marker of injured neurons (ATF3). These results suggest that this animal model of CMT using an adenovirus may provide information regarding CMT as well as a useful strategy for the treatment of neuropathic pain.


Assuntos
Doença de Charcot-Marie-Tooth/diagnóstico , Doença de Charcot-Marie-Tooth/fisiopatologia , Modelos Animais de Doenças , Glicina-tRNA Ligase/genética , Neuralgia/diagnóstico , Neuralgia/fisiopatologia , Animais , Glicina-tRNA Ligase/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutagênese Sítio-Dirigida , Mutação/genética
14.
ACS Appl Mater Interfaces ; 6(12): 9061-70, 2014 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-24844154

RESUMO

Diagnostic sensing device using exhaled breath of human have critical advantages due to the noninvasive diagnosis and high potential for portable device with simple analysis process. Here, we report ultrafast as well as highly sensitive bumpy WO3 hemitube nanostructure assisted by O2 plasma surface modification with functionalization of graphene-based material for the detection of acetone (CH3COCH3) and hydrogen sulfide (H2S) which are biomarkers for the diagnosis of diabetes and halitosis, respectively. 0.1 wt % graphene oxide (GO)- and 0.1 wt % thin layered graphite (GR)- WO3 hemitube composites showed response times of 11.5 ± 2.5 s and 13.5 ± 3.4 s to 1 ppm acetone as well as 12.5 ± 1.9 s and 10.0 ± 1.6 s to 1 ppm of H2S, respectively. In addition, low limits of detection (LOD) of 100 ppb (Rair/Rgas = 1.7 for acetone and Rair/Rgas = 3.3 for H2S at 300 °C) were achieved. The superior sensing properties were ascribed to the electronic sensitization of graphene based materials by modulating space charged layers at the interfaces between n-type WO3 hemitubes and p-type graphene based materials, as identified by Kelvin Probe Force Microscopy (KPFM). Rapid response and superior sensitivity of the proposed sensing materials following cyclic thermal aging demonstrates good potential for real-time exhaled breath diagnosis of diseases.


Assuntos
Acetona , Testes Respiratórios , Diabetes Mellitus/diagnóstico , Halitose/diagnóstico , Sulfeto de Hidrogênio , Acetona/isolamento & purificação , Técnicas Biossensoriais , Grafite/química , Humanos , Sulfeto de Hidrogênio/isolamento & purificação , Limite de Detecção , Nanoestruturas/química , Óxidos/química , Tungstênio/química
15.
ACS Appl Mater Interfaces ; 6(4): 2588-97, 2014 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-24456186

RESUMO

Sensitive detection of acetone and hydrogen sulfide levels in exhaled human breath, serving as breath markers for some diseases such as diabetes and halitosis, may offer useful information for early diagnosis of these diseases. Exhaled breath analyzers using semiconductor metal oxide (SMO) gas sensors have attracted much attention because they offer low cost fabrication, miniaturization, and integration into portable devices for noninvasive medical diagnosis. However, SMO gas sensors often display cross sensitivity to interfering species. Therefore, selective real-time detection of specific disease markers is a major challenge that must be overcome to ensure reliable breath analysis. In this work, we report on highly sensitive and selective acetone and hydrogen sulfide detection achieved by sensitizing electrospun SnO2 nanofibers with reduced graphene oxide (RGO) nanosheets. SnO2 nanofibers mixed with a small amount (0.01 wt %) of RGO nanosheets exhibited sensitive response to hydrogen sulfide (Rair/Rgas = 34 at 5 ppm) at 200 °C, whereas sensitive acetone detection (Rair/Rgas = 10 at 5 ppm) was achieved by increasing the RGO loading to 5 wt % and raising the operation temperature to 350 °C. The detection limit of these sensors is predicted to be as low as 1 ppm for hydrogen sulfide and 100 ppb for acetone, respectively. These concentrations are much lower than in the exhaled breath of healthy people. This demonstrates that optimization of the RGO loading and the operation temperature of RGO-SnO2 nanocomposite gas sensors enables highly sensitive and selective detection of breath markers for the diagnosis of diabetes and halitosis.


