Biomedical subjects

T Takehara

Publications and source records attributed to T Takehara.

At least 109 records · Page 6Linked to original sources

PubMed1992

Proteolytic activation of a single-chain precursor of hepatocyte growth factor by extracellular serine-protease.

Hepatocyte growth factor (HGF) is biosynthesized as a single-chain precursor (pro-HGF) and is proteolytically processed to a two-chain mature form. When MRC-5 fibroblasts were pulse-radiolabeled under serum-free conditions, pro-HGF was the predominant molecular form of HGF in the culture medium. CHO cells transfected with an expression plasmid containing a full-size human HGF cDNA produced pro-HGF when these cells were cultured in serum-free medium. These findings suggest that HGF is secreted as a pro-form, which is then converted to a two-chain form by extracellular protease. Single-chain HGF exhibited mitogenic activity on cultured hepatocytes, with a potency similar to that of mature HGF, but this activity was remarkably inhibited by leupeptin. We postulate that inactive pro-HGF is converted to an active two-chain form by a leupeptin-sensitive serine-protease expressed by hepatocytes. Neither plasminogen activators nor plasmin showed any processing activity of pro-HGF in vitro.

Animals↗

PubMed1992

Detection of hepatitis C virus RNA in serum of patients with chronic hepatitis C treated with interferon-alpha.

We tested serial serum samples for hepatitis C virus RNA from patients undergoing treatment for chronic hepatitis C with interferon-alpha using an assay that combined reverse transcription and polymerase chain reaction. The subjects studied were 20 patients with chronic hepatitis who had serum antibody to hepatitis C virus (anti-C100-3). Before therapy, hepatitis C virus RNA was detected in 18 (90%) and 20 (100%) patients using primer sets derived from the NS3 region or the 5'-noncoding region of hepatitis C virus, respectively. Hepatitis C virus RNA became undetectable in all patients whose ALT level fell into the normal range during therapy. However, hepatitis C virus RNA reappeared in all patients whose ALT levels rose again after therapy, usually before the relapse. In patients whose ALT levels did not become normal, hepatitis C virus RNA did not disappear during therapy. Thus therapy with interferon-alpha appears to be beneficial in chronic hepatitis C because of its suppressive effects on hepatitis C virus replication. Detection of hepatitis C virus RNA in serum is useful for evaluating the antiviral effect of interferon.

Adult↗

PubMed1992

Detection of the minus strand of hepatitis C virus RNA by reverse transcription and polymerase chain reaction: implications for hepatitis C virus replication in infected tissue.

The combination of reverse transcription and polymerase chain reaction is a very powerful tool for the detection of hepatitis C virus RNA in sera of patients with hepatitis C virus infection. However, when studying the presence of this virus in tissue using polymerase chain reaction, it may be difficult to distinguish between blood viral particles adhering to the tissue and viral RNA contained within the tissue. Because hepatitis C virus has a single-stranded RNA of positive polarity, a minus-strand RNA is expected to be found in hepatitis C virus-replicating tissues as a template for the synthesis of genomic RNA. To see whether the detection of the minus strand of hepatitis C virus RNA by polymerase chain reaction can be used for the determination of hepatitis C virus-replicating tissues, we examined the presence of the minus strand of hepatitis C virus RNA in the plasma, peripheral blood mononuclear cells and liver specimens of patients with hepatitis C virus infection. The plus-strand RNA was detected in the plasma, peripheral blood mononuclear cells and the liver specimens, but the minus-strand RNA was only detected in the liver. These results suggest that hepatitis C virus replicates in the liver but not in peripheral blood mononuclear cells. This detection method for the minus strand of hepatitis C virus RNA should be useful for determining hepatitis C virus replication in tissues other than liver tissue.

Adult↗

PubMed1992

Improved serodiagnosis of chronic hepatitis C in Japan by a second-generation enzyme-linked immunosorbent assay.

