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1.
Transfus Med ; 26(5): 365-372, 2016 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-27350440

RESUMEN

BACKGROUND AND OBJECTIVES: The effect of leukoreduction and storage periods on the accumulation of bioactive lysophospholipids and substances in human autologous blood (AB units) has not been fully investigated. MATERIALS AND METHODS: The accumulation of bioactive lysophospholipids such as sphingosine 1-phosphate (S1P) and lysophosphatidylserine (LysoPS) in AB units during the storage was investigated. The time-dependent changes and the effect of the filtration in pre-storage leuckoreduction (LR) and unmodified samples derived from 46 AB units were analysed. Additionally, the changes of lysophospholipids and platelet releasate, namely ß-thromboglobulin (ß-TG), induced by exposure of whole blood (WB) or platelet-rich plasma (PRP) to the filter material were analysed. RESULTS: LysoPS, but not S1P levels, time-dependently and significantly increased in both unmodified and LR samples. LysoPS significantly decreased in LR compared with unmodified samples, whereas S1P increased in LR compared with unmodified samples. In addition, exposure of WB and/or PRP to the filter material in vitro resulted in increased levels of S1P, LysoPS and ß-TG. CONCLUSIONS: LR effectively reduced the accumulation of LysoPS in AB units. On the other hand, it increased concentrations of S1P due to platelet activation by exposure to the filter material. These suggest that increases of S1P levels in LR and LysoPS in the unmodified samples were mainly caused by the leukocytes and/or platelets and that LR was effective in inhibiting the accumulation of LysoPS.


Asunto(s)
Conservación de la Sangre , Transfusión de Sangre Autóloga , Procedimientos de Reducción del Leucocitos , Lisofosfolípidos/sangre , Esfingosina/análogos & derivados , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Esfingosina/sangre
2.
Tissue Antigens ; 80(4): 336-40, 2012 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-22776008

RESUMEN

Human neutrophil antigens (HNAs) play an important role in a variety of clinical conditions including immune-mediated neutropenia, non-hemolytic transfusion reactions, and transfusion-related acute lung injury. The aim of this study was to investigate the frequency distribution of HNAs-1 to -5 among the Japanese population. We analyzed samples from 570 healthy Japanese by molecular and serologic techniques to estimate the gene frequencies of HNAs-1 to -5. DNA samples were obtained and typed for the HNA-1 (n = 523), -3 (n = 570), -4 (n = 570), and -5 (n = 508), by molecular techniques. The HNA-1 genotype was determined by using a commercial polymerase chain reaction-reverse sequence-specific oligonucleotide probes (PCR-rSSOP) kit. The HNA-3 to -5 genotypes were determined by the PCR-sequence specific primer (PCR-SSP), previously described, with a small modification. The HNA-2a phenotype was determined in 301 donors by granulocyte immunofluorescence test. In Japanese, the gene frequencies of HNA-1a, -1b, and -1c were 0.623, 0.377, and 0.000, respectively. The frequency of HNA-2a phenotype was 0.987, and the gene frequencies of HNA-3a and -3b were 0.654 and 0.346, respectively. HNA-4a and -4b were found at 1.000 and 0.000, respectively, and HNA-5a and -5b at 0.840 and 0.160, respectively. We describe, for the first time, the frequencies of all HNAs (HNA-1 to -5) among the Japanese population. This study will be helpful for the prediction of the risk of alloimmunization to HNA, especially to determine the risk of HNA alloantibody production by transfusion of HNA incompatible blood and feto-maternal incompatibility.


Asunto(s)
Pueblo Asiatico/genética , Frecuencia de los Genes , Isoantígenos/genética , Neutrófilos/metabolismo , Alelos , Cartilla de ADN , Femenino , Genotipo , Humanos , Isoantígenos/clasificación , Isoantígenos/inmunología , Masculino , Tipificación Molecular , Neutrófilos/citología , Neutrófilos/inmunología , Fenotipo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
4.
Biochim Biophys Acta ; 1003(1): 84-90, 1989 May 15.
Artículo en Inglés | MEDLINE | ID: mdl-2469474

