Biomedical subjects

T Hanafusa

Publications and source records attributed to T Hanafusa.

At least 145 records · Page 8Linked to original sources

PubMed1992

Integration of SCP1, a giant linear plasmid, into the Streptomyces coelicolor chromosome.

SCP1, coding for the methylenomycin biosynthetic genes in Streptomyces coelicolor, is a giant linear plasmid of 350 kb. Extensive physical characterization revealed that SCP1 has unusually long terminal inverted repeats (TIR) of about 80 kb on both ends and an insertion sequence, IS466, at the end of the right TIR (TIR-R), and the 5'-ends are attached to a terminal protein. In the NF strain S. coelicolor 2612, SCP1 is integrated into the chromosome at the 9-o'clock position. Analysis of the two junctions between the SCP1 DNA and the chromosomal DNA revealed that the left junction had an almost intact left terminus of SCP1, while the right junction was composed of IS466, completely deleting TIR-R. Based on these results, we presented a possible formation mechanism of the NF strain, which is characterized by integration of SCP1 into the chromosome via an interaction of the target site and the combined ends of the racket-frame structure of SCP1 followed by deletion of TIR-R. We also hypothesized that this type of integration of a giant linear plasmid might be involved in the origin and distribution of the chromosomal antibiotic biosynthetic gene clusters in microorganisms.

Base Sequence↗

PubMed1992

The structure of an integrated copy of the giant linear plasmid SCP1 in the chromosome of Streptomyces coelicolor 2612.

In NF strain 2612 of Streptomyces coelicolor, a giant linear plasmid SCP1 is integrated into the chromosome at the 9 o'clock position. To characterize the integrated structure of SCP1, cloning and sequence analysis of the two junctions between the SCP1 DNA and the chromosomal DNA was carried out. The left junction was revealed to retain an almost intact left terminus of SCP1. On the other hand, the right junction was composed of IS466, deleting completely the right terminal inverted repeat of SCP1. This junction might have been formed by recombination of two IS466 elements, one present at the end of the right terminal inverted repeat of SCP1 and one on the chromosome. Based on these results, we have proposed a model for the integration of SCP1 into the chromosome. The unique conjugal transfer of NF strains and the origin of the chromosomal antibiotic biosynthetic genes in Streptomyces species are also discussed in relation to this model.

Base Sequence↗

PubMed1992

Retrovirus gag protein p30 in the islets of non-obese diabetic mice: relevance for pathogenesis of diabetes mellitus.

We investigated the presence of retroviral protein in the pancreatic islets of non-obese diabetic mice to prove that the virus-like particle observed specifically in the pancreatic Beta cell of these mice was retrovirus. Western blot analysis probed with anti-retrovirus antibody demonstrated the existence of retroviral gag (group specific antigen) protein p30 in the islets of female non-obese diabetic mice. Islets of non-obese diabetic mice which were treated with cyclophosphamide, known to accelerate the development of insulitis and diabetes mellitus, have shown both a significantly increased number of retrovirus-like particles (type C) and enhanced expression of gag protein p30, compared to those of mice not treated with cyclophosphamide. These results confirmed the presence of type C retrovirus in non-obese diabetic mouse Beta cells and suggest a role for retrovirus in the development of insulitis and diabetes in these mice.

Animals↗

PubMed1992

Treatment of hepatocellular carcinoma by transcatheter hepatic arterial injection of radioactive iodized oil solution.

After 12 days of culture, VX2 carcinoma cells were inoculated into the liver of 16 rabbits; 14 days later, 131I-labeled iodized oil ([131I]-Lp) suspended in lipiodol was injected into the hepatic artery. Selective accumulation of the contrast material in the tumor for an extended time was evident on X-rays and hepatic scintiphotographs. The antitumor effect was remarkable. [131I]-Lp agents warrant further examination for their clinical usefulness. Internal radiation therapy by transcatheter hepatic arterial injection of [131I]-Lp (group A) was evaluated in 9 patients with hepatocellular carcinoma (HCC, tumor stage III or IV) associated with liver cirrhosis (LC) and compared with combination therapy of Lp-TAE (group B) in 18 patients with HCC (tumor stage III or IV) associated with LC. In group A, serum AFP levels dropped rapidly in eight of the nine patients who had an elevated initial level of more than 500 ng/ml. The average reduction in tumor size was 50% in eight cases as determined by computed tomography. Histological examination of one resected liver specimen at 3 months after the third injection of [131I]-Lp revealed microscopic features highly suggestive of a radiation effect in the [131I]-Lp-containing area. The 1-year survival value for patients with HCC was estimated at 49.0% using the Kaplan-Meier method. The survival of patients treated with internal radiation therapy tended to be better than that of those treated with Lp-TAE (P = 0.119).