To clarify the role of hepatitis C virus (HCV) infection in chronic liver disease, sera from Japanese patients which were negative by the original anti-HCV assay (Ortho) were subjected to a second-generation anti-HCV assay based on a combination of structural (C22) and nonstructural (C200) recombinant HCV proteins. Of 29 patients with chronic non-A, non-B hepatitis, 20 (69%) were anti-HCV-positive by the second-generation enzyme-linked immunosorbent assay (ELISA) and also positive by the reverse transcription-polymerase chain reaction (RT-PCR) which detects the HCV genome. Of 41 chronic hepatitis B virus carriers, 3 (7%) were positive by the second-generation ELISA; 1 of 3 was positive by RT-PCR. The HCV genome was detected in all cases positive for anti-HCV with high titers. Of 59 healthy subjects negative by the second-generation ELISA, none were positive by RT-PCR. These findings indicate that HCV is a major causative agent of chronic non-A, non-B hepatitis in Japan and that second-generation ELISA is specific and a more sensitive diagnostic assay for chronic hepatitis C.

Base Sequence↗

PubMed1992

Effects of alpha-interferon on gamma-interferon production of peripheral blood mononuclear cells in hepatitis B virus carriers.

We studied gamma-interferon production of phytohemagglutinin-stimulated peripheral blood mononuclear cells in response to alpha-interferon in hepatitis B virus carriers and healthy individuals. The magnitude of gamma-interferon production was significantly higher in patients with anti-HBe antibody than in patients with HBe antigen and healthy individuals. Furthermore, alpha-interferon augmented the production of gamma-interferon of peripheral blood mononuclear cells from patients with active liver injury [serum alanine aminotransferase (ALT), greater than 40 U/L], but not that from patients with inactive liver injury (serum ALT, less than 40 U/L) or healthy individuals. These results suggested that alpha-interferon could enhance the cellular immune response against hepatitis B virus by augmenting the endogenous production of gamma-interferon in patients with active liver injury, implying that the responsiveness to alpha-interferon might be responsible for liver cell injury.

Adult↗

PubMed1992

HCV RNA and antibody to HCV core protein in Japanese patients with chronic liver disease.

We evaluated the prevalence of hepatitis C virus (HCV) RNA and antibody (anti-HCVcore) to the putative HCV core protein in Japanese patients with chronic liver disease. Sera were screened by solid-phase enzyme immunoassay with a recombinant HCV core protein and by the reverse transcription-polymerase chain reaction (RT-PCR) test which directly detects the HCV genome. Anti-HCV core was detected with high titers in 95% (69/73) of chronic non-A, non-B hepatitis, and 94% (65/69) of anti-HCVcore-positive patients had the genome. Anti-HCVcore was also found with lower titers in 24% (10/41) of chronic hepatitis B virus carriers, and three of them had the genome. Only one (3%) of the 35 patients negative for anti-HCVcore tested positive to RT-PCR. These findings indicate the overwhelming prevalence of HCV infection in Japanese patients with chronic non-A, non-B hepatitis and a close relationship between the presence of anti-HCVcore and chronic hepatitis C in this population.

Adult↗

PubMed1992

Three cases of posttransfusion hepatitis C treated with interferon-alpha. Confirmation of a carrier state by detection of hepatitis C virus RNA after interferon therapy.

Interferon therapy is useful for decreasing the serum ALT level and improving liver histology in patients with chronic non-A, non-B hepatitis. This study examined the effect of interferon therapy in acute cases of posttransfusion hepatitis C. We report on three cases in which interferon alpha was administered at 100-220.5 million units. HCV RNA became undetectable during interferon administration and the ALT level declined to the normal range. However, after the cessation of the therapy, the ALT level began to fluctuate and HCV RNA reappeared in two patients. We concluded that interferon therapy for the acute phase of posttransfusion hepatitis is useful for suppressing viral replication and quickly improving the ALT level, but it can not always prevent the development of chronic hepatitis. Furthermore, there was a close correlation between the profile of HCV RNA and that of the ALT level, indicating that the replication of HCV plays an important role in liver injury.