RESUMEN

Several monoclonal antibodies to human A-I apolipoprotein were produced after immunising mice with pure delipidated apoA-I. These monoclonal antibodies were characterised for their ability to react with whole lipoproteins, apolipoproteins and fragments of apoA-I generated by cleavage with cyanogen bromide. The data suggest that production of monoclonal antibodies using apoA-I as antigen was influenced by two major epitopes subsequently localised to cyanogen bromide fragments 1 and 3, and have been designated antibodies 1----5 A-IB and 6----10 A-IB, respectively. Cyanogen bromide fragments were first purified to homogeneity before screening by competitive displacement or immunoblotting procedures. Definitive characterisation of one antibody series (1----5 A-IB) depended ultimately on Western blotting following isoelectric focusing of purified apoA-I fragments. This technique identified the epitope for these antibodies to fragment 1, an identification not fully concluded from competitive displacement studies. These studies have also revealed the presence of microheterogeneity in fragment 1 (as well as in fragment 4) of apoA-I, suggesting that structural variations in several regions may account for the polymorphism observed in this apolipoprotein.


Asunto(s)
Anticuerpos Monoclonales/biosíntesis , Apolipoproteínas A/inmunología , Animales , Apolipoproteína A-I , Unión Competitiva , Bromuro de Cianógeno , Ensayo de Inmunoadsorción Enzimática , Epítopos/análisis , Femenino , Humanos , Ratones , Ratones Endogámicos BALB C , Fragmentos de Péptidos/inmunología
5.
Biochim Biophys Acta ; 1165(1): 61-7, 1992 Nov 11.
Artículo en Inglés | MEDLINE | ID: mdl-1420349

RESUMEN

A new apolipoprotein complex designated as the apo(AII-E2-AII) complex was identified in the lipoprotein fractions of human plasma with apoE phenotypes containing apoE2 (E4/E2, E3/E2, and E2/E2). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by an immunoblotting assay using anti-apoE or anti-apoAII antibodies, established that the apo(AII-E2-AII) complex, with a molecular weight of 58,000, was identical to the complex consisting of apoE and apoAII, and that it also dissociated following reduction with beta-mercaptoethanol. This new complex was also demonstrated to be distinct from the apo(E-AII) complex and apoE monomer by isoelectric focusing, in the samples that were not treated with beta-mercaptoethanol. In apoE phenotype E3/E2, the apo(AII-E2-AII) complex was primarily included in the high-density lipoprotein (HDL, 1.063 < d < 1.21 g/ml) fraction, but was also observed in a small quantity in the very-low-density lipoprotein (VLDL, d < 1.006 g/ml) fraction. For further characterization, the apo(AII-E2-AII) complex was isolated by preparative SDS-PAGE, and no contamination of apo(E-AII) complex and apoE monomer was detected by immunoblotting assay using an anti-apoE antibody. It was confirmed by an enzyme-linked immunosorbent assay (ELISA) system that a molecular ratio between apoAII monomer and apoE in the isolated apo(AII-E2-AII) complex was approx. 2, when the apo(E-AII) complex was used as a standard with the ratio of 1:1. It indicates that the apo(AII-E2-AII) complex is formed from two molecules of apoAII monomer and one molecule of apoE.(ABSTRACT TRUNCATED AT 250 WORDS)


Asunto(s)
Apolipoproteína A-II/análisis , Apolipoproteínas E/sangre , Lipoproteínas/sangre , Apolipoproteína E2 , Ácido Ditionitrobenzoico/farmacología , Humanos , Immunoblotting , Focalización Isoeléctrica , Mercaptoetanol/farmacología
6.
J Thromb Haemost ; 3(5): 983-90, 2005 May.
Artículo en Inglés | MEDLINE | ID: mdl-15869595

RESUMEN

BACKGROUND AND OBJECTIVES: Analysis of dysfibrinogens has improved our understanding of molecular defects and their effects on the function of intact fibrinogen. To eliminate the influence of plasma heterozygous molecules, we synthesized and analyzed recombinant-variant fibrinogens. METHODS: We synthesized two recombinant-variant fibrinogens with a single amino acid substitution at the 15Gly residue in the Bbeta-chain: namely, Bbeta15Cys and Bbeta15Ala. RESULTS: Western blotting analysis of purified fibrinogen revealed the existence of a small amount of a dimeric form only for Bbeta15Cys fibrinogen. For Bbeta15Cys fibrinogen, functional analysis indicated (a) no thrombin-catalyzed fibrinopeptide B (FPB) release and (b) markedly impaired lateral aggregation in thrombin- and reptilase-catalyzed fibrin polymerizations. For Bbeta15Ala fibrinogen, such analysis indicated slight impairments of both thrombin-catalyzed FPB release and lateral aggregation in thrombin-catalyzed fibrin polymerization, but nearly normal lateral aggregation in reptilase-catalyzed fibrin polymerization. These impaired lateral aggregations were accompanied by thinner fibrin fiber diameters (determined by scanning electron microscopy of the corresponding fibrin clots). CONCLUSION: We conclude that a region adjacent to Bbeta15Gly plays important roles in lateral aggregation not only in desA fibrin polymerization, but also in desAB fibrin polymerization, and we speculate that the marked functional differences between Bbeta15A and Bbeta15C fibrinogens in FPB release and fibrin polymerization might not only be due to the presence of a substituted cysteine residue in Bbeta15C fibrinogen, but also to the existence of disulfide-bonded forms. Finally, our data indicate that the Bbeta15Gly residue plays important roles in FPB release and lateral aggregation of protofibrils.


