PubMed Health⌕ Search

Biomedical subjects

R Horiuchi

Publications and source records attributed to R Horiuchi.

At least 55 records · Page 3Linked to original sources
PubMed1989

Purification and characterization of 55-kDa protein with 3,5,3'-triiodo-L-thyronine-binding activity and protein disulfide-isomerase activity from beef liver membrane.

Beef liver membranes were shown to have different kinds of 3,5,3'-triiodo-L-thyronine binding proteins including the 55-kDa protein which had been reported to have this activity in many cells by affinity labelling with N-bromoacetyl-3,5,3'-[125I]triiodo-L-thyronine. In order to characterize the molecular features of these binding proteins, the 55-kDa protein was purified from a beef liver membrane fraction abundant in the plasma membrane. The protein was solubilized with 0.5% Chaps and purified by chromatography on gel filtration, hydroxyapatite, and Mono Q anion-exchange columns. The purity was confirmed with reversed-phase HPLC and SDS/PAGE. Consequently, 0.4% of the total proteins in the membrane fraction was recovered as the 55-kDa protein. One fourth of the amino acid composition of this protein was Glx (14.6%) plus Asx (11.7%) and the pI of this protein was 4.5. The purified protein has triiodothyronine-binding activity with a Kd of 57 nM which is similar to the high-affinity binding site of the membranes. The anti-(55-kDa protein) sera specifically recognized the 55-kDa protein of beef, rat and human cells. The immunoglobulin G fraction of the anti-(55-kDa protein) sera inhibited triiodothyronine binding to the beef liver membrane fraction. The purified protein also showed the activity of protein disulfide-isomerase (EC 5.3.4.1) as determined by reactivating scrambled ribonuclease. These data strongly suggested that the multi-functional 55-kDa protein which has triiodothyronine-binding activity and the activity of protein disulfide-isomerase, which is also reported to be the beta subunit of prolyl-4-hydroxylase, glycosylation-site-binding protein of oligosaccharyl transferase and iodothyronine 5'-monodeionidase, could be significant in the action of triiodothyronine towards the target cells.

Amino Acids↗
PubMed1989

Two erbA homologs encoding proteins with different T3 binding capacities are transcribed from opposite DNA strands of the same genetic locus.

Two erbA homologs, termed ear-1 and ear-7, are present in the human genome on chromosome 17. The two genes reside in the same genetic locus with overlapping exons and are transcribed from opposite DNA strands. In addition, the ear-7 mRNA is alternatively spliced to generate two protein isoforms, namely the ear71 and ear72 proteins. Nucleotide sequence analysis predicts that the ear71 protein is a human counterpart of the chicken c-erbA protein, a molecule closely related or identical to thyroid hormone receptor. Indeed, Scatchard analysis of proteins synthesized in vitro indicated very high affinity binding of T3 to the ear71 protein but not to the ear72 protein. Interestingly, the ear-1 gene product showed low, but appreciable, binding to T3, although its authentic ligand remains to be clarified.

Amino Acid Sequence↗
PubMed1989

Inhibitory effect of dibutyryl cyclic adenosine monophosphate on the induction of alkaline phosphatase in human fetal liver cell line.

When human fetal liver cells (HuL-1-317), cultured continuously in a serum-free medium, were incubated with a combination of prednisolone, butyrate and a hypertonic concentration of NaCl at 37 degrees C, alkaline phosphatase activity increased. However, the addition of dibutyryl adenosine cyclic monophosphate (Bt2cAMP) to these agents inhibited the increase in alkaline phosphatase activity in a dose-dependent manner: the inhibitory effect of Bt2cAMP was significant at 0.05 mM, but disappeared at 0.01 mM. Both cycloheximide and actinomycin D inhibited the increase in alkaline phosphatase activity with the combination described above. Western blotting showed that this enzyme activity increase was a consequence of greater biosynthesis of enzyme molecules in HuL-1-317 cells, and that Bt2cAMP regulated the synthesis of enzyme molecules. We conclude that the changes in alkaline phosphatase activity under various conditions are based on the changes in the number of enzyme molecules in HuL-1-317 cells.

