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Biomedical subjects

S Okinaga

Publications and source records attributed to S Okinaga.

At least 73 records · Page 4Linked to original sources
PubMed1989

Functional analysis of the cDNA encoding human tyrosinase precursor.

The DNA segment harboring the promoter region and the exon 1 of the human tyrosinase gene has been cloned and characterized. Sequence analysis reveals the amino-terminal half of tyrosinase molecule including a signal peptide, of which six amino acid residues are not represented in the tyrosinase cDNA, pHT gamma 1 [Shibahara et al. (1988) Tohoku J. Exp. Med. 156, 403-414]. We therefore constructed the expression plasmid containing the human tyrosinase precursor cDNA, and introduced it into mouse amelanotic melanoma cells. Both tyrosine hydroxylase and dopa oxidase activities were expressed only in the cells transfected with such a full-length cDNA, providing direct evidence that tyrosinase actually possesses a dual catalytic activity.

Amino Acid Sequence↗
PubMed1989

Transcriptional control of the rat heme oxygenase gene by a nuclear protein that interacts with adenovirus 2 major late promoter.

Induction of rat heme oxygenase is mediated by at least two factors: its substrate heme and heat shock (Shibahara, S., Müller, R.M., and Taguchi, H. (1987) J. Biol. Chem. 262, 12889-12892). We have identified the cis-acting element of the rat heme oxygenase gene (HO gene), that was specifically bound by a nuclear protein prepared from rat glioma cells. We have termed this protein as heme oxygenase transcription factor (HOTF), whose estimated molecular weight is about 40,000. The element identified is CCACCACGTGACTCGAG (-51/-35) located just upstream of TATA-like sequence. The functional studies indicate that this sequence is required for the accurate and efficient initiation of transcription from the HO gene promoter. Since the HOTF-binding element is similar to the upstream promoter sequence (UPS) of the adenovirus 2 major late promoter (Ad2MLP), we examined whether rat HOTF is homologous to the upstream stimulatory factor (USF) or major late transcription factor previously identified in HeLa cells, which interacts with the UPS and activates the transcription from the Ad2MLP. HOTF specifically binds to the UPS of Ad2MLP, whereas HOTF failed to form a stable complex with the mutated HOTF-binding element lacking a GTGA sequence (-44/-42 of the HO gene), the sequence of which is identical to a core sequence of the USF-binding sites. Moreover, we show that like USF, HOTF is heat stable. These results suggest that HOTF may be homologous to USF or the major late transcription factor. Since USF is present in uninfected cells, it is conceivable that the expression of the HO gene is regulated at least in part by USF.

Adenoviruses, Human↗
PubMed1989

Development of a direct microplate enzyme immunoassay for the determination of pregnanediol-3 alpha-glucuronide in urine.

A sensitive direct enzyme immunoassay for urine pregnanediol-3 alpha-glucuronide was developed. The assay system involves the use of an antiserum against pregnanediol-3 alpha-glucuronide and an enzyme-labelled antigen chemically prepared by linking beta-D-galactosidase to 20 alpha-hydroxy-5 beta-pregnane 3(O-carboxymethyl)oxime. Free from antibody-bound antigen was separated by a solid-phase double antibody method, using a microplate coupled with goat anti-rabbit gamma-globulin. This solid-phase enzyme immunoassay for urine pregnanediol-3 alpha-glucuronide was validated in terms of specificity, accuracy and sensitivity. When urine samples were assayed for pregnanediol-3 alpha-glucuronide, the results obtained by the solid phase enzyme immunoassay and conventional radioimmunoassay methods agreed well (n = 30, r = 0.922). This assay system has an advantage over radioimmunoassay, because it does not require the use of radioisotopes. The procedure of this method is very simple, since it does not require purification steps of the biological samples.

Female↗
PubMed1989

Sex-specific effects on the fetal neuroendocrine system during acute stress in late pregnancy of rat and the influence of a simultaneous treatment by tyrosine.

