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2.
Am J Vet Res ; : 1-9, 2024 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-38942061

RESUMO

OBJECTIVE: Referencing growing concerns over the recruitment and retention of faculty in academic veterinary medicine, the authors hypothesized that among surveyed veterinary residents and early-career faculty, work-life balance and workplace climate and culture are stronger motivators than financial considerations, regardless of demographic factors such as gender, race/ethnicity, and area of specialization. SAMPLE: 541 participants were included in data analysis. METHODS: A mixed methods approach was utilized, incorporating both quantitative data and qualitative, free-text responses to better understand veterinary career choices by contextualizing factors associated with academic medicine. RESULTS: Factors underpinning career-related decision-making were ranked by level of importance as (1) workplace environment/culture, (2) personal well-being/work-life balance, (3) salary and bonuses, (4) geographic location, (5) facilities and resources, (6) benefits, and (7) schedule flexibility. Desires for workload balance, schedule flexibility, support from leadership, and mentorship and collaboration were among the top themes of qualitative responses for both residents and early career faculty respondents. Factors influencing career decision-making for resident and early-career faculty are varied. Workplace environment, work-life balance, and schedule flexibility are areas that academic institutions can address and continue to improve and that are likely to positively impact entry into academia and the desire to stay. CLINICAL RELEVANCE: This study sought to understand factors related to career decision-making and interest in academic veterinary medicine among residents and early-career faculty. Understanding these factors can support efforts to recruit and retain faculty in academic veterinary medicine.

3.
Rev Sci Instrum ; 91(9): 095104, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-33003818

RESUMO

We report on the frozen-spin polarized hydrogen-deuteride (HD) targets for photoproduction experiments at SPring-8/LEPS. Pure HD gas with a small amount of ortho-H2 (∼0.1%) and a very small amount of para-D2 (∼0.001%) was liquefied and solidified by liquid helium. The temperature of the produced solid HD was reduced to about 30 mK with a dilution refrigerator. A magnetic field (17 T) was applied to the HD to grow the polarization with the static method. After the aging of the HD at low temperatures in the presence of a high-magnetic field strength for three months, the polarization froze. Almost all ortho-H2 molecules were converted to para-H2 molecules. Most remaining para-D2 molecules were converted to ortho-D2 molecules. The para-H2 and ortho-D2 molecules exhibited weak spin interactions with the HD. If the concentrations of the ortho-H2 and para-D2 were reduced appropriately at the beginning of the aging process, the aging time can be shortened. We have developed a new nuclear magnetic resonance (NMR) system to measure the relaxation times (T1) of the 1H and 2H nuclei with two frequency sweeps at the respective frequencies of 726 MHz and 111 MHz and succeeded in the monitoring of the polarization build-up at decreasing temperatures from 600 mK to 30 mK at 17 T. Automatic NMR measurements with the frequency sweeps enabled us to omit the use of a manual tuning circuit and to remove magnetic field sweeps with eddy current heat. This technique enables us to optimize the concentration of the ortho-H2 and to efficiently polarize the HD target within a shortened aging time.

4.
J Interferon Cytokine Res ; 21(10): 835-41, 2001 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-11710996

RESUMO

Here we report on the antiviral effects of two commercially available natural interferon-alpha (IFN-alpha) preparations, their subtype compositions, and the effects of combinations of pairs of the subtypes on virally infected cells. Our results show that the antiviral effects of these preparations depend on the target cell and on the infecting virus. The component subtypes vary with the preparations, and combinations of pairs of IFN-alpha subtypes may have synergistic or competitive effects. Our results suggest that optimal preparations of synergistically acting subtypes may provide more therapeutic benefit to patients.


Assuntos
Antivirais/química , Antivirais/farmacologia , Interferon-alfa/química , Interferon-alfa/farmacologia , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Sinergismo Farmacológico , Células HeLa , Humanos , Concentração Inibidora 50 , Interferon Tipo I/farmacologia , Testes de Sensibilidade Microbiana , Isoformas de Proteínas/farmacologia , Proteínas Recombinantes , Vírus da Estomatite Vesicular Indiana/efeitos dos fármacos
5.
J Interferon Cytokine Res ; 21(12): 1129-36, 2001 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-11798471