Adult↗

PubMed1992

Detection of hepatitis C virus RNA in chronic non-A, non-B liver disease.

Serum samples were tested for detection of hepatitis C virus (HCV) RNA from 156 patients with chronic non-A, non-B liver disease. HCV RNA was detected in 121 (93.8%) of 129 patients positive for anti-C100-3 but was also found in 15 (55.6%) of 27 patients negative for anti-C100-3. The rate of positivity for HCV RNA was not significantly different among various stages of liver diseases. These results showed that HCV continues to replicate even in advanced liver disease and that it seems to be related to half of the cases of chronic non-A, non-B liver disease negative for anti-C100-3.

Adult↗

PubMed1992

Enhanced expression of HLA class I by inhibited replication of hepatitis B virus.

HLA class I display on hepatitis B virus (HBV)-infected hepatocytes is important for limiting HBV infection. However, the effect of HBV replication on HLA class I expression on host cells has not been determined. Since acyclovir is known to inhibit HBV replication of the novel cell line HB611, which was transfected with HBV genome using human hepatoblastoma cells as the recipient and continuously replicates HBV DNA, we analyzed HLA class I expression on acyclovir-treated HB611 by quantitative flow cytometry. The results demonstrated that acyclovir treatment clearly increases the level of HLA class I on HB611, and suggested that HBV replication inhibits expression of HLA class I on infected hepatocytes. This effect of HBV replication on the host cell may be a means by which HBV evades immune surveillance to maintain chronic infection.

Acyclovir↗

PubMed1992

Cell density-dependent regulation of albumin synthesis and DNA synthesis in rat hepatocytes by hepatocyte growth factor.

Hepatocyte growth factor (HGF), a potent mitogen for mature hepatocytes, has been considered to act as a hepatotropic factor for liver regeneration. We examined the effect of HGF on albumin synthesis and DNA synthesis of adult rat hepatocytes cultured at various cell densities. HGF stimulated albumin synthesis of hepatocytes by 40-60% when they were cultured at higher cell densities such that there was tight cell-cell contact. But at lower cell densities HGF failed to stimulate albumin synthesis. In contrast, the stimulatory effect of HGF on DNA synthesis of hepatocytes was more potent at lower than at higher cell densities: HGF did not stimulate DNA synthesis of hepatocytes cultured at confluent cell density. Thus, HGF seems to stimulate both albumin synthesis and DNA synthesis of hepatocytes, in a reciprocal relationship depending on cell density. When the effects of various cytokines were examined, epidermal growth factor, transforming growth factor-alpha, and acidic fibroblast growth factor also stimulated albumin synthesis by 20-30%. However, transforming growth factor-beta 1, basic fibroblast growth factor, and interleukin-1 beta had no effect on albumin synthesis, while interleukin-6 inhibited it by 42%. Thus HGF was the most potent in stimulating albumin synthesis in these cytokines. Since HGF is markedly increased in the liver or plasma following various liver insults, HGF may be involved in liver regeneration through the potential to stimulate both cell growth and liver-specific functions such as albumin synthesis in a cell density-dependent manner.

Albumins↗

PubMed1992

Construction of mutants of Actinobacillus actinomycetemcomitans defective in serotype b-specific polysaccharide antigen by insertion of transposon Tn916.