Asunto(s)
Fibrinógeno/química , Fibrinopéptido B/química , Proteínas Recombinantes/química , Alanina/química , Animales , Batroxobina/química , Western Blotting , Catálisis , Cromatografía Líquida de Alta Presión , Cisteína/química , Dimerización , Disulfuros/química , Electroforesis en Gel de Poliacrilamida , Fibrina/química , Fibrina/ultraestructura , Glicina/química , Heterocigoto , Humanos , Immunoblotting , Cinética , Microscopía Electrónica de Rastreo , Mutagénesis , Unión Proteica , Venenos de Serpiente , Trombina/química , Factores de Tiempo
7.
J Thromb Haemost ; 2(3): 468-75, 2004 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-15009465

RESUMEN

BACKGROUND AND OBJECTIVES: Analysis of dysfibrinogens has provided useful information aiding our understanding of molecular defects in fibrin polymerization. We have already reported impaired fibrin polymerization in a variant fibrinogen (gammaArg275Cys), the Cys being located in the D:D interface. Since this substitution occurred in a heterozygous individual, interpretation of the functional analysis was complicated. We tried to resolve this complication by synthesizing a recombinant variant fibrinogen. METHODS: A variant gamma-chain expression plasmid was transfected into Chinese hamster ovary cells expressing normal human fibrinogen Aalpha- and Bbeta-chains. The recombinant variant fibrinogen (gamma275C) was purified using an immunoaffinity column, and we compared its structure and functions with those of normal recombinant fibrinogen (gamma275R) and plasma variant fibrinogen. RESULTS: Mass analyses showed the existence of disulfide-linked Cys in both patient and recombinant variant fibrinogens. Functional analyses indicated that both fibrin polymerization and gamma-gamma dimer formation were markedly impaired in the variant fibrinogen. The impairments were much more pronounced in gamma275C than in plasma variant fibrinogen. In addition, scanning electron microscopic observation of fibrin clots made from gamma275C revealed less dense fibrin fiber bundles and larger fiber diameter than in those made from gamma275R, and also the existence of many aberrant fibrin fibers with tapered ends. CONCLUSIONS: These results indicate that gammaArg275 has an important residue affecting the structure and function of the gamma-chain C-terminal domain. However, the variant D:D interface can interact with that of the normal fibrinogen existing in a heterozygous patient with dysfibrinogenemia.


Asunto(s)
Disulfuros/metabolismo , Fibrinógeno/química , Fibrinógeno/metabolismo , Sustitución de Aminoácidos , Arginina , Secuencia de Bases , Sitios de Unión , Catálisis , Cisteína , Cartilla de ADN , Factor XIIIa/metabolismo , Fibrina/química , Fibrina/metabolismo , Fibrina/ultraestructura , Fibrinógeno/ultraestructura , Humanos , Cinética , Microscopía Electrónica de Rastreo , Peso Molecular , Mutagénesis Sitio-Dirigida , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Trombina/metabolismo
8.
J Thromb Haemost ; 2(8): 1359-67, 2004 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-15304042