RESUMO

Interferon-alpha (IFN-alpha) has a number of therapeutic applications in the treatment of various human cancers and diseases of viral origin. IFN-alpha includes several subtypes, and little has been reported on the biologic properties of the individual subtypes. Here, we report on the individual antitumor effects of five IFN-alpha subtypes, alpha1, alpha2, alpha5, alpha8, and alpha10, against six renal cell carcinoma (RCC) cell lines in vitro. Among the subtypes, IFN-alpha8 most potently inhibited cell proliferation and delayed the G(1)/S transition. Synergistic induction of apoptosis was shown in two of the RCC cell lines when treated with the combination of IFN-alpha and IFN-gamma rather than with either IFN-alpha or IFN-gamma alone. IFN-alpha8 was most effective in the induction of apoptosis when combined with IFN-gamma. In addition, IFN-alpha8 had the strongest ability to upregulate HLA class II antigen expression in the subtypes examined. These data indicate that subtypes of IFN-alpha have disparate antitumor effects in vitro, and in vitro distinctions among the IFN-alpha subtypes should be appreciated more in clinical application.


Assuntos
Antineoplásicos/farmacologia , Carcinoma de Células Renais/tratamento farmacológico , Interferon-alfa/farmacologia , Neoplasias Renais/tratamento farmacológico , Apoptose , Carcinoma de Células Renais/imunologia , Carcinoma de Células Renais/patologia , Ciclo Celular , Divisão Celular/efeitos dos fármacos , DNA de Neoplasias/análise , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Citometria de Fluxo , Fluoruracila/farmacologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Interferon gama/farmacologia , Neoplasias Renais/imunologia , Neoplasias Renais/patologia , Isoformas de Proteínas/farmacologia , Células Tumorais Cultivadas
6.
Neurosurgery ; 16(1): 79-82, 1985 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-3974817

RESUMO

A case of flaccid paraplegia due to a metastatic spinal epidural melanoma is reported. Symptoms occurred approximately 9 years after the enucleation of the left eye. A solid melanotic mass was removed almost totally. Three years after the operation, the patient had not regained the motility of the lower limbs. The need for long term follow-up and management of patients with such problems is discussed, and the pertinent literature is reviewed.


Assuntos
Neoplasias da Coroide/patologia , Melanoma/secundário , Neoplasias da Medula Espinal/secundário , Corioide/patologia , Espaço Epidural , Humanos , Masculino , Melanoma/complicações , Melanoma/patologia , Pessoa de Meia-Idade , Paraplegia/etiologia , Medula Espinal/patologia , Neoplasias da Medula Espinal/complicações , Neoplasias da Medula Espinal/patologia , Fatores de Tempo
7.
Neurosurgery ; 16(2): 252-6, 1985 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-3974840

RESUMO

An unusual case of intradiploic epidermoid tumor is reported. In this case, a soft mass located over the parietal bone was noted shortly after birth. The mass gradually enlarged. The tumor underwent malignant change after several excisions and repeated inflammations. The patient was successfully treated with extensive excision, radiotherapy, and chemotherapy. Compiling a summary of reported cases with a tabulation of pertinent information, we discuss the incidence, pathogenesis, prognosis, and therapy of malignant intradiploic epidermoid.


Assuntos
Carcinoma de Células Escamosas/patologia , Cisto Epidérmico/patologia , Neoplasias Cranianas/patologia , Cisto Epidérmico/cirurgia , Humanos , Masculino , Pessoa de Meia-Idade , Reoperação , Neoplasias Cranianas/cirurgia
8.
Oncol Rep ; 6(6): 1299-302, 1999.
Artigo em Inglês | MEDLINE | ID: mdl-10523701

RESUMO

Eight locally advanced breast cancer patients were treated with neoadjuvant intraarterial high-dose chemotherapy (epirubicin) plus MPA combined with autologous PBSCT. All patients completed the scheduled treatment, and there were no toxic deaths. Patients were treated with an escalating dose of epirubicin (370-480 mg) and cyclophosphamide (0-6000 mg). The rate of clinical response was 100%. The rate of good histologic response was 87.5% in the main tumor and 75% in diseased lymph nodes. The 2-year survival rate was 100%. Six patients were disease-free at the time of writing. This treatment resulted in higher rates of clinical and histologic response when compared with standard-dose intraarterial chemotherapy.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Neoplasias da Mama/terapia , Epirubicina/administração & dosagem , Transplante de Células-Tronco Hematopoéticas , Acetato de Medroxiprogesterona/administração & dosagem , Adulto , Neoplasias da Mama/mortalidade , Neoplasias da Mama/fisiopatologia , Terapia Combinada , Feminino , Humanos , Pessoa de Meia-Idade , Análise de Sobrevida , Transplante Autólogo
9.
J Exp Clin Cancer Res ; 20(3): 419-28, 2001 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-11718224

