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Biomedical subjects

K Kuzushima

Publications and source records attributed to K Kuzushima.

At least 19 recordsLinked to original sources
PubMed1998

Polymerase chain reaction-proved herpes simplex encephalitis in children.

OBJECTIVE: To investigate the clinical features in PCR-proved herpes simplex encephalitis (HSE) in children, excluding neonates. METHODS: We studied the clinical manifestations and laboratory findings of 24 children in whom the diagnosis of herpes infection was confirmed by the PCR assay and compared them with those of 38 children with central nervous system infections other than HSE. RESULTS: There were no significant differences between groups in the percentage with fever or convulsions, the initial neurologic symptoms or the level of consciousness. Analysis of cerebrospinal fluid showed no significant differences in the cell count or concentration of protein and glucose. Computerized tomography of the brain identified localized abnormalities in 18 (75%) of the 24 HSE patients and in 10 (31%) of the 32 non-HSE patients (P = 0.001). Periodic lateralized epileptiform discharges, abnormal findings on electroencephalography, were detected in 8 (36%) of 22 HSE patients and in none of the non-HSE patients (P = 0.0001). The rates of moderate to severe morbidity and death were significantly higher in the HSE patients than in the non-HSE patients. Of the 9 HSE patients with a Glasgow Coma Scale score > or = 11, all patients recovered completely. HSE patients younger than 3 years of age were more likely to develop severe sequelae or to die of the disorder than older patients (P = 0.02). CONCLUSIONS: There were no specific clinical characteristics of HSE patients. The results of electroencephalography and computerized tomography were helpful, but not confirmatory, in diagnosing HSE. The Glasgow Coma Scale score and age significantly influenced the mortality and morbidity in the HSE patients.

Adolescent↗
PubMed1997

[Adoptive immune therapy using EBV-specific CTL].

The Epstein-Barr virus (EBV) has been implicated in the etiology of many lymphoid and other malignancies. Understanding of the immune control of the virus appears to be very important to establish strategy to overcome those disease. Recent advance of T cell biology and the culture technic have made it possible to apply adoptive immune transfer of EBV-specific cytotoxic T lymphocytes (CTL) in clinical settings. This review summarizes immunotherapy using EBV-specific CTL for the treatment of EBV-associated lymphoproliferative disorders in patients receiving bone marrow transplantation and for a patient with severe chronic active EBV infection. Its future application is discussed.

Herpesviridae Infections↗
PubMed1996

Establishment of anti-Epstein-Barr virus (EBV) cellular immunity by adoptive transfer of virus-specific cytotoxic T lymphocytes from an HLA-matched sibling to a patient with severe chronic active EBV infection.

We describe an experience of a specific immune transfer treatment in a patient with chronic active EBV infection. The patient had low anti-EBV T cell-mediated cytotoxic activity in his peripheral blood mononuclear cells (PBMC), which may have been the primary cause of the disease. An EBV-specific cytotoxic T lymphocyte (CTL) line was established from PBMC obtained from the patient's sister whose human leucocyte antigens (HLA) are identical to patient's. The patient received three courses of intravenously administered CTL at 3-week intervals. The number of the cells was increased with each course of treatment. After infusion of the T cell line, anti-EBV CTL activity was detected in the patient's PBMC. CTL activity increased markedly after the second course of immune transfer therapy. The amount of EBV DNA in the patient's plasma showed transient but repeated decreases. Serum levels of tumour necrosis factor-alpha (TNF-alpha), which had elevated before treatment, began to decrease after initiation of treatment. No adverse effects were directly associated with CTL infusions. Despite having previously received a pneumococcal vaccine and prophylactic antibiotics, the patient died of infection caused by Streptococcus pneumoniae bacteraemia 27 days after the third infusion. Although the long-term efficacy and safety of this therapy remains to be established, our findings suggest that adoptive transfer of CTL specific for EBV obtained from an HLA-matched donor might be a promising treatment for patients with chronic active EBV infection.

Child↗
PubMed1995

The role of self peptides in the allogeneic cross-reactivity of CTLs.

