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H G Drexler

Publications and source records attributed to H G Drexler.

At least 91 records · Page 5Linked to original sources
PubMed1997

A model system in haematology and immunology: the human monocytic cell line MONO-MAC-1.

MONO-MAC-1 is a human cell line with properties of blood monocytes, which can be used as a model system to study monocytic functions in vitro. In the present study, we prepared a karyotype of MONO-MAC-1, analysed the growth behaviour, determined the presence of differentiation-associated antigens and studied the expression and secretion of several cytokines upon stimulation with 12-O-tetradecanoyl phorbol 13-acetate (TPA) and lipopolysaccharide (LPS). The MONO-MAC-1 cells have a near diploid karyotype and contain several recurrent chromosomal rearrangements, in particular the translocation (9;11) commonly found in AML-M5. Stimulation with TPA or LPS induced changes in morphology and gene expression, especially an increase in the level of the differentiation marker CD14 and the production of monocyte-related cytokines. Both biomodulators alone were sufficient to promote TNF alpha release; however, the combination of TPA and LPS resulted in a synergistic increase of TNF alpha secretion. Northern blot analysis indicated that upregulated production of TNF alpha was due to induced synthesis of mRNA. The mRNA accumulation peaked approximately 2 h after stimulation and maximum levels of TNF alpha were found in the supernatants after 4-8 h of culture. The MONO-MAC-1 cells could not be restimulated with the same inducer to release TNF alpha when a 48 h pre-treatment was carried out with LPS or TPA. LPS induced the release of granulocyte colony-stimulating factor (G-CSF), while TPA failed to do so. Vice versa, secretion of macrophage CSF (M-CSF) could be induced by TPA, but not by LPS. However, LPS enhanced the TPA-induced M-CSF production. Similarly, incubation of MONO-MAC-1, simultaneously with TPA and LPS, led to granulocyte macrophage CSF (GM-CSF) and interleukin-1beta (IL-1beta)secretion, while both stimulators alone had almost no (TPA) or only a weak (LPS) effect on the secretion of GM-CSF and IL-1beta. Our results demonstrate that MONO-MAC-1 is a unique cell line with distinct monocytic features; certain monocytic properties can be upregulated by activation of intracellular signalling pathway(s). We suggest that, besides the LPS receptor CD14, activation of PKC participates in these process, especially in the production and secretion of cytokines by MONO-MAC-1 cells.

Cell Line↗
PubMed1997

The human bladder carcinoma cell line 5637 constitutively secretes functional cytokines.

We have studied cytokine secretion by the human bladder carcinoma cell line 5637 using growth factor-dependent cell line bioassays and specific ELISA. The 5637 cell line-conditioned medium (5637 CM) stimulated proliferation of human growth factor-dependent leukemia cell lines in a dose-dependent fashion. Quantitation by specific ELISA and by bioassays detected high amounts of G-CSF and GM-CSF and smaller quantities of IL-1beta, M-CSF and SCF in 5637 CM; the concentration of IL-3 was below the detection level of the ELISA, if present at all. The G-CSF and GM-CSF activities secreted by 5637 cells could be inhibited specifically by neutralizing anti-G-CSF and anti-GM-CSF antibodies. In conclusion, 5637 bladder carcinoma cells constitutively produce and secrete several functionally active cytokines; 5637 CM is a valuable, reliable and inexpensive source for cytokines, for instance for the culture of growth factor-responsive or -dependent hematopoietic primary cells and cell lines.

Carcinoma↗
PubMed1997

Expression of tie receptor tyrosine kinase in human leukemia cell lines.

