[Induction of leukemic cell differentiation].
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Cell Differentiation↗
Biomedical subjects
M Hozumi
Publications and source records attributed to M Hozumi.
Inhibition by interleukin 4 of leukemia inhibitory factor-, interleukin 6-, and dexamethasone-induced differentiation of mouse myeloid leukemia cells: role of c-myc and junB proto-oncogenes.
Interleukin 4 (IL-4) inhibited the differentiation of mouse myeloid leukemia M1 cells induced by leukemia inhibitory factor (LIF), interleukin 6, or dexamethasone and conversely enhanced the induction of M1 cell differentiation by 1 alpha,25-dihydroxyvitamin D3. IL-4 blocked LIF-induced differentiation of M1 cells when it was added to the culture medium within 10 h after LIF, but IL-4 did not block differentiation when it was added 12 h after LIF. These results indicate that IL-4 inhibited a critical intermediate step in myeloid leukemia cell differentiation. LIF markedly stimulated the expression of junB mRNA within 2 h but suppressed the expression of c-myb and c-myc after 2- and 12-h treatment, respectively. IL-4 did not significantly affect LIF-induced junB expression or suppression of c-myb expression. However, it interfered significantly with the LIF-induced suppression of c-myc gene expression. Similar results were obtained when interleukin 6 was used to induce differentiation of M1 cells. Dexamethasone and 1 alpha,25-dihydroxyvitamin D3 did not induce junB gene expression but suppressed the expression of c-myb and c-myc. IL-4 also interfered with dexamethasone-induced suppression of c-myc gene expression. On the other hand, IL-4 enhanced 1 alpha,25-dihydroxyvitamin D3-induced down-regulation of c-myc gene expression, consistent with its enhancement of differentiation. These results indicate that the change in c-myc expression induced by IL-4 in M1 cells is closely associated with the effect of IL-4 on the induction of differentiation of M1 cells.
Animals↗
Effects of herbimycin A and its derivatives on growth and differentiation of Ph1-positive acute lymphoid leukemia cell lines.
The molecular basis of the Philadelphia chromosome (Ph1) is a structurally altered c-abl (bcr/abl) gene which encodes an abnormally large protein with protein tyrosine kinase activity. Herbimycin A, an inhibitor of tyrosine kinase, preferentially inhibited the growth of Ph1-positive acute lymphoid leukemia (ALL) cell lines, as well as Ph1-positive chronic myeloid leukemia (CML) cell lines. Although noncytotoxic concentrations of herbimycin A induced erythroid differentiation of two CML-derived cell lines, K562 and KU812, in a previous study, the differentiation-inducing effect of herbimycin A on Ph1-positive ALL cell lines was less strong. Herbimycin A enhanced some differentiation-associated properties of one Ph1-positive ALL cell line, L2, but the effect of herbimycin A on the other Ph1-positive ALL cell lines was cytotoxic rather than cytostatic (differentiation-inducing). Several derivatives of herbimycin A were synthesized and their effects on the cell proliferation of Ph1-positive CML and ALL cell lines were examined. The sensitivities of the Ph1-positive cell lines to herbimycin A derivatives were different from the data on the rat kidney cell line infected with Rous sarcoma virus (v-src) derived from a previous study, suggesting bcr/abl kinase may differ in sensitivity from other tyrosine kinases. Moreover, the sensitivities of the ALL cell lines were not the same as those of the CML cell lines. These results suggest that a specific inhibitor of bcr/abl kinase could be an effective antileukemic agent against Ph1-positive CML or ALL.
Antibiotics, Antineoplastic↗
Three different cDNAs encoding mouse D-factor/LIF receptor.
Three cDNAs for mouse differentiation-stimulating factor (D-factor)/leukemia inhibitory factor (LIF) receptor were isolated from a cDNA library prepared from the liver of a pregnant mouse. A probe for screening was prepared by the RT-PCR method using human cDNA sequences as primers. The mouse D-factor receptor cDNA encoded 1,092 amino acids, which had a marked homology with the human counterpart and consisted of signal sequence, extracellular, transmembrane, and cytoplasmic domains. The WSXWS motif found in members of the cytokine receptor family was also present in the extracellular domain of the mouse D-factor receptor. A second form of cDNA that had a 501 bp insertion was isolated. The insertion introduced a stop codon so that the mRNA encoded the soluble receptor lacking transmembrane and intracellular domains. Because the insertion contained polyadenylation signals, two different sizes of mRNA encoding the soluble receptor were produced, depending on whether or not it utilized these signals. Transcripts utilizing these signals were 2.6-3 kb in size, and were very abundantly expressed in the liver. Transcripts that did not use these signals were longer than 5 kb and of similar size to the mRNA for the cellular receptor.