RESUMO

Vascular endothelial growth factor (VEGF)-C, one of several members of the VEGF family, is a relatively specific lymphangiogenic growth factor. VEGF receptor (VEGFR)-3 (or Flt4) is a VEGF-C receptor with expression restricted to lymphatic endothelial cells. Since the mechanisms by which carcinoma cells metastasize to lymph nodes remain unclear, we constructed a VEGF-C transfectant (AZ-VEGF-C) from the AZ521 human gastric carcinoma cell line, which ordinarily shows little nodal metastatic potential and little VEGF-C expression. We orthotopically implanted transfected tumor cells into the stomachs of nude mice. The number of mice developing lymph node metastases and the number of lymph node metastases per mouse with nodal metastases were higher than with implants of mock-transfected control cells. Specifically, percentages of mice with lymph node metastases were 95.5% (21/22) for AZ-VEGF-C and 29.4% (5/17) for controls (P<0.01), while mean numbers of involved lymph nodes were 3.76 for AZ-VEGF-C and 1.00 for controls (P<0.01). No difference was found between AZ-VEGF-C and controls regarding cell growth and chemotactic responses in vitro, or in volumes of tumors arising from implanted cells. When we performed immunohistochemical staining for VEGFR-3 in these tumors to investigate lymphangiogenesis by VEGF-C, the number of vessels stained for VEGFR-3 in tumors and surrounding tissues was higher for AZ-VEGF-C than for controls. VEGFR-3-positive vessels occupied 14.9/1000 of microscopically examined areas for AZ-VEGF-C, but only 1.30/1000 for controls (P<0.001). Our results suggest that VEGF-C is a specific lymphangiogenic growth factor with an important role in lymph node metastasis.


Assuntos
Fatores de Crescimento Endotelial/toxicidade , Metástase Linfática , Neoplasias Gástricas/patologia , Animais , Fatores de Crescimento Endotelial/genética , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas Recombinantes/toxicidade , Transfecção , Transplante Heterólogo , Células Tumorais Cultivadas , Fator C de Crescimento do Endotélio Vascular
10.
J Exp Clin Cancer Res ; 23(3): 513-20, 2004 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-15595644

RESUMO

We established the novel sublines HPC-1H5, HPC-3H4, HPC-4H4, and Panc-1H5, which have a high potential of liver metastasis, and HPC-1P5a, HPC-3P4a, HPC-4P4a, and Panc-1P5a, which have a high potential of peritoneal dissemination, derived from low metastatic HPC-1, HPC-3, HPC-4, and Panc-1cell lines, respectively. To clarify the molecular mechanisms of cancer metastasis and of the different levels of gene expression in a variety of metastatic potentials in pancreatic cancer, we performed a broad analysis of differential gene expression analysis between parental cell lines and metastatic sublines. In comparison with the parental cell lines, 65 and 36 genes were overexpressed and underexpressed in highly liver-metastatic sublines. On the other hand, 43 and 45 genes were overexpressed and underexpressed in highly peritoneal-metastatic sublines. uPAR and Serin protease were overexpressed, and E2A and IGF1R were underexpressed in both metastatic sublines. Hierarchical clustering analysis revealed 22 genes classifying liver, peritoneal metastatic sublines and low-metastatic parental cell lines. These genes might be targeted genes separating those two major metastatic forms after surgery. A greater number of cell line samples and more genes will have to be utilized in future studies in order to understand the involvement of genes in cancer metastasis more thoroughly. However, these results will help to clarify the molecular mechanisms of pancreatic cancer metastasis.