The tie gene encodes a receptor tyrosine kinase that together with its thus far unidentified ligand appears to play a distinct role in the regulatory pathway of early hematopoiesis and angiogenesis. Here, we attempted to define the possible involvement of tie in the pathobiology of hematopoietic malignancies by examining tie mRNA expression in human leukemia and lymphoma cells. We used a large panel of 93 well-characterized human continuous leukemia-lymphoma cell lines as model systems for the various hematopoietic cell lineages. At the Northern blot level, none of the 27 lymphoid leukemia or lymphoma-derived cell lines (originating from four B-precursor leukemia, four B-cell leukemia, four B-cell non-Hodgkin's lymphoma, two myeloma, two Burkitt lymphoma, four T-cell leukemia, five Hodgkin lymphoma, two anaplastic large cell lymphoma) tested expressed tie transcripts, whereas 23/42 (55%) of the myeloid cell lines analyzed expressed tie mRNA: in detail, 15 of 20 (75%) megakaryocytic, five of 11 (45%) erythroid, three of seven (43%) myelocytic and none of four monocytic cell lines were tie mRNA positive. In the reverse transcriptase-polymerase chain reaction analysis, which can detect very low levels of mRNA expression, all 12 myeloid cell lines and 19 of 39 (48%) lymphoid cell lines were positive. In experiments aimed at inducing cellular differentiation over an incubation period of 4 days, the phorbol ester PMA strongly enhanced tie mRNA expression in one erythroid and in one myelocytic cell line, but (like thrombopoietin) down-regulated tie mRNA expression in two megakaryocytic cell lines. Taken together these results indicate that tie is predominantly expressed in leukemia cells derived from the myeloid cell lineages (and here in particular in megakaryoblastic cells) and not in lymphoid leukemia cells. These observations provide some evidence for the hypothesis that tie is a receptor for a regulatory factor involved in normal and plausibly also leukemic hematopoiesis.

Cell Line↗
PubMed1997

Occurrence of TEL-AML1 fusion resulting from (12;21) translocation in human early B-lineage leukemia cell lines.

The recurrent (12;21)(p13;q22) translocation fuses the two genes TEL and AML1 that have previously been cloned from translocation breakpoints in myeloid leukemias. Using mainly reverse transcriptase-polymerase chain reaction (RT-PCR), the TEL-AML1 chimeric transcript has been observed in 22-27% of pediatric patients with acute lymphoblastic leukemia (ALL), in particular in the early B-lineage ALL subtype, making it the most common genetic lesion in these patients. The vast majority of acute myeloid leukemias, other ALL subtypes and even adults with early B-lineage ALL were TEL-AML1-negative. We determined whether the TEL-AML1 fusion gene can also be observed in continuous human leukemia cell lines with an early B-lineage phenotype. Twenty-nine such cell lines established from children (n = 13) or adults (n = 13) with early B-lineage ALL and five cell lines derived from chronic myeloid leukemia in blast crisis or B cell non-Hodgkin's lymphoma were investigated for the occurrence of the TEL-AML1 rearrangement by RT-PCR. While all 13 adult early B-lineage ALL cell lines and the five cell lines from other leukemias or lymphomas were negative, 1/13 pediatric cell lines (cell line REH) was found to be positive for TEL-AML1; though neither reciprocal AML1-TEL, nor normal TEL, mRNA was detectable by RT-PCR in this cell line. These findings agreed with the results of conventional cytogenetic and FISH analysis of REH which was found to carry the der(21) partner only of t(12;21)(p13;q22), probably resulting from a complex translocation, t(4;12;21;16)(q32;p13;q22;q24.3). Hybridization with flanking cosmid clones (179A6 and 148B6), covering exons 1 and 8 respectively of TEL, confirmed a rearrangement accompanying the t(12;21), and showed cryptic deletion of the residual allele resulting from an apparently reciprocal t(5;12)(q31;p13). These findings in REH provide a further example of, and possible cytogenetic mechanism for, the paradigm of TEL-AML1 fusion accompanied by deletion of the residual TEL allele. The low rate of early B-lineage ALL cell lines carrying this translocation contrasts clearly with the relative high frequency of TEL-AML1-positive cases in primary material. It is possible that expression of the fusion product hampers the in vitro growth and establishment in culture of such leukemic cells. Nevertheless, the cell line REH represents a powerful tool for the further molecular characterization of this unique breakpoint and can serve as a positive control in routine PCR reactions.

