RESUMO
Exposure to paraquat is possibly involved with the development of several conditions, including neurodegenerative diseases, such as Parkinson's disease (PD). This condition is mainly characterized by the loss of dopaminergic neurons in the nigrostriatal pathway and the development of classical motor symptoms. Etiology includes exposure to environmental factors, such as the paraquat exposure, and inflammatory diseases may exacerbate paraquat neurotoxicity. The aim of the study was to investigate whether the exposure to paraquat associated with the presence of periodontal disease is able to induce motor and biochemical changes in rats similar to that observed in PD. Adult male Wistar rats were sent to ligature. After 48 h, they were sent to daily treatment paraquat (1 mg/kg/day; 2 mL/kg; intragastric) or vehicle for 4 weeks. Twenty-four hours after the last administration, the open field test was performed. The rats were euthanized and the left hemimandibles and striatum were dissected for the analysis of dopaminergic and inflammatory markers. Only the combination of periodontal disease model plus paraquat exposure induced motor impairments. Remarkably, the paraquat exposure increased the ligature-induced alveolar bone loss in hemimandibles. Moreover, only the combination of periodontal disease and paraquat exposure induced the loss of dopaminergic neurons and astrocyte activation in the striatum.
Assuntos
Herbicidas/toxicidade , Atividade Motora/efeitos dos fármacos , Paraquat/toxicidade , Animais , Masculino , Doenças Periodontais , Ratos , Ratos WistarRESUMO
UDP-N-acetylglucosamine: beta-D-mannoside beta-1,4-N-acetylglucosaminyl-transferase III (GnT-III) is a key enzyme in the branching of asparagine-linked oligosaccharides, which are present in surface membrane proteins of various tissues and in secretory glycoproteins. The activity of GnT-III was assayed in 2 human hepatoblastoma cell lines, Huh6, which was the parental cell line, and HB611, which was established by transfection of 3 tandem copies of the hepatitis B virus genome into Huh6. A significant difference in GnT-III activity was found between Huh6 and HB611 (136 +/- 18.3 pmol/h/mg versus 6.7 +/- 2.4 pmol/h/mg; mean +/- SD, P < 0.001), whereas levels of the glycosyltransferases alpha-3-D-mannoside beta-1,4-N-acetylglucosaminyltransferase IV, alpha-6-D-mannoside beta-1,6-N-acetylglucosaminyltransferase-V, and beta-1,4-galactosyltransferase were almost the same in both cell lines. Northern blot analysis indicated that the decreased activity of GnT-III in HB611 was due to the decreased transcript. When HB611 was treated with interferon-alpha, expression of hepatitis B virus-related mRNA decreased, and the activity of GnT-III increased from 8.5 +/- 3.8 to 22.0 +/- 7.2 pmol/h/mg (mean +/- SD, P < 0.05). This increase was not found in Huh6. Binding capacity with erythrocyte phytohemagglutinin in these cells using fluorescence-activated cell sorter analysis was different, suggesting that the structure of sugar chain on the cell surface might be altered by suppression of GnT-III activity. This is the first report that hepatitis B virus selectively suppressed the GnT-III activity in hepatoblastoma cells.
Assuntos
Vírus da Hepatite B/genética , Hepatoblastoma/enzimologia , Neoplasias Hepáticas/enzimologia , N-Acetilglucosaminiltransferases/biossíntese , Transfecção , Northern Blotting , Configuração de Carboidratos , Sequência de Carboidratos , Linhagem Celular , Expressão Gênica/efeitos dos fármacos , Humanos , Interferon alfa-2 , Interferon-alfa/farmacologia , Dados de Sequência Molecular , N-Acetilglucosaminiltransferases/metabolismo , Oligossacarídeos/biossíntese , RNA Mensageiro/biossíntese , Proteínas Recombinantes , Supressão Genética/efeitos dos fármacos , Células Tumorais CultivadasRESUMO
The LEC (Long-Evans with a cinnamon-like color) rat is a mutant of the Long-Evans strain which develops hereditary hepatitis and hepatoma with age. Activities and mRNA levels of N-acetylglucosaminyltransferase III and V (GnT-III and GnT-V, respectively) were determined during hepatocarcinogenesis in this rat using a LEA (Long-Evans with an agouti color) rat as a control. GnT-III activity in LEC rat liver increased after 30 weeks of age, at the stage of chronic hepatitis, to about 2.5-11.5 times the level in LEC rats aged 1-9 weeks. GnT-V activity in the LEC rat liver increased after 20 weeks of age, at the stage of acute hepatitis, to about 1.5-2.5 times the level in LEC rats of 1-9 weeks of age and then remained elevated. Both enzymes showed more dramatic increases in males than in females. The mRNA levels of the enzymes increased in proportion with the enzyme activities. Furthermore, GnT-III and GnT-V mRNAs were highly expressed in both cancer lesion and adjacent tissues. In one case of hepatoma with lymph node metastasis, GnT-III and GnT-V mRNA expression was much higher in the metastatic lesion than in the original cancer. GnT-III and GnT-V levels in the original cancer lesions were similar to those in the cancer lesions of the other LEC rats. These results indicated that expression of GnT-III and GnT-V was induced by chronic liver damage and hepatocarcinogenic changes in the LEC rats.
