Biomedical subjects

H Kizaki

Publications and source records attributed to H Kizaki.

At least 37 records · Page 2Linked to original sources

PubMed1997

Inflammation and apoptosis.

Apoptosis is a highly regulated form of programmed cell death defined by distinct morphological and biochemical features; it plays an important role in embryogenesis and the maintenance of cellular and tissue homeostasis in multicellular organisms. Its perturbation has been implicated in a wide range of patho-physiological stages, including autoimmune, bacterial, and viral disease, and degenerative disorders. The critical role of apoptosis in eliminating harmful or injured cells from tissues suggests its participation in inflammatory processes and in the resolution of inflammatory reactions. In this article, we briefly review the molecular mechanism of apoptosis and the role of apoptosis in inflammation.

Animals↗

PubMed1997

Protein synthesis dependent and independent apoptosis of murine splenic T cells.

To elucidate the mechanism of apoptosis in matured T cells, we examined the effect of phorbol esters capable of activating protein kinase C, cAMP agonists, calcium ionophores, and dexamethasone on apoptosis in mouse splenic T cells. All of these agents induced apoptosis in thymocytes. Neither phorbol esters nor calcium ionophores induced apoptosis in the splenic T cells. Dibutyrylcyclic AMP induced apoptosis in splenic T cells but to a lesser extent than that in thymocytes. The results suggest that matured T cells are more resistant to various apoptogenic stimuli than thymocytes. Dexamethasone induced apoptotic cell death in splenic T cells as well as in thymocytes. Cycloheximide, an inhibitor of protein synthesis which completely inhibited thymocyte apoptosis, induced slight apoptosis in splenic T cells, but it inhibited the dexamethasone-induced apoptosis to a level at which cycloheximide induced apoptosis, indicating that at least two pathways of apoptosis exist in peripheral T cells; one protein synthesis dependent and the other independent. Taken together, the results suggest that the apoptosis of peripheral T cells is regulated by a mechanism different from that of thymocytes.

Animals↗

PubMed1996

Molecular cloning of the genes suppressed in RVC lymphoma cells by topoisomerase inhibitors.

Etoposide is a topoisomerase II inhibitor that induces DNA cleavable complex and has been used as an antitumor drug. We isolated two genes that were transcriptionally suppressed at an early stage of incubation in etoposide-treated RVC lymphoma cells, using modified PCR-based subtractive hybridization. Sequencing revealed that one of these genes, which was approximately 1.7 kb and which encoded a protein of 320 amino acids, was identical to hnRNP A1. The other was a novel gene of about 2.2 kb encoding a protein of 469 amino acids. These genes were also down-regulated in the cells incubated with camptothecin, a topoisomerase I inhibitor that induces DNA single strand breaks, but not in those exposed to ICRF-154, a topoisomerase II inhibitor that does not induce DNA cleavable complex formation. These results suggest that the early down-regulation of these genes contributes to the cytotoxicity of the topoisomerase inhibitors that induce DNA cleavage.

Amino Acid Sequence↗

PubMed1996

Cloning and sequence analysis of the pfl gene encoding pyruvate formate-lyase from Streptococcus mutans.

We have isolated a sorbitol-negative mutant of Streptococcus mutans GS-5 following random mutagenesis with plasmid pVA891 clone banks. This mutant did not metabolize sorbitol anaerobically but did so aerobically. A 10-kb chromosomal DNA fragment flanking the pVA891 insertion was deleted in this mutant. The corresponding region from the parental strain GS-5 was then recovered by a marker rescue method with Escherichia coli. The pyruvate formate-lyase gene, pfl, was identified within a 3-kb PstI-XbaI fragment located in the middle of the deleted region of the chromosome, and its inactivation in S. mutans produced the same sorbitol-negative phenotype. Nucleotide sequence analysis of the pfl gene revealed a 2.3-kb open reading frame (ORF) preceded by potential ribosome-binding and promoter-like sequences. The ORF specified a putative protein of 775 amino acid residues with a calculated molecular weight of 87,533. The amino acid sequence deduced from the ORF exhibited significant similarity to that of the E. coli pfl gene.

Acetyltransferases↗

PubMed1996

Establishment and characterization of cell lines, MeMS and MeFS, derived from spontaneous rat mesotheliomas.

New cell lines, designated MeMS and MeFS, were established from the epithelial type of mesotheliomas occurring spontaneously in aged male and female Fischer 344 rats, respectively. Morphologically, MeMS and MeFS expressed the epithelial phenotype. Coexpression of keratin and vimentin in both cell lines was confirmed by the immunocytochemistry, one-dimensional gel electrophoresis and immunoblotting for intermediate filament proteins. Hyaluronic acid synthesis of both MeMS and MeFS was demonstrated by the analysis using high-performance liquid chromatography. MeMS and MeFS seem to be useful models for studying the biological nature of rat mesotheliomas.

