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Biomedical subjects

K Harigaya

Publications and source records attributed to K Harigaya.

At least 73 records · Page 4Linked to original sources
PubMed1990

A case of massive true thymic hyperplasia with non-Hodgkin's lymphoma.

A case of massive true thymic hyperplasia with non-Hodgkin's lymphoma of the mediastinum is reported in a 14-year-old boy. Computed tomographic scan of the chest showed a mass of the anterior mediastinum and conspicuous swelling of the lymph nodes in the upper and lower mediastinum. They were grossly resected. The tumor of the anterior mediastinum was histologically diagnosed as true thymic hyperplasia. All of the mediastinal lymph nodes were diagnosed as non-Hodgkin's lymphoma, diffuse, mixed small and large cell type.

Adolescent↗
PubMed1990

Gap-junctional communication of bone marrow stromal cells is resistant to irradiation in vitro.

Bone marrow is one of the most radiosensitive organs. Irradiation causes a marked decrease in the total number of hematopoietic cells in the bone marrow. The reticular meshwork structure of marrow stromal cells, however, is relatively resistant to irradiation. Unimpaired stromal cell structure has been thought to be a prerequisite for the repopulation of hematopoietic cells during recovery from the effects of irradiation. The reticular framework is maintained by cell adhesion apparatuses such as gap junctions. The in vitro radiobiologic survival values of a cloned stromal cell line, H-1/A, were studied (ñ = 1.8, D0 = 138 cGy). Radiation doses of up to 4000 cGy had no detectable effects on the production of colony-stimulating factor 1. H-1/A cells communicate with each other via gap junctions as determined by the sensitive dye-transfer method. Gap-junctional communication between H-1/A cells was resistant to different levels of irradiation (500 to 10,000 cGy), but it was lost during adipocyte differentiation of H-1/A cells. Marrow stromal cells, which are important in the recovery of hematopoiesis, seemed capable of coordination with each other through gap junctions even when exposed to radiation.

Bone Marrow↗
PubMed1989

Differentiation of human hematopoietic cells increases expression of a gene transferred by a retroviral vector.

To study expression of a retroviral vector in human hematopoietic lineages, two established human hematopoietic cell lines (HL60 and K562) and a human adherent stromal cell line (KM101) were infected with the vector pZIP-SV(X). Expression of the transferred neomycin resistance gene (neor) of pZIP-SV(X) was evaluated as the ability of the cells to form colonies (greater than 50 cells) in an agar assay in the presence of the neomycin analogue, G418. After infection, all three cell lines produced colonies resistant to G418. The level of neor mRNA in separate colonies was analyzed by Northern blot analysis. The neor gene transferred by the vector pZIP-SV(X) was expressed in both human hematopoietic and stromal cell lines. In addition, primary adherent human stromal cells infected with pZIP-SV(X) grew in the presence of G418. To determine if differentiation of hematopoietic cells affects expression of the retroviral vector, HL60 cells infected with pZIP-SV(X) were induced to differentiate, and the level of neor mRNA measured. The amount of neor mRNA increased when HL60 cells were induced to differentiate along the granulocytic pathway. Conversely, when HL60 cells were induced toward monocytoid differentiation (TPA), the level of neor mRNA did not significantly increase. We conclude that the neor gene transferred by a retroviral vector, pZIP-SV(X), is functionally expressed. In addition, expression of the transferred neor gene is regulated during myeloid differentiation of HL60 cells.

Cell Differentiation↗
PubMed1989

Infection of hematopoietic and stromal cells in human continuous bone marrow cultures by a retroviral vector containing the neomycin resistance gene.