Assuntos
Carcinoma Hepatocelular/química , Neoplasias Hepáticas/química , Fígado/química , N-Acetilglucosaminiltransferases/análise , RNA Mensageiro/análise , Fatores Etários , Sequência de Aminoácidos , Animais , Carcinoma Hepatocelular/enzimologia , Feminino , Hepatite Animal/enzimologia , Fígado/enzimologia , Neoplasias Hepáticas/enzimologia , Masculino , Dados de Sequência Molecular , Ratos , Ratos EndogâmicosRESUMO
Changes in oligosaccharide structures alter the biological functions of cancer cells. Alpha1-6 fucosyltransferase (alpha1-6FucT) catalyzes the transfer of fucose to the innermost GlcNAc in N-glycans. Although alpha1-6FucT is barely detected in normal liver, it is enhanced during rat hepatocarcinogenesis and in human hepatoma. To understand the biological meaning of the alpha1-6FucT in hepatoma, especially in terms of metastasis, we established human hepatoma cell lines, which express high levels of alpha1-6FucT by transfection of the alpha1-6FucT gene and investigated intrahepatic metastasis after splenic injection to athymic mice. Tumor formation in the liver was dramatically suppressed in the alpha1-6FucT transfectants (1 of 9 and 1 of 10 in alpha1-6FucT transfectants versus 6 of 9 and 6 of 9 in controls). Although there were no differences in terms of cell invasiveness to a Matrigel or in terms of cytotoxicity to interleukin 2-treated lymphocytes between alpha1-6FucT transfectants and control cells, cell adhesion to mice hepatocytes and nonparenchymal liver cells in culture was significantly inhibited in alpha1-6FucT transfectants, compared to the controls. Attachment of alpha1-6FucT transfectants to a fibronectin-coated dish was decreased compared to controls because alpha5beta1 integrin was more strongly alpha1-6 fucosylated in the alpha1-6FucT transfectants. Two-dimensional electrophoresis followed by lectin blot showed that certain glycoproteins (Mr 50,000-150,000, pI 4.8-5.5) were alpha1-6 fucosylated and might be linked to suppression of intrahepatic metastasis. This is the first demonstration of the biological significance of alpha1-6 fucosylation on N-glycans in hepatoma cells under in vivo conditions.
Assuntos
Carcinoma Hepatocelular/enzimologia , Fucosiltransferases/biossíntese , Neoplasias Hepáticas/enzimologia , Proteínas de Neoplasias/biossíntese , Animais , Caderinas/biossíntese , Caderinas/genética , Sequência de Carboidratos , Carcinoma Hepatocelular/patologia , Adesão Celular , Fucosiltransferases/genética , Glicosilação , Humanos , Injeções , Interleucina-2/farmacologia , Neoplasias Hepáticas/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Dados de Sequência Molecular , Invasividade Neoplásica , Metástase Neoplásica , Proteínas de Neoplasias/genética , Processamento de Proteína Pós-Traducional , Receptores de Fibronectina/análise , Baço , Transfecção , Células Tumorais CultivadasRESUMO
E-cadherin is a central molecule in the process of gastric carcinogenesis and its posttranslational modifications by N-glycosylation have been described to induce a deleterious effect on cell adhesion associated with tumor cell invasion. However, the role that site-specific glycosylation of E-cadherin has in its defective function in gastric cancer cells needs to be determined. Using transgenic mice models and human clinical samples, we demonstrated that N-acetylglucosaminyltransferase V (GnT-V)-mediated glycosylation causes an abnormal pattern of E-cadherin expression in the gastric mucosa. In vitro models further indicated that, among the four potential N-glycosylation sites of E-cadherin, Asn-554 is the key site that is selectively modified with ß1,6 GlcNAc-branched N-glycans catalyzed by GnT-V. This aberrant glycan modification on this specific asparagine site of E-cadherin was demonstrated to affect its critical functions in gastric cancer cells by affecting E-cadherin cellular localization, cis-dimer formation, molecular assembly and stability of the adherens junctions and cell-cell aggregation, which was further observed in human gastric carcinomas. Interestingly, manipulating this site-specific glycosylation, by preventing Asn-554 from receiving the deleterious branched structures, either by a mutation or by silencing GnT-V, resulted in a protective effect on E-cadherin, precluding its functional dysregulation and contributing to tumor suppression.