Amino Acid Sequence↗
Flow-cytometric analysis of in vivo induction of differentiation of WEHI-3B myelomonocytic leukemia cells by recombinant granulocyte colony-stimulating factor.
An animal leukemia model was developed to investigate in vivo induction of differentiation of myeloid leukemia cells. An aneuploid cell line (C15) was isolated from mouse myelomonocytic leukemia WEHI-3B D+ cells. The C15 cells contained twice as much DNA as the parental cells but retained the morphology of myelomonocytic cells and the ability to differentiate into macrophage-like cells in response to all-trans retinoic acid (ATRA) in vitro. When the C15 cells were inoculated into the peritoneal cavity of syngeneic Balb/c mice (10(6) cells/mouse), the mice died of leukemia within 19 days. The DNA content and differentiation antigen (Mac-1) of the cells in the peritoneal cavity were determined by dual-parameter flow cytometry. On day 12 after inoculation, the C15 cells were distinguishable from normal host cells in the peritoneal cavity by their different DNA content. The administration of recombinant human granulocyte colony-stimulating factor (rhG-CSF) (10 micrograms/day) to mice bearing C15 cells induced the leukemia cells to express Mac-1 antigen and to change morphologically into mature granulocytic cells. Because the C15 cells were not responsive to G-CSF in suspension culture in vitro, this result suggests that the cytokine's actions on the cells in vivo and in vitro are different. This experimental model for analyzing in vivo differentiation of leukemia cells will be useful for studying the therapeutic effects of potential differentiation-inducing agents.
Animals↗
Pregnancy associated increase in mRNA for soluble D-factor/LIF receptor in mouse liver.
We examined the distribution of mRNAs for differentiation-stimulating factor (D-factor)/leukemia inhibitory factor (LIF) receptor in various mouse tissues by Northern blotting. A mouse cDNA fragment encoding the D-factor receptor was prepared by the RT-PCR method using human cDNA sequences as primers. The smallest mRNA (3 kb) was present in the liver, but not detectable in other tissues examined. Larger mRNAs (5 and 10 kb) were present in the placenta and the M1 cells, and also detectable in the liver, kidney, heart, lung, brain and embryos. Expression of 3 kb mRNA in the liver increased during pregnancy, being 20 times the initial level on day 15. D-factor receptor cDNAs were isolated from a cDNA library prepared from the liver of a pregnant mouse. Most of the cDNA clones encoded a soluble receptor. A cDNA probe specific for the cellular receptor did not hybridize with 3 kb mRNA in the liver. These results suggest that 3 kb mRNA encodes a soluble D-factor receptor and that the liver is the primary site of synthesis of this soluble receptor.
Amino Acid Sequence↗
Differentiation of human monoblastic leukemia U937 cells induced by inhibitors of myosin light chain kinase and prevention of differentiation by granulocyte-macrophage colony-stimulating factor.
Inhibitors of protein kinase activities are useful for the study of intracellular signal transduction and some of these inhibitors are reported to induce differentiation of human leukemia cells. We examined effects of granulocyte-macrophage colony-stimulating factor (GM-CSF) in combination with several kinase inhibitors on differentiation of human leukemia U937 cells. Nitroblue tetrazolium (NBT)-reducing activity, a typical marker of myelomonocytic differentiation, of U937 cells was induced by genistein and GM-CSF enhanced this activity. GM-CSF also induced the NBT-reducing activity of the cells in combination with 2,5-dihydroxycinnamic acid methyl ester, psi-tectorigenin and staurosporine, although each of them did not induce the activity. Inhibitors of myosin light chain kinase, 1-(5-chloronaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (ML-9) and 1-(5-iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (ML-7), induced in U937 cells NBT-reduction, and lysozyme and alpha-naphthyl acetate esterase activities. GM-CSF inhibited this differentiation and counteracted the anti-proliferation effect of the kinase inhibitors. These results suggest that some protein kinases are involved in differentiation of U937 cells and the kinases inhibited by ML-9 and ML-7 are associated with signal transduction of GM-CSF.