Stability and expression of the bacterial neomycin resistance gene (neor) transferred to human continuous marrow cultures by a retroviral vector [pZIP-NeoSV(X)] was evaluated over 4 weeks. Following infection of long-term human marrow cultures with pZIP-NeoSV(X), 10-15% of the stromal cells demonstrated high replating efficiency in a dose of the neomycin analogue G418 that was toxic to stromal cells from uninfected cultures. In contrast, G418 resistance was detected in less than or equal to 1% of GM-CFUc and CFU-GEMM derived from the same virus-infected compared to control cultures. Infection of human CFU-GEMM enriched 100 X by monoclonal antibody selection with pZIP-NeoSV(X) did not increase the percentage of neor progenitors. Marrow cells from cultures infected with pZIP-NeoSV(X) and a replication competent amphotropic virus transferred the vector and G418 resistance to HeLa cells at a frequency of 1/10(5) for nonadherent and 1/10(4) for adherent cells. Two established human hematopoietic (HL60 and K562) and one stromal cell line (KM101) stably expressed the neor gene. Thus, a higher efficiency of infection and expression of a gene transferred by pZIP-NeoSV(X) to permanent human hematopoietic tumor cell lines and fresh marrow stromal cells contrasts with a lower level of expression in fresh CSF-dependent human hematopoietic stem cells.

Bone Marrow↗
PubMed1989

Novel functions and cellular interaction of human lymphoid stromal cells with lymphoid cell lines in vitro.

Lymphoid stromal (SG) cells have been isolated from the lymph node of a patient with malignant lymphoma, and characterized by positive reaction with a monoclonal antibody against the T-zone stromal cells in human lymph nodes. B-acute lymphoblastic leukemia (BALL) cells showed prominent emperipolesis toward SG cells when they were cocultured, whereas T-acute lymphoblastic leukemia (TALL) cells attached firmly to the surface of SG cells. Autologous peripheral B and T cells behaved, respectively, in the same way as BALL and TALL cells. Both BALL and TALL cells while directly interacting with the SG cells were completely inhibited from incorporating [3H]thymidine, although radioactive grains were observed in 16.4%-12.4% of supernatant BALL and 13.8%-13.0% of supernatant TALL cells in each coculture. Furthermore the media conditioned by SG cells significantly increased the incorporation of [3H]thymidine into the TALL cells as much as 190% of the control. These results indicate that SG cells undergo tissue-specific cellular interactions with B- and T-lymphoid cell lines but not with a myeloid cell line, and they can modify their growth by two distinct mechanisms. SG cells proved to be very useful in studying the effect of the lymphoid microenvironment on the proliferation of lymphocytes in vivo.

Burkitt Lymphoma↗
PubMed1988

Radiosensitivity of permanent human bone marrow stromal cell lines: effect of dose rate.

In contrast to the dose-rate independent X ray killing observed with human bone marrow hematopoietic stem cells, bone marrow adherent stromal cells from the same fresh marrow harvests demonstrate increased radiation resistance at low dose rate (LDR) (5 cGy/min), compared to high dose rate (HDR) irradiation (120-200 cGy/min). Physiologic changes observed in plateau phase bone marrow cells after LDR irradiation in vivo and in vitro suggested that marrow stromal cells might be heterogeneous in LDR irradiation repair. Five permanent clonal bone marrow stromal lines were derived from a single human marrow donor. Each cell line was positive for markers of fibroblasts including: immunohistochemically detectable fibronectin, collagen, acid phosphatase, and nonspecific esterase, and was negative for Factor VIII, alkaline phosphatase, lysozyme and several markers of marrow macrophages. The x-irradiation survival curve of each cell line was determined at LDR and HDR in vitro. Cell lines KM102, KM103, KM104, and KM105 each demonstrated a significant (p less than .05) increase in radioresistance at LDR (D0 = 142, n = 2.9; D0 = 131, n = 2.5; D0 = 145, n = 2.1 and D0 = 127, n = 2.1 respectively) compared to HDR: (D0 = 111, n = 2.1; D0 = 94, n = 3.5; D0 = 99, n = 3.5 and D0 = 95, n = 2.1 respectively). In contrast, cell line KM101 demonstrated no significant change in radiosensitivity relative to dose rate at LDR (D0 = 113, n = 3.3) compared to HDR, D0 = 114, n = 3.3. Cell line KM101 was more supportive than the other lines of cocultivated hemopoietic cells in vitro. Subclones of KM101 and KM104 selected by retroviral vector transfer of the neor gene for growth in the antibiotic neomycin-analogue G418, maintained the stably associated radiobiologic properties of each parent clonal line. These data indicate significant heterogeneity in the LDR irradiation response of clonal stromal cell lines derived from human bone marrow.

