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Biomedical subjects

K Tani

Publications and source records attributed to K Tani.

At least 199 records · Page 11Linked to original sources
PubMed1994

Regulation of recombinant human granulocyte colony-stimulation factor production using herpes simplex virus 1 thymidine kinase gene.

For chronic neutropenic patients requiring long-term injection of recombinant human granulocyte colony-stimulating factor (rhG-CSF), a cellular transplantation system that can produce this cytokine stably and deliver it in a regulatory manner would be advantageous. In this study we aimed at developing a regulation system at cellular level using suicide vectors. We introduced the herpes simplex virus type 1 thymidine kinase (HSV-TK) gene into the rhG-CSF-producing NIH3T3 cells and examined if ganciclovir (GCV) treatment of the cells could control the rhG-CSF production in vitro. The cells transfected with the HSV-TK gene showed a > 100-fold increase in sensitivity to GCV compared with the parent cells, and the median inhibitory dose of GCV to the transfected cells was less than 1.6 microM. The total amount of rhG-CSF production by these cells was strongly suppressed by GCV treatment. This regulatory method may be applicable to cytokine supplement gene therapy.

Ganciclovir↗
PubMed1994

Retrovirus-mediated gene transfer of human pyruvate kinase (PK) cDNA into murine hematopoietic cells: implications for gene therapy of human PK deficiency.

With the advent of recent molecular studies, nonspherocytic hemolytic anemia caused by red blood cell pyruvate kinase (PK) deficiency is now considered to be caused by a structural mutation of the PK-LR gene. Because PK deficiency is a monogenic disorder, the introduction of the normal PK gene into a patient's bone marrow stem cells should cure the disorder. To study the feasibility of gene therapy for PK deficiency, we first constructed the PK retrovirus pMNSM-hPK using human liver-type PK (LPK) cDNA and obtained a producer cell line of E86/AmPK. By using the supernatant of this virus-producer cell, we transduced NIH/3T3 cells, mouse leukemic cells (NFS60, FDCP-2), and human leukemic cells (K562, HEL). The expression of human LPK enzyme activity was ascertained from the retrovirally transduced NIH/3T3 cells. Northern blot analysis demonstrated the expression of the human LPK mRNA in each transduced cell line. Furthermore, bone marrow stem cells (c-kit+, Lin-, Thy-1lo) sorted by fluorescence-activated cell sorting were also transduced by the producer cells in the presence of interleukin-3 and interleukin-6, and were transplanted into lethally irradiated C57BL/6 mice. Polymerase chain reaction analysis demonstrated the expression of human LPK mRNA in both the peripheral blood and hematopoietic organs on day 30 and on day 135 of bone marrow transplantation.

Animals↗
PubMed1994

Human monocyte chemoattractant protein-1 expressed in a baculovirus system.

Human monocyte chemoattractant protein-1 (hMCP-1) was produced using a baculovirus system. The hMCP-1 cDNA was inserted into the genomic DNA of Autographa californica nuclear polyhedrosis virus (AcNPV) using a transfer vector, pJVP10Z. Spodoptera frugiperda insect cells, which were infected with this recombinant virus, secreted recombinant hMCP-1 (re-hMCP-1) at the level of 10-20 micrograms/ml of culture medium. This product was shown to chemoattract monocytes. Three distinct bands of 11, 11.5 and 12 kDa were revealed by immunoblotting analysis, and this heterogeneity was assigned to differences in carbohydrate processing. N-terminal amino-acid sequence analysis of the purified product revealed identity with hMCP-1. Thus, in this system, re-hMCP-1 was produced in large quantities and modified in a manner similar to native hMCP-1.

Amino Acid Sequence↗
PubMed1994

Molecular cloning and characterization of G-CSF induced gene cDNA.