Adolescent↗
PubMed1997

Thrombopoietin supports the continuous growth of cytokine-dependent human leukemia cell lines.

Hematopoiesis is a complex process of regulated cellular proliferation and differentiation from the primitive stem cells to the final fully differentiated cell. The long and extensive search for a factor specifically regulating megakaryocytopoiesis led to the cloning of a hormone, here called thrombopoietin (TPO), that specifically promotes proliferation and differentiation of the megakaryocytic lineage. The availability of recombinant TPO and its imminent clinical use has made a more detailed understanding of its effects on hematopoietic cells more urgent. Normal megakaryocyto- and thrombopoiesis occurs predominantly in the bone marrow, a difficult organ to study in situ, particularly in humans, due to the low numbers of megakaryocytic progenitors and the consequent difficult isolation as pure populations. Thus, we developed an in vitro system which may allow us to address questions regarding the biology of TPO. The acute myeloid leukemia (AML)-derived cell lines HU-3, M-07e, M-MOK and TF-1 have absolute dependence on granulocyte-macrophage colony-stimulating factor (GM-CSF). We cultured these cells long term (> 6 months) in the continuous presence of TPO (omitting GM-CSF). TPO alone supported the maintenance and expansion of these sister cell lines, HU-3/TPO, M-07e/TPO, M-MOK/TPO and TF-1/TPO, that displayed somewhat longer doubling times, a larger cell size, and a higher percentage of polynucleated giant cells and slightly adherent cells than the corresponding countercultures grown with GM-CSF. In the absence of TPO the cells died quickly, within a few days; thus, the TPO-grown cell lines have an absolute dependence on this factor, but could all be switched back to growth with GM-CSF. In comparison with the GM-CSF-treated cells, the receptors for GM-CSF and interleukin-3 (IL-3) were down-regulated and the receptors for stem cell factor (SCF) and TPO were up-regulated in the TPO-exposed cells. A short-term proliferation assay showed a stronger response of the TPO-cell lines to erythropoietin, GM-CSF, IL-3, PIXY-321, SCF and TPO than the GM-CSF-cell lines. Flow cytometric analysis of the GM-CSF-and TPO-cultured lines displayed an up-regulation of the megakaryocytic surface markers CD41, CD42 and CD61, and a down-regulation of the erythroid marker glycophorin A in the latter cell lines, suggesting some differentiation along the megakaryocytic lineage. Thus, in long-term exposure, TPO appears to have both a proliferative and a differentiative effect on responsive cells. Under serum-deprived culture conditions, TPO acted as a survival factor on the TPO-cell lines. Taken together, these findings indicate that the TPO-dependent cell lines represent important biological reagents for further characterization of the biology of TPO and should also provide a great aid for future in vitro experiments aimed at elucidating megakaryocyto- and thrombopoiesis.

Azure Stains↗
PubMed1997

Cytokine response profiles of human myeloid factor-dependent leukemia cell lines.

Research in cytokine biology has grown exponentially in recent years as cytokines (often also termed growth factors) are now known to be involved in a wide range of pathological and physiological processes. Continuous human leukemia cell lines represent powerful tools to investigate these mechanisms. Most cell lines grow autonomously in standard culture media (containing fetal bovine serum) independent of externally added growth stimuli. Over the last 5-10 years a battery of myeloid leukemia-derived cell lines has been established that is constitutively dependent on the addition of cytokines to the culture. Such factor-dependent cell lines die rapidly by apoptosis when deprived of the appropriate growth factor. We determined the cytokine response profiles of 19 absolutely growth factor-dependent leukemia cell lines with myelomonocytic, erythroid or megakaryocytic phenotypes with regard to enhanced or suppressed cellular proliferation. Cells were incubated in liquid culture with optimal concentrations of various recombinant human cytokines known to have effects on the growth of hematopoietic cells. A proliferative or anti-proliferative response to these 41 cytokines was assessed by the short-term 3H-thymidine uptake assay. A proliferative response was considered as positive when the stimulation index (SI) was >2; inhibition was regarded as significant with an SI <0.5. The response profile of each cell line to these 41 cytokines was different and individual. None of the cell lines responded to one or two factors only (minimum to at least five cytokines). Proliferation of most (n = 13-17), but not of all cell lines was significantly enhanced by GM-CSF, IL-3, PIXY-321, SCF and IFN-gamma. TGF-beta1 consistently inhibited proliferation (in 11/19 cell lines). IFN-alpha, IFN-beta, TNF-alpha and TNF-beta had either stimulatory or inhibitory effects. The cell lines responding most often proliferatively (to 15-19 different cytokines) were UCSD/AML1, HU-3, TF-1 and M-07e. In summary, these factor-responsive human leukemia cell lines represent extremely useful model systems for the analysis of cytokine effects on hematopoietic cells. The cytokine response profiles of the individual cell lines provide guidelines for the selection of the appropriate cell culture for such experiments.