Assuntos
Regulação Neoplásica da Expressão Gênica , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Quimiotaxia , Análise por Conglomerados , DNA Complementar/metabolismo , Humanos , Metástase Neoplásica , Neoplasias Peritoneais/metabolismo , Neoplasias Peritoneais/patologia , Neoplasias Peritoneais/secundário , Serina Endopeptidases/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo
11.
J Exp Clin Cancer Res ; 22(2): 321-8, 2003 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-12866584

RESUMO

To investigate the contribution of beta-catenin to the development of gallbladder carcinoma, genetic alteration in beta-catenin gene, ctnnb-1 and subcellular localization of beta-catenin protein were searched. Mutational analysis of exon 3 in ctnnb-1, which encodes the serine/threonine residues for GSK3beta phosphorylation sites, was performed for 21 gallbladder carcinomas affected with/without the pancreaticobiliary malunion, PBM, and 6 non-cancerous tissues affected with PBM. We also analyzed subcellular localization of beta-catenin protein in all cases immunohistochemically. Nucleotide sequencing analysis revealed that none of them carried mutations that altered amino acid residues in the potential GSK3beta phophorylation sites, but one nucleotide substitution was found. We also analyzed subcellular localization of beta-catenin protein in all cases immunohistochemically, and confirmed its accumulation in both the nucleus and cytoplasm in 10 out of 21 cancer tissues, while the non-cancerous tissues which were affected with PBM and histologically diagnosed as hyperplasia or dysplasia displayed intense membranous staining. A significant correlation between cytoplasmic or nuclear beta-catenin immunoreactivity and clinicopathological status of gallbladder carcinomas was found, especially in the poorer histological differentiation grade(p < 0.05). In conclusion our results suggested that beta-catenin alteration might be a minor contributor to the development of gallbladder carcinomas through abnormal Wnt-wingless signalling, however, decreased membranous expression of beta-catenin might be correlated to carcinoma progression through loss of cell adhesive function in E-cadherin-catenin fashion.


Assuntos
Proteínas do Citoesqueleto/genética , Neoplasias da Vesícula Biliar/genética , Mutação , Transativadores/genética , Adulto , Idoso , Sítios de Ligação , Diferenciação Celular , Proteínas do Citoesqueleto/metabolismo , DNA/metabolismo , Análise Mutacional de DNA , Feminino , Neoplasias da Vesícula Biliar/metabolismo , Neoplasias da Vesícula Biliar/patologia , Quinase 3 da Glicogênio Sintase/genética , Glicogênio Sintase Quinase 3 beta , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Fosforilação , Polimorfismo Conformacional de Fita Simples , Serina/química , Transdução de Sinais , Treonina/química , Transativadores/metabolismo , beta Catenina
12.
J Exp Clin Cancer Res ; 21(1): 23-7, 2002 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-12071524

RESUMO

To investigate the contribution of beta-catenin to the development of carcinoma of the ampulla of Vater, genetic alterations of beta-catenin gene, CTNNB-1 were searched. Mutational analysis of exon3 in CTNNB-1, which encodes the serine/threonine residues for GSK-3beta phosphorylation sites, was performed on 21 cases of carcinoma of the ampulla of Vater, by means of polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) followed by nucleotide sequencing. We found one deleted mutation at codon 32 to approximately 65 in one case of carcinoma of the ampulla of Vater. We also analyzed subcellular localization of beta-catenin protein in all cases immunohistochemically, and confirmed its accumulation in the nucleus in four cases including in a CTNNB-1 mutated one. This is the first study to show CTNNB-1 mutation and beta-catenin expression in carcinoma of the ampulla of Vater. These results suggested that abnormal Wnt-wingless signaling and in particular beta-catenin alteration caused accumulation of beta-catenin, which might partially contribute to the development of carcinoma of the ampulla of Vater.


Assuntos
Ampola Hepatopancreática/metabolismo , Neoplasias do Ducto Colédoco/metabolismo , Proteínas do Citoesqueleto/metabolismo , Proteínas de Neoplasias/metabolismo , Transativadores/metabolismo , Idoso , Idoso de 80 Anos ou mais , Sequência de Aminoácidos , Ampola Hepatopancreática/patologia , Sequência de Bases , Neoplasias do Ducto Colédoco/patologia , Proteínas do Citoesqueleto/genética , Análise Mutacional de DNA , Primers do DNA/química , DNA de Neoplasias/análise , Éxons , Humanos , Técnicas Imunoenzimáticas , Pessoa de Meia-Idade , Dados de Sequência Molecular , Mutação/genética , Proteínas de Neoplasias/genética , Reação em Cadeia da Polimerase , Polimorfismo Conformacional de Fita Simples , Transativadores/genética , beta Catenina
13.
J Exp Clin Cancer Res ; 22(4): 623-31, 2003 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-15053306