G-CSF (granulocyte colony-stimulating factor) is known to specifically stimulate the production and the functional activation of neutrophils. To investigate the intracellular signaling pathway of myeloid cells stimulated by G-CSF, we isolated new genes whose expression was induced by G-CSF. First of all, we constructed lambda gt10 cDNA library from G-CSF-stimulated mononuclear cells (MNC) of a chronic myelogenous leukemia (CML) patient (CML-MNC) and screened the cDNA library by a differential hybridization method. The 24 candidate clones which specifically hybridized with G-CSF-stimulated CML-MNC cDNA probes, but not with unstimulated CML-MNC cDNA probes, were obtained after 8 x 10(4) individual clones had been screened. One of these clones, GIG-1 (G-CSF-induced gene-1), was further characterized. The size of the GIG-1 mRNA was about 0.9kb. The GIG-1 mRNA was expressed mainly in the myeloid leukemic cell lines.

Amino Acid Sequence↗
PubMed1994

Production of human granulocyte colony stimulating factor by various kinds of stromal cells in vitro detected by enzyme immunoassay and in situ hybridization.

Production of human granulocyte colony stimulating factor (G-CSF) by stromal cells was studied in vitro. Induction of G-CSF by interleukin 1 (IL-1) and lipopolysaccharide (LPS) was compared using enzyme immunoassay in various kinds of stromal cells. Primary human bone marrow stromal cells, a human bone marrow-derived stromal cell line (KM-102), and peripheral blood monocytes secreted small amounts of G-CSF without stimulation, while vascular endothelial cells and skin fibroblasts secreted G-CSF only when induced by IL-1 or LPS. The production of G-CSF by monocytes was stimulated predominantly by LPS, whereas that by KM-102 cells, endothelial cells, and fibroblasts was induced by IL-1 but much less so by LPS. IL-1 and LPS stimulated similar levels of G-CSF production by primary bone marrow stromal cells which consisted of various types of cells. In situ hybridization for G-CSF mRNA showed that only a small proportion of primary bone marrow stromal cells expressed a large amount of G-CSF mRNA upon stimulation. The positive cells were round or oval in shape, while most of the spindle-shaped stromal cells were negative for specific grains. Although further characterization of positive cells is needed, the results suggest that bone marrow stromal cells are heterogeneous in terms of their capacity for G-CSF production.

3T3 Cells↗
PubMed1994

Richter's syndrome showing pronounced lymphadenopathy in response to administration of granulocyte colony-stimulating factor.

A patient with Richter's syndrome developed rapid generalized lymph node enlargement with a decrease of peripheral blood lymphocytes after recombinant human granulocyte colony-stimulating factor (rhG-CSF) therapy for neutropenia induced by chemotherapy. The lymphadenopathy subsided spontaneously following discontinuation of rhG-CSF medication. Reinstitution of rhG-CSF therapy was followed by the same response as during initial therapy. Histopathologically, the lesions were characteristic of diffuse large cell lymphoma (DLL) with no evidence of myeloid cell involvement. No spontaneous contraction of enlarged lymph nodes followed withdrawal of the second course, but the enlargement subsided with chemotherapy. The patient died of myocardial infarction. All residual tumors examined post mortem presented microscopic features of small lymphocytic lymphoma (SLL), and G-CSF receptor was demonstrated on these neoplastic cells by Northern blot hybridization analysis. This observation indicates that some B cell malignancies may retain G-CSF receptor and respond to G-CSF.

Aged↗
PubMed1994

Recombinant human glycosylated granulocyte colony-stimulating factor (rhG-CSF)-combined regimen for allogeneic bone marrow transplantation in refractory acute myeloid leukemia.