Azepines↗
Pregnancy associated increase in differentiation-stimulating factor (D-factor)/leukemia inhibitory factor (LIF)-binding substance(s) in mouse serum.
Differentiation-stimulating factor (D-factor)/leukemia inhibitory factor (LIF) is known to have multiple biological activities besides induction of differentiation of mouse myeloid leukemia M1 cells. Little is known about how its activities are regulated in vivo, but it has been suggested to play a regulatory role in the mechanisms involved in development of mice. In this study, we found that a single class of D-factor-binding substance is present in normal mouse serum and that it increases transiently in the late stage of pregnancy. It inhibits the induction of differentiation of mouse myeloid leukemic M1 cells by D-factor by blocking the binding of this factor to the cells. It is a heat-labile protein with an apparent molecular weight of 130,000-150,000. The binding of 125I-D-factor to the substance is specific since it was inhibited by excess unlabeled D-factor, but not by interleukin 6 or interferon gamma. The dissociation constant of the binding substance for mouse D-factor in normal mouse serum is 6.6-9.4 nM. In the late stage of pregnancy, the amount of the D-factor-binding substance in the serum apparently increases about 30-fold. These results suggest that the D-factor-binding substance regulates the activity of D-factor during embryonic development of mice.
Animals↗
Genistein exhibits preferential cytotoxicity to a leukemogenic variant but induces differentiation of a non-leukemogenic variant of the mouse monocytic leukemia Mm cell line.
Mouse leukemia Mm-A and Mm-S2 cells are subclones of mouse monocytic leukemia Mm cells, Mm-A cells having much higher leukemogenicity than Mm-S2 cells. The growth-inhibitory effects of several protein kinase inhibitors on leukemogenic Mm-A and non-leukemogenic Mm-S2 cells were examined. Most inhibitors of protein serine/threonine kinases inhibited the growth of Mm-A and Mm-S2 cells similarly, but some protein tyrosine kinase inhibitors exhibited differential inhibitory effects on Mm-A and Mm-S2 cells. Genistein inhibited growth of Mm-A cells more effectively than that of Mm-S2 cells, but another inhibitor of tyrosine kinase, herbimycin A, preferentially inhibited growth of non-leukemogenic Mm-S2 cells. Genistein induced or enhanced several differentiation markers of Mm-S2 cells, such as cell spreading, immunophagocytosis, nitroblue tetrazolium (NBT) reduction and lysozyme activity in a dose-dependent manner, but herbimycin A did not. Genistein was cytotoxic to Mm-A cells rather than inducing cell differentiation. Genistein has effects on several other cellular events as well as inhibition of tyrosine kinases. However, it effectively inhibited protein tyrosine phosphorylation in Mm-A cells and its decrease of tyrosine phosphorylation was closely associated with its inhibition of cell growth. Thus, a genistein-sensitive tyrosine kinase(s) may play an important role in the growth and/or survival of leukemogenic Mm-A cells.
Animals↗
Inhibition of granulocytic differentiation of acute promyelocytic leukemia cells in primary culture by transforming growth factor-beta.
We investigated the effect of transforming growth factor-beta 1 (TGF beta) on the proliferation and differentiation of cultured acute promyelocytic leukemia (APL) cells with the chromosomal t(15;17) translocation obtained from four patients to determine the role of TGF beta on growth and differentiation of APL cells. DNA synthesis, determined by 3H-thymidine uptake, was inhibited in the presence and absence of granulocyte colony-stimulating factor (G-CSF) in a dose-dependent manner by TGF beta in APL cells obtained from three of the four cases. TGF beta and G-CSF did not significantly affect the differentiation of APL cells, but all-trans retinoic acid (RA) induced morphological and functional differentiation in all APL cells tested. G-CSF markedly enhanced RA-induced granulocytic differentiation in APL cells obtained from all four cases. In cells in which TGF beta inhibited DNA synthesis, it also inhibited RA-induced granulocytic differentiation of APL cells and, to a greater degree, granulocytic differentiation induced by RA plus G-CSF. These results suggest that TGF beta is a negative regulator of the proliferation and differentiation of APL cells. The significance of TGF beta as an endogenous regulator in differentiation therapy with RA of APL patients is discussed.
