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Y Kokai

Publications and source records attributed to Y Kokai.

At least 37 records · Page 2Linked to original sources
PubMed1996

Expression of p21(waf-1/cip-1) is significantly induced in the livers of LEC rats with chronic liver injury.

It is reported that hepatocytes isolated from LEC rats with chronic liver injury show reduced growth activity in primary culture. To elucidate the molecular basis of this phenomenon, we examined expression of p21(waf-1/ciP-1) and p27, cyclin-dependent kinase inhibitors, by northern blot analysis. The expression of p21(waf-1/cip-1 ) in the LEC rat liver was 3-fold higher than that of age-matched SD rat liver, while there was no significant difference in p27 expression level. Western blot analysis also revealed a significant increase in p21(waf-1/cip-1) in the nuclear matrix fraction of the LEC rat liver. Immunohistochemically, p21(waf-1/cip-1) was detected in the nuclei of normal LEC rat hepatocytes, but not in those of hepatocellular carcinoma cells, suggesting selective growth of neoplastic hepatocytes.

Animals↗
PubMed1996

Selective inhibition of resistance to hemopoietic allografts but not rejection to a natural killer cell sensitive tumor in transgenic mice for granulocyte colony stimulating factor.

Transplanted allogeneic marrow grafts often fail to engraft in a lethally irradiated host. Resistance to hemopoietic allograft is a complexed phenomenon involving multiple components. To study the involvement of a hemopoietic cytokine, which was known to play a role for stem cell function, we established lines of mice that were transgenic for human granulocyte colony-stimulating factor (hG-CSF). Elevated and constitutive expression was found in sera (1,041 +/- 242 pg/ml) of these transgenic mice regardless of their sexes and ages. Strong neutrophilic granulocytosis correlated with the elevated G-CSF activity in transgenic mice but not in littermate controls, establishing a functional expression of this cytokine. In lethally irradiated mice transgenic for G-CSF, infusion of fully allogeneic marrow cells induced donor-derived spleen colony. Growth of hemopoietic allografts appeared to be similar to those of syngeneic marrow cells, which indicates inhibition of resistance for allogeneic marrow grafts. Because of a positive correlation, involvement of natural killer (NK) cells in resistance of transplanted allografts has been suggested. Inocula of NK-sensitive lymphoma cells were, however, vigorously rejected in the G-CSF-transgenic mice. This observation indicates that G-CSF may play a role in engraftment of transplanted allogeneic marrow grafts and may represent a component of mechanisms of hemopoietic resistance. Furthermore, this result may be an indication that alloresistance and NK cells use different mechanisms to resist each target.

Animals↗
PubMed1996

Galactosemic neuropathy in transgenic mice for human aldose reductase.

We studied the functional consequences of an enhanced polyol pathway activity, elicited with galactose feeding, on the peripheral nerve of transgenic mice expressing human aldose reductase. Nontransgenic littermate mice were used as controls. With a quantitative immunoassay, the expression level of human aldose reductase in the sciatic nerve was 791 +/- 44 ng/mg protein (mean +/- SE), about 25% of that in human sural nerve. When the transgenic mice were fed food containing 30% galactose, significant levels of galactitol accumulated in the sciatic nerve. Galactose feeding of nontransgenic littermate mice led to a 10-fold lower accumulation of galactitol. Galactose feeding for 16 weeks caused a significant and progressive decrease in motor nerve conduction velocity in transgenic mice to 80% of the level of galactose-fed littermate mice, which was not significantly different from that of galactose-free littermate mice. A morphometric analysis of sciatic nerve detected > 10% reduction of mean myelinated fiber size but no alterations of myelinated fiber density in galactose-fed transgenic mice compared with other groups. The functional and structural changes that develop in galactose-fed transgenic mice are similar to those previously reported in diabetic animals. The results of these studies suggest that transgenic mice expressing human aldose reductase may be a useful model not only for defining the role of the polyol pathway in diabetic neuropathy but also for identifying and characterizing effective inhibitors specific for human aldose reductase.