Assuntos
Caderinas/metabolismo , N-Acetilglucosaminiltransferases/metabolismo , Neoplasias Gástricas/metabolismo , Sequência de Aminoácidos , Animais , Asparagina/genética , Caderinas/química , Caderinas/genética , Caderinas/fisiologia , Domínio Catalítico/genética , Linhagem Celular Tumoral , Cães , Mucosa Gástrica/metabolismo , Mucosa Gástrica/patologia , Glicosilação , Células HT29 , Humanos , Células Madin Darby de Rim Canino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , N-Acetilglucosaminiltransferases/antagonistas & inibidores , N-Acetilglucosaminiltransferases/genética , Homologia de Sequência de Aminoácidos , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologiaRESUMO
N-acetylglucosaminyltransferases III (GnT-III) and V (GnT-V) play a pivotal role in the processing of N-linked glycoproteins, and are highly involved in cancer progression and metastasis. Expression of GnT-III and GnT-V in the liver is enhanced during hepatocarcinogenesis, although they are not expressed in the normal liver. Gene expression of GnT-V is regulated by a transcriptional factor, ets-1, which is involved in angiogenesis and invasion of tumor cells. When the formation of the product of GnT-V, GlcNAc-beta1-6 branches, is inhibited by overexpression of GnT-III, lung metastasis of melanoma cells is suppressed. Modification of glycoprotein receptors such as the receptors for epidermal growth factor and nerve growth factor by GnT-III sense transfection changes an intracellular signaling pathway, which may lead to a variety of biological alterations in tumor cells. In this review, we focus on cancer progression and metastasis in relation to GnT-III and GnT-V.
Assuntos
Regulação Neoplásica da Expressão Gênica/genética , N-Acetilglucosaminiltransferases/genética , Neoplasias/enzimologia , Animais , Sequência de Carboidratos , Regulação para Baixo , Regulação da Expressão Gênica , Humanos , Dados de Sequência Molecular , N-Acetilglucosaminiltransferases/química , Metástase Neoplásica , Neoplasias/genética , Neoplasias/patologia , Proteína Proto-Oncogênica c-ets-1 , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-ets , Sequências Reguladoras de Ácido Nucleico , Transdução de Sinais , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transfecção , Células Tumorais CultivadasRESUMO
GDP-L-Fuc:N-acetyl-beta-D-glucosaminide alpha1-6-fucosyltransferase (alpha1-6FucT) catalyzes the transfer of fucose from GDP-Fuc to N-linked type complex glycoproteins. This enzyme was purified from a human fibroblast cell line, porcine brain, a human gastric cancer cell line and human blood platelets. cDNA cloning of porcine and human alpha1-6FucT was performed from a porcine brain and gastric cancer cell cDNA libraries, respectively. Their homology is 92.2% at the nucleotide level and 95.7% at the amino acid level. No putative N-glycosylation sites were found in the predicted amino acid sequence. No homology to other fucosyltransferases such as alpha1-2FucT, alpha1-3FucT and alpha1-4FucT was found except for a region consisting of nine amino acids. The alpha1-6FucT gene is located at chromosome 14q24.3, which is also a different location from other fucosyltransferases reported to date. The alpha1-6FucT gene is the oldest gene family in the phylogenic trees among the nine cloned fucosyltransferase genes. alpha1-6FucT is widely expressed in various rat tissues and the expression of alpha1-6FucT in the liver is enhanced during hepatocarcinogenesis of LEC rats which develop hereditary hepatitis and hepatomas. In cases of human liver diseases, alpha1-6FucT is expressed in both hepatoma tissues and their surrounding tissues with chronic liver disease, but not in the case of normal liver. Serum alpha1-6-fucosylated alpha-fetoprotein (AFP) has been employed for an early diagnosis of patients with hepatoma. The mechanisms by which alpha1-6 fucosylation of AFP occurs in the hepatoma is not due to the up-regulation of alpha1-6FucT alone. Interestingly, when the alpha1-6FucT gene is transfected into Hep3B, a human hepatoma cell line, tumor formation in the liver of nude mice after splenic injection is dramatically suppressed. In this review, we focus on alpha1-6FucT and summarize its properties, gene expression and biological significance.