Adult↗
Differentiation of human myeloblastic leukemia ML-1 cells into macrophages by staurosporine, an inhibitor of protein kinase activities.
Protein kinase activities are involved in cellular proliferation and differentiation, and inhibitors of these activities are useful for studying the mechanisms of induction of differentiation. We found that staurosporine, an inhibitor of protein kinase activities, induced morphological differentiation of human myeloblastic leukemia ML-1 cells along myelomonocytic lineage and also induced functional differentiation (increase in nitroblue tetrazolium-reducing and lysozyme activities) in the cells. Several other protein kinase inhibitors such as 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H-7), sphingosine, N-(6-aminoethyl)-5-chloro-1-naphthalenesulfonamide and 1-(5-chloronaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (ML-9) did not induce the differentiation of ML-1 cells. Treatment with staurosporine induced formation of granules in ML-1 cells, and the granules showed metachromasia by toluidine blue staining; however, histamine content did not increase. The "metachromatic" ML-1 cells were positive for CD14, indicating that staurosporine induced the differentiation of ML-1 cells into metachromatic monocytes/macrophages, 1 alpha,25-dihydroxyvitamin D3 (VD3) enhanced appearance of metachromatic granules in staurosporine-treated cells. These results suggest that modulation of protein phosphorylation by a staurosporine-sensitive protein kinase(s) may be associated with differentiation of ML-1 leukemia cells.
Alkaloids↗
Inhibition of Abelson oncogene function by erbstatin analogues.
The authors examined the effect of a tyrosine kinase inhibitor, erbstatin, and its analogues on abl oncogene functions. Erbstatin and its stable analogue methyl 2,5-dihydroxycinnamate (2,5-MeC) inhibited the growth of v-ablts-NIH3T3 cells at the permissive temperature (33 degrees C) at lower concentrations than at the non-permissive temperature (39 degrees C). 2,5-MeC inhibited the morphological transformation and the activation of v-abl tyrosine kinase by the temperature shift (39 degrees C to 33 degrees C) more effectively than erbstatin. Previously the authors reported that erbstatin induced erythroid differentiation of K562 human chronic myelogenous leukaemia cells, so they examined the effect of erbstatin analogues on the erythroid differentiation. Among eight erbstatin analogues studied, ethyl 2,5-dihydroxycinnamate induced erythroid differentiation of K562 cells most effectively. Ethyl 2,5-dihydroxycinnamate also inhibited bcr-abl tyrosine kinase. These results indicate that the stable analogues of erbstatin suppress oncogene functions of Abl by inhibiting its tyrosine kinase.
Abelson murine leukemia virus↗
[Development of differentiation therapy].
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Antineoplastic Agents↗
Synthesis of active metabolite(s) from 1 alpha-hydroxyvitamin D3 by human monocytic leukemia cells.
Synthesis of the biologically active metabolite(s) from 1 alpha-hydroxyvitamin D3 (1 alpha(OH)D3) was examined in various types of human leukemia cell lines. Untreated monocytoid leukemia cells (U937 and HEL/S) metabolized 1 alpha (OH)D3 to the active metabolite(s), possibly 1 alpha, 24- and/or 1 alpha, 25-dihydroxyvitamin D3, and these cells were efficiently induced to differentiate by treatment with 1 alpha (OH)D3. However, the other types of leukemia cells did not efficiently metabolize it and were not induced to differentiate by 1 alpha (OH)D3. The possible therapeutic advantage of 1 alpha (OH)D3 in the treatment of monocytic leukemia is discussed.
Humans↗
Herbimycin A, an inhibitor of tyrosine kinase, prolongs survival of mice inoculated with myeloid leukemia C1 cells with high expression of v-abl tyrosine kinase.