RESUMEN

BACKGROUND AND OBJECTIVES: We have previously reported that recombinant gamma 275Cys fibrinogen exhibits a marked impairment of functions as well as aberrant fibrin clot and bundle structures, as compared with wild-type, gamma 275Arg, and plasma fibrinogen from a heterozygous proband. Since gamma Arg275His mutations have also been reported in 10 families, we synthesized recombinant gamma 275His fibrinogen and gamma 275Ala fibrinogen (as a control) and analyzed and compared them with gamma 275Cys and gamma 275Arg. METHODS: A variant gamma-chain expression plasmid was transfected into Chinese hamster ovary cells expressing normal human fibrinogen A alpha- and B beta-chains. After purification of the recombinant variant fibrinogens, we performed functional analyzes for thrombin-catalyzed fibrin polymerization and factor XIIIa (FXIIIa)-catalyzed gamma-gamma dimer formation from fibrin or fibrinogen and also ultrastructural analysis of fibrin clots and bundles. RESULTS: By comparison with both gamma 275His and gamma 275Ala fibrinogens, recombinant gamma 275Cys fibrinogen exhibited a more impaired gamma-gamma dimer formation from fibrin or fibrinogen, a more aberrant fibrin clot structure, and thicker fibers in fibrin bundles. In 1 : 1 mixtures of gamma 275Arg and gamma 275Cys fibrinogens or gamma 275Arg and gamma 275His fibrinogens, thrombin-catalyzed fibrin polymerization and both fibrin clot and fiber structures showed some compensation (as compared with gamma 275Cys or gamma 275His alone). CONCLUSION: These results strongly suggest that an amino acid substitution of gamma 275Arg alone disrupts D:D interactions in thrombin-catalyzed fibrin polymerization and the formation of fibrin bundles and fibrin clots. Moreover, the existence of a subsequent disulfide-linked Cys in gamma 275C fibrinogen augments the impairment caused by a His or Ala substitution.


Asunto(s)
Factor XIIIa/química , Fibrina/química , Fibrinógeno/genética , Trombina/metabolismo , Alanina/química , Animales , Células CHO , Catálisis , Cricetinae , Reactivos de Enlaces Cruzados/farmacología , Electroforesis en Gel de Poliacrilamida , Fibrina/ultraestructura , Fibrinógeno/química , Histidina/química , Humanos , Microscopía Electrónica de Rastreo , Plásmidos/metabolismo , Polímeros/química , Proteínas Recombinantes/química , Proteínas Recombinantes/farmacología , Espectrofotometría , Trombina/química , Factores de Tiempo , Rayos Ultravioleta
9.
Thromb Haemost ; 81(5): 763-6, 1999 May.
Artículo en Inglés | MEDLINE | ID: mdl-10365751

RESUMEN

Fibrinogen Matsumoto III (M-III) is a dysfibrinogen identified in a 66-year-old woman with rectal cancer. The fibrinogen level determined by the thrombin-time method was markedly decreased in preoperative coagulation tests of her plasma. Three fibrinogen polypeptide-chain gene fragments from the proposita were amplified by the polymerase chain reaction method, then sequenced. The triplet CGC encoding the amino acid residue gamma275 was replaced by TGC, resulting in the substitution of Arg->Cys. There have been previous reports of nine families with the same alteration, nine families with an Arg->His variant and one family with an Arg->Ser variant in this residue, which has been shown to be one of the most important amino acids in the 'D:D' interaction site. In addition, there are three silent mutations in the Aalpha-chain gene and two mutations in the intron of the Bbeta-chain and the gamma-chain gene. However, none of these mutations is thought to be the cause of the dysfunctional fibrinogen. The thrombin-catalyzed fibrin polymerization in the presence of 1 mM Ca ions was markedly delayed in purified M-III. Its lag period was longer than those of Matsumoto II (M-II; gamma308Asn->Lys) and Matsumoto I (M-I; gamma364Asp-His). gamma364Asp is one of the most important residues in the polymerization pocket of the 'D:E' interaction site and gamma308Asn is located in the vicinity of a high affinity Ca2+ binding site in the D-domain, gamma311-336. The maximum slope of the polymerization curve for M-III was about 4-fold steeper than that for M-1 but less steep than that for M-II. These results may suggest that the tertiary structure of the polymerization pocket plays a more important role in the lateral aggregation of protofibrils than that of the 'D:D' interaction site.