Aged↗

PubMed1992

A novel oncogene, v-ryk, encoding a truncated receptor tyrosine kinase is transduced into the RPL30 virus without loss of viral sequences.

The RPL viruses are acute oncogenic avian retroviruses isolated from chicken tumors. We carried out a genetic analysis of three of the viruses, RPL25, RPL28, and RPL30. While RPL25 and RPL28 were shown to contain the erbB oncogene, RPL30 appeared to contain a novel protein tyrosine kinase oncogene. This gene, v-ryk, was cloned and sequenced. The v-ryk oncogene contains a 1.39-kb nonretroviral sequence that includes a tyrosine kinase domain which was inserted into the viral envelope protein gp37-coding region and fused in frame with upstream gp37 to generate a P69gp37-ryk fusion oncoprotein. Unlike that of other acutely transforming retroviruses, transduction of the v-ryk gene into RPL30 did not result in deletion of viral sequences. Sequence analysis suggested that v-Ryk is more homologous to receptor-type tyrosine kinases than to nonreceptor-type kinases. By reconstitution of a virus from its cDNA, the v-ryk oncogene has been shown to be fully responsible for the transforming activity of the RPL30 virus. Antibodies specific to v-Ryk immunoprecipitated the v-Ryk oncoprotein from cells transformed by the RPL30 virus. The v-Ryk protein was shown to be first synthesized as a 150-kDa precursor and then cleaved into the mature 69-kDa gp37-Ryk fusion protein, both parts of which were found to be localized to the membrane fraction. As expected from the sequence of v-Ryk, immunoprecipitates of v-Ryk from RPL30-transformed cells were found to display a protein tyrosine kinase activity in vitro, and the levels of tyrosine-phosphorylated proteins are elevated in v-ryk-transformed cells.

Amino Acid Sequence↗

PubMed1992

Function of DR-positive thyrocytes from patients with Graves' disease: quantitative analysis of thyroid peroxidase content by fluorescent photometry.

A considerable number of thyrocytes in patients with autoimmune thyroiditis ectopically express HLA-DR antigen. Furthermore, it has been reported that interferon-gamma-induced DR-positive thyrocytes in vitro secrete less thyroid hormone in response to TSH stimulation compared with DR-negative ones. However, the function of the intrinsically DR-positive thyrocytes is unknown. To evaluate their function, we stained by immunofluorescence for both DR antigen and thyroid peroxidase (TPO) in thyroid epithelial cells from patients with Graves' disease. We also measured the quantity of DR antigen and TPO using fluorescent photometry. The content of TPO was not significantly reduced in DR-positive thyrocytes compared with that in DR-negative thyrocytes. The TPO content is one measure of thyrocyte function. There was no significant difference between DR-positive and DR-negative thyrocytes. In conclusion, the function of DR-positive thyrocytes in vivo was not suppressed compared with that of DR-negative thyrocytes.

Adult↗

PubMed1992

Interferon-gamma reduces the thyroid peroxidase content of cultured human thyrocytes and inhibits its increase induced by thyrotropin.

To clarify the role of interferon-gamma (IFN gamma) in autoimmune thyroid diseases, we investigated the effects of IFN gamma on the content of thyroid peroxidase (TPO) and the expression of HLA-DR antigens in cultured normal human thyrocytes. The effect of TSH on the action of IFN gamma was investigated. Immunofluorescence staining and photometric analysis showed that IFN gamma not only induced the expression of DR antigen, but also reduced the content of TPO in a concentration-dependent manner. The addition of TSH increased the content of TPO and enhanced the IFN gamma-induced expression of DR antigen. IFN gamma also inhibited the increase in TPO content induced by TSH. Thus, complex interactions appear to exist between IFN gamma and TSH or thyroid-stimulating antibodies in the modulation of hormone secretion and autoimmune phenomena in the thyroid.

