Biomedical subjects

H Kuramoto

Publications and source records attributed to H Kuramoto.

At least 91 records · Page 5Linked to original sources

PubMed1989

Proliferation and responsiveness to estrogen of human endometrial cancer cells under serum-free culture conditions.

Studies of hormonal growth regulation in cultured human endometrial cancer cells are limited by the requirement of exogenous growth factors, usually supplied by addition of serum. The present report provides evidence that estradiol can stimulate proliferation of endometrial cancer cells of the Ishikawa line in the absence of serum or added growth factors. Mitogenic effects of estrogen were demonstrated in two different experimental systems, in cells attached to the substratum of mammalian tissue culture dishes, and in cells forming colonies in soft agar under anchorage-independent conditions. Addition of estradiol to a mixture of serum-free, phenol red-free Dulbecco's minimal essential medium and Ham's F-12 medium, supplemented with L-glutamine and 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid [basal medium: (BM)] significantly increased the proliferation of cells attached to culture dishes. Dose-response experiments revealed maximal estradiol stimulation at 10 nM; significant responses were also observed at 1 nM and at 100 nM concentrations. The mitogenic effect of 10 nM estradiol was comparable to that of 1% charcoal-treated fetal bovine serum and the two effects were additive. The presence of estradiol in serum-free BM resulted in a shortening of the doubling time of exponentially proliferating cells from 38 to 29 h. From the labeling index, measured after exposure to a pulse of [3H]thymidine, and from the mitotic index, both determined in exponentially proliferating cells, the lengths of the S and M phases were calculated to be 11 and 1 h, respectively. From these data it was estimated that estradiol shortened the G1 phase by approximately 40%, from 22 to 13 h. Estradiol doubled the colony formation efficiency of cells plated in BM containing 0.3% agar in the absence of serum as well as in the presence of 1% charcoal-treated fetal bovine serum. The stimulation of colony formation by estradiol was influenced by medium components, since no effects were observed in minimal essential medium. The colony formation efficiency was positively related to the serum concentrations and remained significantly lower in minimal essential medium than in BM at comparable serum levels. The observed positive relationship between colony formation efficiency and cell densities at plating suggests a cooperative mitogenic effect, likely due to autocrine and paracrine action of secreted growth factors. These results define a model to evaluate hormonal growth regulation mediated by autocrine mitogens in human endometrial cancer cells in the absence of interfering exogenous growth factors.(ABSTRACT TRUNCATED AT 400 WORDS)

Cell Aggregation↗

PubMed1989

Stimulatory effects of 4-hydroxytamoxifen on proliferation of human endometrial adenocarcinoma cells (Ishikawa line).

The effects of trans-4-hydroxytamoxifen (OHTam) on proliferation of cells of the Ishikawa human endometrial adenocarcinoma line were studied under serum-free, phenol red-free conditions and compared to those of estradiol. The addition of OHTam (1 microM) to basal medium (BM), consisting of equal parts of Dulbecco's modified Eagle's medium and Ham's F-12 with additional glutamine and 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid, resulted in significant increases in cell numbers relative to controls. These effects were even greater than those obtained with estradiol (10 nM-1 microM) or 1% charcoal-treated fetal bovine serum (ctFBS). Addition of 1% ctFBS to BM containing 1 microM OHTam further increased cell numbers whereas addition of estradiol (10 nM) did not do so. The stimulation of growth was positively correlated with OHTam concentrations in the range of 10 nM to 1 microM. Dissociation of estradiol and OHTam proliferative effects was observed in a variant of Ishikawa cells in which estradiol did not increase proliferation while OHTam had a strong stimulatory effect. The growth-promoting effects of OHTam were also observed in BM containing 5% or 15% ctFBS. In contrast, in parallel experiments in which BM was replaced by minimal essential medium (Eagle's) with Earle's salts, OHTam (1 microM) did not stimulate proliferation under these conditions and acted as an antiestrogen, inhibiting the proliferative effects of estradiol. These results illustrate marked effects of medium composition on proliferation and antiestrogenic actions of OHTam. Alkaline phosphatase activity was strongly stimulated by estradiol (10 nM) but only very weakly affected by OHTam (1 microM); at these concentrations, OHTam inhibited the effect of estradiol, both in serum-free BM and in minimal essential medium plus 15% ctFBS, demonstrating dissociation in its actions on proliferation and on enzymatic activity. These findings suggest that OHTam may stimulate the proliferation of particular clones of endometrial cancer cells in human tumors. They also suggest that OHTam can exert effects not mediated by the estrogen receptor system, or form OHTam-estrogen receptor agonistic complexes unlike those resulting from estradiol-estrogen receptor interactions. Clearly, Ishikawa cells provide a useful model to investigate mechanisms of action of antiestrogens.

