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1.
To study the influence of a biologic environment on cultured human leukemia cells, KG-1, KG-1a, and HL-60 cells were inoculated subcutaneously into newborn nude mice. The cells developed myelosarcomas at the site of inoculation and in lungs and kidneys. KG-1 and HL-60 myelosarcomas were successfully passaged through adult nude mice, whereas KG-1a tumors proliferated only after transplantation into newborn hosts. The human nature of the cells forming myelosarcomas in mice was assessed by chromosomal analyses and detection of cross- reactivity with an antibody to the human leukemia cell line K562. We undertook electron microscopic and cytochemical examinations of the cells proliferating in vitro and in the mice. The granules of KG-1 cells in vivo did not react for acid phosphatase, as observed in vitro, and the HL-60 cells proliferating in mice lost the perinuclear myeloperoxidase (MPO) demonstrated in cultured cells. Although the influence of an in vivo selection of cell subpopulations cannot be ruled out, the enzymatic changes are compatible with induced cell differentiation. Conclusive evidence of differentiation in vivo was observed in the KG-1a cell subline. The undifferentiated KG-1a blasts developed cytoplasmic granules and synthesized MPO during proliferation in vivo. These observations indicate that human leukemia cells from established cell lines proliferate in nude mice and may acquire new differentiated properties in response to the in vivo environment.  相似文献   

2.
Human myeloid leukemia cell lines: a review   总被引:20,自引:7,他引:20  
Koeffler  HP; Golde  DW 《Blood》1980,56(3):344-350
Several human acute myeloid leukemia cell lines were recently established. These lines provide model systems to study the control of differentiation in human myelogenous leukemia and, in a broader framework, the controls of normal myeloid development. The K562 line is composed of undifferentiated blast cells that are rich in glycophorin and may be induced to produce fetal and embryonic hemoglobin in the presence of hemin. The KG-1 cell line is composed predominantly of myeloblasts and promyelocytes. A unique characteristic of the KG-1 cells is their almost complete dependence on colony-stimulating factor for proliferation in soft-gel culture. The HL-60 is a promyelocytic leukemia cell line. In the presence of DMSO, the cells mature into granulocytes. Both the KG-1 and HL-60 cells differentiate into nondividing mononuclear phagocytes when exposed to phorbol esters. Investigations with these cell lines, and selected variants should provide important insights into the cell biology and perhaps therapy of human leukemia.  相似文献   

3.
The regulation of IgG Fc receptor (Fc gamma R) expression by retinoic acid (RA) in human myelomonocytic cells at different stages of maturation was studied. RA suppressed IgG-coated erythrocyte (EA) rosette formation of myelomonocytic cells blocked at relatively late stages of differentiation such as ML-1, U-937, THP-1-T, normal monocytes, and fresh cells of patients with acute myelomonocytic leukemia. However, RA increased the percentage of EA rosetting promyelocytes of HL-60 and of patients with acute promyelocytic leukemia and a part of myeloblasts isolated from acute myelogenous leukemia patients. Other myeloblasts including KG-1a, KG-1, and fresh cells from patients with acute myelogenous leukemia were not affected. A kinetic study using HL-60 and THP-1-T demonstrated that an increase required at least a 48-h exposure and that the maximum decrease required approximately 6 h. The RA effect on both cell lines was dose-dependent. The number of Fc gamma R of HL-60 and THP-1-T treated with RA became very close, although untreated THP-1 had almost 10 times as many as HL-60. Kd for IgG in both THP-1-T and HL-60, either untreated or treated with RA, remained unchanged. These observations indicate that one of the important roles of RA is regulation of Fc gamma R expression in myeloid cells.  相似文献   