Animals↗
PubMed1997

Two acute monocytic leukemia (AML-M5a) cell lines (MOLM-13 and MOLM-14) with interclonal phenotypic heterogeneity showing MLL-AF9 fusion resulting from an occult chromosome insertion, ins(11;9)(q23;p22p23).

We describe two new human leukemia cell lines, MOLM-13 and MOLM-14, established from the peripheral blood of a patient at relapse of acute monocytic leukemia, FAB M5a, which had evolved from myelodysplastic syndrome (MDS). Both cell lines express monocyte-specific esterase (MSE) and MLL-AF9 fusion mRNA. Gene fusion is associated with a minute chromosomal insertion, ins(11;9)(q23;p22p23). MOLM-13 and MOLM-14 are the first cell lines with, and represent the third reported case of, MLL gene rearrangement arising via chromosomal insertion. Both cell lines carry trisomy 8 which was also present during the MDS phase, as well as the most frequent trisomies associated with t(9;11), ie, +6, +13, +19 variously present in different subclones. Despite having these features in common, differences in antigen expression were noted between the two cell lines: that of MOLM-13 being CD34+, CD13-, CD14-, CD15+, CD33+; whereas MOLM-14 was CD4+, CD13+, CD14+, CD15+, CD33+. Differentiation to macrophage-like morphology could be induced in both cell lines after stimulation with INF-gamma alone, or in combination with TNF-alpha, which treatment also induced or upregulated, expression of certain myelomonocyte-associated antigens, including CD13, CD14, CD15, CD64, CD65 and CD87. Together, these data confirm that both cell lines are likely to be novel in vitro models for studying monocytic differentiation and leukemogenesis.

Adult↗
PubMed1997

A novel ALL-L3 cell line, BALM-16, lacking expression of immunoglobulin chains derived from a patient with hypercalcemia.

A human acute lymphoblastic leukemia (ALL) cell line, BALM-16, was established from the peripheral blood specimen of a patient with B cell ALL L3 type (ALL-L3) in relapse. As with the original leukemia cells, the established line was negative for both cell surface and cytoplasmic immunoglobulin (Ig) chains. Absence of Ig expression was confirmed by Western blotting. Southern blot analysis demonstrated homozygous deletion of the C kappa gene, germ line configuration of the C lambda and rearrangement of IgJH genes. Cytogenetic analysis of both leukemic bone marrow and BALM-16 cells showed the t(8;22)(q24;q11) abnormality which is specifically associated with ALL-L3 and Burkitt lymphoma. The patient's serum showed hypercalcemia, prompting further investigation of the established cell lines which showed parathyroid hormone-related peptide (PTHrP) mRNA detected by reverse-transcriptase polymerase chain reaction. However, PTHrP production was not detected in the culture supernatant. The established cell line, BALM-16, could provide a useful material for analyzing the lack of Ig expression and of clarifying the pathogenesis of this type of B cell malignancy.

Adult↗
PubMed1997

Identity of original and late passage Dami megakaryocytes with HEL erythroleukemia cells shown by combined cytogenetics and DNA fingerprinting.

