Biomedical subjects

M Takayanagi

Publications and source records attributed to M Takayanagi.

At least 109 records · Page 6Linked to original sources

PubMed1993

Comparison of envelope and precore/core variants of hepatitis B virus (HBV) during chronic HBV infection.

We analyzed entire pre-C/C and pre-S/S coding genes of hepatitis B virus (HBV) in serial serum samples from four chronic HBV carriers with 4-5 years of follow-up. Two patients with chronic active hepatitis became seronegative for HB e antigen (HBeAg), but the hepatitis did not subside, while the other two were persistent asymptomatic carriers with normal aminotransferase values. DNAs amplified by PCR were cloned and sequenced (subtype adr). After HBeAg became negative, one patient had 96-183 bp deletions in 4/6 clones for C gene. In addition, both patients had 129-183 bp deletions in 3/6 and 2/5 clones for pre-S1 gene, respectively. Divergence rate of deduced amino acid for both pre-C/C and pre-S/S regions from the adr subtype was significantly higher in patients with chronic hepatitis than in asymptomatic carriers. Furthermore, the divergence rate for pre-S/S region was usually greater in asymptomatic carriers as well as chronic hepatitis patients compared with that for pre-C/C region. However, no significant difference was found in the rate of amino acid divergence for the entire HBV genes between the serial samples in all patients studied here. These results suggest that active hepatitis induces variation of HBV gene and defective virus is often selected along with the disappearance of HBeAg. In addition, the fact that patients with active liver disease possess greater numbers of mutant clones than asymptomatic carriers suggests that viral mutants are being immune-selected.

Adult↗

PubMed1993

Dynamics of genome change in the E2/NS1 region of hepatitis C virus in vivo.

To substantiate and extend the quasispecies model of hepatitis C virus (HCV), we made a pairwise comparison in the nucleotide and deduced amino acid sequences for multiple recombinant clones of the E2/NS1 region, which derived from each blood sample taken from five patients of subtype II or subtype III infection at different stages. Sequence heterogeneity among the clones was generally high. The heterogeneity, however, changed temporally and appeared to be significantly lowered after interferon therapy. The temporal fluctuation involved selection of particular amino acids at particular positions, which had represented only a minor fraction or had been absent in the previous clones. Evolution of defective viruses was featured by most of the cases, the proportion of which also fluctuated temporally and was extremely high at a certain stage in one of the patients. A comparison was also made among the clones from different stages of each patient and revealed highly divergent clones. An extreme case of subtype III infection showed sequence differences well beyond those among chronologically and geographically different isolates of subtype I or subtype II, which were defined by the same calculation of available sequence data. These features of HCV genome suggest that the virus could circulate as an extremely heterogeneous population including defective viruses and that this heterogeneity lends itself to selection pressures including interferon therapy and host immune response. Our results also indicate an absolute need of population based approach in HCV genetics.

Aged↗

PubMed1993

A pilot study of ribavirin and interferon beta for the treatment of chronic hepatitis C.

BACKGROUND: Chronic hepatitis C is a common and often progressive liver disease for which interferon alfa therapy widely spreads, but the beneficial response is frequently transient. Ribavirin is a nucleoside analog with a broad spectrum of antiviral action, and we investigated the efficacy of it in patients with chronic active hepatitis C. METHODS: We conducted a pilot study of oral ribavirin in patients with chronic active hepatitis C. Twenty-seven patients with hepatitis C virus RNA were randomly assigned to receive either 0.8-1.0 g of ribavirin daily or 3 MU of interferon beta three times weekly or combination of the two for 24 weeks. RESULTS: Ribavirin was tolerated well, and all completed the treatment schedule. Ribavirin decreased aminotransferase levels in all instances, and the mean value at termination decreased to half of the baseline level (P < 0.01), but the enzyme level increased after cessation of therapy in most cases. Ribavirin suppressed amounts of hepatitis C virus RNA in 4 of 9 patients, and 1 became negative during follow-up. Interferon alone (P < 0.05) or with ribavirin (P < 0.01) significantly decreased the viral population, resulting in sustained loss of viremia with normal enzyme levels in 2 of 9 and 3 of 9 patients, respectively, in each therapy during follow-up. CONCLUSIONS: These results indicate that ribavirin has a beneficial effect in some patients with chronic hepatitis C, although the antiviral effect is less than interferon beta. Large-scale trials are needed to determine whether the combination of interferon and ribavirin is of more benefit than interferon alone.

Adult↗

PubMed1993

A 15N-1H nuclear magnetic resonance study on the interaction between isoleucine tRNA and isoleucyl-tRNA synthetase from Escherichia coli.