Mutants of Actinobacillus actinomycetemcomitans strain Y4 defective in the capsular-like serotype b-specific polysaccharide antigen (SPA) were constructed by inserting the transposon Tn916. Southern blot analysis suggested that the transposon was inserted into a variety of different sites on the chromosome. Whole cells from two mutants (strains ST1 and ST2) lacked reactivity with a monoclonal antibody to SPA of A. actinomycetemcomitans Y4 (mAb S5) in enzyme-linked immunosorbent assay, but those from another nine mutants (e.g. strains ST3 and ST5) reacted very weakly with mAb S5. Immunodiffusion tests showed that mAb S5 or rabbit antiserum against whole cells of strain Y4 produced a fused precipitin band with purified SPA and autoclaved extract from strain Y4, but no precipitin band with autoclaved extracts from these four mutants. The hydrolysate of autoclaved extract from strain Y4 contained equal amounts of rhamnose and fucose, component sugars of SPA. The hydrolysates of autoclaved extracts from strains ST1 and ST2 contained a trace amount of rhamnose, but not fucose. Those of autoclaved extracts from strains ST3 and ST5 contained a trace amount of fucose, but not rhamnose. All of these SPA-defective mutants reacted with a mAb to lipopolysaccharide of strain Y4. The cell hydrophobicity of SPA-defective mutants was higher than that of the parent strain. These mutant clones will be useful for analysing the gene complex responsible for the synthesis of SPA of A. actinomycetemcomitans and the regulation of expression of the polysaccharide.

Aggregatibacter actinomycetemcomitans↗

PubMed1992

Mechanism of water-insoluble glucan synthesis in Streptococcus sobrinus.

Synthesis of water-insoluble glucan (IG) by 1,3-alpha-D-glucan synthase from Streptococcus sobrinus was examined using methylation analysis. The purified enzyme was incubated with sucrose and dextran T2000 (DT2000) for a given time and only IG was harvested by centrifugation. The remaining supernatant was incubated again, and IG was obtained. By repeating the above method using the residual supernatant, we obtained 5 varieties of IG precipitated in different periods. These IGs were methylated and examined using gas chromatograph mass spectrometry. The DT2000 water-insolubilized in the early reaction stage tended to have a highly ramified structure, with 1,3-alpha-D-glucan on a 1,6-alpha-D-glucan chain as the main chain. On the contrary, the DT2000 water-insolubilized in the late stage tended to have sparse side chains of 1,3-alpha-D-glucan which elongated with incubation. Specifically, the greater the number of side chains, the sooner the DT2000 was insolubilized. These results suggest that water-insolubilization of the water-soluble glucan not only depends on the increase of the ratio of 1,3-alpha-glucoside linkages to 1,6-alpha-glucoside linkages but also on the degree of branching of the 1,3,6-alpha-branched glucoside linkages.

Glucans↗

PubMed1992

Oral health conditions and denture treatment needs in institutionalized elderly people in Japan.

Oral health conditions, including dental conditions, temporomandibular joint (TMJ) conditions, denture status, and oral hygiene status, were assessed on 1908 institutionalized elderly people 65 yr of age and older at 29 of the 30 existing institutions in the city of Kitakyushu, Japan. The percentage of edentulous people was 27% in the 65-74-yr-old group, and increased with age to 56% in the 85 yr and older group. In dentate persons, the mean number of remaining teeth and DF teeth were 13.4 and 8.6, 9.5 and 6.8, and 8.4 and 6.5 in the groups aged 65-74 yr old, 75-84 yr old, and 85 yr and older, respectively. 81% of all the persons examined had no unusual symptoms in their TMJ. A clicking sound was the most frequent symptom (17%). Hygiene of both their teeth and dentures was very poor. Of all the subjects, 36% needed new full and/or partial denture(s), and 41% needed only repair. When the institutionalized elderly people were compared according to their general health condition, no clear differences were observed in percentage of edentulousness, mean number of remaining teeth and DF teeth, and TMJ conditions. However, a higher level of both untreated teeth and denture treatment needs, and poorer oral hygiene, was found in elderly people having poor general health than was observed in those with better health.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗

PubMed1992

Hepatitis B core antigen-specific interferon gamma production of peripheral blood mononuclear cells during acute exacerbation of chronic hepatitis B.

