Biomedical subjects

H Shiku

Publications and source records attributed to H Shiku.

At least 163 records · Page 9Linked to original sources

PubMed1996

Plasma tissue factor and tissue factor pathway inhibitor levels in patients with disseminated intravascular coagulation.

We measured the plasma levels of tissue factor (TF) and tissue factor pathway inhibitor (TFPI) in patients with disseminated intravascular coagulation (DIC) to examine the relationship between TFPI and vascular endothelial cell injury. Plasma TF (273 +/- 90 pg/ml) and TFPI (252 +/- 125 ng/ml) levels were significantly increased in patients with DIC compared with non-DIC patients. Plasma TF antigen level was significantly increased in pre-DIC patients (285 +/- 85 pg/ml), while the plasma TFPI level (152 +/- 54 ng/ml) was not markedly increased in such a state. The plasma TF/TFPI ratio was high in the pre-DIC patients (2.10 +/- 0.90), and low in the DIC patients (1.40 +/- 0.87) and healthy volunteers (0.84 +/- 0.26). There was no significant difference between the DIC patients with a good outcome and those with a poor outcome in terms of plasma TF levels, although the plasma TFPI level in the DIC patients with a good outcome (289 +/- 133 ng/ml) was significantly higher than that in those with a poor outcome (187 +/- 75 ng/ml). During the clinical course of DIC, plasma TF antigen was increased first, and an increase of the plasma TFPI level followed the increase in plasma TF level. These findings suggest that plasma TFPI is released from vascular endothelial cells and it may reflect vascular endothelial cell injury. It is conceivable that TF and TFPI may play an important role in the onset of DIC.

Anticoagulants↗

PubMed1996

Site restricted and neuron dominant expression of alpha 2,8sialyltransferase gene in the adult mouse brain and retina.

Gene expression of the alpha 2,8sialyltransferase (alpha 2,8S-T) responsible for GD3 synthesis in the adult mouse brain and retina was analysed by reverse transcription-polymerase chain reaction/Southern blotting (RT-PCR/Southern) and in situ hybridization. Among various portions of the brain, high levels of 9.5 kb mRNA were observed in the retina and midbrain. Results of RT-PCR/Southern did not necessarily correlate with the enzyme activities in the individual sites. In situ hybridization analysis revealed that this gene was characteristically expressed in the inner segment of photoreceptor cells, some nuclei in the midbrain, cranial nerve nuclei in the pons-medulla, Purkinje cells in the cerebellum, pyramidal cells of the hippocampus and granular cells of the dentate gyrus. In the retina, the alpha 2,8S-T gene was broadly expressed over the layers during development, and retained high expression levels in the photoreceptor cells of adult mice consistent with high expression of GD3. Destruction of neurons in the hippocampus and dentate gyrus by injection of kainic acid and colchicine respectively resulted in the disappearance of the hybridization signal, suggesting that the alpha 2,8S-T gene was mainly expressed by neurons.

Aging↗

PubMed1996

Characterization of the isoenzymes of cyclic nucleotide phosphodiesterase in human platelets and the effects of E4021.

In extracts of human platelets, three isoenzymes of cyclic nucleotide phosphodiesterase (PDE), namely, PDE2, PDE3, and PDE5, were identified; activities of PDE1 and PDE4 were not detected. In human platelets, the cGMP-hydrolytic activity was about six times higher than the cAMP-hydrolytic activity, and PDE5 and PDE3 are the major phosphodiesterase isoenzymes that hydrolyze cGMP and cAMP, respectively. PDE5 exhibited organ-specific expression in humans, and platelets were among the tissues richest in PDE5. A novel inhibitor of PDE5, sodium 1-[6-chloro-4-(3,4-methylenedioxybenzyl)aminoquinazolin-2-yl ] piperidine-4-carboxylate sesquihydrate (E4021), was a potent and highly selective inhibitor of human platelet PDE5. However, E4021 (up to 10 microM) did not inhibit 9,11-epithio-11,12-methano-thromboxane A2-induced platelet aggregation, in vitro. E4021 plus SIN-1 (3-morpholino-sydnonimine), at concentrations that had little effect individually, inhibited aggregation. These results suggest the unique distribution of phosphodiesterase isoenzymes in human platelets and the PDE5 inhibitors might be useful as a new class of antiplatelet drugs.

3',5'-Cyclic-GMP Phosphodiesterases↗

PubMed1996

A possible change in doubling time of haemopoietic progenitor cells with stem cell development.