Imino 15N and 1H resonances of Escherichia coli tRNA(lIle) were observed in the absence and presence of E coli isoleucyl-tRNA synthetase. Upon complex formation of tRNA(lIle) with isoleucyl-tRNA synthetase, some imino 15N-1H resonances disappeared, and some others were significantly broadened and/or shifted in the 1H chemical shift, while the others were observed at the same 15N-1H chemical shifts. It was indicated that the binding of tRNA(lIle) with IleRS affect the following four regions: the anticodon stem, the junction of the acceptor and T stems, the middle of the D stem, and the region where the tertiary base pair connects the T, D, and extra loops. This result is consistent with those of chemical footprinting and site-directed mutagenesis studies. Taken together, these three independent results reveal the recognition mechanism of tRNA(lIle) by IleRS: IleRS recognizes all the identity determinants distributed throughout the tRNA(lIle) molecule, which induces changes in the secondary and tertiary structures of tRNA(lIle).

Base Sequence↗

PubMed1993

Effect of recombinant human transforming growth factor beta 1 on immune responses in patients with chronic hepatitis B.

Studies were undertaken to examine the effect of recombinant human transforming growth factor beta 1 (rTGF-beta 1) on cellular and humoral immune responses of peripheral blood mononuclear cells (PBMC) from patients with chronic hepatitis B. The addition of TGF-beta 1 caused a significant dose-dependent inhibition of hepatitis B (HB) core Ag-stimulated interferon-gamma and antibody to HB core Ag production and proliferation of PBMC from chronic hepatitis patients and HB-immune donors. TGF-beta 1 also induced a significant reduction in pokeweed mitogen-stimulated IgG and IgM production, as well as phytohemagglutinin p-stimulated proliferative response of PBMC. The degree of inhibition of TGF-beta 1 did not differ between antigen-specific and -nonspecific cellular and humoral immune responses, and between control individuals and patients. Pretreatment study with TGF-beta 1 showed that the activities of T cells, B cells and monocytes were similarly inhibited. Further, TGF-beta 1 inhibited activities of HLA class I antigen-matched cytotoxic T cells from patients with chronic hepatitis B for HBV DNA-transfected HepG2 cells in a 51Cr release assay. The results suggest that TGF-beta 1 may play a role in the regulation of antigen-dependent and -independent immune responses in patients with chronic hepatitis B.

Adult↗

PubMed1993

Difference of the hormonal profile at the periovulatory phase in pregnant and nonpregnant cycles of infertile women with unexplained etiology.

Ten women with unexplained infertility, who conceived by intrauterine insemination (IUI) or timed intercourse under controlled ovarian hyperstimulation, were analyzed to investigate the difference of hormonal environment at the periovulatory phase. Forty-two cycles were divided into three groups: the pregnant cycle group, the high luteinizing hormone (LH) group (LH > or = 60 mIU/ml on day -1: the day of human chorionic gonadotropin administration) of nonpregnant cycles and the low LH group (LH < 60 mIU/ml on day -1) of nonpregnant cycles. Progesterone (P4)/estradiol (E2) ratios of the high LH group on day -1 and day 0 were significantly higher than those of each corresponding day in the pregnant cycle. On the other hand, P4/E2 ratio of the low LH group on day +1 was significantly lower. P4/E2 ratio but absolute level of each hormone showed a high indicative value for the estimation of abnormal hormonal environment. It is concluded that the P4/E2 ratio at the periovulatory phase is a good parameter to estimate the efficacy of the method for ovulation induction.

Adult↗

PubMed1993

Molecular cloning and sequence analysis of the cDNA for human mitochondrial short-chain enoyl-CoA hydratase.

Short chain enoyl-CoA hydratase (SCEH) catalyzes the second step of the mitochondrial fatty acid beta-oxidation spiral. We isolated cDNA clones for human SCEH to facilitate investigation of the enzyme structure of the gene and to examine the genetic background of Reye's syndrome and sudden infant death. Oligo(dT)-primed and random primed human liver cDNA libraries in lambda gt11 were screened using the entire sequence of the rat SCEH cDNA as a probe. Three positive clones covered the full-length cDNA sequence with an open reading frame encoding a precursor polypeptide of 290 amino acid residues, and deduced relative molecular mass (31,280) with a putative N-terminal presequence of 29 residues, a 5'-untranslated sequence of 21 bp and a 3'-untranslated sequence of 391 bp. Comparison with the rat SCEH cDNA showed that the deduced amino acid sequence of the human SCEH precursor is 84% identical to that of the rat enzyme precursor. Northern blot analysis gave a single mRNA species of 1.6 kb in the human liver, fibroblast and muscle.