Alkaline Phosphatase↗
PubMed1989

Uptake of 3,5,3'-L-tri-iodothyronine in human erythrocytes.

The mechanisms of 3,5,3'-L-tri-iodothyronine (T3) uptake into human erythrocytes were examined. Purified membranes of human erythrocytes were shown to have two classes of T3-binding sites with one being a high-affinity site (dissociation constant, 59.2 +/- 17.8 nmol/l; maximum binding capacity, 344.3 +/- 95.5 fmol/micrograms protein). Furthermore, it was shown that there were two pathways for T3 uptake in human erythrocytes; one was saturable, stereospecific (T3 much greater than thyroxine greater than 3,5,3'-D-tri-iodothyronine), energy-dependent and dominant at 15 degrees C; the other was not displaced by unlabelled T3 and was energy-independent but did not occur by passive diffusion. The former pathway which, it is suggested, is a receptor-mediated transport pathway, was inhibited by monodansylcadaverine, phloretin or oligomycin at 15 or 37 degrees C, but the latter pathway was not inhibited by these inhibitors. Our results strongly suggest that uptake of T3 by the energy-independent pathway became predominant over the energy-dependent pathway at 37 degrees C and accounted for 83% of total T3 uptake of human erythrocytes.

Binding, Competitive↗
PubMed1989

Epidemiological investigation on cleft palate patterns and sexual difference.

Sexual difference in severity of the cleft palate was studied in 179 patients. An appreciable correlation was found between the severity of cleft and sex: more females showing higher levels of severity, and no sexual difference existing in less severe cases. The results of our current investigations suggested close correlation between the pathogenic factors and sex in the above patients with cleft palate.

Cleft Palate↗
PubMed1989

[Epidemiological investigation of cleft lip and/or palate. II. Incidence of cleft lip and palate among Japanese babies in Gifu prefecture 1986 to 1987].

Since 1981 we have been conducting investigations in Aichi Prefecture where our institution is located in close cooperation with Medical Association of Obstetrics-Gynecologists and member physicians of the Midwives Association. In addition, we surveyed 111 institutions in Gifu Prefecture for the patient who was delivered from January 1, 1986 to December 31, 1987 to learn the incidence and type classification of cleft lip and/or palate in the general population Gifu. Consequently, we acquired the date of 20667 newborns. It was found as a result that 35 babies had these abnormalities and that ratio of the birth of such newborns was one for 590.5 deliveries. Among 35 cleft infants 12 (34.3%) with cleft lip and palate, 5 (14.3%) cleft palate. Looking at the estimates from the results of our investigation, the annual number of newborns with these disease in the Gifu Prefecture was 41.6 to 41.9 in 1986, 33.5 to 33.6 in 1987 with 95 percent confidence limits.

Cleft Lip↗
PubMed1988

Dualistic effects of thyroid hormone on a human hepatoma cell line: inhibition of thyroxine-binding globulin synthesis and stimulation of alpha 1-acid glycoprotein synthesis.