Adenoma↗

PubMed1992

Transthyretin (prealbumin) in the pancreas and sera of newly diagnosed type I (insulin-dependent) diabetic patients.

We investigated transthyretin (TTR) in the pancreases and sera of 10 newly diagnosed type I diabetic patients by immunohistochemistry and nephelometry. In the type I diabetic pancreases, glucagon-positive A-cells showed strong immunoreactivity for TTR, the intensity and distribution pattern of which corresponded to those in normal subjects. Morphometric analysis revealed that the amount of strongly TTR-positive A-cells was not significantly different from that in normal subjects. On the contrary, insulin-positive B-cells, which normally show uneven and weak TTR immunoreactivity, decreased in number, and only a few residual B-cells showed faint immunoreactivity. Neither somatostatin cells nor pancreatic polypeptide cells were positive for TTR. The serum TTR concentration showed a significant decrease in type I diabetic patients compared with that in normal subjects (P less than 0.005). These data suggest that the synthesis or storage of TTR in A-cells is not affected, but that in B-cells is impaired in type I diabetes. The decrease in serum TTR might be one of the features of metabolic disorders in type I diabetes.

Adult↗

PubMed1992

Detection by western blotting of an antibody to the hepatitis C virus E1 envelope protein in sera of patients with chronic liver disease.

We detected an antibody to HCV envelope protein (E1) in sera of patients with HCV-related chronic liver diseases (20 patients with chronic hepatitis and 5 patients with liver cirrhosis) by Western blotting using the fusion protein of E1 envelope protein and beta-galactosidase as an antigen. The antibody to HCV E1 (anti-HCV E1) was detected in 8 (42%) of 19 patients positive for HCV-RNA (16 were positive and 3 were negative for antibody to C100-3) and in 1 (17%) of 6 patients negative for HCV-RNA but positive for antibody to C100-3. HCV-RNA was detected in 8 (89%) of 9 anti-HCV E1 positive sera. The value of alanine aminotransferase was significantly higher in patients positive for anti-HCV E1 than in patients negative for the antibody. Although an antibody to the envelope protein of HCV is suspected to be one of the candidates of virus-neutralizing antibodies, our results suggest this hypothesis appears to be unlikely.

Adolescent↗

PubMed1992

Cell-to-cell communication in cultured rat thyroid monolayer cells is inhibited dose-dependently by methimazole.

We performed monolayer culture of rat thyrocytes and studied gap junctional communication by measuring intercellular fluorescence redistribution after photobleaching. Cell-to-cell communication among thyrocytes gradually developed during the culture. This communication was demonstrated in approximately 60 percent of the cells cultured for 8 days, while it could not be detected in the remaining 40 percent of the cells even after longer culture. When thyrocytes were cultured in the presence of methimazole, the fluorescence recovery after photobleaching was inhibited dose-dependently. Thyroid-stimulating hormone did not affect the fluorescence recovery. We provided, for the first time, that methimazole inhibited the development of cell-to-cell communication of thyrocytes dose-dependently.

Animals↗

PubMed1991

HLA-DQA11 contributes to resistance and A13 confers susceptibility to type 1 (insulin-dependent) diabetes mellitus in Japanese subjects.

In this study HLA-DQA1 and TNF genes in addition to HLA-DQB1 gene were investigated at DNA level for elucidation of the genetic backgrounds of Type 1 (insulin-dependent) diabetes mellitus in Japanese subjects. DNA, amplified by polymerase chain reaction, was subjected to allele specific oligonucleotide dot blot analysis, restriction fragment length polymorphism analysis or DNA sequencing. Polymorphism of the TNF gene to NcoI did not correlate with Type 1 diabetes in Japanese patients. DQw1.2 had a protective effect against the disease, the DQA1*1 allele was significantly decreased and DQA1*3 allele was significantly increased. Seventeen out of twenty-two Type 1 diabetic patients (77%) were homozygous for DQA1*3 and five out of twenty-two (23%) heterozygous. The DQA1*3 gene of Type 1 diabetic patients had a normal nucleotide sequence. Furthermore, DQA1*3 was found unexpectedly in two patients without DR4 or DR9. These data indicate that DQA1 gene confers susceptibility and resistance to Type 1 diabetes in Japanese subjects.

