RESUMO
Genetic abnormalities in mitochondrial complex assembling factors are associated with leukoencephalopathy. We present a 1-year-old girl with consciousness disturbance after a respiratory infection. Brain MRI revealed leukoencephalopathy with bilaterally symmetrical hyperintensity in the substantia nigra, medial thalamic nuclei, and basal nuclei, as well as cavities in the cerebral white matter and corpus callosum. Lactate levels in the spinal fluid were high, while magnetic resonance spectroscopy of the cerebral white matter and basal nuclei showed high peak lactate levels, suggesting mitochondrial dysfunction. The respiratory enzyme activity of complex I was reduced to 17% to 21% in skeletal muscle. Whole exome sequencing identified compound heterozygous variations in NDUFAF3, involved in the assembly of mitochondrial complex I (c.342_343insGTG:p.117Valdup, c.505C > A:p.Pro169Thr). Two-dimensional, blue-native polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate-PAGE revealed reductions in Q-module (NDUFS2, NDUFS3, and NDUFA9) and P-module (NDUFB10 and NDUFB11) subunits, indicating disruption of mitochondrial complex I assembly. Our report expands the spectrum of clinical phenotypes associated with pathogenic variants of NDUFAF3.
Assuntos
Predisposição Genética para Doença , Leucoencefalopatias/genética , Mitocôndrias/genética , Proteínas Mitocondriais/genética , Complexo I de Transporte de Elétrons/genética , Feminino , Humanos , Lactente , Leucoencefalopatias/patologia , Mitocôndrias/patologia , Mutação , Sequenciamento do ExomaRESUMO
Medical radiation exposure increases the likelihood of cataract formation. A personal dosimeter was attached to the left temple of 77 anaesthetists during 45 endovascular aortic aneurysm repairs and 32 interventional neuroradiology procedures. Compared with interventional neuroradiology, the median (IQR [range]) total radiation dose emitted by fluoroscopic equipment was significantly lower during endovascular aortic aneurysm repair (4175 (3127-5091 [644-9761]) mGy than interventional neuroradiology (1420 (613-2424 [165-10,840]) mGy, p < 0.001). However, radiation exposure to the anaesthetist's temple was significantly greater during endovascular aortic aneurysm repair (15 (6-41 [1-109]) µSv) than interventional neuroradiology (4 (2-8 [0-67]) µSv, p < 0.001). These data suggest that anaesthetists at our institution would have to deliver anaesthesia for ~1300 endovascular aortic aneurysm repairs and ~5000 interventional neuroradiology cases annually to exceed the general occupational limits, and ~10,000 endovascular aortic aneurysm repairs and ~37,500 interventional neuroradiology cases to exceed the ocular exposure limits recommended by the International Commission on Radiological Protection. Nevertheless, anaesthetists should be aware of the risk of ocular radiation exposure, and reduce this by limiting the time of exposure, increasing the distance from the source of radiation, and shielding.
Assuntos
Anestesiologia , Procedimentos Endovasculares/efeitos adversos , Olho/efeitos da radiação , Corpo Clínico Hospitalar , Exposição Ocupacional/análise , Anestesia Geral , Aneurisma Aórtico/diagnóstico por imagem , Aneurisma Aórtico/cirurgia , Fluoroscopia/efeitos adversos , Humanos , Aneurisma Intracraniano/diagnóstico por imagem , Aneurisma Intracraniano/cirurgia , Malformações Arteriovenosas Intracranianas/diagnóstico por imagem , Malformações Arteriovenosas Intracranianas/cirurgia , Japão , Neurorradiografia/efeitos adversos , Doses de Radiação , Monitoramento de Radiação/métodos , Radiografia Intervencionista/efeitos adversosRESUMO