We separated haemopoietic progenitors derived from marrow cells of 5-fluorouracil (5-FU)-treated mice into three groups, based on the stages of stem cell development and studied doubling time, using a serum-free clonal culture system. Stage I progenitors were those present in primary marrow cells from 5-FU-treated mice. Stages II and III progenitors were early and late progenies in culture of stage I progenitors respectively. The morphological analysis of colonies derived from stage I, II and III progenitors demonstrated an association of progression of stages with loss of multipotentiality. The doubling time of haemopoietic progenitors was estimated by sequential analysis of colony formation and studies of growth fraction. The time required for haemopoietic progenitors to double shortened as their stage of development progressed. Alteration in one doubling time of haemopoietic progenitors at progressive stages of stem cell development was seen in cultures supported by various combinations of growth factors, including interleukin-3 (IL-3), IL-11. and steel factor (SF), Cell-cycle analysis suggested that reduction of the doubling time of haemopoietic progenitors is probably due to a decrease in the time spent in the G1 phase of the cell cycle. Our results suggest that in early haemopoiesis the doubling time of haemopoietic progenitors may change with stem cell development.

Animals↗

PubMed1996

Activity of the ligand for c-mpl, thrombopoietin, in early haemopoiesis.

We examined the role of the ligand for c-mpl. thrombopoietin (TPO). in murine early haemopoiesis. using a serum-free culture system. TPO in combination with the ligand for c-kit (SF) or interleukin-3 (IL-3) supported colony formation by marrow cells of 5-fluorouracil (5-FU)-treated mice whereas TPO alone yielded no colony. When blast cell colonies grown in the presence of TPO plus SF or TPO plus IL-3 were individually replated in suspension cultures containing serum and several growth factors, various combinations of myeloid lineages were seen, indicating that the progenitors supported by TPO plus SF or TPO plus IL-3 are multipotential. Delayed addition experiments demonstrated that TPO has the potential to effectively support the survival of haemopoietic progenitors. We then studied the effects of TPO on proliferative kinetics of cycling progenitors. TPO hastened IL-3-dependent growth of progenitors by shortening the time required for cell cycling. These results suggest that TPO as a single factor, can support the survival of haemopoietic progenitors and TPO synergizes with SF or IL-3 to act on early multipotential haemopoietic progenitors.

Animals↗

PubMed1996

Heterogeneity in the expression pattern of two ganglioside synthase genes during mouse brain development.

Gangliosides are synthesized by sequential catalytic reaction of multiple glycosyltransferases. GM2/GD2 synthase and GD3 synthase are key enzymes for ganglioside synthesis, because their relative activities regulate the main profiles of ganglioside expression. Mouse GD3 synthase (EC 2.4.99.8) cDNA was cloned by eukaryotic expression cloning, and its mRNA expression as well as that of GM2/GD2 synthase gene during the development of the mouse CNS was analyzed by using northern blotting, reverse transcription-polymerase chain reaction, and in situ hybridization. When brain tissue was analyzed as a whole mass, a typical pattern corresponding to the reported findings obtained by biochemical analyses was observed, i.e., high expression of GD3 synthase gene in the early stage and gradual increase of GM2/GD2 synthase gene expression in the late stage of the development. However, the results of in situ hybridization of these two genes revealed that the expression kinetics of these two genes were heterogeneous among various sites in the brain under development. These findings suggest that various expression patterns of the two genes reflect differences in the course of the development of individual sites, and also different ganglioside components are required in individual portions of the brain for development and maintenance of the function.

Amino Acid Sequence↗

PubMed1996

Myeloid antigen, CD13, CD14, and/or CD33 expression is restricted to certain lymphoid neoplasms.

The authors examined the expression of myeloid antigens (MyAg): CD11b, CD13, CD14, CD15, and CD33 in 249 adults with lymphoid neoplasms using flow cytometric analysis. In this study, acute leukemia that was myeloperoxidase negative by light microscopy and had at least one lymphoid antigen was defined as acute lymphoblastic leukemia (ALL). The patients were classified as follows: 6 with unclassified ALL, 35 early B precursor ALL, 32 T-ALL, 25 B-cell chronic lymphocytic leukemia (B-CLL) and its variants, 24 B-cell non-Hodgkin's lymphoma (B-NHL), 7 plasma cell disorders, 8 T-CLL, 2 adult T-cell leukemia, and 10 T-NHL. CD11b and CD15 were present in a wide range of lymphoid disorders irrespective of B/T lineage and maturity. Unclassified ALL and phenotypically immature ALL frequently expressed CD13 and CD33, and occasionally expressed CD14. Among early B precursor ALL, CD13, and/or CD33 were significantly associated with the presence of stem cell marker CD34 and the chromosomal abnormality t(9;22). In addition, ALL with deletion of chromosome 7 commonly expressed CD13 and CD33. Taken together, CD13 and/or CD33 positive ALL may originate from a multipotential stem cell. Among mature neoplasms, CD14 was frequently, and CD13 and CD33 were occasionally expressed in B-cell, but not T-cell tumors. These results suggest that CD13, CD14, and CD33 are preferentially expressed in two types of lymphoid neoplasms, namely undifferentiated ALL and mature B-cell lymphoproliferative disorders.