Recombinant human glycosylated G-CSF (rhG-CSF) may stimulate proliferation of myeloid leukemia cells and thereby increase their susceptibility to anti-cancer agents. By in vitro colony assay, the rhG-CSF-responsive NFS-60 leukemic cell clones are more effectively killed by Ara C in the presence of rhG-CSF than in the absence of rhG-CSF, while the killing of the rhG-CSF-unresponsive HL-60 cell clones is unaffected by rhG-CSF. Leukemia cell colony forming units (L-CFU) derived from most AML patients demonstrate similar results to those of the NFS-60 cell clone when treated in vitro. Encouraged by these in vitro results, we used rhG-CSF as a component of a conditioning regimen for 15 relapsed AML patients who were receiving allogeneic BMT. The patients were conditioned with total body irradiation (TBI) and high-dose Ara C. rhG-CSF was infused continuously at a dose of 5 micrograms/kg/day from 24 h before the beginning of TBI to the end of Ara C therapy. Proliferation of the leukemia cells in vivo in response to rhG-CSF was confirmed in 7 of 14 patients tested and the combined use of rhG-CSF had no additional adverse effects. After BMT, four patients died of non-leukemic causes and three patients had leukemic relapse: the other eight patients have remained disease-free for 200-1600 (median 417) days. The actuarial probabilities of relapse and disease-free survival (DFS) at 4.4 years after BMT were 43.2% and 41.7%, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Acute Disease↗
PubMed1994

[Proliferation of pulmonary fibroblasts obtained from rats with bleomycin-induced pulmonary fibrosis and effects of rat rIL-1 alpha on this proliferation].

To clarify the mechanism of pulmonary fibrosis, the growth rate of fibroblasts obtained from bleomycin (BLM)-treated rats was compared with that of fibroblasts obtained from control rats. Proliferation of fibroblasts obtained from rats at 4 days after BLM instillation (BRF) was significantly more rapid than that of fibroblasts obtained from rats at 4 days after saline instillation (CRF), as assessed by cell counting and 3H-TdR incorporation. CRF and BRF were separated by the method of discontinuous Percoll gradients. Three fibroblast fractions of specific gravity (S.G.), S.G. < 1.036, 1.036 < or = S.G. < 1.050, 1.050 < or = S.G. < 1.062, were obtained. Percentages of the densest fibroblast fraction were 46.2 +/- 5.2 in BRF and 6.4 +/- 2.8 in CRF. The densest fibroblasts in BRF proliferated more rapidly than the least dense fibroblasts. Rat rIL-1 alpha suppressed the proliferation of CRF and BRF, dose dependently. These results suggest that an increase in the densest fibroblasts in the lung might correlate with BLM-induced pulmonary fibrosis and that IL-1 alpha suppresses the proliferation of pulmonary fibroblasts.

Animals↗
PubMed1994

A marked decrease in defensin mRNA in the only case of congenital neutrophil-specific granule deficiency reported in Japan.

A deficiency of defensins (human neutrophil peptides, HNP) has been previously demonstrated in two individuals with congenital neutrophil-specific granule deficiency (SGD), but its genetic basis is not well understood. We have studied another case of SGD, the only case reported in Japan, to elucidate the molecular basis of defensin deficiency. Using Western blot analysis, we showed for the first time that HNP-4, a novel human defensin, was also deficient in the patient's neutrophils. Northern blot analysis demonstrated that the defensin mRNA was absent in the bone marrow cells of the patient. Limited Southern blot analysis, using a defensin cDNA probe, did not reveal any differences in fragment patterns of the genomic DNA between the patient and the control. We propose that the defensin deficiency in our patient is a consequence of a marked decrease in defensin mRNA in neutrophil precursors. Our findings were consistent with a previous speculation suggesting that the SGD defect occurs at the level of transcription.

Adult↗
PubMed1994

[A case of aseptic meningoencephalitis in a patient with secondary Sjögren syndrome with systemic lupus erythematosus].

A 26-year-old woman, who had been treated with prednisolone 30 mg/day for secondary Sjögren syndrome (SjS) with systemic lupus erythematosus (SLE), was complicated with aseptic meningoencephalitis (AME). Cerebrospinal fluid (CSF) cell counts were 26/3, total protein 126 mg/dl, glucose 41 mg/dl. The CSF, blood, nasopharyngeal swab were cultured for bacteria, mycobacteria, viruses and fungi, but no evidence for an infectious etiology was obtained in this patient. The pair sera study for viruses titer were performed, but no significant titer elevation was observed. Head magnetic resonance imaging showed remarkable atrophy of bilateral occipital lobe and dilatation of posterior horn of bilateral ventricles. The high 2-5AS enzyme activity in the CSF suggested indirectly that the cause of this meningoencephalitis was virus infection. In this case microbiological studies were negative, so central nervous system SLE and/or central nervous system SjS were not completely neglected.

