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Biomedical subjects

J Kato

Publications and source records attributed to J Kato.

At least 325 records · Page 18Linked to original sources
PubMed1994

Inhibition of cyclin-dependent kinases by purine analogues.

While testing purines related to the non-specific protein kinase inhibitors N6-dimethylaminopurine and N6-(delta 2-isopentenyl)adenine as potential inhibitors of the p34cdc2/cyclin B kinase, we discovered a compound with high specificity, 2-(2-hydroxyethylamino)-6- benzylamino-9-methylpurine (olomoucine). Kinetic analysis of kinase inhibition reveals that olomoucine behaves as a competitive inhibitor for ATP and as a non-competitive inhibitor for histone H1 (linear inhibition for both substrates). The kinase specificity of this inhibition was investigated for 35 highly purified kinases (including p34cdk4/cyclin D1, p40cdk6/cyclin D3, cAMP-dependent and cGMP-dependent kinases, eight protein kinase C isoforms, calmodulin-dependent kinase II, myosin light-chain kinase, mitogen-activated S6 kinase, casein kinase 2, double-stranded RNA-activated protein kinase, AMP-stimulated kinase, eight tyrosine kinases). Most kinases are not significantly inhibited. Only the cell-cycle regulating p34cdc2/cyclin B, p33cdk2/cyclin A and p33cdk2/cyclin E kinases, the brain p33cdk5/p35 kinase and the ERK1/MAP-kinase (and its starfish homologue p44mpk) are substantially inhibited by olomoucine (IC50 values are 7, 7, 7, 3 and 25 microM, respectively). The cdk4/cyclin D1 and cdk6/cyclin D3 kinases are not significantly sensitive to olomoucine (IC50 values greater than 1 mM and 150 microM, respectively). N6-(delta 2-Isopentenyl)adenine is confirmed as a general kinase inhibitor with IC50 values of 50-100 microM for many kinases. The purine specificity of cyclin-dependent kinase inhibition was investigated: among 81 purine derivatives tested, only C2, N6 and N9-substituted purines exert a strong inhibitory effect on the p34cdc2/cyclin B kinase. An essentially similar sensitivity to this olomoucine family of compounds was observed for the brain-specific cdk5/p35 kinase. Structure/activity relationship studies allow speculation on the interactions of olomoucine and its analogues with the kinase catalytic subunit. Olomoucine inhibits in vitro M-phase-promoting factor activity in metaphase-arrested Xenopus egg extracts, inhibits in vitro DNA synthesis in Xenopus interphase egg extracts and inhibits the licensing factor, an essential replication factor ensuring that DNA is replicated only once in each cell cycle. Olomoucine inhibits the starfish oocyte G2/M transition in vivo. Through its unique selectivity olomoucine provides an anti-mitotic reagent that may preferentially inhibit certain steps of the cell cycle.

Animals↗
PubMed1994

[Gene expression and function of progesterone receptor isoforms].

Progesterone receptor is unique in terms of the existence of two isoforms of A and B. The isoforms are expressed by two distinct promoters in the peripheral as well as the brain. The 5'-untranslated region and the coding region around ATG 1 of rat progesterone receptor form B as well as the ligand binding domain are cloned. The function of the forms A and B is still unclear. However, it has been reported that the form A is a hormone-dependent transdominant repressor of the B function under certain circumstance of the cell. This model is very interesting to elucidate the molecular mechanism of progesterone action.

Animals↗
PubMed1994

Structural analyses of DNA fragments integrated by illegitimate recombination in Schizosaccharomyces pombe.

In order to elucidate the mechanisms of illegitimate recombination in eukaryotes, we have studied the structure of DNA fragments integrated by illegitimate recombination into the genome of fission yeast. Nonhomologous recombination was rarely identified when a long region of homology with the chromosomal leu1+ gene was present in the introduced leu1::ura4+ DNA fragment; but a decrease in length of homology leads to an increase in the ratio of non-homologous to homologous recombination events. The introduced DNA fragments were integrated into different sites in the chromosomes by nonhomologous recombination. The results suggested that there are multiple modes of integration; most events simply involve both ends of the fragments, while in other cases, fragments were integrated in a more complicated manner, probably via circularization or multimerization. To analyze the mechanism of the major type of integration, DNA fragments containing the recombination junctions of three recombinants were amplified by inverted polymerase chain reaction (IPCR) and their nucleotide sequences were determined. There was no obvious homology between introduced DNA and chromosomal DNA at these recombination sites. Furthermore it was found that each terminal region of the introduced DNA was deleted, but that there were no or very small deletions in the target sites of chromosomal DNA. Two models are proposed to explain the mechanism of nonhomologous integration.