BACKGROUND: To compare the hemodynamic and biological effects of high-adsorption continuous veno-venous hemofiltration (CVVH) with standard CVVH in septic shock. METHODS: In a randomized cross-over clinical trial twelve patients with septic shock and multiple organ failure were enrolled at a tertiary intensive care unit. Patients were allocated to either 9 hours of high-adsorption hemofiltration (CVVH with 3 hourly filter change using AN69 hemofilters - 3FCVVH) or 9 hours of standard hemofiltration (CVVH without filter change - 1F-CVVH). RESULTS: Changes in hemodynamic variables, dose of noradrenaline required to maintain a mean arterial pressure greater than 75 mmHg and plasma concentrations of cytokines (IL-6, IL-8, IL-10 and IL-18) were measured. A 9-hour period of 3F-CVVH was associated with greater reduction in noradrenaline dose than a similar period of 1F-CVVH (median reduction: 16 vs. 3.5 microg/min, p=0.036; median percentage reduction: 48.1% vs. 17.5%, p=0.028). Unlike 1F-CVVH, 3F-CVVH was associated with a reduction in the plasma concentration of IL-6, IL-10 and IL-18 at 9 hours and a significant decrease 30 minutes after additional filter changes (IL-6: p<0.01, p<0.01; IL-10: p=0.03, p=0.016 and IL-18: p=0.016, p<0.01, respectively). Both, 3F-CVVH and 1F-CVVH were associated with decreased plasma concentrations of IL-8 at 9 hours (p<0.01, p<0.01, respectively). In a confirmatory ex-vivo experiment IL-6 concentrations substantially decreased during 3F-CVVH (at baseline 511 pg/mL and at end: 21 pg/mL) whereas IL-6 concentrations increased in control blood (at baseline 511 pg/mL and at end: 932 pg/mL). CONCLUSIONS: High-adsorption CVVH appears more effective than standard CVVH in decreasing noradrenaline requirements and plasma concentrations of cytokines in septic shock patients.
Assuntos
Hemofiltração , Choque Séptico/terapia , Injúria Renal Aguda/complicações , Injúria Renal Aguda/terapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Pressão Sanguínea , Feminino , Frequência Cardíaca , Humanos , Interleucina-10/sangue , Interleucina-6/sangue , Interleucina-8/sangue , Masculino , Pessoa de Meia-Idade , Insuficiência de Múltiplos Órgãos , Norepinefrina , Diálise Renal , Choque Séptico/sangue , Choque Séptico/fisiopatologiaRESUMO
Using a large, international cohort, we sought to determine the effect of initial technique of renal replacement therapy (RRT) on the outcome of acute renal failure (ARF) in the intensive care unit (ICU). We enrolled 1218 patients treated with continuous RRT (CRRT) or intermittent RRT (IRRT) for ARF in 54 ICUs in 23 countries. We obtained demographic, biochemical and clinical data and followed patients to either death or hospital discharge. Information was analyzed to assess the independent impact of treatment choice on survival and renal recovery. Patients treated first with CRRT (N=1006, 82.6%) required vasopressor drugs and mechanical ventilation more frequently compared to those receiving IRRT (N=212, 17.4%), (p<0.0001). Unadjusted hospital survival was lower (35.8% vs. 51.9%, p<0.0001). However, unadjusted dialysis-independence at hospital discharge was higher after CRRT (85.5% vs. 66.2%, p<0.0001). Multivariable logistic regression showed that choice of CRRT was not an independent predictor of hospital survival or dialysis-free hospital survival. However, the choice of CRRT was a predictor of dialysis independence at hospital discharge among survivors (OR: 3.333, 95% CI: 1.845 - 6.024, p<0.0001). Further adjustment using a propensity score did not significantly change these results. We conclude that worldwide, the choice of CRRT as initial therapy is not a predictor of hospital survival or dialysis-free hospital survival but is an independent predictor of renal recovery among survivors.