RESUMO

To elucidate metastasis mechanisms, we established a Panc-1H5 subline with a highly liver metastatic cell line and a Panc-1P4a with a highly peritoneal metastatic cell line, which were sequentially selected from the parental pancreatic cancer cell line Panc-1. Using these three cell lines, we investigated several biological properties and mRNA levels of differentially-expressed genes involved in cancer metastasis with a cDNA macroarray. The tumorigenicity, motile activity, adhesive activity and cytokine production of metastatic sublines were higher than those of parental Panc-1 cells. Particularly, in Panc-1H5 cells, adhesive activity to the extracellular matrix and angiogenetic factors increased, whereas in Panc-1P4a cells, motile activity was extremely enhanced compared with Panc-1 cells. Histopathological findings for the three cell lines were the same. In cDNA macroarray analysis of Panc-1H5 cells, 11 genes were up-regulated and 20 genes were down-regulated compared with parental Panc-1 cells. In Panc-1P4a cells, 7 genes were up-regulated and 13 genes were down-regulated compared with parental Panc-1 cells. This study provides a demonstration of global gene expression analysis of pancreatic cancer cells with liver and peritoneal metastasis and these results provide new insight into the study of human pancreatic cancer metastasis.


Assuntos
Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Neoplasias Pancreáticas/patologia , Neoplasias Peritoneais/genética , Neoplasias Peritoneais/patologia , Animais , Adesão Celular , Diferenciação Celular , Citocinas/biossíntese , Feminino , Humanos , Neoplasias Hepáticas/secundário , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Metástase Neoplásica/genética , Metástase Neoplásica/patologia , Transplante de Neoplasias , Neoplasias Peritoneais/secundário , Células Tumorais Cultivadas
14.
Surg Neurol ; 23(4): 417-20, 1985 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-3975835

RESUMO

A patient is described who had a history of sudden vomiting and rapidly becoming deeply unconscious. There was no history of head trauma. The cause turned out to be an acute spontaneous subdural hematoma from arterial rupture. Comparable cases in the literature are reviewed and the etiological possibilities are discussed.


Assuntos
Hematoma Subdural/etiologia , Adulto , Artérias Cerebrais , Hematoma Subdural/diagnóstico por imagem , Humanos , Masculino , Radiografia
15.
Surg Neurol ; 23(6): 597-604, 1985 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-3887626

RESUMO

A case is reported of intracranial granuloma caused by Aspergillus fumigatus involving the anterior cranial fossa and the frontal lobe. In this case, clinical symptoms developed about 5 years before the diagnosis was made. The final diagnosis was made by a craniotomy. The patient was treated with an extensive excision and chemotherapy, but finally he failed to respond to these treatments. We compile a summary of reported cases with a tabulation of pertinent information and discuss the pathogenesis, prognosis, and difficulty in treating this infection.


Assuntos
Aspergilose/complicações , Lobo Frontal , Granuloma/etiologia , Crânio , Aspergillus fumigatus , Doenças Ósseas/diagnóstico por imagem , Doenças Ósseas/etiologia , Doenças Ósseas/patologia , Doenças Ósseas/cirurgia , Encefalopatias/diagnóstico por imagem , Encefalopatias/etiologia , Encefalopatias/patologia , Encefalopatias/cirurgia , Angiografia Cerebral , Traumatismos Craniocerebrais/complicações , Granuloma/diagnóstico por imagem , Granuloma/patologia , Granuloma/cirurgia , Humanos , Masculino , Pessoa de Meia-Idade , Reoperação , Tomografia Computadorizada por Raios X
16.
Surg Neurol ; 24(1): 95-100, 1985 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-4012568

RESUMO

A 21-year-old Japanese man with basilar impression with an anomalous configuration of the odontoid process and many other vertebral anomalies is reported. We thought that posterior decompression alone would be hazardous; therefore, in one session, the odontoid tip and anterior arch of the atlas were removed transorally, and posterior fixation between the occipital squama and the lamina of C-3 using acrylic plastic was performed. This treatment resulted in marked clinical improvement and required only a short hospital stay.