Aging↗

PubMed1995

Effects of protein tyrosine kinase inhibitors with different modes of action on topoisomerase activity and death of IL-2-dependent CTLL-2 cells.

We studied the effects of protein tyrosine kinase inhibitors with different modes of action on topoisomerase activity and cell death in CTLL-2 cells, whose growth is IL-2-dependent. The Flavonoids genistein, biochanin A, and apigenin inhibited topoisomerase II to the same extent as etoposide, a specific inhibitor of the enzyme. Methyl 2,5-dihydroxycinnamate (2,5-MeC) also inhibited topoisomerase II, but was less potent than genistein. Herbimycin A and staurosporine did not inhibit topoisomerase II. None of the inhibitors of protein tyrosine kinases examined inhibited topoisomerase I activity. All the inhibitors induced cell death with internucleosomal DNA fragmentation in the presence of IL-2. Genistein, biochanin A, and apigenin induced DNA fragmentation and cell death early in the incubation period and did not alter the profiles of phosphotyrosine proteins in either the lysate or pelleted fractions, indicating that the early cell death was induced by the inhibition of topoisomerase II activity rather than by the inhibition of protein tyrosine kinase activity. 2,5-MeC similarly induced early cell death and DNA fragmentation, but to a lesser extent than genistein presumably due to the inhibition of topoisomerase II activity. Herbimycin A induced a slow increase in DNA fragmentation and cell death, accompanied by a decrease in phosphotyrosine proteins in the pelleted fraction, suggesting that the inhibition of protein tyrosine phosphorylation, presumably of the nuclear proteins, is related to cell death and DNA fragmentation. Staurosporine-induced DNA fragmentation appeared to be due to mechanism(s) other than the inhibition of topoisomerases and protein tyrosine kinases, since it neither altered the profiles of phosphotyrosine proteins nor inhibited topoisomerase activity.

Animals↗

PubMed1995

Establishment and characterization of a cell line (NB-YK) derived from a transplantable rat nephroblastoma.

A new cell line, designated NB-YK, was established from a transplantable rat nephroblastoma (NB-Y) which was derived from a spontaneous nephroblastoma in an aged Fischer 344 rat. NB-YK grew in a piling-up and noncohesive pattern on the plastic surface and formed colonies in a soft agar. The main cell type of NB-YK represented morphology of mesenchymal phenotype and most of the cells contained several secretory granules in their cytoplasm. Immunocytochemically the cells were positive for vimentin, cytokeratin, and laminin. Coexpression of vimentin and cytokeratin in the cells was confirmed by the one-dimensional gel electrophoresis and immunoblotting for intermediate filament proteins. NB-YK cells were tumorigenic and produced fibrosarcoma-like tumor when inoculated subcutaneously or intraperitoneally into syngeneic rats and nude mice. NB-YK seems to be a useful model for studying biological properties of nephroblastoma.

Animals↗

PubMed1995

Biphasic effect of staurosporine on thymocyte apoptosis.

When mouse thymocytes were incubated with staurosporine at low doses (1-100 nM), apoptosis was induced dose- and time-dependently. Staurosporine-induced apoptosis was dependent on macromolecular synthesis, and it was also dependent on protein phosphorylation sensitive to 1-(5-isoquinolinesulfonyl-2-methylpiperazine dihydrochloride (H-7). Whereas, staurosporine at high doses (above 500 nM) did not induce significant DNA fragmentation, rather it inhibited the DNA fragmentation induced by 12-O-tetradecanoyl-13-acetate, A23187, and dibutyrylcyclic AMP, as H-7 did. K252a, a derivative of staurosporine, induced apoptosis, which was inhibited by H-7, even at high doses. These results indicate that staurosporine had a biphasic effect on thymocyte apoptosis, a stimulatory effect at low concentration, and an inhibitory effect at high concentration. K252a had only the former action.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1995

[Molecular mechanisms of apoptosis in the immune system].

Apoptosis is a controlled death process under regulation by multiple death and survival genes. In the immune systems, apoptosis has important roles in establishment of the mature immune system (embryological programmed cell death), in dynamic maintenance of functional cell number (physiological programmed cell death) and in elimination of unwanted cells (biochemical programmed cell death). The process of apoptosis is regulated by multiple genes and their products depending on different stimuli. The current knowledge on the molecular events of apoptotic cell death, particularly in thymocytes, is reviewed.