Assuntos
Injúria Renal Aguda/terapia , Estado Terminal , Diálise Renal/métodos , Injúria Renal Aguda/fisiopatologia , Idoso , Causas de Morte , Estudos de Coortes , Cuidados Críticos , Feminino , Seguimentos , Previsões , Humanos , Rim/fisiopatologia , Masculino , Pessoa de Meia-Idade , Alta do Paciente , Estudos Prospectivos , Recuperação de Função Fisiológica/fisiologia , Respiração Artificial , Taxa de Sobrevida , Resultado do Tratamento , Vasoconstritores/uso terapêuticoRESUMO
We examined 28 cases of surgically resected gastric cancer, excluding the diffuse type, for loss of heterozygosity (LOH) on 12 chromosomal arms using polymorphic DNA markers. LOH on chromosome 18q was detected in 61% (14 of 23) of the cases by the probes OLVIIA8, OLVIIE10, p15-65, SAM 1.1, and OS-4, and a putative common region showing LOH included the locus of the DCC tumor suppressor gene. LOH on chromosome 17p was also frequently found (8 of 19 or 42% of the cases) by the probes p10-3 and pHF12-1, and in 5 of these 6 cases the LOH on chromosome 17p was accompanied by LOH on chromosome 18q. On the other hand, the incidence of LOH was 30% or less using probes pHRnES, pHF12-65, p-c-mybE2.6, NJ3 3.2, pHF12-8, pHINS6.0, p9D11, hp2-alpha, pCMM6, and P1A5 on chromosomes 1q, 5, 6q, 7q, 9, 11p, 13q, 16q, 20, and 22q, respectively. LOH on chromosome 18q was frequent irrespective of the depth of tumor invasion, whereas the incidence of LOH on chromosome 17p was higher in the cases in which the tumor invaded beyond the muscularis propria than in those in which tumor invasion was limited to the submucosa and muscularis propria. These results suggest that LOH on chromosome 18q occurs at an earlier stage than LOH on chromosome 17p and that the inactivation of tumor suppressor genes located on chromosome 17p and 18q (e.g., the p53 and DCC genes) is critically involved in the development of the majority of gastric cancers. While alteration of the p53 gene is observed in various human cancers, that of the DCC gene is considered to occur more selectively in gastrointestinal cancers.
Assuntos
Deleção Cromossômica , Cromossomos Humanos Par 17 , Cromossomos Humanos Par 18 , Genes Supressores de Tumor , Neoplasias Gástricas/genética , Mapeamento Cromossômico , Neoplasias Gastrointestinais/genética , Genes p53 , Marcadores Genéticos , Heterozigoto , Humanos , Invasividade Neoplásica/genética , Neoplasias Gástricas/patologia , Neoplasias Gástricas/cirurgiaRESUMO
Hyperparathyroidism is the first manifestation in a majority of multiple endocrine neoplasia (MEN1) patients. To discriminate between sporadic and hereditary parathyroid tumors and characterize MEN1 somatic mutations, we examined MEN1 gene mutations in patients who had undergone surgery for sporadic parathyroid tumors. DNA was extracted from fresh frozen parathyroid tumor specimens from 112 patients as well as from peripheral blood leukocytes from 64 of the 112 patients. Sequence analysis was performed to examine exons 2-10 of the MEN1 gene for mutations. Loss of heterozygosity (LOH) was also examined by an analysis of codon 418 and 541, which lie within a polymorphic region of MEN1. Somatic MEN1 mutations were found in 25 of the 112 patients (22%). Two patients had two point mutations (508del33 and Y341X and 363insT and 1767delT, respectively). A total of 27 mutations were characterized, 20 of which have not been reported previously. There were 7 nonsense mutations, 10 frameshift mutations, 2 splice site deletions, 5 missense mutations, and 3 in-frame mutations. Nineteen mutations (70%) predicted truncation of the menin protein. Germ-line MEN1 mutations were found in 3 of 64 patients (5%) who had no family history of endocrine tumors associated with MEN1, and these patients were identified as MEN1 gene probands. LOH at the MEN1 locus was detected in three parathyroid tumors showing germ-line mutation. LOH was significantly frequent in parathyroid tumors with somatic MEN1 mutations (15 of 22 tumors, 68%) but not in those without germ-line or somatic MEN1 mutations (14 of 51 tumors, 28%; P = 0.0011). Our findings suggest that alterations of both alleles of the MEN1 gene may be associated not only with endocrine tumors of affected MEN1 patients but also with sporadic parathyroid tumors. Germ-line MEN1 gene analysis can distinguish heritable from nonheritable parathyroid tumors, and MEN1 gene evaluation of patients with apparently sporadic parathyroid tumor is recommended before parathyroid surgery.