4.
Amatruda  TT d; Bohman  R; Ranyard  J; Koeffler  HP 《Blood》1987,69(4):1225-1236
We examined the expression of HLA-DR and HLA-DQ antigens and mRNA from myeloid and lymphoid cells obtained from normal volunteers and established cell lines. Cytofluorometric analysis and immunoprecipitation were performed using murine monoclonal antibodies specific for HLA-DR (L-243) and HLA-DQ (Leu 10). The expression of mRNA for HLA-DR and HLA-DQ chains was determined by Northern blot and RNA dot-blot analysis. Lymphoid cell lines expressed both HLA-DR and HLA-DQ antigens, with consistently higher levels of expression of DR. Myeloid cell lines of early myeloblast or bipotent (myeloid-erythroid) phenotype (KG-1, KG-1a, HEL) expressed HLA-DR at high levels, whereas cell lines manifesting a greater degree of myeloid maturation (ML-3, HL- 60, U937) expressed DR at low or undetectable levels. The HLA-DQ antigen was expressed at low levels on the surface of KG-1 and KG-1a cells but was not detectable on other myeloid cell lines. The expression of mRNA for HLA-DR and HLA-DQ chains paralleled the pattern of expression of the respective antigens. The HL-60 and U-937 cells stimulated to differentiate in vitro to macrophages with 1,25 dihydroxyvitamin D3 [1,25(OH)2D3] were induced to express detectable levels of HLA-DR antigens. Exposure to gamma-interferon (gamma-IFN) increased the expression of HLA-DR antigens by all myeloid cell lines. Induction of differentiation in vitro with either 1,25(OH)2D3 or dimethyl sulfoxide potentiated this effect of gamma-IFN. Expression of the HLA-DQ antigens was increased on KG-1 myeloblasts after exposure to gamma-IFN. HLA-DQ expression could not be detected on other myeloid cell lines after exposure to gamma-IFN, nor was HLA-DQ expression stimulated by gamma-IFN after HL-60 and U-937 cells were induced to differentiate to macrophagelike cells in vitro. These results provide additional evidence that expression of the HLA-DR and HLA-DQ genes may be independently regulated in human myeloid cells.  相似文献   

5.
A series of monoclonal antibodies recognizing myeloid differentiation antigens were prepared by immunizing Balb/c mice with HL-60 cells. Hybrids secreting antibodies reactive with HL-60 cells but unreactive with peripheral blood mononuclear cells were isolated and further cloned. One clone was found to produce an IgG2a antibody recognizing an 85,000-dalton molecular weight surface glycoprotein, and a second clone was found to produce an IgM antibody recognizing a heat-stable determinant present on a glycolipid. We have termed these antigens Pro- Im1 and Pro-Im2, respectively (Pro for using HL-60 promyelocytes as an immunogen and Im for the presence of these antigens on immature cells). alpha Pro-Im1 and alpha Pro-Im2 were used to investigate the surface expression and tissue distribution of these two antigens. Pro-Im1 and Pro-Im2 were found to be brightly expressed on a fraction of fetal liver hematopoietic and bone marrow cells. Both antibodies mediated complement-dependent inhibition of CFU-GM, BFU-E, and CFU-GEMM formation assayed by soft agar colony and burst formation, indicating the expression of these antigens by early hematopoietic precursor cells. This was further confirmed by the induction of HL-60 cells by TPA to differentiate into more mature monocytes and macrophages, accompanied by the loss of both antigens. Pro-Im1 and Pro-Im2 were absent from peripheral blood monocytes, erythrocytes, and platelets, but Pro-Im2 was expressed on granulocytes. Both antigens were absent from thymocytes and peripheral T cells. Cytofluorographic analysis suggested their absence from peripheral blood B cells but that both were expressed on a minority of tissue B cells. Analysis of 150 cases of various myeloid and lymphoid malignancies demonstrated Pro-Im1 and Pro-Im2 expression on myeloblasts and promyelocytes from some acute myelogenous leukemias as well as some B cell malignancies, suggesting that these antigens are shared by early hematopoietic cells and a subset of B cells.  相似文献   

6.
Direct differentiation of myeloid leukemia blasts into antigen-presenting dendritic cells (DCs) for use as cellular vaccines is unique in that identification of tumor-specific antigens may not be necessary because the antigens should already be endogenously expressed. We hypothesized that signaling through protein kinase C (PKC) is required for differentiation of HL-60 promyeloblasts into DCs upon stimulation with calcium ionophore A23187. To demonstrate the inhibitory effect of PKC blockade, we pretreated HL-60 myeloid blasts with the protein kinase inhibitor bisindolylmaleimide I (Bis-1) for 24 hours and then treated the cells with calcium ionophore A23187 for an additional 24 hours. Controls consisted of HL-60 blasts treated with A23187, Bis-1 alone, or media. We noted that blasts cultured in media, Bis-1, or Bis-1 then A23187 did not develop the morphologic and phenotypic DC characteristics, up-regulate Rel B, or activate allogeneic T-cells. Our findings suggested that PKC blockade inhibits morphologic, phenotypic, and functional differentiation of HL-60 promyeloblasts into antigen-presenting DCs. Our findings supported the role of PKC as an obligatory pathway for calcium ionophore A23187-induced differentiation of HL-60 myeloblasts into antigen-presenting DCs.  相似文献   