Adenocarcinoma↗

PubMed1989

Effects of serine and glycine on proliferation of an Ishikawa cell variant.

The proliferation of cells on an Ishikawa human endometrial adenocarcinoma line variant (Ishikawa-Var I) is markedly influenced by the medium used to culture them, viz. MEM vs BM (basal medium; DMEM/Ham's F12, 1/1, with additional glutamine and HETES), under serum-free conditions. Components of BM which are not present in MEM were systematically tested in order to identify those that might account for these differences. Cells were cultured for various periods of time, up to 8 days, in serum-free MEM to which the components to be tested were added. Cell population densities were evaluated using a fluorometric DNA assay when the cells were grown in multiwell plates, or by cell counting when the cells were cultured in plastic dishes. It was found that addition to MEM of a mixture of the amino acids that this medium lacks, significantly increased cell density. By testing individual amino acids at the concentrations present in BM, it could be demonstrated that addition of serine alone was sufficient to obtain the densities achieved with BM. Glycine, a metabolic precursor of serine, had a similar but smaller effect. None of the other missing compounds of BM was effective. Effects of serine on DNA synthesis were also estimated by measuring incorporation of [3H]thymidine for 1 h after a 24 h culture period in MEM. The effect of serine was similar and additive to that of 1% charcoal-treated fetal bovine serum. A serine concentration dependence studied either with this method or measuring DNA/well after 8 days in culture showed detectable effects at 0.005 mM concentration and maximal responses at about 0.025 mM. These findings are of potential importance in studies on regulatory mechanisms of cell proliferation. A possibility to be explored, for instance, is that serine added to the medium increases intracellular phosphatidylserine concentrations leading to increases in the activity of protein kinase C, a stimulator of cell proliferation in some systems.

Adenocarcinoma↗

PubMed1989

Distribution of enteric nerve cells that project from the small intestine to the coeliac ganglion in the guinea-pig.

The retrograde tracing agent, Fast blue, was injected into the coeliac ganglia of guinea-pigs and 4-7 days later, nerve cell bodies containing this dye were examined in the small intestine. The cell bodies were found in the ganglia of the myenteric plexus but not in submucous ganglia. The labeled cell bodies were large, on average 42 by 19 microns when viewed in whole mounts, with 4-9 fine processes. The cells increased in frequency anally along the small intestine; the number of neurons per unit length of gut in the distal ileum was more than double that near the duodeno-jejunal flexure. At all points along the intestine the nerve cells were more numerous near the mesenteric attachment than opposite this attachment. About half of the neurons showed immunoreactivity for VIP. It is deduced that the neurons that project from the intestine to the coeliac ganglion are likely to be second-order neurons in the afferent limbs of intestino-visceral reflex pathways.

Amidines↗

PubMed1989

Effects of extracorporeal shock wave lithotripsy on urinary excretion of N-acetyl-beta-D-glucosamidase.

We investigated the effect of extracorporeal shock wave lithotripsy (ESWL) on the urinary excretion of N-acetyl-beta-D-glucosamidase (NAG) activities. No significant increases in NAG excretion were recognized between, before and after ESWL. However, there were significant increases in patients with renal calculi who received over 1,000 shock waves. Therefore, we should not neglect renal impairments which could be caused by extracorporeal shock wave exposure.

Acetylglucosaminidase↗

PubMed1989

Gastric emptying in patients with gastric and duodenal ulcer.