Assuntos
Mutação em Linhagem Germinativa , Neoplasia Endócrina Múltipla Tipo 1/genética , Mutação , Neoplasias das Paratireoides/genética , Adulto , Idoso , DNA de Neoplasias/genética , Feminino , Testes Genéticos , Humanos , Perda de Heterozigosidade , Masculino , Pessoa de Meia-IdadeRESUMO
This multicentre, retrospective observational study was conducted from January 2010 to December 2010 to determine the optimal time for discontinuing continuous renal replacement therapy (CRRT) by evaluating factors predictive of successful discontinuation in patients with acute kidney injury. Analysis was performed for patients after CRRT was discontinued because of renal function recovery. Patients were divided into two groups according to the success or failure of CRRT discontinuation. In multivariate logistic regression analysis, urine output at discontinuation, creatinine level and CRRT duration were found to be significant variables (area under the receiver operating characteristic curve for urine output, 0.814). In conclusion, we found that higher urine output, lower creatinine and shorter CRRT duration were significant factors to predict successful discontinuation of CRRT.
Assuntos
Injúria Renal Aguda/terapia , Terapia de Substituição Renal , Idoso , Creatinina/sangue , Feminino , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Fatores de TempoRESUMO
Although loss of heterozygosity (LOH) on chromosome 18q is frequently found in gastric cancer, the clinical significance of this abnormality has not been well documented. We examined LOH on chromosome 18q22-23 in DNA extracted from the tissues of gastric cancer patients using the PCR-based dinucleotide repeat assay with two microsatellite markers, D18S61 and D18S58. We investigated LOH in 100 samples of DNA extracted from formalin-fixed, paraffin-embedded tissues of cohesive-type gastric cancer patients operated on between 1984 and 1993. Thirty-two of 83 informative cases (39%) showed LOH on chromosome 18q22-23 at one or two loci. The LOH correlated significantly with serosal invasion of the tumor (P = 0.004) and hematogenous recurrence (P = 0.035). In 60 cases who were cured, the 5-year survival rate in patients with LOH (54%) was lower than that in patients without LOH (81%; P = 0.019). These results suggest that 18q22-23 LOH in cohesive gastric cancer is associated with tumor progression and a patient's poor prognosis.
Assuntos
Cromossomos Humanos Par 18/genética , DNA de Neoplasias/análise , Perda de Heterozigosidade/genética , Neoplasias Gástricas/genética , Idoso , Progressão da Doença , Feminino , Marcadores Genéticos , Humanos , Masculino , Neoplasias Gástricas/mortalidade , Taxa de SobrevidaRESUMO
The K-sam gene, originally isolated as an amplified gene from the stomach cancer cell line KATO-III, is characterized by its preferential amplification in the undifferentiated type (diffuse type) of stomach cancer and encodes one of the receptors for heparin-binding growth factors or fibroblast growth factors. The K-sam gene has been isolated by different methods and has been designated BEK, TK14, and Cek2. The receptor for keratinocyte growth factor was also found to be encoded by the same gene. To examine the expression of the K-sam protein in stomach cancer, polyclonal antibody pK1-2 was raised against the extracellular domain of the gene product. This antibody detected K-sam proteins by Western blot and flow cytometry analyses in stomach cancer cell lines KATO-III and HSC39, in which the K-sam gene is amplified and overexpressed. By immunohistochemical analysis, 20 of 38 cases of the undifferentiated type of advanced stomach cancer were K-sam positive, whereas none of 11 cases of the differentiated or intestinal type revealed K-sam staining. The K-sam product was observed predominantly in diffusely infiltrative lesions. In one autopsy case, the K-sam protein was detected only focally in the primary tumor, whereas markedly increased staining for the K-sam product was detected diffusely in the metastasized tumor in the lymph node and liver. These results suggest that K-sam overexpression is associated with the malignant phenotype of the undifferentiated type of stomach cancer, such as infiltrative growth and metastasis.