Adolescent↗

PubMed1996

Plasma levels of activated FVII in various diseases.

Plasma activated factor VIIa (FVIIa) levels were measured in various diseases using mutant tissue factor (TF). FVIIa levels in thrombotic patients and patients with idiopathic thrombocytopenic purpura were significantly higher than those in healthy control subjects. The plasma FVIIa levels in thrombotic patients treated with warfarin were similar to those of control subjects. The plasma FVIIa levels in pregnant women and patients with systemic lupus erythematosus, infection or malignancies were high. However, the levels in patients with disseminated intravascular coagulation (DIC) were not significantly increased. DIC patients are in a severe hypercoagulable state, and exhibit severe consumption of coagulation factors. The slightly increased FVIIa level in the DIC patients observed is probably considered to be caused by consumption of coagulation factors. The plasma FVIIa level was poorly correlated with other hemostatic parameters except for protein C in our analysis of all cases. In the analysis of DIC and thrombotic patients treated without warfarin, the plasma FVIIa level was negatively correlated with TF antigen. Plasma FVIIa levels might reflect hypercoagulability in thrombotic diseases, and a normalized FVIIa level in patients with thrombotic diseases should be considered to be associated with DIC.

Autoimmune Diseases↗

PubMed1996

Expression of a CD44 variant containing exons 8 to 10 is a useful independent factor for the prediction of prognosis in colorectal cancer patients.

PURPOSE: To determine the expression of the CD44 variant containing a variant exon 8 to 10 product (CD44v8-10) in colorectal cancer and to evaluate its prognostic value. MATERIALS AND METHODS: CD44v8-10 was studied in resected tumors and normal mucosae obtained from 215 colorectal cancer patients (118 colon cancer and 97 rectal cancer). The expression of CD44v8-10 was analyzed immunohistochemically using the anti-CD44v8-10 monoclonal antibody (mAb) 44-1V. RESULTS: One hundred of 215 cancer tissues expressed CD44v8-10. Positive staining was intense mainly on the cell membranes. There was no significant correlation between expression of CD44v8-10 and histologic type, primary tumor, lymphatic invasion, venous invasion, or peritoneal invasion. There were significant correlations between CD44v8-10 immunoreactivity and both lymph node and hematogenous metastasis. Patients with CD44v8-10-positive tumors had a greater relative risk of death compared with those whose tumors were CD44v8-10-negative. Among 169 patients who underwent curative resection, CD44v8-10 expression correlated with a high recurrence rate. The 5- and 10-year survival rates were 90.3% of patients with CD44v8-10-negative tumors, and 72.1% and 58.0% of those with CD44v8-10-positive tumors, respectively; these differences between the two groups of patients were significant (P < .01). In multivariate analysis using the Cox regression model, CD44v8-10 expression emerged as an independent prognostic indicator. CONCLUSION: The results suggest that CD44v8-10 plays a role in metastasis of colorectal cancer, and tha CD44v8-10 expression may be a biologic marker of prognostic significance.

Antibodies, Monoclonal↗

PubMed1996

PRAD1/Cyclin D1 gene overexpression in mantle cell lymphoma.

The t(11;14) (q13;q32) translocation and its molecular counterpart Bcl-1 rearrangement are consistent features of mantle cell lymphoma (MCL). This translocation activates the PRAD1/cyclin D1 gene that is considered to be the Bcl-1 oncogene. PRAD1/cyclin D1 gene overexpression is closely associated with MCL. The PRAD1/cyclin D1 protein is localized to the nucleus, and the strong correlation between PRAD1/cyclin D1 and MCL is also found in the protein level. This finding indicates that PRAD1/cyclin D1 expression is a highly specific and sensitive molecular marker for MCL. This gene may function as an oncogene in the malignant transformation of cells.

Cyclin D1↗

PubMed1996

[Plasma fibrin monitor level in DIC patients with a hematopoietic malignancy].