It is well-known that prenatal chronic intermittent stress affects the reproductive system of both sexes. Investigating the effects of an acute maternal stress on the fetal neuroendocrine system, parameters such as hypothalamic catecholamines. CRF, GRF, LH-RH, beta-endorphin, hypophysial beta-endorphin and beta-LPH as well as plasma LH, corticosterone and androstenedione were measured. Pregnant rats of Wistar strain were exposed to restraint stress at day 22 of gestation or to forced immobilization at day 20 of gestation, respectively, and were sacrificed before stress and 10, 30, 60, and 120 min after starting stress. A decrease of fetal hypothalamic catecholamines and an increase of LH-RH content of the hypothalamus as well as of plasma catecholamines were observed under stress on day 22 of gestation. On day 20 of gestation hypothalamic beta-endorphin was depleted in male and unchanged in female fetuses under stress. A depletion of hypothalamic CRF was observed in male fetuses, whereas female fetuses showed an increase of hypothalamic CRF. An increase of GRF was found in fetuses of both sexes. Pituitary opioid content increased in fetuses of both sexes initially, but was depleted secondarily in male fetuses. The LH plasma level was markedly reduced in male, the corticosterone level was elevated in fetuses of both sexes as well as the androstenedione level in female fetuses. A simultaneous treatment of mother animals with tyrosine--a catecholamine precursor--prevented the depletion of hypothalamic and pituitary beta-endorphin as well as in part the reduction of plasma LH levels in male fetuses. Hypothalamic GRF content does not increase under tyrosine treatment in male fetuses, whereas in female fetuses the stress-induced increase of GRF content was rather pronounced under tyrosine than attenuated. These results indicate that fetal hypothalamic neurotransmitters and neurohormones (such as LH-RH, CRF, GRF and opioids) are involved in changing circulating hypophysial and adrenal hormones in fetuses exposed to maternal stress in late pregnancy, whereby sex-specific different pathways might be effective in fetal stress processing. The prenatal administration of tyrosine prevented at least in part--those neurohormonal changes which are affecting the sex-specific brain differentiation.

Androstenedione↗
PubMed1989

Contragestive effects of the antigestagenic agent, RU486, in rats and rabbits.

The contragestive effects of the antiprogesterone RU486 in pregnant rats and rabbits were investigated. Administrations of increasing doses of RU486 in intact or castrated pregnant rats induced abortion in a dose dependent manner. RU486 had less effect when administered on the 16th day of gestation. Antiestrogens (tamoxifen, clomiphene citrate), an inhibition of 3 beta-hydroxysteroid dehydrogenase (epostane) showed a synergistic effect when used in combination with RU486. A decrease in serum progesterone levels was observed after RU486 administration. A larger dose of RU486 was necessary to induce abortion in pregnant rabbits compared to rats. The maximum blood concentration was greater and the area under the plasma level curve was larger in rats than in rabbits following administration of RU486.

Abortifacient Agents↗
PubMed1989

The inhibitory effect of synthetic steroids on proestrous gonadotropin release in the rat.

This study was performed to evaluate the inhibitory effect of synthetic steroids on the hypothalamo-pituitary axis. Six synthetic steroids, used for contraception or other gynecologic indications, were injected to female rats on diestrus 2 (2 mg/kg, sc), and the animals were sacrificed in the afternoon of proestrus. Serum levels of LH, FSH and progesterone, the pituitary contents of LH and FSH, and LH-RH in the median eminence were measured by radioimmunoassays. Among the synthetic steroids tested in this study, norethisterone, d-norgestrel, 3-keto-desogestrel and gestrinone completely blocked the gonadotropin surge on proestrus, showing stronger antigonadotropic activity than progesterone. Desogestrel did not affect proestrous gonadotropin release at this dose. The suppressive effect of danazol could be seen only in a larger dose (20 mg/kg). These findings reconfirmed previously reported results, which demonstrated that the hypothalamo-pituitary complex was a major site of action of these steroids.

Animals↗
PubMed1989

Steroid hormone receptors in the uterus and ovary of immature rats treated with gonadotropins.

We administered either saline (group A) or 10 IU of pregnant mare serum gonadotropin (PMS; groups B and C) to female immature rats. Fifty-three hours later, the rats were injected with saline (groups A and B) or 30 IU of human chorionic gonadotropin (hCG; group C). The rats were decapitated 17 h after the last treatment, and the serum levels of progesterone (P4) and estradiol (E2) were measured by specific radioimmunoassays (RIA). The receptor levels of progesterone (PR) and estrogen (ER) in the uterus and ovaries were measured and the dissociation constant (Kd) of PR was obtained. The highest serum level of P4 was found in group C and that of E2 in group B. Cytosol levels of PR and ER in the uterus and ovary of the group B were the highest. It was indicated that the PMS treated-group (B), which had developing follicles in the ovary and the high serum level of E2, showed the highest concentration of ER and PR in both the ovary and the uterus. In the PMS and hCG-treated group (C), the uterine and ovarian steroid receptors decreased probably because of the luteinization and the high serum level of P4. The Kd uterine PR value was less than that of ovarian PR.