Assuntos
Fucosiltransferases/genética , Animais , Plaquetas/enzimologia , Carcinoma Hepatocelular/sangue , Carcinoma Hepatocelular/diagnóstico , Linhagem Celular , Clonagem Molecular , DNA Complementar/isolamento & purificação , Fucosiltransferases/sangue , Fucosiltransferases/isolamento & purificação , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/diagnóstico , Camundongos , Camundongos Nus , Ratos , Ratos Long-Evans , Suínos , Células Tumorais Cultivadas , alfa-Fetoproteínas/metabolismoRESUMO
N-Acetylglucosaminyltransferase V (GnT-V) is an enzyme that catalyzes beta 1-6 branching of N-acetylglucosamine on asparagine-linked oligosaccharides of cell proteins. Metastatic potential of various cancer cells has been shown to correlate with increase of GnT-V activity and concomitant beta 1-6 branching of N-acetylglucosamine. However, protein expression of GnT-V in human cancer tissue and its clinical significance have not yet been demonstrated. To clarify the possible relationship between metastasis and GnT-V in human colorectal cancer, protein expression of GnT-V was studied using surgically resected specimens. We established a monoclonal antibody against GnT-V and performed immunohistochemical analysis of 103 human colorectal cancer cases. Of 103 cases, 26 cases (25.2 %) showed specific expression of GnT-V in colorectal cancer tissues. The expression of GnT-V was significantly correlated with distant metastasis (P < 0.05, chi2 test). Overall 5-year survival rate was 52.8% for GnT-V-positive patients and 81.7% for GnT-V-negative patients (P < 0.01, Log-rank test). We showed direct evidence for the relationship between GnT-V and metastasis in human colorectal cancer. Screening of GnT-V expression in colorectal cancer may provide useful information for prognosis of postoperative patients.
Assuntos
Neoplasias Colorretais/enzimologia , N-Acetilglucosaminiltransferases/metabolismo , Idoso , Sequência de Aminoácidos , Animais , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Feminino , Humanos , Imuno-Histoquímica , Metástase Linfática , Masculino , Camundongos , Pessoa de Meia-Idade , Dados de Sequência Molecular , Metástase Neoplásica , Estadiamento de Neoplasias , Prognóstico , Taxa de SobrevidaRESUMO
Non-retractable cell surface projections and cytoskeleton-mediated functional defects are distinguishing features of both hairy cell leukemia (HCL) and neutrophil actin dysfunction (NAD). These defects in NAD neutrophils are attributed to moderate over-expression of pp52 (LSP1), the F-actin-binding, leukocyte-specific phosphoprotein. Here we report that pp52 is similarly elevated in HCL patient PBMCs. Established HCL cell lines exhibited characteristic morphological features like those of fresh HCL cells and showed elevated pp52 levels. The excess pp52 in these HCL cell lines was selectively associated with the F-actin-rich cytoskeletal arrays in surface projections. Treatments producing radical changes in HCL cell shape also altered pp52 expression and intracellular distribution. Alpha interferon (IFNalpha, used to treat HCL) reduced pp52 levels, normalized intracellular pp52 distribution and reverted HCL cells to rounded B cell morphology. Phorbol ester stimulation rapidly generated hyper-phosphorylated pp52 isoforms which translocated from the cytoskeleton to the cytosol prior to the further elongation of surface spikes. This indicates a direct role for phosphorylation in controlling pp52 interactions with the cytoskeleton. Overall, these findings strongly suggest that elevated pp52 expression and/or selective cytoskeletal association contributes to the distinctive morphology of HCL cells.