Assuntos
Receptores Proteína Tirosina Quinases/análise , Receptores de Fatores de Crescimento de Fibroblastos/análise , Neoplasias Gástricas/química , Sequência de Aminoácidos , Amplificação de Genes , Humanos , Imuno-Histoquímica , Dados de Sequência Molecular , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/imunologia , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos , Receptores de Fatores de Crescimento de Fibroblastos/genética , Receptores de Fatores de Crescimento de Fibroblastos/imunologia , Células Tumorais CultivadasRESUMO
BACKGROUND: The impact of different blood pressure targets is unknown for post cardiac surgery patient in the intensive care unit. We, therefore, investigated the effects of a mean arterial pressure (MAP) target of 65 or 85 mmHg on splanchnic oxygenation, metabolic function, cytokine regulation and gastric tonometry after cardiopulmonary bypass. METHODS: Sixteen patients were randomized to the HLH group (high-low-high) where MAP of 85-65-85 mmHg was targeted or the LHL group where MAP 65-85-65 mmHg was targeted with norepinephrine infusion. RESULTS: MAP targets were achieved in all patients at all timepoints (64 ± 3, 84 ± 4; 65 ± 5, LHL group; vs. 84 ± 3; 66 ± 2; 85 ± 5 mmHg, HLH group). At corresponding timepoints, hepatic venous saturation was 41 ± 15%; 58 ± 24%; 56 ± 21% in the LHL group vs. 50 ± 19%; 43 ± 20%; 41 ± 18% in the HLH group (P<0.05). No changes were observed in cardiac output, global or trans-splanchnic lactate levels and cytokine levels or in gastric tonometry CO2. CONCLUSION: Achieving a MAP target of 85 mmHg by means of norepinephrine infusion after CPB appears safe for the splanchnic circulation.
Assuntos
Pressão Sanguínea , Ponte Cardiopulmonar , Circulação Esplâncnica , Idoso , Idoso de 80 Anos ou mais , Pressão Arterial , Estudos de Coortes , Estudos Cross-Over , Citocinas/sangue , Feminino , Humanos , Masculino , Manometria , Pessoa de Meia-Idade , Norepinefrina/uso terapêutico , Oxigênio/sangue , Projetos Piloto , Circulação Esplâncnica/efeitos dos fármacos , Estômago/efeitos dos fármacos , Vasoconstritores/uso terapêuticoRESUMO
The region preceding putative transmembrane segment M1 of the glutamate receptor (GluR) channel is well conserved among subunits and has been proposed to constitute a part of the agonist binding site. The functional significance of this region was examined by introducing point mutations into charged residues of the alpha 1 subunit of the mouse alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)-selective GluR channel. The dose-response relationships of the mutant receptors were studied after expression in Xenopus oocytes by injection of the mutant alpha 1 subunit-specific mRNA together with the wild-type alpha 2-subunit-specific mRNA. Variable changes in the EC50 values for different agonists were found for the replacement of glutamic acid 398 by lysine and for the replacement of lysine 445 by glutamic acid. These residues may be involved in selective interaction of the GluR channel with agonists.
Assuntos
Ácido Ibotênico/análogos & derivados , Receptores de Neurotransmissores/metabolismo , Sequência de Aminoácidos , Animais , Sítios de Ligação , Clonagem Molecular , Glutamatos/metabolismo , Ácido Ibotênico/metabolismo , Ácido Caínico/metabolismo , Camundongos , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Oócitos , Ácido Quisquálico/metabolismo , Receptores de Glutamato , Receptores de Neurotransmissores/genética , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/metabolismo , Homologia de Sequência do Ácido Nucleico , Xenopus laevis , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol PropiônicoRESUMO
To develop an assay system that allows the N-methyl-D-aspartate (NMDA) receptor subtype-selective antagonistic potency of drugs, we have established Chinese hamster ovary cell lines expressing the four NMDA receptor subtypes (GluRepsilon1/zeta1-GluRepsilon4/zeta1) heat-indelibly. Using these clonal cells, we found that a novel antagonist, (1S,2R)-1-phenyl-2[(S)-1-aminopropyl]-N,N-diethylcyclopropanecarboxamide, was less selective for the GluRepsilon1/zeta1: the IC(50) values for the GluRepsilon1/zeta1-GluRepsilon4/zeta1 were 41.7, 13.3, 12.6 and 11.5 microM, respectively, while two well-known antagonists, DL-2-amino-5-phosphonovaleric acid and ifenprodil, showed the known potency and selectivity for each subtype. Thus, the established clonal cells are of use in characterizing the pharmacological properties of drugs that act on NMDA receptors.