Aldehyde Reductase↗
PubMed1996

Lack of CD54 expression and mutation of p53 gene relate to the prognosis of childhood Burkitt's lymphoma.

Expression of the intercellular adhesion molecule-1 (CD54) as well as the mutations of p53 gene were studied in childhood Burkitt's lymphoma (BL). Expression of CD54 was identified in 6 of 15 fresh BL cases. Mutations of p53 gene, analyzed by polymerase chain reaction-single stranded chain polymorphism followed by sequencing, were found in 5 of 14 cases examined. Interestingly, all the cases with p53 mutation were CD54 negative. This high frequency of p53 mutation in the CD54 negative group prompted us to analyze the clinical features of these cases. Six of 15 cases died within 21 months after initiation of therapy and five of these were CD54 negative. In addition, four of these had p53 mutation. These results suggest that the lack of CD54 by BL cells may provide the background for the mutation of p53 gene to occur which could result in the transformation to a more aggressive phenotype.

Adolescent↗
PubMed1996

Overexpression of the granulocyte colony-stimulating factor gene leads to osteoporosis in mice.

Although granulocyte colony-stimulating factor (G-CSF) was originally isolated as an activity for the growth and differentiation of cells in granulocytic lineage, it has been gradually accepted that G-CSF may have a function on a wide variety of cells besides granulocytes. To elucidate the function of G-CSF on bone cells in vivo, we examined the bone tissue of transgenic mice that overexpress G-CSF. Transgenic mice express human G-CSF at an elevated level (1041 +/- 242 pg/ml in sera) under the direction of SRalpha promoter. We performed radiographic, routine histologic, and histomorphometric analyses of the bone tissue and serum biochemical assay. Nontransgenic littermates were examined as age-matched, wild-type controls in all experiments. Radiographic analysis revealed cortical thinning accompanied by enlarged bone marrow cavities in both vertebral bodies and long bones. Histologically, a decreased number and thickness of trabecular bones and cortical thinning were observed in lumber vertebrae as well as in femur specimens. The enlarged bone marrow cavities exhibited an increased number of mature neutrophilic granulocytes without apparent changes in other types of cells. The static and dynamic parameters reflecting bone resorption were found to be significantly increased in the transgenic mice. By contrast, no significant differences were detected in the parameters reflecting bone formation. Transgenic mice and littermate controls had similar serum calcium, phosphorous, and alkaline phosphatase levels. However, the serum osteocalcin level was significantly higher in transgenic mice. These findings indicate that G-CSF-expressing transgenic mice developed osteoporosis because of increased osteoclastic activity. Collectively, G-CSF could have a negative influence on bone homeostasis in vivo.

Animals↗
PubMed1996

Elevation of lymphocyte and hematopoietic stem cell numbers in mice transgenic for human granulocyte CSF.

Transgenic mice expressing human granulocyte colony stimulating factor (hG-CSF) were bred to establish an experimental model for studying the biologic function of hG-CSF. The mice produced approximately 1000 pg/ml of hG-CSF in sera and expressed hG-CSF mRNA in all of the tissues examined. Granulocytosis and elevation of hematopoietic precursor number were observed in hematopoietic tissues as expected, but lymphocytosis with 5-fold increase in peripheral blood and 2-fold increase in spleen was also evident. Hematopoietic stem cells capable of reconstituting lethally irradiated recipient mice were abundant in the circulation of these transgenic mice. No disadvantageous tissue damage was observed when various organs were analyzed. As shown in this study, G-CSF has a stimulating effect on lymphopoiesis when it is expressed in a certain condition. The lymphocytosis, as well as the high level of hematopoietic stem cell mobilization into circulation, indicates a wider range of potential G-CSF activity than expected. The mice transgenic for hG-CSF described herein will contribute greatly to the understanding of the regulatory mechanism in hematopoiesis.

Animals↗
PubMed1995

Presence of a closely related subgroup in the aldo-ketoreductase family of the mouse.