During routine authentication checks, we noticed untoward karyotypic similarities between late-passage stocks of the Dami megakaryocyte and HEL erythroleukemia cell lines. Genetic identity of Dami with HEL was demonstrated by DNA fingerprinting with a (gtg)5 multilocus probe and confirmed for earlier passages of Dami deposited by its originators with the ATCC. As initial passage stocks of Dami are no longer available for comparison, we investigated whether cross-contamination by HEL was more likely to have occurred during Dami's establishment or subsequently, by karyotyping currently available stocks of both cell lines for comparison with that originally reported for Dami. We found that the karyotype of current stocks of Dami overwhelmingly resembles that described in the original report, having retained at least 16/18 structural chromosome rearrangements as described therein, cf. 12/20 shared by current stocks of Dami and HEL. HEL's antecedence is shown by the retention of normal homologs of chromosomes, 6, 18, and 21--all rearranged in Dami. These results confirm the identity of current and early passage stocks of Dami and their common origin by cross-contamination with HEL which had occurred by July 1987, a year prior to publication. Thus, most, if not all, studies using Dami are likely to have employed HEL instead and may require reappraisal.

Chromosome Aberrations↗
PubMed1997

Interferon-gamma induced proliferation of human myeloid leukaemia cell lines.

Interferon-gamma (IFN-gamma) is a pleiotropic cytokine involved in the regulation of various phases of immune and inflammatory responses; it also has anti-viral and anti-proliferative activity. Using continuous human leukaemia cell lines as model systems, we found that IFN-gamma stimulated the proliferation of leukaemic myeloid cells; this effect was specifically neutralized by an anti-IFN-gamma monoclonal antibody (McAb). No proliferative response was seen in autonomously growing cell lines; however, 11/19 constitutively growth factor-dependent cell lines showed a significant response in short-term proliferation assays upon incubation with IFN-gamma. The stimulation indices ranged from 2 to 37 compared with the untreated control cells; the EC50 values for these cell lines were in the range of 0.1-0.6 ng/ml IFN-gamma. Flow cytometric analysis demonstrated heterogeneity in the expression of the IFN-gamma receptor, as it was found on 37-97% of the cells per cell line. The effects of IFN-gamma on proliferation triggered by a spectrum of 10 other cytokines were variable, and both stimulation and attenuation of the proliferative responses were seen in different cell lines. Under serum-free culture conditions, IFN-gamma acted as a survival factor suppressing apoptosis. As has been described for other functional processes triggered by IFN-gamma, the proliferation-inducing activity of IFN-gamma also led to activation of the signal transducing element STAT 1. Thus, IFN-gamma can induce myeloid leukaemia cells to proliferate and can modulate their proliferative response to other cytokines. Therefore IFN-gamma may be a pathologically relevant ligand for leukaemic cell proliferation in vivo. In physiological settings, IFN-gamma might be a bifunctional regulator of haemopoietic cell proliferation, depending on other differential co-signals from the micro-environment.

Adult↗
PubMed1997

Expression and function of CD95 (FAS/APO-1) in leukaemia-lymphoma tumour lines.