Assuntos
Proteínas de Ligação ao Cálcio/biossíntese , Citoesqueleto/metabolismo , Leucemia de Células Pilosas/metabolismo , Actinas/metabolismo , Linfócitos B/efeitos dos fármacos , Linfócitos B/metabolismo , Western Blotting , Linfoma de Burkitt/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Tamanho Celular/efeitos dos fármacos , Citosol/metabolismo , Humanos , Interferon-alfa/farmacologia , Líquido Intracelular/metabolismo , Leucemia de Células Pilosas/patologia , Proteínas dos Microfilamentos , Microscopia Confocal , Neutrófilos/metabolismo , Neutrófilos/patologia , Fosfoproteínas/biossíntese , Fosforilação/efeitos dos fármacos , Acetato de Tetradecanoilforbol/farmacologia , Células Tumorais Cultivadas/efeitos dos fármacosRESUMO
BACKGROUND: Our previous study demonstrated that Helicobacter pylori eradication was less effective in smokers than in non-smokers. Cetraxate is an anti-ulcer drug that increases gastric mucosal blood flow. AIM: To evaluate the effect of cetraxate combined with new triple therapy for the eradication of H. pylori in smokers. METHODS: This study had a single-centre, double-blind, randomized non-placebo design. A total of 106 consecutive H. pylori-positive smoking patients were randomly allocated to one of two regimens: one group received omeprazole (20 mg), amoxycillin (1500 mg), and clarithromycin (600 mg) for 7 days (OAC, n=55). The other group recieved OAC plus cetraxate (600 mg) for 7 days (OAC + CET, n=51). The success of H. pylori eradication was evaluated by histology and the 13C-urea breath test at 4 weeks after completion of treatment. RESULTS: By intention-to-treat analysis, the H. pylori eradication rate was 55% in the OAC group and 92% in the OAC + CET group (P<0.01). By per protocol analysis, the H. pylori eradication rate was 58% in the OAC group and 94% in the OAC + CET group (P<0.01). CONCLUSION: Cetraxate combined with new triple therapy increases the eradication of H. pylori in smokers.
Assuntos
Amoxicilina/administração & dosagem , Antibacterianos/administração & dosagem , Antiulcerosos/administração & dosagem , Claritromicina/administração & dosagem , Mucosa Gástrica/efeitos dos fármacos , Infecções por Helicobacter/tratamento farmacológico , Helicobacter pylori/efeitos dos fármacos , Omeprazol/administração & dosagem , Penicilinas/administração & dosagem , Fumar , Ácido Tranexâmico/análogos & derivados , Adulto , Idoso , Método Duplo-Cego , Quimioterapia Combinada , Feminino , Mucosa Gástrica/irrigação sanguínea , Mucosa Gástrica/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Fluxo Sanguíneo Regional , Ácido Tranexâmico/administração & dosagem , Ácido Tranexâmico/farmacologia , Resultado do TratamentoRESUMO
OBJECTIVES: Transport ventilators with inspiratory triggering functions and pressure support-control modes have recently become commercially available. We evaluated these ventilators in comparison with a standard ICU ventilator. STUDY DESIGN: Laboratory study with a mechanical lung model. METHODS: We compared the performance of four transport ventilators (model 740, Mallinckrodt, Pleasanton, CA; TBird, Bird Products Corp, Palm Springs, CA; LTV1000, Pulmonetic Systems, Colton, CA; Esprit, Respironics, Vista, CA) with a standard ICU ventilator (model 7200ae; Mallinckrodt) using a test lung that simulated spontaneous breathing (compliance, 46.8 mL/cm H(2)O; resistance, 5 cm H(2)O/L/s). The settings of ventilators were positive end-expiratory pressure (PEEP) of 0 or 5 cm H(2)O, and pressure support (PS) of 0 or 10 cm H(2)O. The settings of the test lung were inspiratory time of 1 s, respiratory rate of 10/min, peak inspiratory flow of 40, 60, and 80 L/min. To evaluate inspiratory function at each setting, we measured the inspiratory delay time (DT), inspiratory trigger pressure (P-I), and the time for airway pressure to rise from the baseline pressure to 90% of the end-inspiratory pressure (T(90%)); for expiratory function, supraplateau expiratory pressure (P-E) and the time constant (taue) for pressure decrease during exhalation were evaluated. Oxygen requirement was assessed as the time required to empty a 3.5-L oxygen tank. RESULTS: For inspiratory triggering, four transport ventilators had DT < 100 ms, which is considered clinically satisfactory, in all the settings except for PS 0 cm H(2)O, PEEP 0 cm H(2)O, and inspiratory flow of 80 L/min with LTV1000. P-I increased only in LTV1000 when PEEP was increased from 0 to 5 cm H(2)O. taue for the transport ventilators was > 50% shorter than for the ICU ventilator except for PS 0 cm H(2)O and PEEP 5 cm H(2)O with TBird. Oxygen requirement was lowest for the Esprit, followed by the 740, LTV1000, and TBird. CONCLUSION: The newer Food and Drug Administration-approved transport ventilators have performance indexes comparable to the ventilator currently used in ICUs and can probably be recommended for clinical use.