Bone Marrow↗
PubMed1988

A burst-promoting activity derived from the human bone marrow stromal cell line KM-102 is identical to the granulocyte-macrophage colony-stimulating factor.

Recently, several human bone marrow stromal cell lines have established and produced hematopoietic growth factors. One of these factors, a burst-promoting activity (BPA), was purified from 6 liters of serum-free conditioned medium cultured from stromal cell line KM-102, which was stimulated by phorbol myristate acetate (PMA) and calcium ionophore A23187. This stimulation induced 60 times more production of BPA than the unstimulated control culture. BPA was purified 4000-fold by sequential fractionation using ammonium sulfate precipitation, anion-exchange and lentil lectin affinity chromatographies, high performance gel filtration chromatography, and reversed phase high performance liquid chromatography. Purified BPA gave a single broad band of protein with a molecular weight of approximately 18 kd, as assessed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The concentration required for half maximal growth of early erythroid colonies was estimated as 10 pg/ml or 0.6 pM. At a higher concentration (125 pg/ml) this factor also stimulates the growth of granulocyte, macrophage, and eosinophil colonies in agar culture. The profile of amino acid composition is very similar to that of the human granulocyte-macrophage colony-stimulating factor (GM-CSF) deduced from its complementary DNA sequence. The result of amino-terminal sequence analysis strongly suggests that the purified material consists of GM-CSF and tetrapeptide-deleted GM-CSF. Moreover, antibody against GM-CSF completely neutralized the biological activities of this factor. These results indicate that the human bone marrow stromal cell line secretes GM-CSF as a burst-promoting activity and GM-CSF may play a significant role in the interaction between stem cells and stromal cells in the hematopoietic microenvironment.

Amino Acid Sequence↗
PubMed1987

Effect of direct cell-to-cell interaction between the KM-102 clonal human marrow stromal cell line and the HL-60 myeloid leukemic cell line on the differentiation and proliferation of the HL-60 line.

Hematopoietic cellular interaction was investigated in a coculture of the human clonal marrow stromal line, KM-102, and the myeloid leukemia cell line, HL-60. In the coculture, a large number of HL-60 cells remained in the supernatant but some of them became firmly attached to KM-102 cells. The attached HL-60 cells showed little positive reaction in the NBT test when the culture was supplemented with 10(-9) to 10(-7) M 1 alpha, 25-dihydroxyvitamin D3. In contrast, differentiation in the supernatant HL-60 cells was strikingly responsive to the agent in a dose-dependent way. Furthermore, the complete inhibition was observed in the incorporation of [3H]thymidine into the attached HL-60 cells with autoradiography, but 23.6% of the supernatant cells moderately incorporated [3H] thymidine into their nuclei. There was no attachment between HL-60 cells and stromal cells from human thymus and lymph node, or between lymphocytic leukemia cells and marrow stromal cells. These results indicate that there is direct cellular interaction between myeloid leukemic cells and marrow stromal cells which modulates the proliferation and differentiation of the myeloid leukemic cells. This modulation by marrow stromal cells is more strongly affected by this interaction than by exogenously added differentiation-inducing agents. Apparently marrow stromal cells produce a definitive milieu for the proliferation and differentiation of myeloid leukemic cells.

Bone Marrow Cells↗
PubMed1986

Conversion to adipocytes of a clonal bone marrow preadipocyte line (H-1/A) and fatty acid composition of the resultant adipocytes.