Cross-linkage of the CD95 (FAS/APO-1) antigen is responsible for the induction of programmed cell death or apoptosis in a variety of normal and malignant cells of the haemopoietic system. In order to evaluate predominant expression of the CD95 gene in a cell lineage-specific manner, we have determined the CD95 expression patterns in cell lines of myeloid, T-, pre-B- or B-cell origin as well as those established from Hodgkin's disease (HD). Our results reveal constitutive transcriptional activation of the CD95 gene in all cell lines derived from the lymphoid and myeloid lineages. Despite the ubiquitous expression of CD95 transcripts in haemopoietic cells, the corresponding protein was undetectable in 2/5 cell lines derived from Burkitt lymphomas and 6/16 leukaemia cell lines of the megakaryocytic or monocytic lineage. In an effort to identify apoptosis-resistant cell lines resulting from mutations in the death-signalling domain of CD9 5 or from defects in the apoptotic pathway or in survival programmes, we applied a CD95-mediated apoptosis assay. However, 21/38 CD95-expressing cell lines were sensitive upon induction with an anti-CD95 antibody whereas the remaining cell lines (predominantly of myeloid derivation) were resistant to antibody-induced cell death. Resistance to CD95-mediated apoptosis was not due to mutations within the CD95 open reading frame as confirmed by a combined reverse transcription PCR sequencing method. Five myeloid out of 13 tumour lines with the apoptosis-resistance phenotype analysed showed programmed cell death, when protein synthesis was blocked by treatment with cycloheximide prior to CD95-mediated induction. These data suggest an active cellular mechanism for the maintenance of an apoptosis-resistant phenotype. Elucidating the steps in such an active process of resistance to apoptosis might be expected to provide new approaches for therapeutic intervention in certain tumours.

Apoptosis↗
PubMed1997

In vitro culture studies of childhood myelodysplastic syndrome: establishment of the cell line MUTZ-1.

Myelodysplastic syndrome (MDS) in childhood is considered to be very rare and detailed pathobiological data are scarce. More biological information regarding MDS in children is clearly needed and in vitro culture studies provide one possibility for gaining further pathophysiological insights into this malignancy. Here, we incubated bone marrow samples from 30 children with MDS in liquid suspension culture in order to grow the transformed cells in vitro. In most cultures, the hematopoietic cells died quickly and only fibroblastic (stromal) background layers proliferated temporarily; several normal Epstein-Barr virus (EBV)-transformed B-lymphoblastoid cell lines (B-LCL) were established. Only in one instance, albeit from the peripheral blood and not from the bone marrow, could we establish a cell line, termed MUTZ-1, from the malignant cells of a 5-year-old girl with MDS (FAB subtype refractory anemia with excess of blasts). The MDS arose from a pre-existing Fanconi anemia and progressed quickly to an acute myeloid leukemia (FAB M2). Despite positivity for EBV, MUTZ-1 is not an EBV + B-LCL and further characterization of MUTZ-1 confirmed the derivation from the transformed clonal cells. Immunophenotyping showed a pre B-cell surface marker profile (CD10+ CD19+ cytoplasmic IgM+); receptor gene rearrangement analyses underlined the clonal B-cell nature of MUTZ-1 cells. MUTZ-1 cells exhibit a highly rearranged, unstable karyotype with a high frequency of spontaneous chromatid breaks and exchanges; del(5q) and additional rearrangements involving chromosome 5 [der(15)t(5;15)] were detected. The present data and results from a few other MDS-derived cell lines suggest that the transforming event in MDS seems to occur in an immature pluripotent progenitor cell. The new MDS-derived continuous cell line MUTZ-1 provides a useful in vitro model system for studies on the pathogenetic events leading to MDS.

Adolescent↗
PubMed1996

Chromosomal breakage correlates with delayed lethality in normal and ataxia telangiectasia cell lines treated with bleomycin.

Cytogenetic damage and cytotoxicity produced by a range of acute treatments with bleomycin (BLM) were investigated in three B-lymphoblastoid cell lines: EM-OC, PA-AT and CP-NC; established from heterozygous, and homozygous, ataxia telangiectasia (AT) and normal donors, respectively. The following endpoints were studied: (1) the maximum (initial) yields of different classes of chromosomal aberrations (CA) produced in G2-phase; (2) the percentage distributions of cells within bins with different numbers of CA; (3) short-term (0-48 h) viability and proliferation; and (4) (delayed) lethality, as measured by a modified limiting-dilution assay. While only slight losses of short-term viability and proliferation were detectable after standard BLM pulse-treatment at 25 or 100 micrograms/ml (results for latter dose given in parentheses), both regimes subsequently led to intense lethality as follows: CP-NC, 79.3% (95.1%); EM-OC, 90.4% (96.7%) and PA-AT, 98.4% (99.9%). Relative lethality after the respective BLM treatments increased 6.2 x (48.3 x) among PA-AT cells more than in controls-resembling the corresponding ratios for chromosome breaks (csb) of 7.0 x (30.9 x), more than those for chromatid breaks (ctb) of 3.1 x (2.7 x). EM-OC exhibited slightly increased relative lethality after the respective BLM treatments at 2.15 x (1.45), and increased aberration sensitivities for csb of 3.0 x (14.0), while the corresponding ratios for ctb were actually lower at 0.66 x (0.50 x). Significant correlations were observed between lethality and both, the yields of most types of chromosomal aberrations (excepting unambiguous exchanges) and also, the percentages of cells bearing CA.