This study presents data relevant to understanding the molecular and structural basis for the cross-reactivity of many CTLs on multiple MHC targets. Five anti-H-2Kb alloreactive CTL clones derived from B6.C-H-2bm1 (bm1), B6.C-H-2bm8 (bm8), and B6.C-H-2bm11 (bm11) mice and a Sendai virus-specific H-2Kb-restricted CTL clone were studied. Self peptides extracted from Kb molecules were fractionated by HPLC and tested for their ability to be recognized on RMA/s (H-2b) cells by those clones. For each alloreactive clone, a single dominant peptide peak was found to sensitize target cells. In addition to recognizing peptides presented by the Kb molecule, the five alloreactive clones and the one Sendai virus-specific clone all showed cross-reactivities on a panel of Kbm mutant cells in a peptide-dependent manner. Two CTL clones, one alloreactive and one virus specific, cross-recognized Kbm targets by each responding to a unique self peptide in the context of the mutant MHC molecules. Our data underscore the prevalent idea that TCR-alpha beta have an inherent structural capability to react with several peptide/MHC structural patterns other than the original peptide/MHC pattern that might have been used to select that TCR. The high incidence of cross-reactivity seems to reflect a feature of the mechanism of positive selection in the thymus and the need for T cells in the repertoire to have an expanded capability for responding to a wide variety of foreign Ags.

Animals↗
PubMed1995

Intact antigen presentation for Epstein-Barr virus (EBV)-specific CTL by a lymphoblastoid cell line established from a patient with severe chronic active EBV infection.

Severe chronic active Epstein-Barr virus (EBV) infection is a lymphoproliferative disease characterized by extremely high antibody titers to EBV, fever, lymphadenopathy, hepatosplenomegaly, and pancytopenia, without any prior immunological abnormality. A spontaneous lymphoblastoid cell line was established from a 4-year-old boy with severe chronic active EBV infection. Immunofluorescence and Western blotting analyses showed that the cell line was of B cell origin and expressed Epstein-Barr nuclear antigens 1, 2 3a, 3b and 3c, and latent membrane protein 1, which are reported to be targets for EBV-specific cytotoxic T lymphocytes (CTL). The cytotoxicity of peripheral blood mononuclear cells derived from the patient and his HLA-identical sister was assayed against the cell line. The cell line was recognized and killed by anti-EBV CTL derived from the HLA-identical sister, but the patient's peripheral blood mononuclear cells had no cytotoxicity. We conclude that antigen presentation in the EBV-infected cells from the patient is intact and sufficient for generation of an EBV-specific CTL response. These observations suggest that severe chronic active EBV infection may not be caused by impaired EBV-antigen presentation of the infected cells but by impaired cellular immune responses to the virus. Our results also suggest the therapeutic possibility that this disease may be treated by adoptive transfer of EBV-specific CTL or bone marrow transplantation from an HLA-matched donor whose immune response to EBV is intact.

Animals↗
PubMed1995

Detection and quantification of virus DNA in plasma of patients with Epstein-Barr virus-associated diseases.

Epstein-Barr virus (EBV) causes various diseases, such as infectious mononucleosis (IM), fatal IM, EBV-associated hemophagocytic syndrome (EBVAHS), and chronic active EBV infection (CAEBV). In the present study, cell-free EBV DNA was detected in the plasma of patients with EBV-associated diseases by PCR assay. The patients included 20 patients with IM, 2 patients with fatal IM, 4 patients with EBVAHS, 4 patients with CAEBV, and 38 healthy children (20 EBV seropositive and 18 EBV seronegative). In patients with IM, plasma samples were positive for EBV DNA in all patients (100%) in the acute phase and in 44% of the patients in the convalescent phase, but plasma samples from the 38 healthy control children were negative (0%) for EBV DNA. Quantitative PCR assay revealed that plasma from patients with IM contained the highest amount of virus DNA within 7 days following the onset of disease (mean, 6 x 10(4) copies per ml). The EBV DNA concentration decreased thereafter as the patients recovered. Plasma from patients with fatal IM contained more than 100 times more copies of EBV DNA (3 x 10(7) copies per ml) than plasma from patients with IM. Plasma from patients with the acute phase of EBVAHS contained 10 times more copies of EBV DNA (5 x 10(5) copies per ml) than plasma from IM, and then patients with the number of copies decreased similarly in both groups of patients in the convalescent phase (2 x 10(4) copies per ml). The amount of virus DNA in patients with CAEBV (6 x 10(4) copies per ml) was similar to that noted in patients with IM; however, it became higher (1 x 10(6) copies per ml) when the patients' clinical status deteriorated. These data suggest that the presence of cell-free EBV DNA in plasma is a common phenomenon in patients with EBV-associated diseases. The concentration of EBV DNA in plasma seems to be higher in patients with the more severe clinical categories of EBV diseases.