Assuntos
Respiração com Pressão Positiva/instrumentação , Trabalho Respiratório/fisiologia , Desenho de Equipamento , Humanos , Unidades de Terapia Intensiva/provisão & distribuição , Modelos BiológicosRESUMO
Recent studies on the transcriptional factor ets-1 and carcinoma have shown that ets-1 is linked to carcinoma progression, including tumor invasion and metastasis. We studied the clinical significance of ets-1 in human hepatocellular carcinoma (HCC) by using immunohistochemical staining methods. In 99 HCC cases, the levels of ets-1 expression were analyzed in comparison with various clinicopathologic parameters, such as TNM stage, intrahepatic metastasis, histologic differentiation, and prognosis. Expression of ets-1 was scarcely detected in normal liver but markedly enhanced in noncancerous lesions adjacent to HCC lesions. In HCC lesions, ets-1 expression was observed with high incidence, although the average labeling index (LI) was lower than in noncancerous lesions. However, unexpectedly, the average LI in HCC was lower in cases of high TNM stage, poor differentiation, portal invasion, intrahepatic metastasis, large tumor size, and high Ki-67 LI. Furthermore, cases with high ets-1 expression showed better outcomes for disease-free survival than those with low ets-1 expression by univariate and multivariate analyses. These findings strongly suggest that, unlike in other neoplasms, ets-1 has a crucial role in hepatocarcinogenesis and HCC progression, especially during their early phases.
Assuntos
Carcinoma Hepatocelular/metabolismo , Expressão Gênica , Neoplasias Hepáticas/metabolismo , Proteínas Proto-Oncogênicas/genética , Fatores de Transcrição/genética , Northern Blotting , Carcinoma Hepatocelular/patologia , Feminino , Humanos , Antígeno Ki-67/análise , Neoplasias Hepáticas/patologia , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Invasividade Neoplásica , Metástase Neoplásica , Estadiamento de Neoplasias , Prognóstico , Proteína Proto-Oncogênica c-ets-1 , Proteínas Proto-Oncogênicas c-ets , RNA Mensageiro/análiseRESUMO
GDP-L-Fuc:N-acetyl-beta-D-glucosaminide:alpha1-6 fucosyltransferase (alpha1-6 FucT), which catalyzes the transfer of fucose from GDP-Fuc to N-linked type complex glycopeptides, was purified from a culture supernatant of human gastric cancer cell line MKN45. The purification procedures included chromatographies on Q-Sepharose Fast Flow, synthetic GDP-hexanolamine-Sepharose, and GnGn-bi-Asn-Sepharose columns. SDS-PAGE of the purified enzyme gave a major band corresponding to an apparent molecular mass of 60 kDa. The enzyme was recovered in a 12% final yield with an approximately 4,600-fold increase in specific activity. The pH optimum was 7.5, and the enzyme was fully active in the presence of 5 mM EDTA and did not require divalent cations, Mg2+ and Ca2+. Oligonucleotide primers designed from partial amino acid sequences were used to amplify and clone alpha1-6 FucT cDNA from a cDNA library of MKN45 cells. The cDNA encodes 575 amino acids in length, and contains the predicted N-terminal and internal amino acid sequences derived on lysyl endopeptidase digestion. The homology to porcine brain alpha1-6 FucT is 92.2% at the nucleotide level and 95.7% at the amino acid level. No putative N-glycosylation sites were found in the predicted amino acid sequence of the human MKN45 cell enzyme or that of porcine brain. Thus, the enzyme is distinct from other fucosyltransferases which catalyze alpha1-2, alpha1-3, and alpha1-4 fucose addition.
Assuntos
Fucosiltransferases/genética , Fucosiltransferases/isolamento & purificação , Neoplasias Gástricas/enzimologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Encéfalo/enzimologia , Cromatografia por Troca Iônica , Clonagem Molecular , DNA Complementar , Humanos , Dados de Sequência Molecular , Neoplasias Gástricas/patologia , Suínos , Células Tumorais CultivadasRESUMO
An assay method for GDP-L-Fuc:N-acetyl-beta-D-glucosaminide alpha 1-6fucosyltransferase (alpha 1-6FucT; EC 2.4.1.68) activity has been developed, involving a fluorescent pyridylaminated substrate. A glycopeptide derived from bovine gamma-globulin was coupled with 4-(2-pyridylamino)butylamine (PABA) through the peptide bond, and the following substrate was obtained. [equation: see text] The substrate and guanosine diphospho-fucopyranoside (GDP-Fuc) were incubated with a crude enzyme extract for 2 h, and then the enzymatic product was separated by reversed phase HPLC. Quantitation of the product involved measurement of the fluorescence intensity of the fucosylated pyridylaminated sugar. The structures of both synthesized GnGn-bi-Asn-PABA (substrate), and synthesized GnGnF-bi-Asn-PABA (product) were analyzed by 1H NMR. The enzymatic product was also analyzed by 1H NMR and was found to have alpha 1-6fucose at the reducing end GlcNAc. This method is highly specific for alpha 1-6FucT and is applicable for various experiments, including purification and cell culture ones.