Antibiotics, Antineoplastic↗
PubMed1996

Expression of the LMP1 oncoprotein in the EBV negative Hodgkin's disease cell line L-428 is associated with Reed-Sternberg cell morphology.

The latent membrane protein 1 (LMP1) oncogene is one of the major proteins synthesized by the Epstein-Barr virus (EBV) and is expressed in Reed-Sternberg (RS) cells of EBV-associated Hodgkin's disease (HD). We have studied the effect of this oncoprotein on the formation of RS cells in the EBV-negative HD cell lines L-428 and KM-H2 as well as on the formation of multinucleated cells in the mononuclear human embryonic kidney cell line 293. LMP1 prototype (B95-8) and its naturally occurring carboxy terminal 30 base pair deletion variant LMP1-del were transfected into the cell lines and cytocentrifuge preparations were analysed after 24, 48, 72, 144, 216, and 240 h. While no oncoprotein expression was seen in the KM-H2 cell lines, expression of LMP1 and LMP1-del was observed in the L-428 and 293 cell lines. In the HD cell line L-428 oncoprotein expression was infrequent but when observed was very strong and preferentially associated with multinucleated RS cell morphology (71% of LMP1 positive cells). This is in contrast with the untransfected or transfected but not expressing cells where intermediate mononuclear elements predominated over multinucleated RS cells (< 3%). Frequent oncoprotein expression was observed in the 293 cells and again was associated with multinuclearity. These LMP1 expressing 293 giant cells showed strong expression of ICAM-1(CD54), not detectable in the untransfected cells. In the LMP1-del transfectants giant cells with more than four nuclei were frequently observed. However, giant cells were much less frequent in 293 cells transfected with the amino terminal deletion variant of LMP1 or the lytic form of LMP1, known to induce low NF-kappa B activation compared to the LMP1 prototype. Therefore, LMP1 mediated NF-kappa B activation appears to be involved in polycaria formation. The strong association of LMP1 expression with multinuclearity in a genetically unstable condition -the L-428 and 293 cells show multiple chromosomal abnormalities-suggests that this oncoprotein including its naturally occurring carboxy terminal deletion variant promote the formation of multinuclear cells, in particular of RS cells in EBV-associated HD.

Blotting, Western↗
PubMed1996

Cohabiting t(12;22) and inv(3) primary rearrangements in an acute myelomonocytic leukemia (FAB M4) cell line.

We describe the cytogenetic characterization of MUTZ-3, the first continuous cell line to be established from acute myelomonocytic leukemia (FAB M4) cells, exhibiting recurrent chromosomal rearrangements associated with this disease category. MUTZ-3 was established from peripheral blood taken at presentation from a 29-year-old male patient and carries the t(12;22)(p13.2;q11.2) associated with acute myelomonocytic leukemia (AMMoL), the inv(3)(q21.2q26.3) associated with multilineage acute myeloid leukemias (AML), and the inv(7)(p 14q35) associated with ataxia telangiectasia (A-T). There was no evidence that the patient was an A-T heterozygote. The breakpoint on chromosome 22 mapped between 5' BCR and D22S39, consistent with the G-banding assignment. Both inversions were translocation-associated and may be further examples of an association previously described in AML FAB M4eo with inv(16). We suggest that the combination of inv(3)/t(3;3) with t(12;22) may represent a new, nonrandom association in AML.