Asunto(s)
Fibrina/metabolismo , Fibrinógenos Anormales/genética , Fibrinógenos Anormales/metabolismo , Neoplasias del Recto/genética , Anciano , Dimerización , Femenino , Humanos , Mutación Puntual , Neoplasias del Recto/sangre , Especificidad por Sustrato
10.
Am J Clin Pathol ; 115(3): 424-9, 2001 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-11242799

RESUMEN

By making comparisons with the usual manual method, we evaluated an automatic fluorescent image analyzer (Image Titer, Tripath Imaging, Burlington NC), the software for which was developed to simplify measuring indirect immunofluorescent antinuclear antibodies (FANAs). In this new system, images of the stained sample are displayed, and it measures the FANA titer and staining pattern using only 1 slide per subject and does not required the staining of a series of diluted samples as does the manual method. This system showed good reproducibility and linearity for 4 types of control serum samples (with homogeneous, speckled, discrete speckled, and nucleolar staining patterns). In 132 serum samples, consistency between the methods was 100% for the FANA staining pattern and 93.9% for the FANA titer. The Image Titer system detected each pattern in samples with 2 mixed patterns. This system should partly reduce labor and lead to results with minimum differences among individuals, including newly trained persons.


Asunto(s)
Anticuerpos Antinucleares/sangre , Autoanálisis/instrumentación , Técnica del Anticuerpo Fluorescente Indirecta/instrumentación , Nucléolo Celular/inmunología , Humanos , Control de Calidad , Reproducibilidad de los Resultados
11.
Clin Chim Acta ; 295(1-2): 77-85, 2000 May.
Artículo en Inglés | MEDLINE | ID: mdl-10767395

RESUMEN

During the past 3 years, we encountered 12 new cases suspected of being dysfibrinogenemias, via plasma coagulation screening tests, which included determination of fibrinogen concentration both by thrombin time and immunologic methods. We performed sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions and immunoblot analysis for these plasma fibrinogens. We identified two cases that were characterized by two distinct gamma-chain bands, similar to previous observations with Matsumoto-II (the substitution of gamma308Asn to Lys). Therefore, in order to identify the gamma308Lys variant easily and rapidly, we established an allele-specific polymerase chain reaction (AS-PCR). AS-PCR results indicated that the two cases were indeed heterozygous for the gamma308Lys variant; ten other cases were negative for this mutation. In conclusion, the ratio of fibrinogen concentrations determined by functional and antigenic methods in combination with the immunoblot analysis made these cases attractive for identifying the gamma308Lys mutation. The AS-PCR method proved to be a useful procedure to identify the gamma308K mutation.


Asunto(s)
Asparagina/química , Pruebas de Coagulación Sanguínea , Fibrinógenos Anormales/metabolismo , Lisina/química , Alelos , Sustitución de Aminoácidos , Secuencia de Bases , Western Blotting , Cartilla de ADN , Electroforesis en Gel de Poliacrilamida , Femenino , Fibrinógenos Anormales/química , Humanos , Masculino , Reacción en Cadena de la Polimerasa
12.
Ann Clin Lab Sci ; 31(2): 163-70, 2001 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-11337906

RESUMEN

In a proband (21-yr-old female), we previously identified an apolipoprotein (apo) E variant, apoE3 (Arg 145-->His), with an isoelectric point midway between apoE3 and apoE2. ApoE gene analysis of 4 of the proband's kin indicated that 3 possess the same variant. All 4 had a high concentration of apoE in plasma, while 3 of 4 had hypertriglyceridemia. In the proband (who had no hypertriglyceridemia), most apoE was distributed in slow-alpha lipoproteins (predominantly in the form of apoE-AII heterodimer) and in larger molecules with apparent molecular weights of 80 and 100 kDa. In the proband's brother (with hypertriglyceridemia), however, most apoE was distributed in slow pre-beta lipoproteins, predominantly in the form of monomeric apoE. In each subject, the concentration of apoE3 variant was significantly higher than that of normal apoE3 in the predominant apoE-rich lipoprotein. The apoE3 variant, which displayed a slightly reduced binding ability to LDL-receptor and heparin, may induce an accumulation of apoE-rich lipoproteins. These observations suggest that the difference in distribution of apoE3 variant in plasma lipoproteins between the proband and her brother (combined with its reduced affinity for the LDL receptor) may provide key insights into the pathogenesis of hypertriglyceridemia.