Aldose reductase (alditol:NAD(P)+ 1-oxidoreductase), an enzyme implicated in the pathogenesis of various diabetic complications, catalyzes the reduction of a variety of aldehydes. From a mouse kidney library, we isolated aldose reductase cDNA that encodes a 316-amino-acid protein with approximately 97% identity to rat lens aldose reductase, approximately 69% identity to the mouse vas deferens protein and also approximately 69% identity to mouse fibroblast growth-factor-1-regulated protein. RNA-blot analysis demonstrated abundant expression of the enzyme transcript in the testis, skeletal muscle and kidney. However, a very low level of the transcript was detected in the sciatic nerve and lens, where abundant expression and involvement of the enzyme in diabetic complications were documented in other animals species. The isolated cDNA was expressed in Escherichia coli and the recombinant protein was purified to homogeneity by affinity chromatography and chromatofocusing. The expressed enzyme demonstrated reductase activity for various aldo sugars but not for the steroids. The enzyme reaction with DL-glyceraldehyde was, however, competitively inhibited by progesterone or 17 alpha-hydroxyprogesterone. The results not only indicate a unique tissue distribution and enzyme attribute of mouse aldose reductase, but also the presence of a closely related subgroup within the aldo-oxidoreductase superfamily in mouse tissues.

Aldehyde Reductase↗
PubMed1995

Acute onset of diabetic pathological changes in transgenic mice with human aldose reductase cDNA.

To investigate the role of human aldose reductase (hAR) in the pathogenesis of diabetic complications, we generated transgenic mice carrying hAR cDNA driven by the murine MHC class I molecule promoter (hAR-Tg). Northern and Western blot analyses and immunoassay of hAR revealed that both hAR mRNA and the protein were expressed in all tissues tested. Thrombosis in renal vessels and fibrinous deposits in Bowman's capsule were observed in 6-week-old hAR-Tg mice fed a normal diet. Ingestion of a 30% glucose diet for 5 days caused sorbitol concentrations in the liver, kidney, and muscle of hAR-Tg mice to be elevated significantly. Seven-week-old hAR-Tg mice fed a 20% galactose diet for 7 days developed cataracts and occlusion of the retinochoroidal vessels, in addition to pathological changes in the kidney. Despite an elevated aldose reductase level in hAR-Tg mice and their intake of an aldose diet, no histopathological changes were found in other tissues, including the brain, lungs, heart, thymus, spleen, intestine, liver, muscle, spinal cord, or sciatic nerve. Results suggest that target organs of diabetic complications, such as the kidney, lens, and retina are sensitive to damage associated with a high level of AR expression, but other organs are not; the susceptibility of each organ to diabetic complications is determined by not only hAR but also other factors.

Aldehyde Reductase↗
PubMed1995

Novel monoclonal antibody reactive with thrombin-sensitive 74-kDa glycoproteins present on platelets and megakaryocytes both from mouse and rat.

A monoclonal antibody (designated 1C2) that reacts only with mouse platelets and megakaryocytes among hematopoietic cells was established by immunizing mouse platelets to an Armenian hamster. 1C2 reactive mouse molecule (1C2 antigen) was a surface glycoprotein with molecular weight of 74 kDa. Side by side comparison revealed that 4A5, a rat monoclonal antibody against mouse platelet, immunoprecipitated the identical molecule to 1C2 antigen. Of particular interest, 1C2 also labeled rat tissues with an identical pattern to that of mouse tissues and recognized a 74-kDa protein from rat platelets. Reactivity of 1C2 to mouse and rat platelets decreased when they were treated with thrombin. Following thrombin treatment of mouse platelets, 1C2 reactive 69-kDa protein appeared in the supernatants. Mouse and rat 1C2 antigens purified on 1C2-coated beads were cleaved by thrombin to generate 69-kDa fragments, establishing that 1C2 antigen is a direct substrate for thrombin. 1C2 is the first antibody to platelets and megakaryocytes of mouse and rat whose reactive molecule is well characterized, i.e., substrate for thrombin. 1C2 can be a useful tool in studying megakaryocytopoiesis and thrombopoiesis in rodent systems.