Adult↗
PubMed1994

[A case of sarcoidosis with elevated SCC antigen in the serum and broncho-alveolar lavage fluid which was difficult to distinguish from lung cancer].

A case of sarcoidosis with a high level of SCC antigen in the serum and broncho-alveolar lavage fluid (BALF) which was difficult to distinguish from lung cancer is reported. A 66-year-old female was admitted with a fever, sputum and coughing. Chest X-ray on admission showed bilateral hilar swelling and infiltrative shadows in the left lower lung. Chest CT showed bilateral lymph node enlargement and a localized nodular mass. The level of SCC antigen in the serum and BALF was high. Initially she was suspected of having lung cancer, but she was subsequently diagnosed as having sarcoidosis from a bronchial biopsy specimen, BALF and gallium scintigraphy. It was difficult to distinguish this case showing lymph node enlargement and mass shadow from lung cancer. Although this is a rarely reported case of sarcoidosis with a high level of SCC antigen in the serum and BALF, it is possible that SCC antigen may be elevated in patients with benign inflammatory lung disease due to various interactions of cytokines.

Aged↗
PubMed1993

Stepwise assembly of functionally active transport vesicles.

Budding of COP-coated vesicles (the likely carriers of newly synthesized proteins from the endoplasmic reticulum through the Golgi stack) from Golgi cisternae requires ADP-ribosylation factor (ARF), coatomer proteins from the cytosol, GTP, and fatty acyl-coenzyme A (CoA). The assembly of coated buds on the membranes requires coatomer, ARF, and GTP. When palmitoyl-CoA is added, membrane fission occurs at the coated bud, releasing coated vesicles. We show that COP-coated vesicles can be generated stepwise in vitro and isolated in a functionally active form, demonstrating that the minimal set of cytosolic components required for their formation as well as principal steps in their assembly have been identified.

ADP-Ribosylation Factors↗
PubMed1993

Binding of membrane-anchored macrophage colony-stimulating factor (M-CSF) to its receptor mediates specific adhesion between stromal cells and M-CSF receptor-bearing hematopoietic cells.

To explore the biologic significance of the membrane-anchored form of macrophage colony-stimulating factor (M-CSF), we examined whether interaction between membrane-bound M-CSF and its receptor mediates intercellular adhesion as well as cell proliferation and differentiation. Human M-CSF receptors were expressed on a murine interleukin-2 (IL-3)-dependent cell line, FDC-P2, by DNA transfection with the c-fms gene (FDC-P2-MCSFR). A human bone marrow-derived stromal cell line, KM102, was used in the cell adhesion assay. The expression of membrane-bound M-CSF on KM102 cells was demonstrated by flow cytometry and immunoblot analysis. After the incubation of parent and transformed FDC-P2 cells on confluent KM102 cells, nonadherent cells were removed and the cells attached to KM102 cells were examined microscopically. Almost all parent FDC-P2 cells were nonadherent, whereas a significant number of FDC-P2-MCSFR cells bound to KM102 cells. The addition of anti-M-CSF or anti-M-CSF receptor neutralizing antibodies dose-dependently inhibited the binding of [3H]-thymidine-labeled FDC-P2-MCSFR cells to KM102 cells. An excess amount of M-CSF also inhibited the binding. On the other hand, the addition of antibodies against some representative adhesion molecules (vitronectin receptor, Pgp-1/CD44, and VLA-4) did not inhibit the adhesion between FDC-P2-MCSFR cells and KM102 cells. These results support the idea that membrane-anchored M-CSF and its receptor may function as mediators of cell-cell adhesion.

Animals↗
PubMed1993

Changes in hematopoiesis-supporting ability of C3H10T1/2 mouse embryo fibroblasts during differentiation.