Base Sequence↗
PubMed1994

Expression and extracellular release of transferrin receptors during peripheral erythroid progenitor cell differentiation in liquid culture.

The expression and extracellular release of transferrin receptor (TR) was investigated by in vitro model system of erythroid differentiation. Human peripheral blood mononuclear cells were cultured with interleukin-3 (IL-3) for 7 days, and with erythropoietin (EPO) for an additional 8 days. After EPO stimulation, IL-3-stimulated blastic cells were serially differentiated into mature erythrocytes. [3H]-thymidine incorporation of cultured cells increased linearly from day 0 to 5, followed by a decrease. Flow cytometric analysis showed an increase of TR expression from day 0 to 5, followed by a slight decrease. By metabolic labeling with [35S]methionine and immunoprecipitation, the cell lysate exhibited a 95-kD band corresponding to the intact TR on sodium dodecyl sulfate-polyacrylamide gel electrophoresis/autoradiography at day 5, when polychromatic erythroblasts had their peak. The culture supernatant solubilized by tween-20 exhibited a 95-kD and an 85-kD band on days 5 and 8, which corresponded to the intact and the truncated forms of TR, respectively. The 95-kD band was more intense at day 5 than at day 8. The reverse transcriptase-polymerase chain reaction assay showed that the receptor-mRNA expression was parallel to receptor synthesis. Thus, the synthesis and expression of TR on erythrocytes is associated mainly with cell proliferation in the early phase, and with both cell proliferation and hemoglobin production in the middle to late phases of maturation. Concomitantly, the extracellular release of TR from erythrocytes occurs in the middle to late phases of maturation. These data suggest that polychromatic erythroblasts release soluble TR as both intact and truncated forms and may be an important source of serum TR implicated as an index for erythropoietic activity in the marrow.

Cell Differentiation↗
PubMed1994

Gene expression of progesterone receptor isoforms in the rat brain.

Progesterone receptors (PRs) are known to exist in two forms: a larger molecular, form B, and a smaller one, form A. Rat PR cDNA corresponding to the region around the translation-initiation site (ATGB) of the putative PR form B mRNA was cloned, together with cloning of steroid-binding domain of the PR forms A and B. An imperfect "estrogen responsive element," GGTCG*** TGACT, was located around ATGB of the rat PR cDNA. The distribution of PR mRNA-containing neurons was mapped in the female adult rat brain by in situ hybridization, which was largely in agreement with that of PR proteins. Differential intracerebral distribution of the mRNAs, measured by the quantitative RT-PCR Southern blotting assay, was found between the total (A+B) and form B mRNA levels, indicating possible distinct mechanisms responsible for regulation of the expression of the PR mRNAs. A region-specific and stage-related gene expression of form B seems predominantly to be "turned on" first around birth, followed by that of form A around Days 8-12. The postnatal developmental pattern of PR form B mRNA in the cerebral cortex resembled that of the PR proteins. Noninducibility of the cortical PR by estrogen might be ascribable to the predominance of form B mRNA, which was reported to have little or no estrogen inducibility. A missmatch existed between PR from A and its mRNA levels, suggesting some impairment of the synthesis of form A. Thyroid hormone may be a regulator of gene expression of the receptor in the developing cortex.

Amino Acid Sequence↗
PubMed1994

Extracellular proteolytic activity of Cryptococcus neoformans.

Eight strains of Cryptococcus neoformans var. neoformans isolated from AIDS patients in the Infectious Disease Institute, University of Turin, Italy, were examined for growth and extracellular proteolytic activity in culture with solid and liquid media. All of the strains grew well on Yeast Carbon Base (YCB) agar medium supplemented with both 0.1% (w/v) bovine serum albumin (BSA) and 0.01% (w/v) polypeptone (Pp), and produced a clear proteolytic zone around their colonies, whereas they exhibited less growth and proteolytic activity on YCB medium supplemented with BSA alone. Strain #8 with a strong proteolytic activity was cultured in three different liquid media. Its growth was limited in YCB medium supplemented with 0.1% BSA, but was moderate in that with 0.01% Pp. Enhanced growth was supported by the addition of both BSA and Pp to the YCB medium. The relative value of the final cellular yields obtained with the above YCB-0.1% BSA, YCB-0.01% Pp and YCB-0.1% BSA-0.01% Pp media was approximately 1:10:20. In the culture with YCB medium containing both BSA and Pp, a rapid decrease in the amount of BSA was demonstrated by a spectrophotometric assay and gel electrophoresis of the culture supernatant after the log-to-stationary phase. The proteolytic activity in the culture supernatant became detectable after the log phase when tested with skim milk agarose plates. These results allowed us to conclude that Cr. neoformans var. neoformans is able to secrete protease and to utilize protein as a source of nitrogen.