Alleles↗

PubMed1991

Alpha 1-adrenergic regulation of thyrotropin-stimulated release of 3, 5, 3' -triiodothyronine and thyroxine from perifused mouse thyroid.

The effect of methoxamine, a specific alpha 1-adrenergic agonist, on the release of T3, T4 and cAMP from perifused mouse thyroid was studied to clarify the role of the alpha 1-adrenergic receptor in the regulation of thyroid hormone secretion. TSH-stimulated T3 and T4 release was inhibited significantly by methoxamine. With regard to cAMP release, methoxamine inhibited TSH-stimulated cAMP release in the presence of 4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone but did not inhibit TSH-stimulated cAMP release in the presence of 3-isobutyl-1-methylxanthine. Methoxamine did significantly suppress TSH-stimulated release of T3 and T4 in the presence of each phosphodiesterase inhibitor. Depletion of Ca2+ in the perifusion buffer abolished completely the inhibitory effect of methoxamine on TSH-stimulated T3 and T4 release. The present study suggests that activation of the alpha 1-adrenergic receptor inhibits TSH-stimulated T3 and T4 secretion through a Ca(2+)-dependent mechanism in the mouse thyroid gland.

1-Methyl-3-isobutylxanthine↗

PubMed1991

Nucleotide sequence analysis of the unusually long terminal inverted repeats of a giant linear plasmid, SCP1.

SCP1 is a giant linear plasmid of 350 kb coding for the methylenomycin biosynthetic genes in Streptomyces coelicolor. The unusually long terminal inverted repeats present on both ends of SCP1 were analyzed on the nucleotide sequence level. Analysis of six clones containing the terminal 0.35-kb XbaI fragment revealed a slight heterogeneity in the nucleotide sequences of the SCP1 ends. Moreover, it was indicated that this fragment contained seven palindromic inverted repeats and a GT-rich region in the 5'-end strand. The size of the terminal inverted repeats was determined to be 81 kb by the cloning and sequencing of their end-points. An insertion sequence, IS466 was shown to be present just at the end of the right terminal inverted repeat.

Anti-Bacterial Agents↗

PubMed1991

Two types of autoantibodies to adrenal medullary cells in type 1 (insulin-dependent) diabetic patients: prevalence, properties and implications.

Complement-fixing adrenal medullary antibodies were examined in sera from 170 (114 Type 1 and 56 Type 2) diabetic patients and normal subjects by indirect immunofluorescence methods. Two types of antibodies were detected; one showed a homogeneous immunofluorescence pattern (homogeneous-type) and the other a spotty pattern (spotty-type) in the cytoplasm of adrenal medullary cells. Both antibodies were IgG class and adrenal medulla-specific. The prevalence of the homogeneous-type was significantly higher in Type 1 diabetic patients with disease duration under 1 year (36%) than in those with duration of 1 year or more (1.1%), in Type 2 diabetic patients (1.8%) or in normal subjects (0%; P less than 0.01). Conversely, the prevalence of the spotty-type was not significantly different among all subjects examined (3.6-4.5%). The epitope for the homogeneous-type is likely to be a glycoconjugate since binding of this antibody was abolished after periodate oxidation. The epitope for spotty-type antibody is considered to be a peptide since it was trypsin sensitive. Patients who were positive for the homogeneous-type were also positive for islet cell antibodies, although their antibody titers were not correlated. We conclude that (1) adrenal medullary antibodies are of homogeneous-type or spotty-type and the antigenic determinants of these antibodies are different, and (2) the prevalence of the homogeneous-type is significantly higher in newly diagnosed Type 1 diabetic patients and its presence is associated with that of islet cell antibodies.

Adolescent↗

PubMed1991

HLA-DP and susceptibility to insulin-dependent diabetes mellitus in Japanese.