Asunto(s)
Apolipoproteínas E/genética , Variación Genética , Hipertrigliceridemia/genética , Adulto , Apolipoproteínas E/sangre , Apolipoproteínas E/química , Cromatografía de Afinidad , Electroforesis en Gel de Agar , Electroforesis en Gel Bidimensional , Electroforesis en Gel de Poliacrilamida , Femenino , Humanos , Focalización Isoeléctrica , Punto Isoeléctrico , Masculino , Fenotipo , Receptores de LDL/metabolismo
13.
Ann Clin Lab Sci ; 27(5): 351-7, 1997.
Artículo en Inglés | MEDLINE | ID: mdl-9303174

RESUMEN

Plasma lipids and apolipoproteins were determined in 19 children with acute lymphoblastic leukemia (ALL) or malignant lymphoma (ML) who were treated by L-asparaginase with prednisolone and vincristine. Extreme hypertriglyceridemia, i.e., over 10,000 mg/l of the maximum serum triglyceride concentration, was induced in 8 patients; these concentrations were not over 10,000 mg/l in the remaining 11 patients. The possibility was raised that the apolipoprotein E (apoE) isoform apoE4 (epsilon 4) participated in the induction of extreme hypertriglyceridemia, since the frequency of the apoE4/E3 phenotype in the patients with extreme hypertriglyceridemia was higher compared to those in the patients without extreme hypertriglyceridemia and control subjects (n = 248). The acute and severe hypertriglyceridemia was induced at 8 to 14 days after the end of the L-asparaginase therapy, with an earlier remarkable increase in the apoCIII/apoCII ratio and an extreme decrease of fibrinogen concentrations (a marker of the protein productivity of the liver). It is well known that apoCII and apoCIII have possible functions as an activator and an inhibitor of lipoprotein lipase (LPL), respectively. The extreme increase in the apoCIII/apoCII ratio could be one of the reasons for the accumulation of triglyceride-rich lipoproteins in plasma.


Asunto(s)
Antineoplásicos/efectos adversos , Asparaginasa/efectos adversos , Hipertrigliceridemia/inducido químicamente , Linfoma/tratamiento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamiento farmacológico , Adolescente , Antineoplásicos/administración & dosificación , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Apolipoproteínas/sangre , Apolipoproteínas C/sangre , Apolipoproteínas E/sangre , Asparaginasa/administración & dosificación , Electroforesis de las Proteínas Sanguíneas , Niño , Preescolar , Colesterol/sangre , Electroforesis en Gel de Agar , Fibrinógeno/análisis , Fibrinógeno/metabolismo , Humanos , Lactante , Lipoproteínas/sangre , Prednisolona/administración & dosificación , Triglicéridos/sangre , Vincristina/administración & dosificación
14.
Jpn J Infect Dis ; 54(1): 17-22, 2001 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-11326124

RESUMEN

In July, 1999, an outbreak of vancomycin-resistant Enterococcus faecalis (VREF) with the vanB genotype occurred for the first time in Japan at Hokushin General Hospital, Nakano City, Nagano Prefecture. Four VREF strains were isolated from the clinical specimens of four inpatients, and 16 VREF strains were isolated by the screening of asymptomatic carriers and by surveillance of the hospital environment. All of the isolates possessed vanB genes. In a pulsed-field gel electrophoresis analysis, 19 out of 20 VREF isolates exhibited the indistinguishable restriction endonuclease digestion patterns of the chromosomal DNA. Additional investigation by Southern hybridization using the vanB probe implied that the vanB gene of these 19 isolates was encoded on a 110-kb plasmid. These findings indicate that the outbreak was principally caused by a single clone. The restriction endonuclease digestion patterns of the remaining single isolate was different from those of the other isolates. The vanB gene was encoded on the chromosome.


Asunto(s)
Proteínas Bacterianas/genética , Brotes de Enfermedades , Enterococcus faecalis/efectos de los fármacos , Infecciones por Bacterias Grampositivas/epidemiología , Resistencia a la Vancomicina , Anciano , Anciano de 80 o más Años , Southern Blotting , Enzimas de Restricción del ADN , Farmacorresistencia Microbiana , Electroforesis en Gel de Campo Pulsado , Enterococcus faecalis/genética , Enterococcus faecalis/aislamiento & purificación , Femenino , Genotipo , Infecciones por Bacterias Grampositivas/microbiología , Humanos , Japón/epidemiología , Plásmidos
15.
Rinsho Byori ; 48(11): 1006-13, 2000 Nov.
Artículo en Japonés | MEDLINE | ID: mdl-11132553