Cryptococcus neoformans↗
PubMed1994

Regulation of asialoglycoprotein receptor synthesis by inflammation-related cytokines in HepG2 cells.

The asialoglycoprotein receptor (AGPR) is responsible for the catabolism of acute phase proteins. The effects of inflammation-related cytokines on the expression of AGPR were investigated in HepG2 cells derived from a human hepatoblastoma cell line. Binding studies, using a [125I]-labeled asialo-orosomucoid ligand, revealed that AGPR numbers on cell surfaces were increased by interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor (TNF). In cells treated with IL-1, IL-6, or TNF, immunohistochemical staining with an anti-AGPR monoclonal antibody demonstrated augmented expression. Pulse labeling analysis, using [35S]-labeled methionine, showed newly synthesized AGPR in both the precursor and the mature forms. When IL-1, IL-6, and TNF were added to the culture medium, total synthesis of AGPR (sum of the mature and precursor forms) was increased. In addition, the relative proportion of the synthesized precursor form of AGPR was higher in cytokine-treated than in untreated cells, suggesting that these cytokines augment the synthesis of AGPR, particularly in the stage prior to glycosylation.

Acute-Phase Proteins↗
PubMed1994

Ethanol inhibits asialoglycoprotein receptor synthesis but augments its mRNA expression in a human hepatoma cell line.

The effect of ethanol on the expression of asialoglycoprotein receptor protein and its mRNA was studied in a human hepatoma cell line, HepG2. The number of asialoglycoprotein receptors on the cell surface was decreased to 60% of the control level, without a loss in affinity, by incubating the cells with 100 mM ethanol. The decrease in cell surface asialoglycoprotein receptors was paralleled by a decrease in total receptor numbers, including intracellular and surface receptors. The internalization of asialoglycoprotein was also diminished, to 44% of that in control cells. The decreases in cell surface receptors and total receptor numbers were partially restored by 2 mM 4-methylpyrazole, suggesting that ethanol metabolites participated in the diminution of asialoglycoprotein receptor expression. However, the steady-state expression of asialoglycoprotein receptor mRNA was increased in ethanol-treated cells and further augmented by a longer ethanol exposure. These paradoxical results, i.e., the decrease of asialoglycoprotein receptor protein and the increase of its mRNA expression, suggest that the reduction in the asialoglycoprotein receptor protein is a post-transcriptional event and that a possible feedback regulatory mechanism may control asialoglycoprotein receptor gene transcription and/or impair the degradation of its mRNA.

Alcohol Dehydrogenase↗
PubMed1994

Induction of Fos protein in the rat trigeminal nucleus complex during an experimental tooth movement.

The induction and temporal changes of Fos protein in the rat spinal trigeminal nucleus complex during experimental tooth movement were studied immunohistochemically. Separating elastics were unilaterally inserted between the upper molars. The animals were perfused at 0, 1, 2, 4, 8, 12 and 24 h thereafter, and the brains containing the mesencephalic trigeminal nucleus and the spinal trigeminal nucleus complex were then removed. Cells showing Fos immunoreactivity were observed in the superficial layers of the subnucleus caudalis on the ipsilateral side except at 0 and 24 h. The numbers of the immunoreactive cells peaked 2 and 4 h after the insertion. On the contralateral side, only a few immunoreactive cells were observed in the subnucleus caudalis of the 1-, 2- and 4-h groups. The subnucleus interporalis and the subnucleus oralis of the spinal trigeminal nucleus complex, the principal trigeminal nucleus and the mesencephalic trigeminal nucleus of the experimental animals and all the trigeminal nuclei of the control animals contained no immunopositive cells. Thus experimental tooth movement induced a Fos protein in the superficial layers of the subnucleus caudalis of the spinal trigeminal nucleus complex. The subnucleus caudalis may modulate pain induced by tooth movement.