Adult↗
PubMed1996

Expression of thrombopoietin and thrombopoietin receptor MPL in human leukemia-lymphoma and solid tumor cell lines.

Thrombopoietin (TPO) is the major regulator of platelet production in vivo and is the ligand for the MPL receptor. In an effort to determine the distribution of TPO and MPL in the different hematopoietic cell types and in various types of tissue, we examined the mRNA expression of this ligand-receptor pair in two series of human leukemia-lymphoma cell lines and of solid tumor cancer cell lines using northern blot and reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. At the northern blot mRNA level, 8/15 (53%) megakaryocytic and 3/11 (27%) erythroid leukemia cell lines expressed MPL mRNA; except for one positive monocytic cell line, the remaining 78 pre B-cell, B-cell, plasma cell, T-cell, NK cell, myeloid, monocytic and Hodgkin/anaplastic large cell lymphoma (ALCL)-derived cell lines were negative. No MPL message was detected in any of the 23 solid tumor cell lines established from 21 different tumors. In order to examine whether a low level of MPL expression could be detected, 51 leukemia cell lines were investigated with the RT-PCR technique. By this technique, MPL message was seen in many more cell types: 13/26 (50%) of non-erythromegakaryocytic cell lines and in nearly all megakaryocytic (14/15, 93%) and erythroid (10/11, 91%) cell lines. Thus, the highest expression of MPL clearly occurs in cells with megakaryocytic differentiation; furthermore, expression of MPL appears to be restricted to hematopoietic cell types. TPO mRNA expression was examined by RT-PCR and found in 9/11 (82%) of the solid tumor cell lines (derived from colon, endometrium, kidney, liver, ovary, retinoblastoma and urinary bladder cancers). Among the leukemia-lymphoma cell lines, TPO mRNA was detected by RT-PCR in most plasma cell, myeloid, megakaryocytic and erythroid cell lines, but not in pre B-cell, B-cell or T-/NK-cell lines. The results reported here extend the observations of MPL and TPO expression in normal cells to the whole spectrum of hematological cell types and to an array of different tissue types, both exemplified by their malignant counterparts.

Blotting, Northern↗
PubMed1996

MUTZ-3, a monocytic model cell line for interleukin-4 and lipopolysaccharide studies.

The human monocytic cell lines MUTZ-3 and MONO-MAC-6 express the lipopolysaccharide (LPS) receptor CD14. Paralleling the situation in peripheral blood monocytes (PBMo), recombinant human interleukin-4 (IL-4) down-regulated the expression of CD14 on the cell surface of MUTZ-3, but not that of MONO-MAC-6 cells. In addition, preincubation with IL-4 prevented the LPS-induced up-regulation of IL-1 beta mRNA levels in MUTZ-3, but not in MONO-MAC-6 cells. We examined whether the differential responsiveness of the cell lines was due to the missing expression of the IL-4 receptor (IL-4R) alpha or gamma c chain in MONO-MAC-6 cells. Flow cytometric and immunoprecipitation analysis revealed expression of both IL-4R chains in both cell lines. In addition, short-term stimulation with IL-4 induced tyrosine-phosphorylation of the gamma c chain. As both cell lines also expressed signal transducer and activator of transcription 6 (STAT 6), our data suggested that the differential reaction patterns of MUTZ-3 and MONO-MAC-6 cells were not due to a generally defective IL-4R complex. Interestingly, long-term (48 hr) treatment with LPS rendered MONO-MAC-6 cells sensitive to IL-4. LPS up-regulated expression of monocyte-specific esterase (MSE) mRNA as well as CD14 protein in MONO-MAC-6 cells; both effects were inhibited by IL-4. This stimulation was not paralleled by an increase of IL-4R mRNA or protein expression supporting the above hypothesis of a constitutively present and active IL-4R. We discuss possible causes for the differential reaction patterns of MUTZ-3 and MONO-MAC-6 cells to IL-4.

Antigens, CD↗