Assuntos
Células CHO , Ciclopropanos/farmacologia , Antagonistas de Aminoácidos Excitatórios/farmacologia , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Receptores de N-Metil-D-Aspartato/metabolismo , Valina/análogos & derivados , Animais , Cálcio/metabolismo , Cricetinae , Eletrofisiologia , Regulação da Expressão Gênica/fisiologia , Ácido Glutâmico/farmacologia , Glicina/farmacologia , Temperatura Alta , Piperidinas/farmacologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Receptores de N-Metil-D-Aspartato/genética , Valina/farmacologiaRESUMO
BACKGROUND: Chronic liver diseases may alter trace element contents in the brain. Among these trace elements, manganese is a ubiquitous transition metal excreted by the liver into the bile. Blood concentrations of manganese are elevated in patients with biliary atresia who have undergone hepatic portoenterostomy. The present study investigated the effects of liver transplantation on manganese deposition in the brain in such patients. METHODS: The signal intensity of the globus pallidus was calculated as an index defined as the percentile ratio of signal intensity in the globus pallidus to the subcortical frontal white-matter in sagittal T1-weighted magnetic resonance imaging planes. RESULTS: Brain magnetic resonance imaging revealed hyperintense signals in the globus pallidus due to manganese deposition in biliary atresia patients. Few neurologic symptoms related to manganese intoxication were observed. However, one 23-year-old female with biliary atresia had depressive symptoms and dyskinesia; she improved after oral administration of the dopamine precursor, L-DOPA. Manganese deposition disappeared in two patients after living-related reduced-size hepatic transplantation. CONCLUSIONS: Manganese accumulates in the brain during cholestasis associated with biliary atresia and disappears after hepatic transplantation. Manganese deposition is likely to be subclinical and reversible but may be associated with some age-related neurologic symptoms.
Assuntos
Atresia Biliar/metabolismo , Globo Pálido/metabolismo , Manganês/metabolismo , Adulto , Atresia Biliar/sangue , Atresia Biliar/diagnóstico , Atresia Biliar/cirurgia , Criança , Pré-Escolar , Feminino , Globo Pálido/patologia , Humanos , Lactente , Transplante de Fígado , Doadores Vivos , Imageamento por Ressonância Magnética , Masculino , Manganês/sangue , Período Pós-Operatório , Valores de Referência , Análise de RegressãoRESUMO
BACKGROUND: We previously reported that pretransplant donor-specific blood transfusion (DST) induces CD45RC-CD4+ T cells, Th2-like effector cells, and prolongs rat hepatic allograft survival. Our study investigated the effects of posttransplant DST on rat hepatic allograft survival. METHODS: Three days after transplantation, LEW (RT1(1)) recipient rats with ACI (RT1a) livers were injected i.v. with freshly heparinized donor-specific blood. The time kinetics of CD45RC-CD4+ and CD45RC+CD4+ T cell subsets in hepatic infiltrates were examined. RESULTS: Posttransplant DST significantly prolonged rat hepatic allograft survival. Interferon (IFN)-gamma, interleukin (IL)-12, and IL-18 mRNA levels in hepatic allografts of untreated recipients were significantly greater than in recipients treated with posttransplant DST. However, hepatic allografts of recipients treated with posttransplant DST showed significantly higher IL-4, IL-10, and transforming growth factor (TGF)-beta mRNA levels than untreated recipients. The ratio of CD45RC-CD4+ T cells to CD45RC+CD4+ T cells was significantly higher in hepatic allografts treated with posttransplant DST than in untreated animals. Immunostaining with anti-rat dendritic cell (OX-62) monoclonal antibody revealed that OX-62+ cells were distributed to the splenic red pulp of animals treated with posttransplant DST and to the splenic white pulp in untreated animals. Most OX62+ cells isolated from the spleen of recipients treated with posttransplant DST expressed donor RT1Ba class II major histocompatibility complex antigens, suggesting that OX-62+ cells were of donor origin. CONCLUSION: Posttransplant DST was associated with persistent infiltration of CD45RC-CD4+ T cells, Th2-like effector cells, in rat hepatic allografts, causing immunologic unresponsiveness and establishment of microchimerism in the spleen.