Animals↗
PubMed1994

Gastroesophageal reflux occurring after repair of congenital diaphragmatic hernia.

Over the past 2 decades, 110 patients with congenital diaphragmatic hernia (CDH) were treated in the authors' hospital. Eighty-six survived; of these, 10 patients (11.6%) had gastroesophageal reflux (GER) after repair of CDH. Seven occurred in the past 5 years, during which time advanced intensive care including extracorporeal membrane oxygenation (ECMO) was used. Vomiting started within 4 weeks after repair of CDH in eight cases, and hiatal hernia was demonstrated in six cases. Three patients responded to conservative therapy; the other seven required antireflux surgery. Several factors are believed to be possible causes of the development of GER in CDH cases. Among them, slow pulmonary expansion of the affected side was thought to be the most important. Namely, in a case of CDH associated with severe hypoplastic lung, the esophagus may be deviated to the affected side before the lung is expanded. After expansion, the abdominal esophagus shortens, and GER or a hiatal hernia can occur in severe cases. There were seven such patients in our series of 10. With the increase in the survival rate of CDH cases associated with severe hypoplastic lung, the number of such patients also may increase. Therefore, some additional procedure to prevent the lower esophagus from sliding will be necessary in the repair of diaphragmatic hernia.

Female↗
PubMed1994

Effects of hypothyroidism on the gene expression of progesterone receptors in the neonatal rat brain.

The effects of neonatal hypothyroidism on the gene expression of progesterone receptors (PRs) in the 8-day-old female rat brain were examined. The levels of PR ('A'+'B')-, PR ('B')-, estrogen receptor (ER)- and beta-actin mRNAs in the cerebral cortex and hypothalamus-preoptic area of propylthiouracil-treated rats (PTU group) or untreated rats (control group) were analyzed using a quantitative reverse transcription-polymerase chain reaction-Southern blotting assay. When the levels of PR mRNAs were calibrated by respective levels of beta-actin mRNA, the levels of PR ('A'+'B')- and PR ('B') mRNAs in the cerebral cortex of the PTU group were markedly less than those in the control group with no significant changes in the levels of PR messages in the hypothalamus-preoptic area of both groups. No significant difference in the calibrated levels of ER mRNA between both groups was found in these tissues. These results on the PR-, and ER mRNAs were essentially similar to those on the levels of PR- and ER proteins previously reported suggesting that thyroid hormones affect the transcriptional machinery of PR in the developing brain, in a region-specific manner.

Animals↗
PubMed1994

Distribution and postnatal changes of aromatase mRNA in the female rat brain.

Distribution and postnatal changes of aromatase mRNA in the Wistar strain female rats brain were investigated to elucidate the mechanism of the region-specific and stage-related regulations of brain aromatase activity. Total RNAs extracted from the hypothalamus-preoptic area (HPOA), amygdala, cerebral cortex, cerebellum and anterior hypophysis were subjected to a quantitative reverse transcription-polymerase chain reaction-Southern blotting (RT-PCR-SB) assay. The levels of aromatase mRNA were as follows: amygdala > cerebral cortex not equal to HPOA >> cerebellum not equal to anterior hypophysis. These results roughly paralleled the distribution of the aromatase activity and the aromatase protein reported previously, with the exception of that of the cerebral cortex. The postnatal developmental patterns of aromatase mRNA in the HPOA and cerebral cortex were further studied. The levels of aromatase mRNA in the HPOA tended to be high around birth (day 0) and to decrease at days 4-8 followed by no remarkable change to an adult. The developmental pattern in the HPOA was essentially similar to that of aromatase activity. On the other hand, in the cerebral cortex where very low or no aromatase activity was reported throughout developmental stages, aromatase mRNA was low but detectable level around day 0, thereafter gradually increased to an adult level. From these results, it is inferred that synthesis of the aromatase protein by the level of the mRNA seems to mainly regulate the aromatase activity in a region-specific and stage-related fashion in most parts of the rat brain, except for the cerebral cortex where the posttranscriptional regulation may play an important role.

Animals↗
PubMed1994

Androgen receptor mRNA in the rat ovary and uterus.