To investigate the functional change of stromal cells along with differentiation, we used a differentiation-inducible mouse embryo fibroblast cell line, C3H10T1/2 (10T1/2). Stably determined preadipocyte and myoblast cell lines were established after a brief exposure of 10T1/2 cells to 5-azacytidine. These cell lines terminally differentiated into adipocytes and myotubes, respectively, under appropriate conditions. The hematopoiesis-supporting ability of each 10T1/2-derived cell line was examined by coculture with FACS-sorted murine hematopoietic stem cells (Thy-1lo c-kit+ Lin-). The number of granulocyte-macrophage progenitors (CFU-GM) was slightly reduced after 7 days of culture with parent 10T1/2 fibroblasts, whereas a marked increase in CFU-GM number was observed when the cells were cultured on preadipocytes. Mature adipocytes and myogenically determined cell lines, on the other hand, did not support CFU-GM growth. Further, Northern analysis showed that the preadipocyte cell line acquired the ability to produce a significant amount of stem cell factor (SCF), interleukin-6 (IL-6), leukemia inhibitory factor, and macrophage colony-stimulating factor mRNAs in response to IL-1 or lipopolysaccharide stimulation. Terminal adipocytic differentiation resulted in reduced ability to express these cytokine mRNAs. Similarly, highest IL-6 activity was detected in the supernatant of preadipocyte culture, whereas adipocytes did not secrete IL-6 even after IL-1 stimulation. Interestingly, hematopoiesis-nonsupporting myoblasts and myotubes also expressed abundant SCF mRNA, suggesting that SCF, per se, may not be sufficient for stem cell growth and survival. The 10T1/2-derived cell lines could provide a valuable tool to aid in the analysis of stromal cell development and the search for novel stromal factors.

Adipose Tissue↗
PubMed1993

Loss of heterozygosity on chromosome arm 17p in small cell lung carcinomas, but not in neurofibromas, in a patient with von Recklinghausen neurofibromatosis.

BACKGROUND: It has been suggested that the genetic abnormality responsible for von Recklinghausen neurofibromatosis (NF1) increases a patient's risk of various kinds of malignancies. The incidence of small cell lung carcinoma (SCLC) as a complication of NF1, however, is rare. To clarify the relationship between NF1 and SCLC, possible loss of heterozygosity of chromosome 17 in a patient with SCLC combined with NF1 was analyzed. METHODS: Possible loss of heterozygosity for chromosome 17 was analyzed by a molecular genetic approach using several chromosome 17-specific polymorphic DNA markers. RESULTS: In both primary tumor and metastatic tumors of SCLC, loss of heterozygosity was detected on chromosome arm 17p, but not on 17q. Loss of heterozygosity, however, was detected on neither 17p nor 17q in neurofibromas and normal tissue. CONCLUSIONS: The formation of SCLC may result from several genetic alterations, including inactivation of tumor-suppressor gene on chromosome 17p, most likely P53, although it still is unknown whether or not a mutation of the NF1 gene on 17q was involved in the development of SCLC in this patient.

Alleles↗
PubMed1993

Polyclonal B cell activation and antigen-specific stimulation in mice with chronic graft-versus-host reaction.

Primary chronic graft-versus-host reaction (GVHR) was induced in (C57BL/10 x DBA/2)F1 (BDF1) mice via the transfer of parental DBA/2 derived lymphocytes. A significant increase in the absolute number of IgM- and IgG-secreting cells was found in recipient BDF1 mice at 3 weeks post-transfer. The increase included B cells reactive with both self- and non-self-antigens, without being skewed toward the production of autoantibodies. Later in the disease process (approximately 6-8 weeks post-transfer), we detected an increased number of B cells that secreted antibodies reactive with a select subset of autoantigens in about 30% of BDF1 recipients (3 of 9 mice). These findings suggest that not only antigen-specific stimulation, which was detected in a late stage, but also initial polyclonal activation may account for chronic GVHR.

Animals↗