Conversion to adipocytes and fatty acid composition were investigated in a clonal bone marrow preadipocyte line (H-1/A). The growing cells exhibited a fibroblastic appearance. After the cessation of growth, triacylglyceride (TG) synthesis in the cells increased as they incorporated precursor from the growth medium and became adipocytes. Hydrocortisone and insulin accelerated the TG synthesis in H-1/A cells in a dose-dependent manner when they were cultured in the growth medium containing 10% horse serum. The rate of conversion to adipocytes was reduced as the concentration of horse serum was decreased, and this reduction was not influenced by the addition of insulin and/or hydrocortisone. These results suggest that conversion to adipocytes of H-1/A cells is primarily dependent on some component(s) of the serum. Conversion to adipocytes of the cells may involve a process of differentiation since the conversion was completely inhibited when the cells were cultured in the presence of bromodeoxyuridine. Fatty acid composition was significantly different between adipose H-1/A cells and adipocytes derived from other marrow preadipocyte line MC3T3-G2/PA6 cells. Unsaturated fatty acids accounted for 76% of the fatty acid composition of adipose H-1/A cells; in contrast, saturated fatty acids constituted 65% of the fatty acid composition of the adipose MC3T3-G2/PA6 cells. These results suggest that there is a heterogeneity of preadipocytes in bone marrow. These two preadipocyte lines thus provide a useful tool for the study of marrow adipocytes and can also be used to analyze the hematopoietic microenvironment through studies of the effect of these cells on hematopoietic cell proliferation.

Adipose Tissue↗
PubMed1985

Generation of functional clonal cell lines from human bone marrow stroma.

Five clonal human bone marrow stromal cell lines were isolated from the adherent cell populations in long-term liquid cultures after transfection with the recombinant plasmid pSV3gpt. All the cell-line feeder layers and their conditioned media stimulated the proliferation of committed granulomonocytic stem cells (CFUc) from human bone marrow. The size and number of early erythroid stem cell (BFUe)-derived colonies were significantly increased when in the presence of 10% conditioned medium from the cell lines. Furthermore, a substantial number of mixed colonies with erythroid components were observed in the cultures in the presence of erythropoietin and conditioned medium. These findings suggest that the human bone marrow stromal cell lines obtained after transfection with pSV3gpt may be extremely useful in identifying the hematopoietic factors derived from the hematopoietic microenvironment and in analyzing their mechanism of action.

Bone Marrow↗
PubMed1985

Correlation between production of colony-stimulating activity (CSA) and adipose conversion in a murine marrow-derived preadipocyte line (H-1/A).

The correlation between adipose conversion of cloned H-1 cells (H-1/A) and their production of colony-stimulating activity (CSA) was examined. The production of CSA from H-1/A cells declined after adipose conversion, although H-1/A cells are active producers of CSA during their fibrocytic stage. The addition of 2 X 10(-5) M 5-bromo-2'-deoxyuridine to the cultures almost completely inhibited adipose conversion and there was no reduction of CSA levels after 9 days of culture. On the other hand, the addition of 10(-6) M hydrocortisone sodium succinate to the culture markedly enhanced adipose conversion, and a greater reduction in the CSA level was observed in the supernatants than in the control cultures after 12 days of culture. Indomethacin had no effect on the production of CSA or on adipose conversion. Furthermore, there were no significant differences between the CSA levels of nondialyzed supernatants and dialyzed supernatants from the control cultures during the entire course of the experiment. Supernatants during the adipocyte stage of H-1/A cultures did not inhibit the CSA derived from the fibrocytic stage. There were no differences in colonies in agar cultures stimulated by supernatants derived from cultures that had undergone either of the above treatments. These results suggest that the reduction of CSA is not due to the production of inhibitors, but that the production of CSA declines after adipose conversion of H-1/A cells. Preadipocytes in bone marrow therefore appear to contribute to granulopoiesis during the fibrocytic stage and are hematopoietically inactive when they convert to adipocytes.

Adipose Tissue↗
PubMed1984

Influence of a fibroblastoid cell line derived from murine bone marrow (H-1 cells) on stem cell proliferation.

A murine fibroblastoid cell line (H-1) with properties similar to those of adventitial reticular cells can support granulopoiesis and the development of mononuclear phagocytes in vitro. In the current study the effect of these cells on stem cell maintenance in vitro was assessed. The H-1 cells were unable to support CFUs replication in liquid culture, while treatment of some stem cells with H-1 conditioned medium appeared to inhibit their proliferation.

Animals↗