In order to examine the action of thyroid hormone on the secretory proteins of the liver, we investigated the effects of thyroid hormones on the synthesis of T4-binding globulin (TBG), alpha 1-acid glycoprotein (AGP), and albumin in a human hepatoblastoma cell line, Hep G2. Hep G2 cells grown to be confluent in medium with 10% fetal calf serum were further cultured in serum-free medium for 4 days, and followed by treatment with hormones for 2 days changing the medium every 24 h. On day 2 (the second 24 h of hormone treatment), about 30% of TBG accumulation was inhibited by 10(-12) M T3 and 50% was inhibited by 10(-8) M T3, although no change was observed on day 1 (the first 24 h of hormone treatment). This inhibitory effect of T3 closely resembled the effect of T3 on [35S]methionine-labeled TBG synthesis by the cells incubated for 3 h after 42 h of pretreatment. About 30-55% of the newly synthesized [35S]TBG immunoprecipitated with anti-TBG serum was inhibited by 10(-8) M T3. These results showed that thyroid hormone inhibited TBG synthesis in Hep G2 cells. On the other hand, T3 stimulated the accumulation of AGP in the media on day 1 (140% of control by 10(-8) M T3), and the effect increased drastically on day 2 (250% of control by 10(-8) M T3). No effect of T3 on albumin accumulation or total protein synthesis was seen. The concentrations of T4 which had significant effects on TBG and AGP accumulation were 10 and 10(3) times higher than those of T3, respectively. In conclusion, thyroid hormone has dualistic effects on the secretory proteins synthesized by a human hepatoblastoma cell line: physiological concentrations of thyroid hormones decrease the synthesis of TBG, but increase the synthesis of AGP.

Carcinoma, Hepatocellular↗
PubMed1987

[Effects of estrogen and endocrine therapeutic agents on the estrogen receptor, progesterone receptor and DNA synthesis in MCF-7 human breast cancer cells using the whole cell uptake method].

In order to investigate the mechanisms of the endocrine therapeutic agents and the applicability of combined endocrine therapies for breast cancers, we studied the effects of estrogen and the endocrine therapeutic agents on estrogen receptor (ER), progesterone receptor (PgR), and DNA synthesis in MCF-7 human breast cancer cells, which is known to be sensitive to estrogen. ER and PgR of MCF-7 cells were determined by whole cell uptake method. In brief, intact MCF-7 cells cultured in the multi-well plates were incubated with various concentrations of tritiated estradiol (E2) or promegestone (R5020) at 37 degrees C for 1 hour. The characteristics of the hormone binding were analyzed by Scatchard plots. MCF-7 cells had single class of ER (Kd: 2.1 +/- 0.2 X 10(-10) M, MBC: 9.0 +/- 1.5 X 10(3) sites/cell) and PgR (Kd: 7.2 +/- 1.1 X 10(-10) M, MBC: 3.1 +/- 0.5 X 10(4) sites/cell). When cultured in the presence of 10(-8) M or 10(-6) M of E2, tamoxifen (TAM), R5020 or medroxyprogesterone acetate (MPA) for 48 hrs, the numbers of binding sites of ER and PgR altered, but the affinities of either of them did not change. E2(10(-8) or 10(-6) M) increased about twice the number of PgR. Although treatment of 10(-8) M TAM, a non-steroidal antiestrogen, slightly increased the number of PgR, 10(-6) M TAM significantly decreased the number of PgR. Both of R5020 (10(-8) or 10(-6) M) and MPA (10(-8) or 10(-6) M), synthetic progestins, decreased the number of ER dose-dependently. On the other hand, E2 (10(-8) and 10(-6) M) and R5020 (10(-8) M) enhanced DNA synthesis, but 10(-6) M TAM or MPA inhibited DNA synthesis. The effects of single, sequential and coincidental treatment of these agents were compared. The sequential treatment of 10(-8) M E2 for 36 hrs and followed 10(-6) M MPA for 36 hrs ("10(-8) M E2----10(-6) M MPA") and "10(-6) M TAM----10(-6) M MPA" inhibited DNA synthesis of MCF-7 cells more efficiently than 10(-6) M TAM alone or 10(-6) M MPA alone for 72 hrs. However, "10(-6) M MPA----10(-6) M TAM" inhibited DNA synthesis less than "10(-6) M TAM----10(-6) M MPA".(ABSTRACT TRUNCATED AT 400 WORDS)

Breast Neoplasms↗
PubMed1987

Sequence of membrane-associated thyroid hormone binding protein from bovine liver: its identity with protein disulphide isomerase.