Apoptosis↗

PubMed1995

Staurosporine-induced thymocyte apoptosis is inhibited by herbimycin A, a specific inhibitor of protein tyrosine kinase.

When mouse thymocytes were incubated with staurosporine at low doses (1-100 nM), increased tyrosine phosphorylation of proteins with a molecular weight of 31-34 kD in the detergent-soluble fraction was observed, together with an increase in internucleosomal DNA fragmentation. Incubation with staurosporine for 6 h was enough to trigger and progress of apoptosis. Herbimycin A, a specific inhibitor of protein tyrosine kinases, and cycloheximide, an inhibitor of protein synthesis, reduced the amount of the phosphotyrosine proteins, this being followed by the inhibition of DNA fragmentation and cell death, suggesting a close relationship between protein tyrosine phosphorylation and the induction of apoptosis by staurosporine.

Alkaloids↗

PubMed1994

Multiplication of Clostridium botulinum in dead honey-bees and bee pupae, a likely source of heavy contamination of honey.

Multiplication of Clostridium botulinum in honey-bees was examined to explain the heavy contamination of honey which may occur with this pathogen. When dead bees were inoculated with C. botulinum spores at levels of 10(2)-10(3) and incubated aerobically for 10 days, the organisms increased to 10(4)-10(5). When botulinum spores were inoculated together with Bacillus alvei, the growth of most strains was significantly enhanced (10(5)-10(7)). Similar results were obtained in bee pupae, but not in bee larvae. The heavy contamination of honey with botulinum spores that we have sometimes encountered may have been caused by contamination from dead bees in which C. botulinum had proliferated.

Animals↗

PubMed1994

bis(2,6-dioxopiperaxine) derivatives, topoisomerase II inhibitors which do not form a DNA cleavable complex, induce thymocyte apoptosis.

Internucleosomal DNA fragmentation and cell death were induced dose- and time-dependently by incubation of mouse thymocytes with bis(2,6-dioxopiperazine) derivatives, ICRF-154 and MST-16, inhibitors of topoisomerase II, which do not induce cleavable complex formation. The process was inhibited by actinomycin D and cycloheximide, indicating that the process was an active apoptotic process. Bis(2,6-dioxopiperazine) derivatives have been known to inhibit the etoposide-induced DNA cleavage, but ICRF-154 did not inhibit etoposide-induced apoptosis in thymocytes at 6 h incubation, suggesting that DNA cleavage is not essential for induction of apoptosis by topoisomerase II inhibitors. The alteration of DNA helicity induced by a subtle inhibition of topoisomerase II activity may have an important role in the induction of apoptosis in thymocytes, since topoisomerase II is a major component of the nuclear matrix that can regulate gene expression.

Animals↗

PubMed1994

[Apoptosis and diseases].

Apoptosis, a form of cell death defined by morphological and biochemical characteristics, is involved in many important physiological phenomena including embryogenesis, organ involution, maintenance of homeostasis and biodefense systems of the body. Impaired regulation of apoptosis may play a part in the etiology of malformation and disfunction of various organs causing autoimmune diseases, immunodeficiency diseases or neurodegenerative diseases. Apoptosis is also related to cancer. Elucidation of the mechanism of apoptosis may provide new insights into the pathophysiology of a variety of diseases, and its specific regulation may lead to exciting new therapies for these diseases.

Animals↗

PubMed1993

Isolation and sequence analysis of the pmi gene encoding phosphomannose isomerase of Streptococcus mutans.

A gene encoding a phosphomannose isomerase from Streptococcus mutans GS-5 was identified immediately downstream from the fructokinase gene, scrK. Nucleotide sequence analysis of this region revealed an open reading frame (ORF) specifying a putative protein of 316 amino acids. The gene cloned in Escherichia coli expressed strong phosphomannose isomerase activity. The deduced amino acid sequence of the pmi gene has no significant similarity with any of the previously reported phosphomannose isomerase enzymes. Insertional inactivation of the upstream gene, scrK, in S. mutans also drastically reduced phosphomannose isomerase activity and the ability of the organism to utilize mannose as a sole carbon source. These results suggest that the S. mutans pmi gene constitutes an operon with the scrK gene.

Amino Acid Sequence↗

PubMed1993

Topoisomerase inhibitors induce apoptosis in thymocytes.