Assuntos
Transfusão de Sangue , Transplante de Fígado/imunologia , Animais , Formação de Anticorpos/fisiologia , Linfócitos T CD4-Positivos/imunologia , Separação Celular , Corantes , Citometria de Fluxo , Sobrevivência de Enxerto/imunologia , Imuno-Histoquímica , Antígenos Comuns de Leucócito/fisiologia , Masculino , Ratos , Ratos Endogâmicos ACI , Ratos Endogâmicos Lew , Baço/química , Fatores de Tempo , Quimeras de Transplante/imunologia , Transplante Homólogo/imunologiaRESUMO
We report on a father and his son with supernumerary nipples. No male-to-male transmission has previously been described with this trait. This observation confirms that this trait is inherited in an autosomal dominant fashion.
Assuntos
Mamilos/anormalidades , Pré-Escolar , Genes Dominantes , Humanos , Masculino , Fatores Sexuais , SíndromeRESUMO
OBJECTIVE: Constitutively activating mutations of the thyrotropin receptor (TSHR) have been found in the majority of autonomously functioning thyroid nodules (AFTNs) in European patients. The reported frequency of these mutations varies among reports but amounts to 50-80%. To date, only one such mutation responsible for AFTNs has been identified in the Japanese population and the pathogenic role of such mutations in Japanese AFTNs has been questioned. In the present study, we evaluated the frequency of activating mutations in the TSHR and G(alpha)s in 10 Japanese AFTNs. DESIGN: Genomic DNA was extracted from fresh frozen tissue. The TSHR and the almost entire sequence of the gene coding for the alpha subunit of Gs have been amplified and sequenced. RESULTS: In sequence analysis, four mutations in the TSHR (T632A, I486M, M453T and L512R) were found. To complete our analysis, we searched mutations in the gene coding for the alpha subunit of Gs, in the samples negative for TSHR mutations. In one case a mutation (R201H) affecting GTPase activity was found. CONCLUSIONS: If we focus on the solitary nodules, we obtain the same mutation proportion as in European patients (70%). The absence of TSHR and G(alpha)s mutations in a significant proportion of autonomous adenomas in multinodular goiters suggests that other causes may also play a role in the genesis of these lesions.
Assuntos
Mutação , Receptores da Tireotropina/genética , Nódulo da Glândula Tireoide/genética , DNA/análise , Feminino , Subunidades alfa Gs de Proteínas de Ligação ao GTP/genética , Humanos , Japão , Masculino , Análise de Sequência de DNARESUMO
To develop a drug screening system, we introduced expression vectors carrying the mouse N-methyl-D-aspartate (NMDA) receptor channel epsilon1 and zeta1 subunit cDNAs under the promoter of the Drosophila heat shock protein hsp70 into Chinese hamster ovary (CHO) cells. We selected clonal cell lines by means of RNA blot hybridization and fura-2 fluorometry. One of these cell lines, ZE1-1, optimally expressed the epsilon1 and zeta1 subunit mRNAs when induced by an incubation at 43 degrees C for 2 h. Heated ZE1-1 cells exhibited the NMDA-induced intracellular Ca2+ elevation, whereas unheated they showed no such response. NMDA and L-glutamate, but not alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) and kainate, induced an increase in the intracellular Ca2+ concentration. The response to the agonists was marginal in the absence of glycine, and diminished by Mg2+ and NMDA receptor antagonists. Furthermore, exposure to agonists of ZE1-1 cells expressing the epsilon1/zeta1 NMDA receptor channel resulted in the release of lactate dehydrogenase (LDH) activity in the culture medium indicating agonist-induced cell death. NMDA receptor antagonists inhibited the LDH activity release. These results suggest that ZE1-1 cells will provide a useful screening system for novel drugs acting on the epsilon1/zeta1 NMDA receptor channel.