7.
Nauseef  WM 《Blood》1986,67(4):865-872
The biosynthesis and processing of myeloperoxidase (MPO), a cationic enzyme present in the azurophilic granules of human polymorphonuclear leukocytes (PMNs), were studied in the human promyelocytic leukemia cell line, HL-60. HL-60 cells produce large quantities of enzymatically active MPO that has the same electrophoretic behavior as MPO isolated from normal PMNs. Mature MPO is a glycoprotein of approximately 150,000 molecular weight (mol wt) composed of two heavy-light protomers (alpha 2 beta 2) with subunits of 59,000 and 13,500 mol wt, respectively, under reducing conditions. The primary translation product of MPO messenger RNA (mRNA) isolated from HL-60 cells was a single polypeptide of mol wt 80,000. In HL-60 cells labeled with [35S]-methionine, the labeled MPO isolated by immunoprecipitation had a mol wt of 89,000. Treatment of this 89-kilodalton (kDa) species with endoglycosidase H produced a 79-kDa peptide, suggesting that the 89-kDa protein contained high-mannose side chains. The 89-kDa species had no detectable peroxidase activity. During chase experiments some of the 89-kDa peptide was processed to smaller species of mol wt 39,000, 59,000, and 13,500, although a fraction of the 89-kDa peptide remained unprocessed after a chase of 100 hours. In addition, a small amount of the 89-kDa peptide appeared in the medium without any of the processed smaller peptides. These studies suggest that the primary translation product in MPO biosynthesis is an 80-kDa peptide that undergoes cotranslational cleavage of the signal peptide and glycosylation to produce an 89-kDa pro-MPO, that pro-MPO is a single polypeptide containing the alpha and beta subunits of MPO and contains endoglycosidase H-susceptible high- mannose side chains, and that posttranslational modification of pro-MPO results in targeting to the lysosome and proteolytic maturation of pro- MPO to active enzyme. In light of the previous observation that MPO- deficient and normal PMNs contain an 89-kDa protein immunochemically related to MPO, these studies on MPO biosynthesis indirectly support the hypothesis that defective posttranslation processing by pro-MPO may underlie hereditary MPO deficiency.  相似文献   

8.
The human leukemic cell lines, K562, KG-1, and HL-60, and the blast subclones, KG-1a and HL-60 blast, were utilized to relate differences in nonhistone protein antigens to stages of myeloid cell differentiation. Chromatin proteins were separated on SDS- polyacrylamide gels, transferred electrophoretically to nitrocellulose sheets, and visualized by the peroxidase-antiperoxidase method of Sternberger. Screening with antisera raised against total and dehistonized chromatin and a nuclear extract from these cells revealed quantitative as well as qualitative differences between the cell lines. A decrease in antigen content seemed to parallel progressive stages of myeloid cell development. The results indicate that a number of chromosomal protein antigens are lost or modified during differentiation. An antigen(s) of approximately 55,000 molecular weight was found in HL-60 chromatin, but was not present in its less differentiated subclone or in the other lines representative of earlier stage cells. Upon the induction of HL-60 cells to mature to end stages with 4 microM retinoic acid, a significant increase in the mol wt 55,000 activity was seen. This antigen was detected only with antisera against HL-60 total chromatin and granulocyte nuclei, and it was found only in normal mature granulocytes and in the later stage cells of the HL-60 culture. Thus, the antigen appears to be associated with a differentiated myeloid function.  相似文献   

9.
Expression of c-jun during macrophage differentiation of HL-60 cells   总被引:2,自引:0,他引:2  
R Gaynor  K Simon  P Koeffler 《Blood》1991,77(12):2618-2623
  相似文献   