Amino Acid Sequence↗

PubMed1993

Pregnancy complicated by the May-Hegglin anomaly.

May-Hegglin anomaly is a prepotent autosomal hereditary disease characterized by giant thrombocytes, thrombocytopenia and continuous appearance of inclusion bodies (Döhle like corpuscle) in the cytoplasm of granulocytes. In this report we report our experience of a case whose pregnancy was complicated by this disease. This anomaly is rare and there are only 15 families reported in this country. The only clinical symptom is the tendency to bleed, but since this is not rare, it may be necessary to consider examination of peripheral blood in every case of thrombocytopenia. If this disease complicates pregnancy, it is important to plan the mode of delivery to consider both the maternal and fetal prognosis.

Adult↗

PubMed1993

Tertiary structural analysis of Escherichia coli lysine tRNA.

It has been suggested that Escherichia coli lysine tRNA (E. coli tRNA(Lys)) has an unusual anticodon loop structure, judging from the abnormal CD pattern derived from 5-methylaminomethyl-2-thiouridine (mnm5s2U) located at the first position of the anticodon of the tRNA. Here, the structure of E. coli tRNA(Lys) was analysed by chemical modification and nuclear magnetic resonance (NMR) of the imino protons. The chemical probing approach showed that the anticodon arm region forms the ordinary stem-and-loop structure as expected from the standard clover-leaf structure, because N7 of G30, N3 of C40, N7 of A41 and N7 of A42 were protected under a native condition. By using the heteronuclear 15N-1H single quantum coherence (HSQC) spectra of 15N-labeled E.coli tRNA(Lys) and the analyses of the nuclear Overhauser effects (NOE) between the imino protons of non labeled E.coli tRNA(Lys), the imino protons in the acceptor stem was assigned, which indicates that the acceptor stem exists in an usual form. The tertiary base-base interactions was also detected. Thus, E.coli tRNA(Lys) seems to maintain a similar structure to the ordinary L-shaped tertiary structure, as a whole. The fine structure of the anticodon loop is now being analyzed.

Anticodon↗

PubMed1993

Relative immunogenicity of hepatitis B virus-encoded antigens as targets for cytotoxic T-cell response.

To analyse the immunological mechanism of hepatocellular injury in hepatitis B virus (HBV) infection, the immunoreactivity of HBV-encoded antigens as a target for cytotoxic T lymphocyte (CTL) response was examined using recombinant vaccinia virus (RVV) expressing surface protein (S), precore/core protein (PC), and core protein (C) of HBV. C3H/He mice (H-2k) were inoculated with each RVV. Their spleen cells were then harvested and stimulated in vitro with the histocompatible transfectant, which stably expressed hepatitis B surface antigen (HBsAg), hepatitis B e antigen (HBeAg), and hepatitis B core antigen (HBcAg), and used as effectors. As the targets, L cells (H-2k) infected with individual RVV were used. Cytotoxic test was performed with various combinations and ratios of effectors and targets. The reactivity of PC-primed effectors against PC-expressing targets was greatest with 71.4% specific lysis on average at an effector/target ratio of 12.5:1 among all the combinations. C-primed effectors against C-expressing target also revealed rather high cytotoxicity (specific lysis, 40.6% at an E/T ratio of 12.5:1). Furthermore, PC-primed and C-primed effectors showed a cross-reactivity to the targets expressing other nucleocapsid antigen, respectively. S-primed effectors showed less lytic activity against S-expressing targets (specific lysis, 18.4% at an E/T ratio of 12.5:1). The CTL responses were blocked by anti-CD8 and anti-major histocompatibility complex (MHC) class I antibodies, but not by anti-CD4 or anti-MHC class II. These findings suggest that endogenously synthesized nucleocapsid antigen, especially PC, is a dominant target for the MHC class I-restricted CTL in H-2k mice and that this system may work as an efficient model to study immunopathogenesis of HBV infection.

Animals↗

PubMed1993

Pilot study of ribavirin and interferon-beta for chronic hepatitis B.