Acute Disease↗
PubMed1994

Characterization of an incompletely assembled major histocompatibility class I molecule (H-2Kb) associated with unusually long peptides: implications for antigen processing and presentation.

We have identified two forms of a major histocompatibility complex (MHC) class I molecule, H-2Kb, distinguishable by specific antibodies through a study of a genetically engineered mouse cell line that overexpresses these molecules. One form, a complex associated with beta 2-microglobulin (native, beta 2m+ class I), is detectable by conformation-dependent antibodies. The other form, which remains after preclearing cell lysates of native class I, is only poorly, if at all, associated with beta 2-microglobulin (beta 2m- class I) and is detectable by an antiserum against the cytoplasmic tail region of H-2K molecules. Both forms are also present in normal cell lines. The affinity-purified native class I molecules bind short peptides (8 or 9 residues) and assemble tightly with beta 2-microglobulin. In striking contrast, the beta 2m- class I molecules bind peptides that are longer (> 15 residues) than those bound to native class I molecules. This finding is consistent with the recent evidence that peptides longer than 8-10 amino acid residues are transported into the endoplasmic reticulum and suggests the possibility of a control step for peptide presentation by MHC in which the incompletely processed peptides bind to the heavy chain and a selected fraction undergoes final processing and presentation on the cell surface.

Amino Acid Sequence↗
PubMed1993

Patterns of hepatitis B virus DNA integration in liver tissue of children with chronic infections.

Although an integration of hepatitis B virus (HBV) DNA in children with chronic HBV infection has been documented at early phases of the disease, the incidence of this process is not known. Therefore we examined nine liver DNA specimens from chronic HBV carriers ages 5-14 years and one sample from a neonate delivered of a carrier mother, in order to determine the HBV DNA patterns of these patients at different ages and phases of chronic infection. The integrated HBV DNA was detected by Southern blot hybridization and analyzed by molecular cloning. Southern blot showed a smear pattern of HBV DNA integration in four of six chronic hepatitis patients (ages 5-14), as well as in one asymptomatic carrier (age 12). Multiple and random integrations occurred during chronic infections in childhood. The neonate did not, however, show signs of any integrations, suggesting that integration starts after HBV multiplication. A band pattern that suggested clonal growth of integrated liver cells was found in a chronic active hepatitis patient (age 9) and in one of two hepatocellular carcinoma patients (age 11). Molecular cloning in two cases with chronic active hepatitis showed that the HBV genome structure was preserved in five of six HBV DNA inserts. Our findings confirm that HBV DNA integration can occur at early stages of chronic HBV infection. In Japanese children, the process of integration seems to be common regardless of HBeAg/anti-HBe status.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1992

Quantitation of cytomegalovirus DNA in lung tissue of bone marrow transplant recipients.

Five bone marrow transplant recipients who died of respiratory failure were retrospectively analyzed with polymerase chain reaction (PCR) assay for pulmonary cytomegalovirus (CMV) infection. Two patients had CMV interstitial pneumonitis according to the virus isolation and the histologic and immunofluorescent examinations of the lungs, while the other three patients had non-CMV diseases (ie, idiopathic interstitial pneumonitis, pulmonary aspergillosis, or Streptococcus mitis septicemia). Cytomegalovirus DNA was amplified from the postmortem lung tissue with PCR. The PCR assay showed apparent PCR signals specific to CMV DNA in the two patients with CMV pneumonitis. In contrast, CMV DNA was hardly detectable or undetectable in the three patients without CMV disease. With quantitative PCR assay the initial CMV copy number in the lung tissue of the two patients with CMV pneumonitis was more than 10(4) copies/micrograms DNA and was over 1,000-fold more than that of the three patients without CMV pneumonitis. These results show that quantitative PCR assay could be useful as a diagnostic measure for pulmonary CMV infection.