Human leukocyte antigen (HLA) genes are candidates for susceptibility genes in insulin-dependent diabetes mellitus (IDDM). Recently, the association of DR and DQ with IDDM has been reported, but the role of HLA-DP genes remains uncertain. To address the question, we analyzed the DPB1 gene of 20 Japanese IDDM patients and 30 control subjects using a combination of polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis (PCR-RFLP method). DPB1*0501 was the most frequent allele both in Japanese patients and control subjects. There was no appreciable association between IDDM and the DPB1 allele in Japanese. The absence of association between IDDM and DP, in spite of the known association between this disease and both DR and DQ, suggests that the HLA locus (loci) telomeric to DP encodes susceptibility to IDDM.

Alleles↗

PubMed1991

Interferon-gamma inhibits thyroid-stimulating hormone-induced morphological changes and induces the expression of major histocompatibility complex class II antigen in thyroid follicles in suspension culture.

The effects of interferon-gamma (IFN gamma) on the morphology of thyroid follicles and the expression of major histocompatibility complex (MHC) class II antigens were examined. The thyroid follicles were suspended in RPMI-1640 containing 10% fetal calf serum with or without IFN gamma (200 U/ml). After culture for 5 days, follicles were incubated in the presence of TSH (10 mU/ml) for 1 h and fixed for electron microscopic and immunohistochemical examination. Regardless of the presence of IFN gamma, suspended follicles became inverted within 5 days. However, MHC class II antigens were expressed only in inverted follicles cultured with IFN gamma. In inverted follicles cultured without IFN gamma, TSH stimulation induced remarkable morphological changes, such as elongation of microvilli and an appearance of pseudopods. On the other hand, the follicles cultured with IFN gamma showed poor response to TSH. Thus, IFN gamma induced the expression of MHC class II antigens of cultured thyroid follicles and inhibited TSH-induced morphological changes in the cells.

Animals↗

PubMed1991

Inhibitory effects of ionophore A23187 on the release of thyroid hormone and colloid reabsorption in mouse thyroid glands.

The effect of the ionophore A23187 on a. the release of thyroid hormone from perifused mouse thyroid glands and b. the morphological changes in follicular epithelial cells was evaluated. A23187 at a concentration of 5 mumol/l significantly inhibited both the TSH- and the forskolin-stimulated release of T3 and T4. In the presence of 3-isobutyl-1-methylxanthine or (4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone) RO 20-1724, A23187 did not affect the forskolin-stimulated release of cAMP, but did inhibit the release of T3 and T4 stimulated by forskolin. Light and electron microscopic evaluation of the follicular epithelial cells of mouse thyroid tissues following 1-h stimulation with forskolin showed numerous pseudopods engulfing luminal colloid of various size and the presence of reabsorbed colloid droplets in the apical cytoplasm. Quantitative electron microscopic analysis revealed that the addition of the ionophore A23187 reduced the number of reabsorbed colloid droplets to one eighth in follicular epithelial cells. These observations suggest that the increase in intracellular Ca2+ induced by the ionophore A23187 inhibits the TSH-stimulated thyroid hormone release independently of the cAMP level, and that the suppression of thyroid hormone release may be due to an inhibition of colloid reabsorption.

1-Methyl-3-isobutylxanthine↗

PubMed1991

Human recombinant interleukin 1 inhibits TSH-stimulated morphological changes in thyroid follicles cultured as semi-organs.

To study the effects of human recombinant interleukin-1 on thyrocytes, we cultured thyroid follicles as semi-organs, each consisting of approximately 10-20 follicles, in the presence or absence of IL-1 alpha or beta. Semi-organ culture reproduces the in vivo environment well. After culture for 2 or 4 days, the follicles were incubated with TSH (10 U/l) for 4 h and fixed for light and electron microscopical examinations. Regardless of the presence or absence of IL-1, follicular structure, polarity, and luminal colloid did not change during culture. In thyroid epithelial cells cultured without IL-1, TSH markedly induced elongation of microvilli and formation of reabsorbed colloid droplets. On the other hand, both IL-1 alpha and beta inhibited these TSH-stimulated changes. The degree of inhibition correlated with the concentration of exposure to IL-1. We conclude that IL-1 inhibits TSH-stimulated morphological changes in thyroid follicles cultured as semi-organs, depending on the concentration of IL-1.

Animals↗