Herbimycin A, a benzoquinonoid ansamycin antibiotic, reduces intracellular phosphorylation by some tyrosine kinases, including v-abl. The mouse megakaryoblastic cell line C1 expresses v-abl protein at high levels. Herbimycin A at about 20 ng/ml caused 50% inhibition of growth of C1 cells but at 100 ng/ml scarcely affected the growth of another mouse leukemia cell line, M1 cells, or of normal bone marrow cells. Injection of 10(6) C1 cells into nude mice resulted in death of all the mice within 30 days. Administration of herbimycin A significantly enhanced the survival of mice inoculated with C1 cells but scarcely affected the survival of mice inoculated with M1 cells. These results suggest that herbimycin A and/or related compounds may be useful for treatment of some types of leukemia in which tyrosine kinase activity is implicated as a determinant of the oncogenic state.
Animals↗
Identity of a differentiation inhibiting factor for mouse myeloid leukemia cells with NM23/nucleoside diphosphate kinase.
Mouse myeloid leukemic line M1 cells can be induced to differentiate into the monocyte/macrophage pathway by various inducers. The induction of differentiation of M1 cells can be inhibited by protein inhibitors termed differentiation inhibiting factors (I-factors) in a cell lysate and conditioned medium of differentiation resistant M1 cells. Production of the I-factor activity in resistant M1 cells is well associated with development of resistance of M1 cells to differentiation inducers. We have now purified one of the I-factors from conditioned medium of differentiation resistant M1 cells. The purified I-factor has a relative molecular mass of approximately 16000-17000 Da (16K I-factor). The amino acid sequence of all fragments of the 16K I-factor we have found are identical with Nm23/nucleoside diphosphate kinase (EC2.7.4.6) protein involved in tumor metastasis. The findings indicate that the I-factor, a candidate suppressor protein for differentiation of leukemic cells, is Nm23/nucleoside diphosphate kinase protein.
Amino Acid Sequence↗
Leukemia inhibitory factor/differentiation-stimulating factor (LIF/D-factor): regulation of its production and possible roles in bone metabolism.
Leukemia inhibitory factor/differentiation-stimulating factor (LIF/D-factor), expression of its mRNA, and possible roles in bone metabolism were studied in murine primary and clonal osteoblast-like cells. Local bone-resorbing factors such as IL-1, TNF alpha, and LPS strongly induced expression of LIF/D-factor mRNA in both clonal MC3T3-E1 cells and primary osteoblast-like cells. Neither parathyroid hormone nor 1 alpha,25-dihydroxyvitamin D3 stimulated expression of LIF/D-factor mRNA. LIF/D-factor per se did not stimulate expression of its own mRNA. Appreciable amounts of LIF/D-factor were detected in synovial fluids from rheumatoid arthritis (RA) patients but not in those with osteoarthritis (OA). Simultaneous treatment with LIF/D-factor, IL-1, and IL-6 at the concentrations found in synovial fluids from RA patients greatly enhanced bone resorption, though these cytokines did not stimulate bone resorption when separately applied. This suggests that LIF/D-factor produced by osteoblasts is in concert with other bone-resorbing cytokines such as IL-1 and IL-6 involved in the bone resorption seen in the joints of RA patients. LIF/D-factor specifically bound to MC3T3-E1 cells with an apparent dissociation constant of 161 pM and 1,100 binding sites/cell. LIF/D-factor dose-dependently suppressed incorporation of [3H]thymidine into MC3T3-E1 cells. In addition, it potentiated the alkaline phosphatase activity induced by retinoic acid, though LIF/D-factor alone had no effect on enzyme activity. These results suggest that LIF/D-factor is involved in not only osteoclastic bone resorption but also osteoblast differentiation in conjugation with other osteotropic factors.
Acid Phosphatase↗
Prolongation by differentiation-stimulating factor/leukemia inhibitory factor of the survival time of mice implanted with mouse myeloid leukemia cells.
Mouse myeloid leukemic M1 cells can be induced to differentiate into macrophages by differentiation-stimulating factor (D-factor)/leukemia inhibitory factor (LIF). We examined the effect of D-factor on the survival times of syngeneic mice implanted with two different clones (T-22 and R-4) of M1 cells. D-factor induced differentiation and suppressed DNA synthesis of sensitive T-22 cells but not resistant R-4 cells in vitro. For in vivo experiments, we used recombinant mouse D-factor (rmD-factor) produced in mammalian cells, which is glycosylated and is more stable in vitro and in vivo than unglycosylated rmD-factor produced in Escherichia coli. Treatment with rmD-factor prolonged the survival times of mice implanted with T-22 cells but not R-4 cells.
Animals↗