RESUMEN

It is beyond doubt that clinical examination is one of the essential issues in(living-related) liver transplantation as well as other clinical cases. To support liver transplantation, our laboratory has prepared efficient systems and has been trying to improve these systems as follows. 1) We need to construct a system to be able to comply with clinical requests. Thus, we need to know what kinds of laboratory examinations are likely to be required before, during, and after transplant surgery. In general, common laboratory examinations are sufficient to support liver transplantation. The most important thing is whether all the clinical examinations required can be assayed anytime. However, only general biochemical tests and complete blood count are provided outside standard laboratory hours by the medical technologist on duty because of staffing limitation and differences in each specialty. Therefore, we recently introduced a 24-hour on-call system in addition to the system described above. Actually, five persons in charge from each of the five groups divided by each specialty carry the pocket-bells in turns. 2) We supply a report to support the diagnosis and treatment. The report should include opinions and suggestions. A supplementary examination would be recommended if considered necessary. 3) To supply effective comments, we must improve our abilities to understand pathologic findings obtained from laboratory data. Especially, timely biopsy for the diagnosis of rejection depends on a proper interpretation of laboratory tests. Therefore, we need to investigate past cases after liver transplantation using statistical estimations and advanced examinations.


Asunto(s)
Técnicas de Laboratorio Clínico , Trasplante de Hígado , Atención Perioperativa , Servicios Médicos de Urgencia , Humanos , Laboratorios de Hospital , Grupo de Atención al Paciente
16.
Rinsho Byori ; 47(11): 997-1004, 1999 Nov.
Artículo en Japonés | MEDLINE | ID: mdl-10590676

RESUMEN

Genetic technology is finding active application today in the field of clinical laboratory medicine. Genetic examinations are divided into following three main classes: 1) examination for infectious disease according to the detection of the gene derived from bacteria or viruses, 2) examination for inherited disease according to molecular analysis of the genetic variation, 3) examination for oncogene according to molecular analysis of genetic abnormalities. At present, the main genetic examination in a large number of laboratories is for infectious disease because of its relatively simplified technique and high demand. The division of genetics is not a new independent section of clinical laboratory, but rather an ultramodern and powerful tool for existing divisions, such as biochemistry, serology, hematology, microbiology, and pathology. Genetic technology quickly provides results with high sensitivity and reliability, and plays a role at the core of the clinical laboratory. We should remember that the genetic technology is a great present given to clinical laboratories, however, it will eventually change into only one of the routine examinations according to the method of used. Examinations utilized in the clinical laboratory must be well established and standardized. Genetic examinations are no exception to that rule. These tests require a remarkably high precision since the results have an extraordinarily important meaning. There are more than 8,000 inherited diseases for instance. It is difficult to cover all examinations for those 8,000 in one laboratory. We need a network of laboratories that possess a genetic division, so that the examinations for as many inherited disease as possible can be comprehensively offered.


Asunto(s)
Técnicas de Laboratorio Clínico , Enfermedades Genéticas Congénitas/diagnóstico , Técnicas Genéticas , Neoplasias/diagnóstico , Oncogenes , Virosis/diagnóstico , ADN Viral/análisis , Humanos
17.
Rinsho Byori ; 46(7): 689-94, 1998 Jul.
Artículo en Japonés | MEDLINE | ID: mdl-9721537

RESUMEN

To make diagnosis arteriosclerosis directly by biochemical markers is not easy, but to identify risk factors by biochemical markers is useful. Lipoprotein disorder is one such risk factor. Low density lipoproteins (LDL), remnants and small LDL were high risks of coronary disease in Japanese. Moreover, those incidences were significantly higher in diabetes mellitus, especially with nephropathy, and latter two lipoproteins frequently coexisted. Oxidizability of small LDL was the highest among LDLs, indicating that small LDL promotes atherosclerosis by forming oxidized lipids, which enhance complicated lesion of atherosclerosis. The mechanism by which the remnant is retained remains unknown. We measured LPL mass in preheparin serum. Preheparin LPL mass was negatively correlated with triglyceride, and positively correlated with high density lipoprotein cholesterol. Further more, preheparin LPL mass was lower in remnant-positive persons, indicating that preheparin LPL mass might be involved in remnant clearance. Understanding the role and catabolism of LPL itself requires further study.