Animals↗
PubMed1995

Non-phenotypic detection of osteopetrotic (op/op) mutation by using PCR-SSCP analysis.

Mice homozygous for recessive mutation osteopetrosis (op/op) on chromosome 3 provide a unique model to study the mechanism of haematopoiesis in conjunction with bone formation. Based on the DNA sequence data recently reported, we established a PCR-SSCP (polymerase chain reaction--single strand conformation polymorphism) assay which identifies an amplified fragment having an insertional point mutation present in macrophage colony-stimulating factor (M-CSF) gene of op/op mice. With this assay, three genotypes, op/op, +/op, and +/+ can be distinguished. Although heterozygous (+/op) and normal (+/+) mice could not be discriminated phenotypically, we could generate op/op mutant mice starting from a single heterozygous (+/op) mouse using only the PCR-SSCP aided screening method. This assay will permit introduction of the op mutant into any strain to generate a new animal model to study the cytokine network and haematopoiesis.

Animals↗
PubMed1994

Clinical significance of growth hormone-binding protein measurements in children.

To investigate the role of growth hormone (GH) and its downstream axis in normal growth and growth disorders, we measured serum GH-binding protein (GHBP) levels in children by ligand-mediated immunofunctional assay (LIFA). Samples were taken from 512 healthy children of normal stature, 146 healthy neonates, 153 short but otherwise normal children, and 27 patients with GH deficiency. Serum GHBP showed no significant diurnal variation. Serum GHBP levels were low in neonates, acutely rose within 6 months, and gradually increased toward midpuberty in normal children (r = 0.2, P < 0.01, assessed among subjects over 1 year old). Neither significant sexual dimorphism nor rapid pubertal changes were observed. In normal short children and patients with GH deficiency, GHBP was lower than normal, but not significant. These data suggest that one of the etiologies for growth disorder in children could be resistance to, or low efficacy for, utilization of GH.

Adolescent↗
PubMed1993

[Flow cytometric analysis of the DNA content of sclerosing hemangioma of the lung].

DNA ploidy and DNA content of two surgically resected sclerosing hemangioma of the lungs were examined by flow cytometry. Cell kinetics were analyzed by using DNA index and the proliferation index. Both cases were diploid, and their proliferation indices were 22.91 and 25.53, respectively, suggesting that they are benign tumors.

DNA, Neoplasm↗
PubMed1992

Expression pattern of SR alpha promoter in human embryonal carcinoma and transgenic tissues in mice.

Human embryonal carcinoma is thought to be a counterpart of mouse embryonal carcinoma, which has provided useful information for studying early molecular events in murine embryogenesis. A major practical problem in the use of human embryonal carcinoma for molecular pathological studies is the lack of an efficient expression system for foreign genes. The SR alpha promoter is a fusion promoter containing the SV 40 early promoter and the R segment and part of the U5 sequence of the long terminal repeat derived from human T-cell leukemia virus type I. We analyzed the expression pattern of the SR alpha promoter in human and mouse embryonal carcinoma lines and transgenic mouse tissues. Efficient and stable expression was detected in all cell lines tested, and tissues from all mice of four independent transgenic lines carrying the SR alpha-CAT vector showed a detectable level of CAT expression. These data suggest that the SR alpha promoter is useful for studies of both human embryonal carcinoma and transgenic mouse tissue. Using this expression system, we are now able to label human embryonal carcinomas with various genes, for example beta galactosidase, and follow their fate at the single-cell level in nude mice, where xenotransplanted human embryonal carcinoma expresses differentiation capability.

Animals↗
PubMed1991

Progesterone augments proliferation induced by epidermal growth factor in a feline mammary adenocarcinoma cell line.