Gastric emptying was measured in patients with proximal gastric, distal gastric and duodenal ulcers using a radioisotopic method. Gastric emptying was rapid in patients with proximal gastric ulcer, and was considered to be due to accelerated proximal evacuation. However, emptying was delayed in patients with distal gastric ulcer, and was considered to be due to reduced emptying in the antrum. On the other hand, rapid emptying seen in duodenal ulcer patients was considered to be due to accelerated emptying in both the proximal stomach and the antrum. Gastric emptying in the healing stage was closer to that in healthy subjects than in patients with active-stage ulcer.

Duodenal Ulcer↗

PubMed1989

[Responsiveness and mechanisms of action of steroid hormones in human endometrial adenocarcinoma cells].

The responsiveness and action mechanisms of steroid hormones and epidermal growth factor on human endometrial carcinoma cells are analyzed by using in vitro culture system. 1) The Ishikawa cells, derived from a well differentiated endometrial adenocarcinoma and possess ER and PR, are shown to respond to estrogens by increasing a variety of parameters, viz cell proliferation, PR levels, ALP and DNA polymerase activities. 2) ER and PR of those cells are localized in the nuclei by immunocytochemical staining using the monoclonal antibodies against to ER and PR, confirming the correctness of Gorski and Greene's one step theory involving the action mechanisms of steroid hormones. 3) Progestins reduced the ER level and stimulate E2DH activities and glycogen content, which are completely abolished by anti-progestin (RU486), suggesting that PR of those cells should be functional. 4) These responses to steroid hormones of Ishikawa cells are synergistically enhanced or appeared earlier by addition of EGF. 5) The main metabolite of E2 incubated with Ishikawa cells is E2-3-sulfate instead of E1, indicate that the higher estrogenic status may be persisted in endometrial cancer tissues.

Adenocarcinoma↗

PubMed1989

[Statistical analysis of trophoblastic disease in Kanagawa prefecture].

Two thousand one hundred and thirty-five cases of trophoblastic disease were registered in Kanagawa Prefecture in 7 years (from 1978 to 1984). The incidence of trophoblastic diseases, hydatidiform mole, partial mole, invasive mole, choriocarcinoma and persistent trophoblastic disease (PTD) was 3.22, 1.67, 1.20, 0.07, 0.06 and 0.20, respectively, per 1,000 live births. The incidence was constant for 7 years, and did not differ significantly from that of 16 prefectures in Japan. In Kanagawa, the incidence of hydatidiform mole was less and that of partial mole was more than those in the 16 prefectures. The incidence of invasive mole and choriocarcinoma was lower and that of PTD was higher than those in the 16 prefectures. Total incidence of invasive mole, choriocarcinoma and PTD was higher in the western part of Japan, and lower in the eastern and northern parts. Kanagawa Prefecture is between these two regions. The distribution by age of hydatidiform mole was high in 25-29 y.o. and over 40 y.o. The distribution by age of partial mole tended to be similar to that of hydatidiform mole, but less remarkable. The distribution by age of invasive mole and choriocarcinoma had 2 peaks of 30-34 y.o. and 45-49 y.o. The distribution by age of PTD was between that of hydatidiform mole and choriocarcinoma.

Adolescent↗

PubMed1989

A modified method for the determination of aluminium-gel in vaccine.

The conventional method of aluminium analysis, described in the Biological Preparation Standard for Animals, was found to give erroneous results in the case of vaccine containing aluminium-gel as an adjuvant. This is because of incomplete decomposition of the gel and unsettled reaction system for the determination. But these problems were settled definitely. Heating on a direct fire for 15 min, by which temperature of the content attained 115-120 degrees C at atmospheric pressure, allowed the gel to complete decomposition and the result came to close the theoretical value. And a constant result by the decomposition itself was obtained with 2 volumes of nitric acid against 1 volume indicated in the standard method. Quantitative reaction of the gel-decomposed solution did hold in range of about 0.75-2.0 micrograms aluminium per ml. Use of acetate buffer solution for dilution of the decomposed solution following by sampling in 2 volumes of the dilution into the reaction mixture made the variation to the least as compared with the water-dilution following by 1 volume sampling indicated in the standard method. By these modified conditions, we succeeded in reducing the relative error and the coefficient of variation to less than +/- 5% and +/- 2% respectively.

Adjuvants, Immunologic↗

PubMed1989

[Diagnosis and treatment of dysplasia of the uterine cervix].