Assuntos
Vértebra Cervical Áxis , Neoplasias Ósseas/complicações , Cerebelo , Bulbo , Síndromes de Compressão Nervosa/etiologia , Processo Odontoide , Osteoma/complicações , Platibasia/complicações , Medula Espinal , Adulto , Vértebra Cervical Áxis/anormalidades , Vértebra Cervical Áxis/diagnóstico por imagem , Vértebra Cervical Áxis/cirurgia , Neoplasias Ósseas/cirurgia , Forame Magno/diagnóstico por imagem , Forame Magno/cirurgia , Humanos , Masculino , Mielografia , Síndromes de Compressão Nervosa/cirurgia , Processo Odontoide/anormalidades , Processo Odontoide/diagnóstico por imagem , Processo Odontoide/cirurgia , Osteoma/cirurgia , Plásticos , Próteses e Implantes , Tomografia
17.
Kansenshogaku Zasshi ; 63(12): 1296-300, 1989 Dec.
Artigo em Japonês | MEDLINE | ID: mdl-2559935

RESUMO

We tried to isolate rotaviruses from travelers with diarrhea arriving at Nagoya International Airport. 1. Ten cases revealed positive for rotavirus out of 334 diarrheal patients tested during a period from 1985 to 1988. 2. Most of the rotavirus positive cases were in their forties or in their fifties. 3. Geographic distribution of the infected area of the cases was not concentrated with specific countries. 4. Diarrhea was the most frequent clinical manifestations of the cases. 5. There was no cluster in seasonal distribution of the cases.


Assuntos
Diarreia/microbiologia , Infecções por Rotavirus/microbiologia , Viagem , Adolescente , Adulto , Fatores Etários , Criança , Diarreia/epidemiologia , Feminino , Humanos , Japão/epidemiologia , Masculino , Pessoa de Meia-Idade , Rotavirus/isolamento & purificação , Infecções por Rotavirus/epidemiologia
18.
J Parasitol ; 99(3): 453-8, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23245337

RESUMO

Neospora caninum is an Apicomplexan parasite that causes repeated abortion and stillbirth in cattle. The aim of this study was to clone the gene encoding the N. caninum orthologue (NcBAG1) of the Toxoplasma gondii bradyzoite-specific protein TgBAG1 and characterize its expression pattern in the parasite. Isolation of the full-length 684-bp gene revealed that it shared 78.3% sequence similarity with TgBAG1. NcBAG1 encodes a predicted protein of 227 amino acids with 80.3% similarity to TgBAG1. A putative signal peptide sequence and an invariant GVL motif characteristic of small heat-shock proteins were identified in the predicted N. caninum amino acid sequence. We expressed the NcBAG1 gene as a recombinant glutathione S-transferase fusion protein (rNcBAG1) in Escherichia coli and used the purified 60 kDa protein to obtain a monoclonal antibody (Mab). rNcBAG1 reacted to Mabs specific for NcBAG1 and TgBAG1. No reaction between the NcBAG1 Mab and N. caninum tachyzoites was observed. Although the predicted molecular mass of NcBAG1 is 25 kDa, Western blot analysis of parasite lysates using the NcBAG1 Mab revealed a cross-reactive protein of approximately 30 kDa. Additionally, immunofluorescence assays using the tachyzoite-specific Mab for NcSAG1 and the bradyzoite-specific Mab for TgBAG1 or NcSAG4 revealed NcBAG1-specific expression in bradyzoites in cultures exposed to sodium nitroprusside, a reagent that increases the frequency of bradyzoites. Interestingly, the NcBAG1 protein was identified in the cytoplasm of the bradyzoite-stage parasites. This preliminary analysis of the NcBAG1 gene will assist investigations into the role of this protein in N. caninum .


Assuntos
Antígenos de Protozoários/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Neospora/genética , Proteínas de Protozoários/genética , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/imunologia , Antígenos de Protozoários/química , Antígenos de Protozoários/metabolismo , Chlorocebus aethiops , Clonagem Molecular , DNA de Protozoário/química , Feminino , Proteínas de Choque Térmico/genética , Camundongos , Camundongos Endogâmicos BALB C , Neospora/imunologia , Neospora/metabolismo , Doadores de Óxido Nítrico/farmacologia , Nitroprussiato/farmacologia , Reação em Cadeia da Polimerase , Proteínas de Protozoários/química , Proteínas de Protozoários/imunologia , Proteínas de Protozoários/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/metabolismo , Especificidade da Espécie , Células Vero
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