Animals↗
PubMed1989

Inhibition of rat ovarian 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD), 17 alpha-hydroxylase and 17,20 lyase by progestins and danazol.

The site of action of synthetic progestins or danazol in the treatment of endometriosis is considered to be mainly the hypothalamo-pituitary level, but the direct action to the uterine endometrium and the ovary is also suggested. We investigated the effect of these synthetic steroids to rat ovarian steroidogenic enzymes. The effect of norethisterone, levonorgestrel, danazol, gestrinone, desogestrel and 3-keto-desogestrel was studied in vitro. The sources of the enzymes were prepared from ovaries of immature rats treated either with pregnant mare serum gonadotropin (PMS) and human chorionic gonadotropin (hCG) for 3 beta-hydroxy steroid dehydrogenase (3 beta-HSD), or with PMS for 17 alpha-hydroxylase and 17,20 lyase. The substrates used were pregnenolone (P5) for 3 beta-HSD, progesterone (P4) for 17 alpha-hydroxylase, and 17 alpha-hydroxy-progesterone (17 alpha-OH-P4) for 17,20 lyase. The substrates were incubated with the enzyme sources and coenzymes, and the products formed were measured. All the steroids inhibited 3 beta-HSD, and the inhibition by gestrinone (Ki = 3.0 microM) and 3-keto-desogestrel (17.5 microM) was particularly marked. Only desogestrel (Ki = 30.3 microM) and danazol (168 microM) inhibited 17 alpha-hydroxylase. All the steroids inhibited 17,20 lyase, and the inhibition by desogestrel (Ki = 0.70 microM), danazol (0.80 microM), and gestrinone (30 microM) was particularly marked.

3-Hydroxysteroid Dehydrogenases↗
PubMed1989

Shoulder arthrodesis by external fixation.

The choice of pin site in shoulder arthrodesis after brachial plexus injury is important for efficient external fixation. The pins are inserted into the coracoid process from the anterior aspect and the scapular spine from the acromion. This insertion method holds the scapula more rigid and allows the patient to lie supine. Moreover, it makes it possible to correct the fixation angle after the operation. In 11 cases of brachial plexus injury, solid bony arthrodesis was obtained within three months.

Adult↗
PubMed1988

Alteration of infection pattern of duck hepatitis B virus by immunomodulatory drugs.

The relationship between host immune state and hepatic inflammation and infection pattern of the Duck hepatitis B virus (DHBV) was investigated by experimental transmission of DHBV into 98 Japanese 7-day-old ducklings that had been pretreated with immunoregulatory drugs including cyclophosphamide, OK 432, and a steroid hormone. Immunosuppressive treatment with cyclophosphamide revealed an extension of the viremic period associated with an absence of inflammatory changes in the liver. Although immunostimulating treatment with OK 432 showed a remarkable accumulation of inflammatory cells in the liver, the viremic period was not shortened. Treatment with a steroid used as a immunosuppressant did not suppress the hepatitis; moreover, it increased viral DNA replication and extended the viremic period. This phenomenon of viral replication seemed to be caused by the direct effects of the steroid. Alteration of DHBV infection by modifying the host immune state is quite similar to that of hepatitis B virus (HBV) in humans. In DHBV infection, the host immune state seemed to have a considerable role in determining the infection pattern and degree of hepatitis activity. DHBV may be a helpful model of HBV for studying host-viral interaction and the immunological mechanism of viral hepatitis.

Adjuvants, Immunologic↗
PubMed1988

The effect of an acute maternal stress on beta-endorphin and growth hormone releasing factor in the rat fetus.

Pregnant rats were injected with saline or L-tyrosine methylester HCl (200 mg/kg) and subjected to an acute forced immobilization stress on day 20 of gestation. At 10, 30, 60, and 120 minutes after the onset of stress, their fetuses were dissected out, and the contents of hypothalamic and pituitary immunoreactive beta-endorphin (IR-beta-EP) and hypothalamic immunoreactive growth hormone-releasing factor (IR-GRF) were determined by specific radioimmunoassays. The maternal stress arose a significant decrease of hypothalamic IR-beta-EP at 30 minutes, while pituitary IR-beta-EP slightly elevated at 30 minutes, then declined at 60 minutes. Hypothalamic IR-GRF showed a gradual increase during the maternal stress. Tyrosine supplementation tended to attenuate stress-induced changes in hypothalamic and pituitary IR-beta-EP, but the response of hypothalamic IR-GRF was less modified by tyrosine. These results showed the functional changes in fetal central beta-EP and GRF under maternal stress in the late gestational life, and suggested that catecholaminergic regulations participate, at least in part, in the fetal neuroendocrine response to maternal stress.