The distribution of androgen receptor messenger RNA (ARmRNA) in the reproductive tissues of adult rats was examined by Northern blot analysis and in situ hybridization using ARcRNA probes corresponding to the androgen binding domain of the receptor. About 10-kilobase rat ARmRNA was observed in all tissues examined in the Northern blot analysis. The amount of ARmRNA in the ovary, uterus and testis was less than that in the prostate. In the in situ hybridization study, extensive labeling was observed in the theca cells of the ovary (proestrous) and the endometrium and endometrial glands of the uterus (proestrous). Moderate labeling was observed in the granulosa cells and stromal cells of the ovary and in the myometrium of the uterus. These results were largely in agreement with the distribution of AR previously reported by ligand binding studies. This present in situ hybridization study may provide a useful tool for the analysis of the regional regulation of AR synthesis in the rat female reproductive tissues.

Animals↗
PubMed1994

Transforming growth factor beta 1 secreted from scirrhous gastric cancer cells is associated with excess collagen deposition in the tissue.

To explore the mechanism of increased collagen deposition in scirrhous carcinoma of the stomach, an attempt was made to define the role of transforming growth factor beta 1 (TGF-beta 1), secreted from tumour cells, as a possible humoral factor which functions in a paracrine manner to stimulate the production of collagen in regional fibroblasts. Immunohistochemical staining revealed that tumour cells in scirrhous carcinomas were generally stained more intensively than those in other types of carcinomas. On Northern blot analysis the tumour cells established from scirrhous carcinoma (KATO-III, OCUM-1 and HSC-39) exhibited relatively strong signals compared with those from non-scirrhous carcinoma (MKN-28 and MKN-45). In the culture media of scirrhous carcinoma cells, the active form of TGF-beta 1 was detected, while in those of the non-scirrhous carcinoma cells the latent form was demonstrated by both colony and radioreceptor assays. The culture medium from KATO-III showed strong stimulating activity of collagen synthesis in fibroblasts, and this activity was partially neutralised by an anti-TGF-beta 1 antibody. These results suggest that tumour cells in scirrhous carcinoma produce more active-form TGF-beta 1 than does non-scirrhous carcinoma and thus is partially responsible for the observed enhanced collagen deposition in the region.

Adenocarcinoma, Scirrhous↗
PubMed1994

Close correlation between the dephosphorylation of p53 and growth suppression by transforming growth factor-beta 1 in nasopharyngeal carcinoma cells transduced with adenovirus early region genes.

The mechanism of growth inhibition by transforming growth factor (TGF)-beta 1 was investigated. We examined the growth inhibitory effects of TGF-beta 1 on human nasopharyngeal carcinoma (KB) cells which constitutively expressed p53. TGF-beta 1 suppressed the DNA synthesis of KB cells in a dose-dependent manner. It had minimal effect on adenovirus-2-transduced KB cells expressing either adenovirus early region 1B (E1B) or 1A (E1A) product, which respectively binds to p53 or Rb product and inhibits its function, and no growth inhibition at all was observed with KB cells expressing both E1B and E1A products. Dephosphorylation of the p53 was promoted by TGF-beta 1 stimulation in KB cells, but not in E1B-producing KB cells, which sequestrate the function of p53. The growth inhibition of KB cells by TGF-beta 1 was significantly reduced by treatment with okadaic acid. These results suggest that p53 transduces the antiproliferative signal of TGF-beta 1 possibly through its dephosphorylation.

Adenovirus E1A Proteins↗
PubMed1994

Antisense oligodeoxynucleotides for IL-2, c-myc and transferrin receptor synchronize mitogen-activated lymphocytes in the G1 phase.

In order to clarify the role and interrelationship of c-myc, interleukin-2 (IL-2), and transferrin receptor (TfR) expressions on PHA-stimulated lymphocyte proliferation, we examined the effects of antisense oligodeoxynucleotides directed against c-myc, TfR, and IL-2 mRNAs on DNA synthesis and cell-cycle phase. Exposure of PHA-stimulated lymphocytes to each antisense oligomer resulted in approximately 75-80% inhibition of DNA synthesis. TfR expression was not inhibited in PHA-stimulated lymphocytes by c-myc or IL-2 antisense oligonucleotides, suggesting that the expression of c-myc, TfR, and of IL-2 is regulated by an independent mechanism. All three antisense oligonucleotides for c-myc, TfR and IL-2 synchronized mitogen-activated lymphocytes to the G1 phase as assessed by morphologic blast formation and cell-cycle phase analysis.

Base Sequence↗