The effects of the inhibitors of topoisomerase I and II, camptothecin and etoposide, as well as novobiocin and adriamycin, on the DNA fragmentation and viability of mouse thymocytes in primary culture were examined. All inhibitors were shown to produce dose-dependent internucleosomal DNA cleavage by resolving isolated DNA by agarose-gel electrophoresis. The DNA fragmentation seemed to precede cell death, determined on the basis of LDH release, by a few hours. Etoposide-induced DNA fragmentation progressively increased after incubation and was enhanced by pretreatment with phorbol 12,13-dibutyrate, a phorbol ester capable of activating protein kinase C, whereas camptothecin-induced DNA fragmentation increased progressively after 12 h incubation and was unaffected by phorbol 12,13-dibutyrate-pretreatment. The process was also energy-dependent and required RNA and protein synthesis and protein phosphorylation, since it was inhibited by sodium azide, actinomycin D, cycloheximide and 1-(5-isoquinoline-sulfonyl)-2-methylpiperazine hydrochloride, a protein kinase inhibitor. DNA fragmentation was also inhibited by zinc ions, suggesting the involvement of a specific endonuclease in DNA cleavage. These phenomena are similar to those detected in thymocytes undergoing apoptosis following exposure to glucocorticoids (Cohen, J.J. and Duke, R.C. (1984) J. Immunol. 132, 38-42). Considering that topoisomerases function in cellular proliferation and differentiation by altering DNA topology, the results suggest that topoisomerases have important roles in T-lymphocyte ontogeny in the thymus and are in part involved in the elimination of autoreactive or harmful cells by an apoptotic process.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1993

An assay method for DNA topoisomerase activity based on separation of relaxed DNA from supercoiled DNA using high-performance liquid chromatography.

A method for assaying the activities of DNA topoisomerases I and II has been developed. The assay for type I and II enzymes is based on monitoring relaxation of supercoiled plasmid DNA in the absence or presence of ATP, respectively. The reaction product, relaxed DNA, was separated from the substrate, supercoiled pBR329 plasmid, by a linear gradient of NaCl using high-performance ion-exchange chromatography with a DEAENPR column. In this method, nanogram amounts of relaxed DNA are detectable within 30 min, indicating that a subtle change in the activity in the cells can be assayed precisely.

Adenosine Triphosphate↗

PubMed1993

Induction of mouse thymocyte apoptosis by inhibitors of tyrosine kinases is associated with dephosphorylation of nuclear proteins.

Incubation of mouse thymocytes with the protein tyrosine kinase inhibitors herbimycin A and methyl-2,5-dihydroxycinnamate induced a decreased and altered profile of nuclear phosphotyrosine proteins in parallel with an increase in internucleosomal DNA fragmentation and cell death dose-dependently. No change in the profile of cytoplasmic phosphotyrosine proteins was observed. DNA fragmentation was dependent on the synthesis of RNA and protein, suggesting that the inhibition of tyrosine phosphorylation of the nuclear proteins induces apoptosis. DNA fragmentation was enhanced by simultaneous incubation with phorbol esters capable of activating protein kinase C. Genistein, another inhibitor of protein tyrosine kinase, induced DNA fragmentation more rapidly than herbimycin A, but there was no predominant alteration of phosphotyrosine proteins in early incubation, suggesting that genistein may induce apoptosis by a mechanism other than direct inhibition of protein tyrosinekinase activity.

Animals↗

PubMed1993

Tumour necrosis factor-alpha enhances cAMP-induced programmed cell death in mouse thymocytes.

During T-lymphocyte differentiation in the thymus, the majority of thymocytes die by apoptosis in situ. This process is characterized by internucleosomal DNA fragmentation and is induced by a number of stimuli including glucocorticoids, calcium ionophore, cAMP and 12-o-tetradecanoylphorbol 13-acetate (TPA). In this study, the effect of cytokines tumour necrosis factor-alpha (TNF-alpha) and interferon gamma (IFN-gamma) on the programmed cell death of thymocytes was examined by measuring DNA fragmentation and LDH release. TNF-alpha and IFN-gamma had no effect on DNA fragmentation in control and TPA, or A23187-treated thymocytes. Both human and murine rTNF-alpha enhanced cAMP-induced programmed cell death dose-dependently, but IFN-gamma had no effect on the process. TNF-alpha did not stimulate cAMP accumulation in control or 2-chloroadenosine-treated thymocytes. TPA markedly stimulated cAMP-induced DNA fragmentation as a result of 6 h incubation, whereas TNF-alpha did not. Thus TNF-alpha did not appear to activate protein kinase C directly. The effect of TNF-alpha was observed in the cell preparations from which adherent cells had been removed, suggesting that cytokines secreted by adherent cells in response to TNF-alpha are not involved in the process. The enhancement of cAMP-induced DNA fragmentation was observed in CD4+CD(8+)-double positive cells, but not in CD4+CD(8-)-single positive cells. The results of the present study indicate that a physiological cytokine, TNF-alpha, may modulate programmed cell death in immature thymocytes in concert with cAMP.

2-Chloroadenosine↗