Ribavirin is a nucleoside analog that inhibits the replication of many DNA and RNA viruses. To evaluate the efficacy of oral ribavirin, we randomly assigned 24 HBeAg-positive patients with chronic active hepatitis to a 12-wk course of treatment with 0.8 to 1.0 gm/ribavirin day, 3 mU interferon-beta three times a week intravenously or a combination of those drugs. Ribavirin, alone and in combination with interferon-beta, decreased hepatitis B virus levels in most patients, and mean serum hepatitis B virus DNA and DNA polymerase levels at the end of treatment were approximately half of baseline levels (p < 0.05). Interferon alone exerted the most inhibitory effect on hepatitis B virus activity (p < 0.01). During ribavirin treatment, changes in serum aminotransferase values varied considerably and the mean values did not change significantly, although interferon alone and the combination of interferon and ribavirin were associated with significant reductions in serum aminotransferase activities. Ribavirin was well tolerated, but we transiently reduced the dosage in two cases because of mild hemolytic anemia, although all patients completed the treatment schedule. The combination of interferon and ribavirin did not appear to result in greater toxicity. During the follow-up period (6 to 9 mo), HBeAg and hepatitis B virus DNA disappeared in one patient treated with ribavirin, in two treated with interferon and in two given the combination. These results indicate that ribavirin suppresses hepatitis B virus replication, although its effect is less than that of interferon, and that it may be useful as adjunctive therapy for chronic hepatitis B.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1993

[Airborne Japanese red cedar allergens studied by the immunoblotting technique--comparison with pollen counts obtained by Durham sampling and effect of weather].

We collected airborne particles with Burkard's sampling tape in Toyama from March to July. According to the immunoblotting technique (Takahashi et al, 1990), the airborne pollen allergens were reacted with anti-Cry j I polyclonal antibody and were stained as blue spots. The areas of these spots were measured with a color image analyzer (CIA) and a densitometer. There were significant correlations between the airborne Cry j I allergen spots (CIA and densitometer) and the pollen counts obtained with a Durham sampler (r = 0.624, 0.555, p < 0.001). The airborne Cry j I allergens however, tended to maintain a higher level than the pollen counts (Durham) in May. We theorized that this discrepancy is caused by cross reactivity with other pollen or crushed cedar pollen particles have allergenicity. There was a negative correlation between the Cry j I spots and rainfall. The pollen counts had a negative correlation with rainfall, and a positive correlation with average wind speed. These results suggest that this method is useful in evaluating fluctuations in airborne Cry j I allergen.

Allergens↗

PubMed1992

Detection of hepatitis C virus by polymerase chain reaction and response to interferon-alpha therapy: relationship to genotypes of hepatitis C virus.

To investigate the relationship between genotypes of hepatitis C virus and response to interferon-alpha therapy, hepatitis C virus RNA was assayed by polymerase chain reaction with three sets of primers and probes in 70 patients with non-A, non-B chronic hepatitis who received interferon-alpha. Twenty-four patients sustained long-term remissions (complete responders). Polymerase chain reaction for 5'-terminal noncoding region detected hepatitis C virus RNA in 94.3% (66 of 70) of the patients. Polymerase chain reaction for nonstructural region 3, in which primers and a probe were synthesized to be identical to hepatitis C virus-J, detected hepatitis C virus RNA in 40 patients. Polymerase chain reaction for nonstructural region 5-in which sequences of primers and a probe were derived from hepatitis C virus-K2, a genotype different from hepatitis C virus-J--detected hepatitis C virus RNA in 17 patients. Only one patient was positive on both nonstructural region 3 and nonstructural region 5 polymerase chain reaction. Nucleotide sequence of clones obtained from 5' terminal noncoding region polymerase chain reaction products of two patients positive on polymerase chain reaction for nonstructural region 3 and negative on polymerase chain reaction for nonstructural region 5 (group 1) corresponded to that of the hepatitis C virus-J group, and those of clones from two patients negative on polymerase chain reaction for nonstructural region 3 and positive on polymerase chain reaction for nonstructural region 5 (group 2) corresponded to that of hepatitis C virus-K2.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1992

Effects of transforming growth factor-beta 1 against the inhibitory action of interferon on DNA synthesis and viral replication in hepatitis B virus DNA-transfected cell.

Studies were undertaken to examine the effects of recombinant human transforming growth factor beta 1 (TGF-beta 1) on DNA synthesis and antiviral actions of interferons (IFNs) in HepG2 cell, a hepatoma cell line, transfected with hepatitis B virus (HBV) DNA. The inhibitory effects of IFN-alpha and -gamma on DNA synthesis of HepG2 cells were enhanced in a dose-dependent manner by a simultaneous addition of TGF-beta. The degree of suppression by the reagents was greater in HBV-nontransfected cells than in transfected cells. Inhibition of DNA synthesis was not due to direct cytotoxic effects of the additives, since the viability of HepG2 cells was comparable in the control and treated cultures as determined by trypan blue exclusion. Treatment of HBV DNA-transfected HepG2 cells with IFNs resulted in decrease in production of HB surface and e antigens, and in the level of HBV DNA, but TGF-beta reversed the IFN-induced antiviral state in HBV DNA-transfected HepG2 cells. TGF-beta had no direct effect on HBV replication. These results indicate that rTGF-beta 1 exerts a differential effect against the inhibitory actions of IFN on DNA synthesis and viral replication in HepG2 cells.