The accuracy of diagnosis and the results of treatment for dysplasia of the uterine cervix are analyzed. Three hundred and thirty-seven cases of mild, 231 of moderate and 200 of severe dysplasias have been treated at Kitasato University Hospital. By cytology 40.4% of mild, 74.5% of moderate and 89.0% of severe dysplasias were judged suspicious or positive, and the accurate cytopathological detection rate was 19.8%, 16.1% and 60.1%, respectively. Abnormal colposcopic findings such as M, P and W were observed in more than 95% of cases with dysplasia. The area of abnormal findings tends to be large as the lesion becomes more severe. Mild dysplasia progressed to cervical cancer in 1.5%, moderate dysplasia in 2.4% and severe dysplasia in 9.4%. Conservative therapy by laser vaporization or conization was used for 31 mild, 96 moderate and 114 severe dysplasias. The cure rate of each dysplasia was 100%, and pregnancy following laser therapy was encountered in 28 cases without abnormal course. It is concluded that colposcopy is more useful in diagnosis of mild and moderate dysplasia than cytology, and conservative laser treatment is an effective method for dysplasia.

Adult↗

PubMed1989

[Mechanism of action of endocrine therapy of endometrial carcinoma].

The mechanism of action of sex steroid hormones on endometrial carcinoma was analyzed by a cell culture system. 1) Estrogenic actions such as enhancement of ALP activity, PR level and cellular growth were confirmed in Ishikawa endometrial carcinoma cells which possess ER. 2) ER and PR are both localized immunocytochemically in the nuclei of Ishikawa cells, confirming the correctness of Gorsky and Greene's theory in endometrial carcinoma. 3) In Ishikawa cells with PR, progestins stimulated E2DH activity and glycogen synthesis and reduced ER level, thus proving the existence of their hormonal action. We cannot, however, rule out the pharmaceutical action of progestin, because a decrease in nucleic acid syntheses and ALP activity, alteration in electron microscopic observation and finally cellular death were observed in endometrial cancer cells without PR. 4) Conversely, Tamoxifen, anti-estrogen, stimulated the cellular growth in serum-free environment of culture. We must therefore be careful in using it clinically.

Adenocarcinoma↗

PubMed1988

Fine-structural localization of neuropeptide tyrosine (NPY)-like immunoreactivity in the neuronal somata of colchicine-pretreated celiac ganglia of rats.

In colchicine-pretreated cells of sympathetic ganglia, intensely NPY-immunoreactive material was localized within vacuoles and vesicles of the disorganized, widely dispersed Golgi apparatus. Intensely positive large granular vesicles, which are known to be one of major storage sites of various peptides in the autonomic nerve endings, were essentially unobserved in the perikaryal cytoplasm. The present finding provides evidence that one pool of NPY-like immunoreactivity is localized in the Golgi apparatus of colchicine-pretreated as well as normal sympathetic ganglion cells. It is also clear that visualization of NPY-immunoreactive somata by colchicine-pretreatment in the sympathetic ganglia is due to the accumulation of the neuropeptide in the disorganized Golgi stacks instead of increased amount of the large granular vesicles containing NPY.

Animals↗

PubMed1988

An immunohistochemical study of cellular and nervous elements in the taste organ of the bullfrog, Rana catesbeiana.