Assuntos
DNA Complementar/genética , Receptores de N-Metil-D-Aspartato/metabolismo , Animais , Células CHO/metabolismo , Células Cultivadas , Cricetinae , Regulação da Expressão Gênica/genética , CamundongosRESUMO
Using a baculovirus expression vector system, the zeta 1 subunit of the mouse N-methyl-D-aspartate (NMDA) receptor channel was expressed in Spodoptera frugiperda insect cells. The peptide corresponding to the C-terminus of the zeta 1 subunit was synthesized by using the multiple antigen peptide (MAP) system, and an antibody to the synthetic peptide was produced. Immunoblotting using the newly developed antibody revealed the major 122-kDa and the minor 104-kDa protein bands. The effect of tunicamycin on the immunoblots and [35S]methionine/[35S]cysteine metabolic radiolabeling suggested that the two bands corresponded to glycosylated and non-N-glycosylated forms, respectively. Membranes prepared from insect cells infected with the recombinant virus had the binding activity of antagonist ligand 5,7-[3-3H]dichlorokynurenate (DCKA) of a glycine recognition domain of the receptor. Both immunofluorescence labeling and the [3H]DCKA binding assays also showed a greater level of expression (Bmax = 51 pmol/mg protein) in the insect cells. The ligand binding characteristics of the receptors expressed in insect cells suggested that the single zeta 1 subunit protein has glycine antagonist binding properties comparable to those of the native NMDA receptor channels. The lack of DCKA-binding activity of the non-N-glycosylated NMDA receptor expressed in the presence of tunicamycin suggested that N-linked oligosaccharide is essentially required for expression of a functional receptor in insect cells. This is the first report describing the importance of N-glycosylation for the acquisition of ligand binding to NMDA receptor channel subunit protein.
Assuntos
Baculoviridae/genética , Receptores de N-Metil-D-Aspartato/biossíntese , Animais , Anticorpos/imunologia , Baculoviridae/metabolismo , Western Blotting , Células Cultivadas , Eletroforese , Imunofluorescência , Expressão Gênica , Cinética , Coelhos , Receptores de Glutamato/genética , Receptores de N-Metil-D-Aspartato/genética , Spodoptera , Tunicamicina/farmacologiaRESUMO
Arginine-481 is located in the putative agonist-binding region preceding the putative transmembrane segment M1 of the alpha1-subunit of the AMPA-selective glutamate receptor (GluR) channel. This amino acid is completely conserved among GluR proteins. A site-directed mutagenesis study using a baculovirus expression system showed that substitution of glutamate, glutamine and lysine for arginine-481 of the recombinant alpha1-subunit protein abolishes binding to [3H]AMPA completely. The present study provides the first direct experimental evidence that the conserved charged arginine-481 residue is essential, directly or indirectly, for the acquisition of ligand-binding activity by the receptor protein.
Assuntos
Arginina/metabolismo , Mutação/genética , Receptores de Glutamato/efeitos dos fármacos , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico/farmacologia , Animais , Sequência de Bases , Canais Iônicos/efeitos dos fármacos , Camundongos , Dados de Sequência Molecular , Ensaio Radioligante , Receptores de Glutamato/metabolismoRESUMO
The organization of the blood and lymphatic microvessels of the gallbladder in the guinea pig is demonstrated by scanning electron microscopy (SEM) of vascular corrosion casts, and SEM of KOH-macerated tissues. In the lamina propria of the gallbladder, there is a dense network of subepithelial capillaries. The network is supplied by the arterioles that come off the arterial plexus located deep in the lamina propria. The network gathers into the postcapillary venules continuous with the collecting venular plexus located immediately below the subepithelial capillary network. The precapillary arterioles are sparsely surrounded by a single layer of circularly oriented extensions of smooth muscle cells. The terminal arterioles are endowed with circularly oriented fusiform smooth muscle cells. The nervous plexus is also noticed along the terminal arterioles. The capillaries are embraced by flat prolongations of pericytes. The postcapillary venules are sparsely surrounded by stellate pericytes and the collecting venules are sparsely surrounded by elongated or branched spindle-shaped, primitive smooth muscle cells which extend their long process in various directions along the vascular wall. The lymphatics are mostly located in the subserosal layer. The tips of the initial lymphatics are closed by endothelial cells, although there are frequently some gaps between them. The thin flaps of the lymphatic endothelial cells overlap or interdigitate with each other. The luminar surfaces of the lymphatics show oval nuclear protrusions, while the abluminal surfaces showed numerous microfolds except for the oval and flat nuclear portions. The lymphatics possess neither smooth muscle cells nor pericytes.