Assuntos
Acetona/análise , Diabetes Mellitus/diagnóstico , Grafite/química , Halitose/diagnóstico , Sulfeto de Hidrogênio/análise , Nanoestruturas , Compostos de Estanho/química , Humanos , Microscopia Eletrônica de Varredura , Óxidos/química , Sensibilidade e Especificidade , Análise Espectral Raman , Difração de Raios X
16.
J Mol Histol ; 45(2): 121-8, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23990368

RESUMO

Charcot-Marie-Tooth disease type 2D is a hereditary axonal and glycyl-tRNA synthetase (GARS)-associated neuropathy that is caused by a mutation in GARS. Here, we report a novel GARS-associated mouse neuropathy model using an adenoviral vector system that contains a neuronal-specific promoter. In this model, we found that wild-type GARS is distributed to peripheral axons, dorsal root ganglion (DRG) cell bodies, central axon terminals, and motor neuron cell bodies. In contrast, GARS containing a G240R mutation was localized in DRG and motor neuron cell bodies, but not axonal regions, in vivo. Thus, our data suggest that the disease-causing G240R mutation may result in a distribution defect of GARS in peripheral nerves in vivo. Furthermore, a distributional defect may be associated with axonal degradation in GARS-associated neuropathies.


Assuntos
Adenoviridae/genética , Doença de Charcot-Marie-Tooth/enzimologia , Animais , Axônios/enzimologia , Doença de Charcot-Marie-Tooth/genética , Doença de Charcot-Marie-Tooth/patologia , Modelos Animais de Doenças , Gânglios Espinais/enzimologia , Gânglios Espinais/patologia , Vetores Genéticos , Glicina-tRNA Ligase/genética , Glicina-tRNA Ligase/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neurônios Motores/enzimologia , Mutação de Sentido Incorreto , Fibras Nervosas Mielinizadas/enzimologia , Especificidade de Órgãos , Nervos Periféricos/enzimologia , Nervos Periféricos/patologia
17.
J Mol Histol ; 44(2): 241-7, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23264110

RESUMO

In in vitro and in vivo systems, understanding localization and the functional role of ATP is essential, but effective methods to monitor ATP in cells and tissues are limited. Although quinacrine dihydrochloride is a well-known fluorescent dye used to detect ATP, it is limited in its use because it shows non-specific nuclear staining both in vitro and in vivo. A commercial luciferin-luciferase bioluminescence assay has also been used to detect ATP, but it can not be easily used in vivo. Thus, to effectively monitor ATP in vivo, we employed a novel two-photon ATP fluorescent probe, acedan-based Zn(DPA). Using the acedan-based Zn(DPA) probe, we show that this probe produces high quality images of ATP in lung, spleen, liver and spinal cord tissues.


Assuntos
Trifosfato de Adenosina/metabolismo , Corantes Fluorescentes/metabolismo , Medições Luminescentes , Microscopia de Fluorescência , Animais , Corantes Fluorescentes/química , Hepatócitos/metabolismo , Medições Luminescentes/métodos , Camundongos , Microscopia de Fluorescência/métodos , Quinacrina/química , Medula Espinal/metabolismo , Baço/metabolismo , Zinco/química
18.
Biochem Biophys Res Commun ; 430(2): 488-93, 2013 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-23237805

RESUMO

The adenosine triphosphate (ATP) plays important roles under physiological and pathological conditions such as traumatic brain injury, neuroinflammation and neuropathic pain. In the present study, we set out to study the role of lysosomal vesicles on ATP release from the dorsal root ganglion neurons. We found that the lysosomal vesicles, which contain the quinacrine-positive fluorescence and express the vesicular nucleotide transporter (VNUT), were localized within the soma and growth cone of the cultured dorsal root ganglion neurons. In addition, the number of the quinacrine staining was decreased by application of lysosomal exocytosis activators, and this decrease was suppressed by the metformin and vacuolin-1, which suppressed lysosomal exocytosis. Thus, these findings suggest that ATP release via the lysosomal exocytosis may be one of the pathways for ATP release in response to stimulation.