To examine the relationship between hepatitis B core antigen-specific interferon gamma production and the liver injury, we measured the sequential change in this production by peripheral blood mononuclear cells of seven patients with chronic hepatitis B. Four patients who experienced acute exacerbation showed increased interferon gamma production when the serum alanine aminotransferase level peaked or during the recovery phase. In the three patients who did not experience acute exacerbation, interferon gamma production gradually decreased in one who had a low peak of alanine aminotransferase but did not show significant change in the other two. Increased production of hepatitis B core antigen-specific interferon gamma at the time of acute exacerbation suggests that interferon gamma induced by hepatitis B core antigen plays a role in hepatocellular injury of patients with chronic hepatitis B.

Acute Disease↗

PubMed1992

Modulation of acute phase protein synthesis in cultured rat hepatocytes by human recombinant hepatocyte growth factor.

Human recombinant hepatocyte growth factor (HGF) added to primary cultures of rat hepatocytes stimulates synthesis of some acute phase proteins, especially alpha-2-macroglobulin. As indicated by changes in mRNA abundance HGF increases alpha-2-macroglobulin production at the pretranslational level. Interleukin-6, the main acute-phase cytokine, does not show synergy with HGF in enhancing synthesis of alpha-2-macroglobulin, and inhibits HGF-induced DNA-synthesis. On the other hand, dexamethasone potentiates the effects of HGF on synthesis of DNA and acute phase proteins by cultured rat hepatocytes.

Acute-Phase Proteins↗

PubMed1991

Deletion of kringle domains or the N-terminal hairpin structure in hepatocyte growth factor results in marked decreases in related biological activities.

To determine the essential domain for biological activity in the hepatocyte growth factor (HGF) molecule, we prepared various mutated recombinant HGFs using site-directed mutagenesis, and examined the effects on DNA synthesis in hepatocytes, scattering of MDCK cells and the antiproliferative activity on HepG2 hepatoma cells. Native HGF and mutant HGFs, in which Gln534 and/or Tyr673 were respectively substituted for His and Ser to coincide with the catalytic triad amino acids in plasmin, markedly stimulated DNA synthesis of hepatocytes and scattering of MDCK cells but inhibited DNA synthesis of HepG2 cells. The mutant HGF deleted with the third or fourth kringle domain resulted in marked decrease of all three biological activities, while deletion of the N-terminal hairpin structure or the first or second kringle domain almost completely inactivated biological activities. We propose that the N-terminal hairpin structure and the first and second kringle domains are essential for biological activities of HGF and possibly for binding to its receptor.

Animals↗

PubMed1991

Scatter factor from human embryonic lung fibroblasts is probably identical to hepatocyte growth factor.

Human embryonic lung fibroblasts (MRC5) produced scatter factor which enhanced motility of Madin-Darby canine kidney (MDCK) epithelial cells and a factor which stimulates DNA synthesis of adult rat hepatocytes in primary culture. These activities were both completely neutralized by antibody against human hepatocyte growth factor (HGF). Human recombinant HGF induced a marked scattering of MDCK cells. Moreover, MRC5 cells highly expressed 6kb mRNA which hybridized with HGF cDNA probe and scatter factor cDNA cloned from the MRC5 cDNA library had the same sequence as that of HGF cDNA from human leukocytes. These results indicate that HGF possesses scatter factor activity and the scatter factor derived from the MRC5 cells is probably identical to HGF.

Animals↗

PubMed1991

The extension of alpha-D-1,3-branch linkages by 1,3-alpha-D-glucan synthase from Streptococcus sobrinus.

In the presence of an acceptor, 1,3-alpha-D-glucan synthase of Streptococcus sobrinus synthesizes water-insoluble glucans from sucrose. Under such conditions, 1,3-alpha-D-glucoside linkages were extended without any change in the glucose-residue number between the 1,3,6-branch points on the acceptor. From these results, the mechanisms of water-insoluble-glucan formation were proposed as follows: (i) the attachment of an acceptor to the glucan binding sites of 1,3-alpha-D-glucan synthase occurs during the initiation of the reaction, and concurrently determines the positions of the branched portions of 1,3,6 on the acceptor, and (ii) the 1,3-alpha-D-glucoside linkage extends from these positions.

Carbohydrate Conformation↗