Animals↗
PubMed1988

Metabolism of round spermatids: gossypol induces uncoupling of respiratory chain and oxidative phosphorylation.

The effect of gossypol on energy metabolism of round spermatids of rats was examined. When spermatids were treated with various concentrations of gossypol for 30 min at 32 degrees C, a biphasic response (stimulation at low concentrations and inhibition at high concentrations) was seen in pyruvate and CO2 production from lactate. At the early period of incubation, gossypol at even high concentrations stimulated CO2 production to an extent similar to that stimulated by 2,4-dinitrophenol (DNP). At longer periods of incubation, however, the rates of CO2 production from lactate dropped to those seen in the rotenone-treated cells. The rates of oxygen consumption were not increased further by DNP when cells were pretreated with gossypol. The adenosine triphosphate (ATP) content in spermatids was reduced markedly, although lactate oxidation to CO2 and mitochondrial respiration were stimulated by gossypol. These results suggest that gossypol probably exerts its effect on spermatids by uncoupling respiratory chain and oxidative phosphorylation.

2,4-Dinitrophenol↗
PubMed1987

A sensitive bridge heterologous enzyme immunoassay of progesterone using geometrical isomers.

A sensitive bridge heterologous enzyme immunoassay of progesterone using geometrical isomers of progesterone 3(O-carboxymethyl)oxime(E/Z) was developed. Isomers were separated by synthesis of N-hydroxysuccinimide esters. Progesterone 3(Z)(O-carboxymethyl)oxime N-hydroxysuccinimide ester bound with beta-galactosidase in an appropriate molar ratio provided a conjugate suitable for an enzyme immunoassay. The antiserum was raised in rabbits by immunizing the animals with the progesterone 3(E)(O-carboxymethyl)oxime-bovine serum albumin conjugate. This bridge heterologous enzyme immunoassay proved to have sufficient sensitivity equivalent to radioimmunoassay and excellent specificity.

Antigens↗
PubMed1987

Small dose of 2-deoxy-D-glucose completely excludes the overgrowth of fibroblasts.

The present work has been undertaken to assess the effects of a small dose of 2-deoxy-D-glucose on proliferation of fibroblasts in culture by morphological criteria. Fibroblasts in culture medium with 2-deoxy-D-glucose proliferated only to a slight degree, but cell confluence was not achieved. They became swollen and the cytoplasm was increasingly occupied by lucent area. These changes became prominent on the 10th day of incubation. When cell mixtures of fibroblasts and TYK-nu (cell line of undifferentiated adenocarcinoma of the ovary) or TYK-nu only were incubated under the influence of 2-deoxy-D-glucose, the degenerative changes aforementioned were observed only in fibroblasts, while no significant changes were ascertained in the cell line of the cancer by morphological criteria. These results suggest that a small dose of 2-deoxy-D-glucose shows considerable promise in specific inhibition of fibroblastic proliferation during the process of establishing the cell line of the cancer.

Adenocarcinoma↗
PubMed1986

Effects of neuroleptics administered to lactating rats on the behavioral development of offspring.

Effects of diazepam (DZP) and haloperidol (HAL) given to nursing mothers from the day after delivery to weaning on the behavioral development of their pups were investigated. Weight gain in the pups of mothers that received these drugs did not differ from that in the control pups. Male and female pups exposed to HAL through the milk showed an increase of aggressive behavior (attacking and pinning play) when examined within the social unit of the colony at 4 weeks of age. Although an age-related decrease in this play was observed in the pups of all groups at 7 weeks of age, HAL exposed male and female pups or DZP exposed female pups showed this play more frequently than control pups. At 5 weeks of age, male and female pups exposed to DZP or HAL showed an increase in central ambulation of open-field, in spite of no change in total ambulation. Female pups exposed to DZP or HAL showed an increase of rearing. Female pups exposed to DZP or HAL took more time to reach the goal and made more errors in the maze test at 13-15 weeks of age. Male pups exposed to DZP or HAL showed a decrease in the number of complete mounts. Female pups exposed to DZP or HAL showed a decrease in the number of complete lordoses and an increase in the number of defensive behavior. These results indicate that neuroleptics administered to the dams appear to modify the functioning development of the brain in their offspring.

Animals↗