10.
An antigen histochemically localized in the nuclei and cytoplasmic granules of normal and leukemic human myeloid cells has been identified as myeloperoxidase (MPO; EC 1.11.1.7). The localization and amount of the enzyme was determined by using a murine monoclonal antibody designated H-43-5 raised against nuclear proteins derived from human promyelocytic HL-60 leukemia cells. The highest amount of nuclear MPO (3.5 micrograms per 10(6) nuclei) was found in granulocytes; less than half of this amount was detected in nuclei from HL-60 cells. Still lower levels were found in nuclei from monocytes and a series of human monomyelocytic leukemia cells. MPO from HL-60 cells was purified by immunoaffinity chromatography and fractionated into three components (forms I, II, and III) by CM-cellulose chromatography. Chromatography of these MPO forms on DNA-Sepharose columns confirmed that all three forms of MPO were tightly bound to DNA with apparent relative affinities in the order of form III greater than form II greater than form I. The affinity of MPO form III for DNA was sufficient to enable the formation and elution of DNA-MPO complexes during size-exclusion chromatography at high ionic strength and neutral pH. This form of MPO was also able to shield DNA from strand scission induced by active oxygen species generated by xanthine oxidase acting aerobically on xanthine. These data suggest that intranuclear MPO may help to protect DNA against damage resulting from oxygen radicals produced during myeloid cell maturation and function.  相似文献   

11.
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13.
We analyzed platelet-associated antigens from a hemophilia B patient with human immunodeficiency virus type 1 (HIV-1)-related thrombocytopenia. Two bands appeared at 31,000 and 37,000 daltons in the platelet lysate after reaction with autologous serum in SDS-PAGE and Western blots. The band at 37,000 daltons was obtained using anti-herpes simplex type 1 (HSV-1) rabbit antiserum. Doublet bands at 36,000 and 37,000 daltons also appeared after reaction with HSV-1 seropositive human serum. The band at 31,000 daltons appeared after reaction with anti-HIV-1 rabbit serum. These results suggest that the platelet-associated antigens in this patient are components of both HSV-1 and HIV-1 antigens. In addition, acyclovir decreased his PAIgG level and increased his platelet count, and zidovudine increased his platelet count. Thus, we concluded that each of the platelet-associated antigens is partially responsible for the thrombocytopenia by causing deposition of immune complexes in this patient.  相似文献   

14.
The 80,000-dalton form of the adenovirus (Ad) terminal protein (pTP) has been purified from Ad-infected HeLa cells. pTP was assayed by its ability to form a covalent complex with dCMP. The protein copurified with an activity that is essential for in vitro Ad DNA replication (Ad protein activity) as well as with a DNA polymerase activity that was distinguished from those of HeLa cell DNA polymerases alpha, beta, and gamma. The Ad protein-associated DNA polymerase activity was detected with activated DNA but not with poly(rA).oligo(dT) as template and was insensitive to aphidicolin and sensitive to N-ethylmaleimide. The Ad protein, DNA polymerase, and pTP-dCMP complex-forming activities sedimented in a glycerol gradient as a single peak with an apparent molecular size of 180,000 daltons. NaDodSO4/polyacrylamide gel analysis of the glycerol gradient fraction showed major bands of 80,000 and 140,000 daltons. The 80,000-dalton band was identified as pTP by comparison of its tryptic peptide map with that of the 55,000-dalton form of the terminal protein, which was purified from Ad virions.  相似文献   

15.
Two monoclonal antibodies (McAb) termed 12A4 and 19C1 have been raised against human factor VIII. In immunoassays 12A4 bound to factor VIII antigen (VIII:Ag) in plasma but not serum whilst 19C1 bound to VIII:Ag in both plasma and serum. Both McAb were shown by immunoblotting to react with the carboxy (C) terminal polypeptide of factor VIII which appeared as a doublet with a molecular weight (Mr) of 77,000/75,000. The C terminal factor VIII polypeptide was detectable by immunoblotting in each of 12 therapeutic factor VIII concentrates, from six different manufacturers, although its level was variable. Factor VIII was visualized in plasma by immunoblotting following its immunoadsorption and elution from agarose-bound monoclonal antibodies. No Mr 77,000/75,000 bands were detectable in plasma obtained from 13 unrelated CRM- haemophiliacs whilst 11 CRM+ haemophilic plasmas from seven kindred were shown to have a C terminal factor VIII polypeptide of normal molecular size.  相似文献   