Adolescent↗
PubMed1992

Prophylactic oral acyclovir in outbreaks of primary herpes simplex virus type 1 infection in a closed community.

Oral acyclovir was given prophylactically to 37 children in the early stages of three outbreaks of herpes simplex virus type 1 (HSV-1) infection and the results were compared with those in untreated control subjects in two other outbreaks. The rates of seroconversion to HSV were significantly reduced in children treated with acyclovir compared with control subjects (91% vs 27%, P less than .001). The incidence of symptomatic disease was also significantly reduced (82% vs 0%, P less than .001). In some children receiving prophylactic acyclovir, anti-HSV antibody titers did not rise despite the presence of replicative HSV on throat swabs just before the start of treatment. Restriction endonuclease analysis of isolated HSV-DNA confirmed that one strain was responsible for the five outbreaks. No resistance to acyclovir was detected during the study, and no adverse effects of treatment were noted. In conclusion, short-term prophylactic acyclovir may limit the spread and reduce clinical manifestations of HSV infections in closed communities, although this use should be restricted to communities where severe symptoms are observed.

Acyclovir↗
PubMed1992

Relapse of herpes simplex encephalitis in children.

The polymerase chain reaction method was used to diagnose herpes simplex encephalitis in children. Initial samples of cerebrospinal fluid from 15 patients with herpes simplex encephalitis were all positive for the herpes simplex virus DNA by polymerase chain reaction assay. In terms of early diagnosis, polymerase chain reaction assay became positive significantly earlier than the detection of intrathecally produced anti-herpes simplex virus antibody using the enzyme-linked immunosorbent assay (4.4 vs 8.9 days after onset; P less than .01). Serial examinations showed that the presence of virus DNA in cerebrospinal fluid continued for 3 to 18 days after the neurologic onset (mean 10.1 days). Four of the 15 patients had a relapse of encephalitis after completing acyclovir therapy. The mean duration of initial acyclovir therapy in the recurrent group was significantly shorter than that in the nonrecurrent group. In recurring cases, herpes simplex virus DNA reappeared temporarily in the cerebrospinal fluid of two patients. These results show that polymerase chain reaction assay is a useful diagnostic tool for the early and noninvasive diagnosis of herpes simplex encephalitis in children. Results also suggest that a comparatively short duration of acyclovir therapy may be related to a relapse of herpes simplex encephalitis in some children.

Acyclovir↗
PubMed1991

Detection of viral DNA in neonatal herpes simplex virus infections: frequent and prolonged presence in serum and cerebrospinal fluid.

Polymerase chain reaction (PCR) assay was used to detect herpes simplex virus (HSV) DNA in mouth, skin, sera, or cerebrospinal fluid (CSF) from seven neonates with HSV infection. In a culture-negative patient, the diagnosis was confirmed by detection of HSV DNA. Serial examinations revealed that HSV DNA remained in the serum and/or CSF from several patients for 1-2 weeks after the beginning of treatment. Next, the results of PCR assay in neonatal HSV infections were compared with those in older children with herpes simplex encephalitis (HSE). HSV DNA was detected in CSF from four neonates with central nervous system involvement and in CSF from all nine children with HSE. Sera were positive for HSV DNA in five of seven neonates, including two cases of localized infections, but in none of the children with HSE. These results suggest that HSV may be spread principally via viremia in neonates. PCR assay could be useful for the confirmative diagnosis of neonatal HSV infections, especially in culture-negative cases.

Base Sequence↗
PubMed1991

Clinical manifestations of primary herpes simplex virus type 1 infection in a closed community.