The cellular and nervous elements of the bullfrog taste organ were examined by immunohistochemical methods using various antibodies. The immunoreactivity for spot 35 protein, a soluble protein isolated from bovine cerebellum, was found in numerous taste cells located at the middle or slightly lower levels within the gustatory cell layer. The immunoreactive cells possessed cytoplasmic processes rising upward the free surface and also issued branched processes to the base of the epithelium. The immunoreaction for spot 35 protein was found diffusely throughout the cytoplasm from the apical to the basal parts of the taste cells. NSE-immunoreactive taste cells were located at the upper or middle levels within the gustatory cell layer in the taste organ. The fact that the cells were smaller in number and size than spot 35 protein-reactive cells and further differed in localization distinguished the NSE-taste cells from the spot 35 protein cells. Serotonin-like immunoreactivity was detectable in the basal cells localized at the base of the taste epithelium. The immunoreactive cells were arranged in a circle at the periphery of the taste organ, each extending a slender process toward the center. The terminal portion of this process spread leaf-like; numerous fine projections protruded from its margin. The serotonin-immunoreactive cells appear to coincide with the monoamine-containing basal cells, which have been previously reported. Substance P-, calcitonin gene-related peptide (CGRP)-, vasoactive intestinal polypeptide (VIP)-, peptide HI (PHI)- or gastrin releasing peptide (GRP)-immunoreactive nerve fibers with varicosities were demonstrated within the taste organ. Some substance P-fibers ran along the bottom of the taste organ epithelium. A few thinner substance P-fibers ascended among the epithelial cells of the organ and terminated closely below the free surface. CGRP-fibers were found to correspond to substance P-fibers from their evidencing a double immunostaining. VIP- and PHI-fibers formed a meshwork in the basal area of the taste epithelium. Abundant substance P- and/or CGRP-fibers formed a meshwork among the ciliated cells located at the periphery of the taste organ. However, PHI- and GRP-fibers were detected less than substance P- and/or CGRP-fibers, though VIP-fibers were rarely present in the same region. Neurofilament protein- or tyrosine hydroxylase-like immunoreactivities were found in thick nerve fibers in the taste organ, whereas no immunoreactivities were present in cellular elements within the taste organ. The relationship between cellular and nervous elements in the taste organ was examined by double immunostainings.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1988

[Establishment and characterization of a new SCC antigen producing cell line (HCS-2) from a carcinoma of the uterine cervix].

The cell line designated HCS-2 established from a squamous cell carcinoma of the uterine cervix has been subcultivated 77 times since Nov. 8, 1983. The cultured cells appear epithelial in shape, with a pavement-like arrangement and grow without contact inhibition. In electron microscopy, the cells are characterized by desmosomal cell contacts and a few tonofilaments. The cells are transplanted subcutaneously to nude mice and produce tumor which resembles the original tumor of large cell non-keratinizing squamous cell carcinoma. The growth rate of subculture has increased gradually, and population doubling time of cells at 17th passage was about 65 hours. The chromosome studies show aneuploidy and chromosomal number was mainly from hypertriploid to hypotetraploid range. The modal number of cells at 33rd passage was 81. Specific marker chromosome is not realized. The production of SCC antigen is detected from the cells and the amount of SCC antigen in cultured media was recorded from 1.5 to 2.0 ng per 1 x 10(4) cells for 48 hours. CEA synthesis is also confirmed immunohistochemically.

Adult↗

PubMed1988

[Cell culture--its application in the study of hormone and endometrial carcinoma and feed-back to clinical medicine].

By statistical study on 135 patients with endometrial carcinoma, it is clarified that the most effective prognostic factor of the cancer is the histological grading. Well differentiated type is best prognostic and possesses hormone receptors. Application of cell culture is one of the most suitable choices in the study of hormone and human endometrial carcinoma. Present paper is to show usefulness of in vitro study by taking example of the above theme. 1) Binding ability of endometrial carcinoma cells to estrogen: Being explained by Gurpide et al. by using HEC-1 cells, the ability is under control of cGMP and cAMP ratio. 2) Responses to estrogen: DNA polymerase alfa of Ishikawa cells which possesses both estrogen and progesterone receptors (ER, PR) is stimulated first showing peak at 18 hours and alkaline phosphatase (ALP) is at 72 hours by E(2)10(-8)M, which is antagonized by OH-tamoxifen. PR level is also enhanced at its maximum after 3 day E2 treatment, and is analyzed by immunocytochemistry with PR mono-clonal antibody as well as biochemical assay. Gorski and Greene's theory that steroid receptor is localized in nuclei is confirmed in endometrial carcinoma. Growth of Ishikawa cells is apparently enhanced in the aspects of shortened cell cycle and unlimited saturation density. 3) Responses to progestogen: Nucleic acid syntheses of HEC-1 are immediately suppressed by progesterone (P) 2.5 microg or more. Electron microscopic findings show appearances of Golgi apparatus and lysosomal granules. Growth suppression is observed in the cell lines regardless of PR positivity. ALP activity of PR-negative HEC-50 cells

Alkaline Phosphatase↗

PubMed1988

[Effects of cepharanthin on leukopenia and thrombocytopenia caused by CDDP-ACR-CPA therapy of ovarian cancer].