DNA, Viral↗

PubMed1992

Immune response of peripheral blood mononuclear cells to antigenic determinants within hepatitis B core antigen in HB virus-infected man.

Residues 72-146 within hepatitis B core Ag (HBcAg) represent T-cell recognition site in HB-virus-infected man. This study was undertaken to define critical residues involved in the immunogenicity of dominant T-cell determinants of HBcAg. For this purpose, p120-131 and its analog (p120-131 [A] containing alanine substitutions at residues 122 and 125, which were identified as epitopic residues in mice, were synthesized. These peptides and recombinant HBcAg were analyzed for their ability to stimulate peripheral blood mononuclear cells (PBMC) from 25 patients with chronic HBV infection and three patients with acute hepatitis B. PBMC from 18 out of 28 patients showed significantly increased IFN-gamma production and proliferative response in the presence of recombinant HBcAg. Eight patients responded to the two peptides, while 12 patients did not. Four patients responded only to p120-131, and four displayed a response only to p120-131 [A]. The responses to the two peptides were similar among HBeAg-positive and anti-HBe-positive patients, and did not depend on disease activity, except for HBeAg-positive asymptomatic carriers in whom there was no response to any additive. These results indicate that immune responses to p120-131 and its analog were similar in our patient groups. The dominant epitopic residues in this region of HBcAg may differ between man and mouse.

Adult↗

PubMed1992

Identification of a common mutation in Finnish patients with nonketotic hyperglycinemia.

Nonketotic hyperglycinemia (NKH) is an autosomal recessive metabolic disorder caused by the defects in the glycine cleavage system (GCS; EC 2.1.2.10), a multienzyme system that consists of four individual components. NKH is a rare disorder in many countries, but with a very high incidence in northern Finland. To understand the genetic background of this high incidence, we examined the GCS in a typical case of NKH at the molecular level. The activity of P protein, a component of the GCS, was not detected in the lymphoblasts of the patient, while P protein mRNA of a normal size and level was present in the cells. Structural analysis of P protein mRNA from the patient revealed a single nucleotide substitution from G to T in the protein coding region, which resulted in an amino acid alteration from Ser564 to Ile564. No P protein activity was detected when the mutant P protein with this amino acid substitution was expressed in COS 7 cells. The patient was homozygous for this mutation. Furthermore, this mutation was present in 70% (14 of 20) of P protein gene alleles in Finnish patients with NKH, whereas it was not found in 20 alleles of non-Finnish patients. The results suggest that this mutation is responsible for the high incidence of NKH in Finland.

Amino Acid Metabolism, Inborn Errors↗

PubMed1992

Neurotrophic effects of epidermal growth factor on cultured brain neurons are blocked by protein kinase inhibitors.

The influences of protein kinase inhibitors, K-252a and staurosporine, on the neurotrophic effects of epidermal growth factor (EGF) were investigated in dissociated cell cultures of the hippocampus and cerebellum of fetal rats. Addition of 1 ng/ml EGF enhanced the survival of cultured neurons of both brain regions. Both K-252a (10-200 nM) and staurosporine (1-100 nM) blocked the survival-promoting effects of EGF in a concentration-dependent manner. These results suggest that activation of protein kinase(s) is involved in the neurotrophic effects of EGF.

Alkaloids↗

PubMed1992

15N and 13C labeling of Escherichia coli tRNAs toward the NMR analysis.

Escherichia coli tRNAs were labeled with stable isotope 15N in vivo. Three species of tRNA, tRNA(Glu), tRNA(Lys) and tRNA(Ile), were purified by an HPLC system and their NMR spectra were observed. In heteronuclear 1H-15N multiple or single quantum coherence (HMQC or HSQC) spectra, the crosspeaks corresponding to NH3 of U and NH1 of G can be distinguished clearly since their 15N chemical shifts are significantly different from each other. Thus, this combination of 15N-labeling and the proton detected heteronuclear experiments are useful for the signal assignment and the conformational analysis of tRNAs. Furthermore, C1'- selective 13C-labeling of nucleotides was examined in vivo in order to resolve the H1' signals of tRNAs. By using a newly constructed E. coli mutant strain, the isotopic enrichments of more than 90% at C1' and of less than 10% for other ribose carbons were achieved.

Carbon Isotopes↗