Asunto(s)
Arteriosclerosis/diagnóstico , Hiperlipidemias/sangre , LDL-Colesterol/sangre , Enfermedad Coronaria/sangre , Humanos , Lipoproteína Lipasa/metabolismo , Lipoproteínas LDL/sangre , Factores de Riesgo
18.
Rinsho Byori ; 42(11): 1172-6, 1994 Nov.
Artículo en Japonés | MEDLINE | ID: mdl-7844889

RESUMEN

We examined a patient with hyper-cholesterolemia with a high level of HDL2 and LDL-cholesterol in serum. The metabolism of lipoproteins in this case was different from that in well-known hyper-high density lipoproteinemia or hyper-low density lipoproteinemia, because the patient had normal levels of cholesterolester transfer protein, lipoprotein lipase and hepatic triglyceride lipase activity. This study describes the characterization of LDL obtained from the patient's serum. LDL from the patient was separated by ultracentrifuge, and analyzed by gradient PAGE. The molecular weight of two LDL from the patient have been estimated to be approximately 1250 and 1450kDa by polyacrylamide gel electrophoresis, and were larger than those of normal individuals and patients with typical hypercholesterolemia (approximately 1150kDa in molecular weight). The LDL from the patient was separated into three fractions by HPLC, and their lipid composition was not significantly different from that of normal LDL. The high level and large size of LDL from the patient may be caused by a reduction in the transfer of cholesterol from LDL to HDL2, or an equilibrium of cholesterol with the increased HDL2.


Asunto(s)
Proteínas Portadoras/sangre , Hipercolesterolemia/sangre , Lipoproteínas HDL/sangre , Lipoproteínas LDL/sangre , Anciano , Cromatografía Líquida de Alta Presión , Electroforesis en Gel de Poliacrilamida , Femenino , Humanos , Lipoproteínas LDL/química , Peso Molecular
19.
Rinsho Byori ; 46(6): 605-10, 1998 Jun.
Artículo en Japonés | MEDLINE | ID: mdl-9691771

RESUMEN

Platelet aggregation, induced by agonist-mediated activation of membrane glycoprotein (GP) IIb/IIIa, and binding of fibrinogen to GPIIb/IIIa, is commonly analyzed using an aggregometer in the clinical laboratories. However, this method has a limitation to get precise results on the samples with small number of platelet (less than 100,000/1) or hyperlipidemia. Recently, flow cytometry has been used to evaluate platelet function due to the detection of fibrinogen binding to activated platelets using fluorescence labeled fibrinogen or anti-fibrinogen antibody. However, the appropriate rule for evaluation of the results has not been established yet. We converted a ratio of fibrinogen binding platelets to a velocity per unit concentration of ADP as follows: a difference of two ratios of fibrinogen binding platelets on neighboring two ADP concentrations was divided by a difference of ADP concentrations. It was considered to be a mean velocity between the two ADP concentrations. We adopted the range of ADP concentration, which gave the maximum velocity, as an index of platelet activation. If the peak of maximum velocity move toward lower or higher ADP concentration, it means hyper- or hypoactivation of the platelets, respectively. The objectivity of this method may make it a useful technique for clinical examination of platelet function.


Asunto(s)
Adenosina Difosfato/sangre , Plaquetas/metabolismo , Fibrinógeno/metabolismo , Citometría de Flujo , Activación Plaquetaria/fisiología , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Valores de Referencia , Reproducibilidad de los Resultados
20.
Rinsho Byori ; 45(12): 1172-6, 1997 Dec.
Artículo en Japonés | MEDLINE | ID: mdl-9437899

RESUMEN

Distribution of apolipoprotein(apo) E4 and E3 in lipoproteins of serum with apoE4/E3 phenotype was analyzed. ApoE was eluted in two major peaks by gel chromatography; peak 1 and 2 corresponding to very- and intermediate-low density lipoprotein (VLDL + IDL) and high density lipoprotein2 (HDL2), respectively. ApoE in peak 1 (VLDL + IDL) consisted of monomers of 34 kDa, complexes with a high molecular weight (apoEs) of 100 kDa and with a small amount of apoE-AII complexes weighing 43 kDa. In contrast, apoE in peak 2 (HDL2) was composed mainly of apoE-AII complexes and apoEs complexes, and a small amount of monomers. Both apoE3 and E4 isoforms were detected in these peaks; E4 was more predominant in peak 1 while E3 was more predominant in peak 2. These findings suggest that different distributions of apoE3 and E4 in lipoprotein particles.


Asunto(s)
Apolipoproteína A-II/sangre , Apolipoproteínas E/sangre , Lipoproteínas/sangre , Apolipoproteína E3 , Apolipoproteínas E/genética , Humanos , Focalización Isoeléctrica , Fenotipo
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