Assuntos
Trifosfato de Adenosina/metabolismo , Exocitose , Gânglios Espinais/metabolismo , Lisossomos/metabolismo , Neurônios Aferentes/metabolismo , Animais , Transporte Biológico , Células Cultivadas , Gânglios Espinais/citologia , Masculino , Neurônios Aferentes/ultraestrutura , Proteínas de Transporte de Nucleotídeos/metabolismo , Ratos , Ratos Sprague-Dawley
19.
Biochem Biophys Res Commun ; 430(2): 852-7, 2013 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-23194661

RESUMO

After nerve injury, Schwann cells proliferate and revert to a phenotype that supports nerve regeneration. This phenotype-changing process can be viewed as Schwann cell dedifferentiation. Here, we investigated the role of extracellular ATP in Schwann cell dedifferentiation and proliferation during Wallerian degeneration. Using several markers of Schwann cell dedifferentiation and proliferation in sciatic explants, we found that extracellular ATP inhibits Schwann cell dedifferentiation and proliferation during Wallerian degeneration. Furthermore, the blockage of lysosomal exocytosis in ATP-treated sciatic explants is sufficient to induce Schwann cell dedifferentiation. Together, these findings suggest that ATP-induced lysosomal exocytosis may be involved in Schwann cell dedifferentiation.


Assuntos
Trifosfato de Adenosina/fisiologia , Desdiferenciação Celular , Exocitose , Modelos Biológicos , Células de Schwann/patologia , Degeneração Walleriana/patologia , Trifosfato de Adenosina/metabolismo , Animais , Proliferação de Células , Glicoproteínas de Membrana Associadas ao Lisossomo/metabolismo , Lisossomos/metabolismo , Masculino , Metformina/farmacologia , Proteínas do Tecido Nervoso , Ratos , Ratos Sprague-Dawley , Receptores de Fatores de Crescimento , Receptores de Fator de Crescimento Neural/genética , Receptores de Fator de Crescimento Neural/metabolismo , Células de Schwann/metabolismo , Neuropatia Ciática/metabolismo , Neuropatia Ciática/patologia , Degeneração Walleriana/metabolismo
20.
Biochem Biophys Res Commun ; 429(3-4): 163-7, 2012 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-23142593

RESUMO

The present study demonstrates that adenosine triphosphate (ATP) is released from Schwann cells through lysosomal exocytosis during Wallerian degeneration and in response to stimulation. In primary Schwann cell cultures, ATP was stored in lysosomal vesicles. ATP could then induce Ca(2+)-dependent lysosomal exocytosis. Among three stimulants of lysosomal exocytosis (glutamate, NH(4)Cl and zymosan), only NH(4)Cl was sufficient to induce ATP release from ex vivo sciatic nerve explants at 3 days in vitro. Lysosomal exocytosis inhibitors (metformin, chlorpromazine and vacuolin-1) reversed the effect of NH(4)Cl-enhanced ATP release, replicating the state of explants treated with NH(4)Cl in the absence of lysosomal exocytosis inhibitors. Furthermore, we observed ATP release through lysosomal exocytosis during Wallerian degeneration in sciatic explant cultures using the recently identified vesicular nucleotide transporter (VNUT). From these experiments, we conclude that the exocytosis of lysosomes in Schwann cells during Wallerian degeneration is Ca(2+)-dependent, and that it induces ATP release from Schwann cells.


Assuntos
Trifosfato de Adenosina/metabolismo , Exocitose/fisiologia , Lisossomos/metabolismo , Células de Schwann/metabolismo , Degeneração Walleriana/metabolismo , Cloreto de Amônio/farmacologia , Animais , Células Cultivadas , Exocitose/efeitos dos fármacos , Masculino , Ratos , Ratos Sprague-Dawley
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