The Plasma level of soluble fibrin monomer (sFM) was measured in 218 patients with a hematopoietic malignancy. Of them, 198 were diagnosed with disseminates intravascular coagulation (DIC), 20 with Pre-DIC, and 20 with Non-DIC. Pre-DIC was retrospectively defined as the condition at least 1 week before the onset of DIC. The plasma levels of sFM, thrombin-anti-thrombin III complex (TAT), plasmin alpha 2-antiplasmin inhibitor complex (PIC), and FDP-D-dimer were significantly higher in patients with DIC than in those with Non-DIC. These levels were significantly higher in patients with Pre-DIC than in those with Non-DIC. Among these hemostatic parameters, the plasma sFM showed the highest sensitivity and specificity for DIC or Pre-DIC. These findings suggests that sFM is the most valuable marker hemostatic for the diagnosis of DIC and Pre-DIC.

Biomarkers↗

PubMed1996

[Expression of nm23/NDP kinase and cancer metastasis].

The nm23 gene was originally identified by differential hybridization between low and highly metastatic murine melanoma lines. The nm23 protein is identical with NDP kinase and there are at least two isotypes in several mammalian species such as human, mice and rats. Reduced expression of nm23/NDP kinase gene has been reported of association with highly metastatic characteristics in several experimental tumors and in certain types of human cancer. Detailed mechanisms of nm23 molecules in metastasis suppression remain unknown. The nm23/NDP kinase appears to participate to multiple cellular functions such as differentiation, proliferation and development. In this article, we reviewed recent reports on nm23/NDP kinase.

Animals↗

PubMed1996

Differential association of protein Ser/Thr phosphatase types 1 and 2A with the cytoskeleton upon platelet activation.

The association of protein Ser/Thr phosphatase type 1(PP1) and type 2A (PP2A) with the cytoskeleton (Triton X-100 insoluble residue) during human platelet activation was investigated. In unstimulated platelets, 40% of total PP1-like activity was present in the Triton-insoluble cytoskeleton, while only 10% of the total PP2A-like activity was present in this fraction. Stimulation with 1 U/ml thrombin produced a 1.8-fold increase in PP1-like activity and a 7-fold increase in PP2A-like activity, respectively, in the cytoskeletal fraction, under aggregating conditions. Immunoblot analysis revealed that thrombin treatment increased association of PP1 catalytic subunit isozymes (PP1 alpha, PP1 gamma, PP1 delta) and PP2A catalytic subunit with the cytoskeleton, with concomitant decrease of these enzymes in Triton-soluble fractions. The amounts of cytoskeleton-associated PP1 and PP2A depended on the dose of thrombin which could activate platelets. Agonist-induced redistribution of PP1 and PP2A into the cytoskeleton was inhibited by OP-41483 (a prostaglandin I2 analog). Interaction of PP2A with cytoskeletal proteins strongly correlates with aggregation, whereas the association of PP1 with cytoskeleton can be detected upon platelet activation, even in the absence of aggregation. Co-extraction of protein kinase C and myosin light chain kinase with the cytoskeleton eventually translocated to the cytoskeleton, but only during aggregation. These results suggest that differential translocation of PP1 and PP2A to the cytoskeleton is involved in platelet activation, and their association with cytoskeletal proteins may regulate phosphorylation levels together with protein kinases in platelets.

Cytoskeleton↗

PubMed1995

Activation of p21WAF1/Cip1 expression by a temperature-sensitive mutant of human p53 does not lead to apoptosis.

The cyclin-dependent kinase inhibitor p21WAF1/Cip1 (p21) inhibits cellular proliferation and has been implicated in p53-dependent apoptosis. In this study, we examined the regulation of p21 in the K562 erythroleukemia cell line using a human temperature-sensitive mutant of p53, 143Ala. We showed that 143Ala at the permissive temperature (32 degrees C) activated the expression of p21. Stable cell lines expressing 143Ala or other p53 mutants were established. We then examined whether elevation of p21 promotes apoptosis and sensitized cells to radiation-induced apoptosis. Our results showed that p21 elevation did not promote or sensitize K562 cells to apoptosis.

Apoptosis↗

PubMed1995

Up-regulation of protein serine/threonine phosphatase type 2C during 1 alpha,25-dihydroxyvitamin D3-induced monocytic differentiation of leukemic HL-60 cells.

Treatment with 20 nM 1 alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3) caused a progressive increase in the activity of Mg(2+)-dependent protein serine/threonine phosphatase type 2C (PP2C) in subcellular fractions of HL-60 cells, whereas PP2C activity was relatively constant throughout all-trans retinoic acid-induced (1 microM) granulocytic differentiation. The increase in PP2C activity appeared to parallel the 1,25(OH)2D3-induced phenotypic and functional changes in HL-60 cells. Immunoblot and Northern blot analysis indicated that the increase in PP2C activity corresponded to the increased expression of PP2C protein, which was preceded by an increase in the level of mRNA for PP2C beta. No mRNA for PP2C alpha was detected in resting or 1,25(OH)2D3-stimulated HL-60 cells. These results suggest that the increased expression of PP2C is related with the 1,25(OH)2D3-induced monocytic differentiation of HL-60 cells.