The clinical features and the molecular epidemiology of primary herpes simplex virus type 1 (HSV-1) infection among children younger than 3 years of age were investigated in day-care nursery. Serial sera were assayed for anti-HSV-1 glycoprotein B antibody by enzyme-linked immunosorbent assay. Serologic examinations revealed 55 cases of primary HSV infection during the observation period. Fifty-one of them (93%) had typical herpetic gingivostomatitis, showing a high rate of clinically overt infection. Four outbreaks of herpetic gingivostomatitis were observed during the observation period. Forty-one children were infected with HSV-1 in the outbreaks. The rates of infection in the susceptible children were 81%, 73%, 78%, and 100%, respectively, in the four outbreaks. Restriction endonuclease analysis of DNA of isolated HSV revealed that only one strain of HSV-1 had been transmitted among children for a long period.

Age Factors↗
PubMed1990

Inhibitory effect of herpes simplex virus infection to target cells on recognition of minor histocompatibility antigens by cytotoxic T lymphocytes.

Target cell lysis by CTL specific for minor histocompatibility Ag (minor HA), which were generated in (C3H/He x BALB/c)F1 mice immunized with A/J mouse spleen cells, was dramatically reduced by infection of HSV to Neuro-2a (A/J mouse origin) cells as target. The reduction was apparent at 5 h after infection of HSV to target cells, when many viral proteins were produced in the cells. Conversely, MHC-restricted HSV-specific CTL-mediated cell lysis increased time dependently. Using an RNA virus, vesicular stomatitis virus, significant reduction of minor specific CTL-mediated target cell lysis was also found. During the time when this reduction of target cell lysis by HSV occurred, the surface expression of class I H-2Dd molecules was maintained, and anti-H-2a allo-MHC-specific CTL lysed HSV-infected Neuro-2a cells as strongly as uninfected Neuro-2a cells. When HSV-infected or uninfected Neuro-2a cells were treated with Brefeldin A that selectively blocks transportation of newly synthesized proteins out of endoplasmic reticulum, both HSV- and minor HA-specific CTL-mediated cell lyses were blocked. These observations demonstrated that minor HA are continuously synthesized and associated with class I molecules at pre-Golgi and transported via trans Golgi system with quick turnover, and that newly synthesized HSV Ag, which are also associated with class I molecules and transported via the same system, should take the place of intrinsic minor HA and be presented on the surface of the cells to be recognized by MHC-restricted CTL.

Animals↗
PubMed1990

Human monoclonal antibodies to human cytomegalovirus derived from peripheral blood lymphocytes of healthy adults.

Immunoglobulin G class human monoclonal antibodies to human cytomegalovirus (HCMV) were produced by the fusion of Epstein-Barr virus-transformed B cells and a murine myeloma cell line. The B cells were derived from the peripheral blood lymphocytes of healthy adult volunteers. Four hybridomas producing HCMV-specific monoclonal antibodies were established and each of four antibodies immunoprecipitated an HCMV-specific protein with a molecular weight of 68 kDa. However, the antibodies differed in some of their properties as characterized by indirect immunofluorescence assay and immunoblotting studies, due to the detection of different epitopes on the reacting antigen. None of the four antibodies had any virus neutralizing activity.

Adult↗
PubMed1990

Human cytomegalovirus infection during childhood: detection of viral DNA in peripheral blood by means of polymerase chain reaction.

Polymerase chain reaction (PCR) technique was applied to detect cytomegalovirus (CMV) DNA. Two pairs of synthetic oligonucleotide primers were used to amplify DNA from the immediate early 1 and the late antigen genes of CMV, respectively. Either primer sets could detect as few as 0.01 plaque-forming unit of CMV strain AD 169 by Southern blot hybridization. Sixteen CMV clinical isolates were examined and all were found to be positive by the both primer sets. The PCR was used to detect CMV DNA in peripheral blood from six children with elevated anti-CMV antibody titers, who showed abnormal liver-function tests. In three immunocompromised patients, all blood samples were positive for CMV DNA. In three immunocompetent young infants with primary CMV infection, CMV DNA was detected from peripheral blood of one patient during acute phase. Presence of CMV DNA in peripheral blood seemed to be related with the extent of CMV infection, and possibly diagnostic for CMV hepatitis.

Child, Preschool↗