Assuntos
Fucosiltransferases/análise , Animais , Encéfalo/enzimologia , Sequência de Carboidratos , Bovinos , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Corantes Fluorescentes/química , Fucosiltransferases/metabolismo , Glicopeptídeos/química , Fígado/enzimologia , Neoplasias Hepáticas Experimentais/enzimologia , Espectroscopia de Ressonância Magnética , Dados de Sequência Molecular , Estrutura Molecular , Ratos , Espectrometria de Fluorescência , Especificidade por Substrato , SuínosRESUMO
Intra-nigral administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyrindine (MPTP) caused a lesion in the substantia nigra, compact part (SNc) and a specific loss of dopamine and its metabolites in the striatum of rats. The animals were then tested in the two-way active avoidance task. MPTP-treated animals presented lower learning scores in the training and test sessions, an effect that was not caused by motor impairment or by a decreased sensitivity to footshock since their reaction time to the footshock (unconditioned stimulus - UCS) was not reduced. These lower scores were also not attributable to lower acoustic sensitivity or to a slowing in the association of the sound cue (conditioned stimulus - CS) with the UCS since the reaction time to the CS in the active avoidance response did not differ between MPTP-treated and control groups. Therefore, these results are more properly attributable to an impairment of the memory acquisition and retention processes. In addition, this study is presented as a model of early Parkinson's Disease amnesia and is discussed in terms of the importance of the nigrostriatal pathway to memory acquisition and storage processes.
Assuntos
1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina , Aprendizagem da Esquiva/efeitos dos fármacos , Rememoração Mental/efeitos dos fármacos , Transtornos Parkinsonianos/induzido quimicamente , Substância Negra/efeitos dos fármacos , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/farmacologia , Animais , Aprendizagem por Associação/efeitos dos fármacos , Aprendizagem por Associação/fisiologia , Aprendizagem da Esquiva/fisiologia , Condicionamento Clássico/efeitos dos fármacos , Condicionamento Clássico/fisiologia , Dopamina/metabolismo , Masculino , Rememoração Mental/fisiologia , Vias Neurais/efeitos dos fármacos , Vias Neurais/fisiopatologia , Transtornos Parkinsonianos/fisiopatologia , Ratos , Ratos Wistar , Retenção Psicológica/efeitos dos fármacos , Retenção Psicológica/fisiologia , Substância Negra/fisiopatologiaRESUMO
Improvements in perioperative care, namely, organ preservation solutions, immunosuppression, and increased experience of surgical, anesthetic, and intensive care teams, have contributed to the success of pancreas transplantation. The objective of this study was to present data on anesthesia for pancreas transplantation alone (PTA) or simultaneous with kidney (SPKT), evaluating crystalloid, albumin and blood component infusions, graft ischemic times, and weights and ages of recipient. We evaluated patients undergoing SPKT (n=73), PTA (n=49), or SPKT with kidney living donor (n=8). Aggressive monitoring and therapy were used to avoid hypoperfusion, optimized with intravenous fluids, vasoative drugs, and correction of metabolic disturbances. Three SPKT patients were not extubated at the end of surgery. There were no other complications related to anesthesia in any patient. Although it is a high-risk surgery, PTA or SPKT is routine in our practice. Adequate perioperative care is important not only for the safety of the procedure but also for graft viability, contributing to a promising long-term treatment of insulin-dependent diabetic patients.