The complementary DNAs of the bovine liver membrane-associated 3,5,3'-triiodo-L-thyronine binding protein with 55 k-dalton (T3BP) were cloned and the nucleotide sequences were determined. Monospecific antibodies against T3BP were used to screen a bovine liver cDNA library in lambda gtll. We analyzed the sequences of two cloned T3BP cDNAs. The clones encoded a polypeptide of 510 amino acid residues, including a signal peptide of 20 amino acid. Northern blot analysis of bovine and human RNA showed that the mRNAs encoding T3BP are 2.7 kilobase in length. Amino acid sequence of N-terminal and other three peptides isolated from purified T3BP were found in the predicted amino acid sequence from the cDNA sequence. The predicted amino acid sequence is closely homologous (93%) with that of rat protein disulphide isomerase (EC 5.3.4.1), which catalyzes the isomerization of the protein disulphide bonds and has been shown to play an important role in post-translational regulation. The results suggest that T3BP and protein disulphide isomerase should be the same protein.

Amino Acid Sequence↗
PubMed1985

Localization of hepatitis B surface and core antigens in human hepatocellular carcinoma by immunoperoxidase methods. Replication of complete virions of carcinoma cells.

The localization of hepatitis B surface antigen (HBsAg) and core antigen (HBcAg) was investigated by an indirect immunoperoxidase method in formalin-fixed, paraffin-embedded liver specimens obtained from 95 Japanese patients with hepatocellular carcinomas. Routine and immune electron microscopic examinations were done in one case. The correlation between expression of hepatitis B virus antigens in the tissue and serum hepatitis B e antigen (HBeAg)/antibody to HBeAg (anti-HBe) status was examined. Hepatitis B surface antigen was detected in the cytoplasm of noncarcinomatous hepatocytes in 28 (29.5%) cases and of carcinoma cells in 11 (11.6%) cases. Hepatitis B core antigen was stained in noncarcinomatous hepatocytes in 13 (13.7%) cases and in carcinoma cells in 4 (4.2%) cases. Hepatitis B core antigen was present mainly in the nuclei, and all HBcAg-positive cases were positive for HBsAg. The routine electron microscopic examination revealed many round particles, 25 to 30 nm in diameter both in the nuclei and in the cytoplasm, and larger particles, 40 to 45 nm in diameter in the cytoplasm of carcinoma cells. Both types of particles had reaction products of HBcAg by immunoelectron microscopic study. Therefore, it was confirmed that the former were cores and the latter were Dane particles. There was a tendency that HBeAg-seropositive cases showed localization of HBcAg in the noncarcinomatous tissue. Among four cases with positive HBcAg in carcinoma cells, two were positive for HBeAg, one was positive for anti-HBe, and the other was negative both for HBeAg and anti-HBe in the sera. The data suggested occasional production of complete hepatitis B viruses of carcinoma cells in anti-HBe-positive as well as in HBeAg-positive hepatocellular carcinomas.

Adult↗
PubMed1983

Efficacy of hepatitis B vaccine in chimpanzees given transfusions of highly infective blood.

The protective efficacy of a hepatitis B vaccine against infections from transfusions of large volumes of highly infective blood in five immunized chimpanzees was assessed. Hepatitis B surface antigen (HBsAg) became positive and antibody to HBsAg (anti-HBs) disappeared soon after transfusion in the five chimpanzees. Two chimpanzees that had HBsAg only on the day of transfusion did not develop infection. However, the remaining three chimpanzees with persistent HBsAg antigenemia for three to four days developed serologic evidence of infection. Two chimpanzees did not have hepatitis and the third had a mild, transient case of acute hepatitis. The hepatitis B vaccine prevented the four immunized chimpanzees from developing illness. The remaining chimpanzee developed hepatitis, but a rapid booster response of anti-HBs owing to the previous vaccination appeared to lighten the severity of the disease and prevent chronicity.

Animals↗