16.
Myeloperoxidase (MPO) cytochemical activity, recognized as a very important hallmark of myeloblasts, is generally negative in chronic myeloid leukemia (CML) blast crisis (BC). Because this finding is unexpected, being not in keeping with the myeloproliferative nature of CML, we tried to ascertain if MPO cytochemical negativity could be an intrinsic property of blasts of CML and hence present in the preblastic phases as well. Myeloperoxidase cytochemistry of peripheral blood blasts in 161 cases of CML, including 103 in chronic phase (CP) and 29 each in accelerated phase (AP) and BC, was assessed and compared with that of 30 cases of acute myeloid leukemia, AML-M2. Blasts of 97 (94.2%) of 103 cases of CP, 28 (96.6%) of 29 cases of AP, and 22 (75.9%) of 29 cases of BC were negative for MPO (<3% MPO-positive blasts). Compared with the strong MPO positivity, both in terms of intensity and proportion, in the AML-M2 cases, the positivity in the CML cases was generally weak and was seen in a small number of blasts (5–15%), except in one case of BC with 20% positive blasts. Absence or, at times, weak MPO cytochemical activity is an intrinsic property of blasts of all phases of CML, and use of the term myeloblast in CML should be understood to refer to a cell with this property. This also explains why MPO cytochemistry, despite its high reputation as a myeloid-lineage marker, generally does not help in CML BC. CML BC should therefore be considered as a possible diagnosis along with acute lymphoblastic leukemia, AML-M0, AML-M7, etc., in the setting of MPO-negative blasts. Similarity between MPO expression pattern in CML, i.e., negative in blasts and positive in the more mature cells, and that during maturation of normal myeloid series of cells shows the deranged myelopoiesis of CML to be undisturbed at least with respect to MPO expression. There is need for a more comprehensive study of blasts of preblastic phases.  相似文献   

17.
Chloroform:methanol (2:1) extracts of bovine surfactant were subjected to LH-20 Sephadex chromatography in order to isolate a 6,000-dalton surfactant protein. The 6,000-dalton protein eluted in the void volume and was shown to be homogeneous by protein sequencing, although SDS gel electrophoresis revealed bands of 6, 14, and 18 kDa. The N-terminal sequence obtained was very hydrophobic, as was the amino acid composition of the 6,000-dalton protein. An antiserum raised against the low molecular weight protein fraction from TA surfactant recognized the 6,000-dalton bovine and human proteins in addition to protein bands at 14,000 and 18,000 daltons. These bands appear to be aggregates of the 6,000-dalton protein. No cross-reactivity of the 6,000-dalton protein antiserum could be demonstrated with the 35,000-dalton surfactant-associated protein. These studies strongly suggest that the 35,000- and 6,000-dalton surfactant proteins do not have a precursor-product relationship.  相似文献   

18.
Apolipoprotein (apo) B-100 and B-48 are prominent apolipoproteins in VLDL, IDL, and chylomicrons. Organ cultures of normal adult human liver were established to ascertain the form of apo B synthesized by hepatocytes in humans. Human liver was minced and incubated in 15 mL methionine-free RPMI-1640 medium with 10% dialyzed fetal calf serum plus 250 microCi 35S-methionine for eight hours at 37 degrees C. Lipoproteins secreted by the liver were isolated by ultracentrifugation and the content of newly synthesized apo B determined by quantitation of radioactivity in the apoB-100 and apoB-48 bands after separation by 3% NaDodSO4 gel electrophoresis. In the eight-hour period, 2.5% to 3.2% of added 35S-methionine was secreted in TCA-precipitable protein of which 0.34% was apo B. Ninety-nine percent of the apo B in VLDL, IDL, and LDL was in the apo B-100 electrophoretic band. No significant radioactivity was detected in the apo B-48 electrophoretic band. Eighty-nine percent of the total radioactivity of apo B-100 was in VLDL with 3% and 8% in IDL and LDL, respectively. These results establish that adult human liver in organ culture synthesizes apo B-100 but not apo B-48.  相似文献   

19.
The human promyelocytic leukemia cell line HL-60 is reactive with an antiserum raised against normal human granulocytes (AGS). Immunoprecipitation with AGS on [35S]methionine-labeled HL-60 cell lysates with subsequent analysis by NaDodSO4/polyacrylamide slab gel electrophoresis shows a major antigenic doublet with molecular weights of 88,000 and 86,000, together with some minor antigens of lower molecular weight. Upon stimulation with dimethyl sulfoxide or 12-O-tetradecanoylphorbol 13-acetate, which induces HL-60 to differentiate to mature granulocytes or monocytes/macrophages, respectively, this antigenic doublet disappears. 12-O-Tetradecanoylphorbol 13-acetate induces the synthesis of an antigen, molecular weight 83,000, reactive with an antimonocyte serum. Neutrophil-specific alloantigens were not detected on HL-60 or its differentiated derivatives.  相似文献   

20.
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