Assuntos
Anestesia/métodos , Transplante de Rim/métodos , Transplante de Pâncreas/métodos , Adulto , Humanos , Doadores Vivos , Monitorização Intraoperatória/métodos , Resultado do TratamentoRESUMO
Pancreas transplantation is a method to restore endogenous insulin secretion in insulin-dependent diabetic patients. Because glycemia >150 mg/dL may harm pancreatic graft beta cells, early glucose control using insulin administration is recommended during transplantation. The aim of this study was to evaluate the benefits of strict glycemic control during pancreas transplantation by comparing two types of insulin and glucose administration: continuous infusion and bolus. Capillary glucose was measured every 30 minutes after anesthetic induction for pancreas transplantation alone or simultaneously with kidney transplantation. Intravenous regular insulin was administered for values >150 mg/dL or glucose for values <100 mg/dL. The following timepoints were evaluated: anesthetic induction, before pancreatic graft reperfusion, and the first 4 minutes after reperfusion. Pancreatic graft ischemia time was significantly lower in the bolus group (P <.02). Immediately after reperfusion, there was a small increase in glycemia with a decrease in subsequent measurements in both groups. No significant difference in glycemia was observed between the groups at any time. Induction values were greater than all other timepoints in both groups. Glycemic control is important; it was successfully obtained with both methods. The trend to decrease glucose after reperfusion suggest early graft function.
Assuntos
Glicemia/metabolismo , Transplante de Pâncreas/métodos , Adulto , Diabetes Mellitus Tipo 1/sangue , Diabetes Mellitus Tipo 1/cirurgia , Feminino , Humanos , Cuidados Intraoperatórios , Masculino , Monitorização IntraoperatóriaRESUMO
PURPOSE: To analyse the main cardiac risk factors responsible for immediate and late outcomes in patients undergoing thoracic surgeries. METHODS: We performed a retrospective analysis of 90 cases of cardiac patients submitted to non-cardiac thoracic surgeries. Surgeries were divided into greater ones and others and the heart diseases into severe and mild disease. We analysed immediate and late complications, and the mortality inside these groups. RESULTS: We found a greater morbi-mortality in the greater surgeries group and a greater late mortality in the severe heart disease group. There were evidences that the degree of the heart disease does not influence immediate outcome. CONCLUSION: The heart disease was not a limiting immediate risk for surgery.
Assuntos
Cardiopatias/complicações , Complicações Intraoperatórias/epidemiologia , Complicações Pós-Operatórias/epidemiologia , Procedimentos Cirúrgicos Torácicos , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Fatores de RiscoRESUMO
Epignathus (pharyngeal teratoma) is a rare disease of newborns associated with a high mortality secondary to airway obstruction in the neonatal period. We report anesthetic management of a newborn with epignathus who underwent tumor resection. He was delivered vaginally at 39 weeks of gestation and Apgar scores were 9 at 1 and 5 min. The tumor originated from the palate, almost filled the oral cavity and protruded through the mouth with its external part 6 x 7 cm in size. He could breathe with the head and mass turned to the left. The excision of the tumor was scheduled on the fifth day of life. Mask ventilation and laryngoscopy were considered impossible. Fiberoptic nasal intubation was successfully performed with topical anesthesia without sedation. Tumor was resected with blood loss of 103 gm. The trachea was extubated on the third postoperative day and the postoperative course was uneventful. For safe management of cases of pharyngeal teratoma, careful preoperative assessment of the airway is most important and sufficient preparation and careful intubation are mandatory to keep airway patent. The perioperative bleeding from the tumor and the airway obstruction by the tumor or its remnant after the excision could also be hazardous to the airway.
Assuntos
Anestesia por Inalação , Neoplasias Faríngeas/cirurgia , Teratoma/cirurgia , Tecnologia de Fibra Óptica , Humanos , Lactente , Intubação Intratraqueal/métodos , MasculinoRESUMO
We evaluated mortality of 2689 patients admitted to the Intensive Care Unit, Osaka University Hospital from January, 1987 to December, 1998. The patients were divided into 3 groups. Group A consisted of 1408 patients who underwent cardiovascular surgery, group B, 1082 patients who underwent other surgical procedures and group C, 199 patients who were transferred from the department of medicine. We studied mortality rate, causes of death, correlation between length of ICU stay and mortality rate, and mortality rate among age groups for 12 years. The main causes of death were cardiac failure and sepsis in group A, and respiratory failure and sepsis in group B and C. Mortality rate in each group showed no significant change for the last 12 years. Those who stayed more than 2 weeks in ICU showed a significantly higher mortality rate (p < 0.0001). Thus, length of ICU stay and mortality rate showed a positive correlation (p < 0.0001). The youngest group (age 0-1) showed a significantly higher mortality rate than other age groups (p < 0.0001). As sepsis was the most important cause of death in all the groups, the prevention and treatment of infection are the most important issue in our ICU to reduce mortality rate.