Effects of cepharanthin (50 mg/day, i.v.) an alkaloid of bis-coclaurin type, were studied on leucopenia and thrombocytopenia caused by CDDP-ACR-CPA (CAP) therapy against ovarian cancer. The study was carried out with 44 therapy courses on 17 patients of ovarian cancer receiving CAP administration. Nineteen of the 44 courescin loaded cepharanthin. The minimum number of leucocytes was found on 15.2 +/- 4.4 and 17.2 +/- 2.1 days after CAP-administration with and without cepharanthin, respectively. Leucocyte count 21 days after CAP-administration compared to that before the administration were 90.1 and 56.8% in cepharanthin and non-cepharanthin groups, respectively, which indicate a significant difference (p less than 0.05). As to the platelet count, cepharanthin group also showed attendant toward earlier recovery of thrombocytopenia than non-cepharanthin group. We conclude that the administration of cepharanthin during CAP therapy promotes the recovery from leucopenia and thrombocytopenia.

Adult↗

PubMed1987

Calcitonin gene-related peptide (CGRP)-like immunoreactivity in scattered chromaffin cells and nerve fibers in the adrenal gland of rats.

The present immunohistochemical study reveals that a small number of chromaffin cells in the rat adrenal medulla exhibit CGRP-like immunoreactivity. All CGRP-immunoreactive cells were found to be chromaffin cells without noradrenaline fluorescence; from combined immunohistochemistry and fluorescence histochemistry we suggest that these are adrenaline cells. In addition, all CGRP-immunoreactive cells simultaneously exhibited NPY-like immunoreactivity. CGRP-chromaffin cells were characterized by abundant chromaffin granules with round cores in which the immunoreactive material was densely localized. These findings suggest the co-existence of CGRP, NPY and adrenaline within the chromaffin granules in a substantial number of chromaffin cells. Thicker and thinner nerve bundles, which included CGRP-immunoreactive nerve fibers, with or without varicosities, penetrated the adrenal capsule. Most of them passed through the cortex and entered the medulla directly, whereas others were distributed in subcapsular regions and among the cortical cells of the zona glomerulosa. Here the CGRP-fibers were in close contact with cortical cells. A few of the fibers supplying the cortex extended further into the medulla. The CGRP-immunoreactive fibers in the medulla were traced among and within small clusters of chromaffin cells and around ganglion cells. The CGRP-fibers were directly apposed to both CGRP-positive and negative chromaffin cells, as well as to ganglion cells. Immunoreactive fibers, which could not be found close to blood vessels, were characterized by the presence of numerous small clear vesicles mixed with a few large granular vesicles. The immunoreactive material was localized in the large granular vesicles and also in the axoplasm.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenal Glands↗

PubMed1987

Estradiol metabolism in Ishikawa endometrial cancer cells.

Estrogen-responsive human cells derived from a specimen of well differentiated endometrial adenocarcinoma (Ishikawa line) were incubated with [3H]estradiol (E2) at various concentrations and the medium was sampled at 3, 6 and 24 h to evaluate the kinetics of removal of the hormone and the formation of unconjugated or sulfated metabolites. The detectable products of metabolism were estrone and the conjugate estradiol-3-sulfate. The latter was identified by high pressure chromatography, before and after acetylation, oxidation, and hydrolysis. The disappearance of [3H]E2 from the medium was found to follow first order kinetics between 3 and 24 h, with half-lives increasing from 4.7 to 53 h as the initial concentrations of the hormone were raised from 10(-8) to 10(-6)M. At the lowest concentration, practically all of the [3H]E2 added to the cultures was converted to estradiol-3-sulfate in 24 h, whereas at 10(-6)M oxidation to estrone was quantitatively more important than sulfation. These results indicate the presence in Ishikawa cells of an estrogen sulfotransferase of low Michaelis constant for E2, and 17 beta-oxidoreductase activity that significantly contributes to the metabolism of E2 only at higher concentrations of substrate.

Adenocarcinoma↗