Steroid hormones and peptide growth factors promote growth and development of normal mammary tissues and some types of breast cancer. Ovarian steroids may influence mammary growth directly or indirectly. The epidermal growth factor (EGF) family of proteins may also regulate mammary growth. These two pathways may function independently of each other or they may act in concert, with steroids inducing transcription of genes that encode growth factors or growth factor receptors. We used a feline mammary adenocarcinoma cell line (K12) to address whether there was an interrelation between progesterone (PGN) and EGF-associated growth pathways. K12 cells responded to EGF by a dose-dependent increase in proliferation. PGN or promegestone (R5020, a synthetic progestagen) alone did not stimulate K12 growth, but when EGF and PGN, or EGF and R5020 were combined, they were synergistic. This synergistic response was abrogated by the PGN receptor antagonist RU486 or by antibodies that blocked binding of EGF to its receptor. K12 cells expressed characteristic double-affinity EGF receptors, as well as p185 (a functionally and structurally related protein, product of the neu gene) on their surface. PGN receptors were also found on intact cells and in cleared cytosols. Stimulation of K12 cells by PGN or by R5020 induced a two- to threefold increase in the number of high-affinity surface EGF receptors after 24 h. Stimulation of these cells by PGN also affected the relative levels of phosphorylation of the EGF receptor and p185 within minutes, but not of other cellular phosphoproteins. Our results show that PGN enhances the EGF-induced growth of K12 cells and suggest that this effect may be mediated at least partly via an increase in the number or function of high-affinity EGF receptors.

Adenocarcinoma↗
PubMed1990

Cloning and expression of human aldose reductase.

The complete amino acid sequence of human retina and muscle aldose reductase was determined by nucleotide analysis of cDNA clones isolated using synthetic oligonucleotide probes based on partial amino acid sequences of purified human psoas muscle aldose reductase. The cDNA sequence differs substantially in the noncoding and coding regions of recently published sequences of this enzyme. The mRNA for aldose reductase was abundantly expressed in HeLa cells, but only scarcely in a neuroblastoma cell line. Recombinant baculovirus containing one of the muscle cDNA clones was constructed and used to infect Spodoptera frugiperda (SF9) cells. A prominent protein with an apparent molecular size of 36 kDa was identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the culture medium as well as in the homogenate of SF9 cells after 2 days of infection. Culture medium or the supernatant fraction of cell homogenates containing this protein had high aldose reductase activity which showed characteristics of the reported human enzyme. These findings indicate that the amino acid sequence reported in this paper represents human retina and muscle aldose reductase and that functional human aldose reductase can be expressed in large amounts in a baculovirus expression system. The result should facilitate refined structural analysis and the development of new specific aldose reductase inhibitors for the treatment of diabetic complications.

Aldehyde Reductase↗
PubMed1990

Characterization of the common acute lymphoblastic leukaemia antigen (CD10) as an activation molecule on mature human B cells.

Distinct expression pattern of CD10 molecules during B cell activation was analysed using in vivo and in vitro systems. By two-colour flowcytometrical analysis, CD10 was found to be expressed at a specific stage of in vivo activating B cells. The expression of CD10 during B cell activation appeared to be unique from that of other activation-related B cell antigens including L29, MA6, OKT9 and OKT10. Although the expression of CD10 was associated with that of the activation-related B cell antigens, CD10+ B cells could be separated in the distinct fractions to those expressing other activation-related B cell antigens when fractionated by cell gravity. In particular, certain CD10+ B cells were detected positive for the resting B cell antigen, L30. In vitro studies revealed that CD10+ B cells arose from CD10- B cells at an early step of B cell activation, and disappeared lately when activated by Staphylococcus aureus Cowan I. Collectively, CD10 was an antigen transiently expressed at an early phase of B cell activation process. Expression of CD10 and other antigens on Burkitt's lymphomas (15 cases) was studied next. All cases were CD10+, and 87% (13 cases) were also L30+. In addition, six of CD10+ L30+ cases were L29+. This observation suggested that Burkitt's lymphomas were phenotypically similar to the B cells at an early phase of activation, those expressing CD10 and L30, simultaneously. The present study has dissected a precise expression pattern of CD10 on mature B cell activation in vitro and in vivo, and could be implicated for the histogenesis of one of the poorly characterized B cell lymphoma, namely Burkitt's lymphoma.

Antigens, CD↗