Calcitriol↗

PubMed1995

The effects of the site-directed removal of N-glycosylation sites from beta-1,4-N-acetylgalactosaminyltransferase on its function.

The amino acid sequence deduced from the cloned human cDNA of beta-1,4-N-acetylgalactosaminyltransferase (GalNAc-T; EC 2.4.1.92) gene predicted three potential sites for N-linked glycosylation. Although many glycosyltransferases isolated contain from 2 to 6 N-glycosylation sites, their significance has not been adequately demonstrated. To clarify the roles of N-glycosylation in GalNAc-T function, we generated a series of mutant cDNAs, in which some or all of the glycosylation recognition sites were eliminated by polymerase chain reaction (PCR)-mediated site-directed mutagenesis. Using transcription/translation in vitro, we confirmed that all potential N-glycosylation sites could be used. Although cell lines transfected with mutant cDNAs showed equivalent levels of GalNAc beta 1-->4(NeuAc alpha 2-->3)Gal beta 1-->4Glc-Cer (GM2) to that of the wild-type, the extracts from mutant cDNA transfectants demonstrated lower enzyme activity than in the wild-type. The decrease in enzyme activity was more evident as the number of deglycosylated sites increased, with about 90% decrease in a totally deglycosylated mutant. The enzyme kinetics analysis revealed no significant change of Km among wild-type and mutant cDNA products. The intracellular localization of GalNAc-T expressed in transfectants with wild-type or mutant cDNAs also showed a similar perinuclear pattern (Golgi pattern). These results suggest that N-linked carbohydrates on GalNAc-T are required for regulating the stability of the enzyme structure.

Animals↗

PubMed1995

Preparation of a murine cell line which stably expresses human T lymphotropic virus type I (HTLV-I) env genome products.

We prepared the murine myeloma cell line NS-1, which stably expressed the human T lymphotropic virus type I (HTLV-I) env gene. The plasmid BCMGEnv was constructed from the episomal vector BCMGSNeo, which was primarily derived from bovine papilloma virus. Transfected env expression was detected by Northern blotting, as well as by flow cytometry using envelope protein-specific monoclonal antibodies (mAb). Expression was detectable for at least seven months. The env transfectants induced syncytium formation which is characteristic of HTLV-I-infected cells, in the human uterine cervical cancer line, HeLa, and the rat cell line, XC. The requirement of envelope proteins for syncytium formation was confirmed by an inhibition assay with envelope protein-specific mAb. Therefore, env transfectants are not only stable, but also have its specific biological function. This system may be useful to analyze the initial steps of viral attachment to the cell surface and to search for the HTLV-I receptor.

Animals↗

PubMed1995

Gene therapy for hepatoma cells using a retrovirus vector carrying herpes simplex virus thymidine kinase gene under the control of human alpha-fetoprotein gene promoter.

The alpha-fetoprotein (AFP) gene is normally expressed in fetal liver and is transcriptionally silent in adult liver but is reactivated in hepatocellular carcinoma. It has been shown that the positive and negative transcriptionally regulatory elements of the human AFP gene, which play an important role in its developmental regulation, exist over the quite extended region (4 kb). We constructed a hybrid gene consisting of herpes simplex virus thymidine kinase (HSV-tk) gene under the control of the 0.3-kb human AFP gene promoter and inserted it into a retroviral vector. When AFP-producing hepatoma cells were infected with this recombinant retrovirus (LNAF0.3TK virus), the cells expressed HSV-tk gene and exhibited increased sensitivity to ganciclovir parallel with the ability of AFP production. On the other hand, the retroviral infection had little effect on ganciclovir-mediated cytotoxicity in AFP-nonproducing hepatoma or non-hepatoma cells. Moreover, the addition of dexamethasone increased the cytotoxicity of aciclovir to the virus-infected, AFP-producing cells through a glucocorticoid-responsive element in the AFP promoter, although aciclovir, by itself, had little cytotoxicity. These results demonstrate that the AFP promoter sequence alone can provide enough tumor-specific activity for therapeutic gene expression and induce selective growth inhibition by ganciclovir in the virus-infected, AFP-producing human hepatoma cells. In addition, it is possible that expression of the therapeutic gene is modulated by administration of dexamethasone or other agents that alter AFP promoter activity after gene transduction.

3T3 Cells↗