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M Rosenzwajg

Publications and source records attributed to M Rosenzwajg.

14 recordsLinked to original sources
PubMed2000

CD13/N-aminopeptidase is involved in the development of dendritic cells and macrophages from cord blood CD34(+) cells.

Expression of CD13/N-aminopeptidase may reflect cell activation and growth. We examined its role regarding cell growth in cultures of cord blood CD34(+) cells with stem cell factor/Flt-3 ligand/granulocyte-macrophage colony-stimulating factor/tumor necrosis factor-alpha. Indeed, 82% +/- 6% of cells from culture day 5 were CD13(hi), 25% +/- 8% of which were still Lin-. About 50% of CD13(hi)Lin- cells, which comprise progenitors of dendritic cells (DC), monocytes/macrophages and granulocytes, and 30% of CD13(lo)Lin- cells were CD34(+). Sorted CD34(+)CD13(hi)Lin- cells, cultured further for 7 days with the same cytokines, expanded 31-fold and CD34(-)CD13(hi)Lin- cells 7-fold, but CD34(+)CD13(lo)Lin- and CD34(-)CD13(lo)Lin- cells did not grow. Thus, cell growth correlated with CD13 expression, all the more so that cells were CD34(+). Actinonin, the most potent N-aminopeptidase inhibitor, was used to engage CD13 on sorted CD13(hi)Lin- cells and on culture day-7 bulk cells. In both cases, this resulted in reversible cell growth arrest, with 30% to 60% fewer cells in the G2/S-M phase than in controls. Interestingly, similar effects were noted with CD13 monoclonal antibody TUK1, which does not inhibit N-aminopeptidase activity, but not with N-aminopeptidase-blocking antibodies WM15 and F23. All cycling cells appeared susceptible to actinonin, which induced cell apoptosis at the same time as Bcl-2 was downregulated and caspase-3 activity increased, but finally percentages and yields of DC and macrophage precursors were affected more than those of granulocytic cells. Thus, through engagement of N-aminopeptidase enzymatic site but possibly also of an independent determinant, CD13 plays a role in the growth of DC/macrophage progenitors and precursors. (Blood. 2000;95:453-460)

Antigens, CD34↗
PubMed1999

Human herpes virus 8 (HHV8) serology in allogeneic bone marrow transplant recipients.

Human herpes virus 8 (HHV8) may be sexually transmitted, but transmission via blood cells has not yet been excluded. We used a modified immunofluorescence assay to detect Ab to HHV8 latency-associated nuclear Ag in sera of 200 allogeneic BMT recipients and their related donors. In control subjects, Ab were found in 85% of patients with AIDS-related Kaposi sarcoma (n = 52), 34% of HIV-1 infected subjects without Kaposi sarcoma (n = 56) and 9. 5% of blood donors (n = 42). Among BMT donors, 14.5% were HHV8+, while 10% of recipients were positive before, and 18% after BMT. In the 176 HHV8-negative recipients at BMT, there was no relationship between post-BMT seroconversion, which occurred in 26 cases (15%), and the donor's serological status. Of note, 10 HHV8+ recipients before BMT became negative post-BMT. Outcome of BMT was not influenced by prior HHV8 seropositivity, seroconversion or seroreversion of recipients. That HHV8 seropositivity among blood donors from the Paris area was comparable to that of BMT donors and recipients before BMT indicates that these patients had not been at risk of HHV8 by blood products received before BMT, although post-BMT HHV8 seroconversion probably corresponded to contamination by blood transfusions rather than by the BMT.

Acquired Immunodeficiency Syndrome↗
PubMed1998

[Dendritic cells: a complex cellular system].

Dendritic cells (DC) are the most potent antigen-presenting cells. Thus, ex vivo antigen-pulsed DC are a potentially powerful tool to induce in vivo immunity against tumor-associated or viral antigens. Therefore, culture methods to generate high numbers of DC from bone marrow or blood CD34+ hematopoietic progenitor cells have recently been developed. These methods, which use different combinations of growth factor--mainly granulocyte/macrophage colony-stimulating factor (GM-CSF), tumor necrosis factor (TNF)-alpha and interleukin (IL)-4--make the characterization of DC obtained from CD34+ cells of different origins easier and allow to assess whether DC relate to a unique or distinct differentiation pathways. Monocytes and even macrophages can also directly differentiate into DC in the presence of GM-CSF and IL-4. This has to be reconciled with evidence supporting earlier branching off of the macrophage and DC lineages, and raises questions as to the identity of the latter lineage. Apart from DC of myeloid origin, DC may also originate from lymphoid progenitors. Because the capacity of DC to capture, process and present antigens is known to vary according to their differentiation stage, and lymphoid DC might behave differently from lymphoid DC in this respect, the definition of which type of DC to use for immunotherapy must be more precise, in order to avoid detrimental side effects or results. From a practical point of view, it is also necessary to define the most appropriate cytokine combinations and schedules thereof to optimize proliferation and differentiation of DC from different origins. These conditions should then be applied to generated DC for their efficient and safe use for clinical immunotherapy.

Cell Culture Techniques↗
PubMed1998

The influence of interleukin (IL)-4, IL-13, and Flt3 ligand on human dendritic cell differentiation from cord blood CD34+ progenitor cells.

Culturing cord blood CD34+ cells with granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumor necrosis factor (TNF)-alpha for 12 days, and stem cell factor (SCF) for 5 days, resulted in a 40- +/- 26-fold expansion in cell numbers, with 38 +/- 20% dendritic cells (DCs). Interleukin (IL)-4 and IL-13, which share properties, were examined first. Adding either one to the former baseline condition beginning on day 0 halved cell growth while the percentage of DCs increased to 60-70%, resulting in unchanged DC yields. Delaying use of IL-4 or IL-13 to day 5 led to 25-fold cell expansion with approximately 80% DC, the yield of which was then twofold over that of baseline control cultures, while numbers of other cells decreased. IL-4 and IL-13 had no additive or antagonistic effect on DC generation. The effect of Flt3 ligand (FL), known to enhance proliferation of hematopoietic progenitors induced by other growth factors, was examined next. FL added alone induced DC in the same manner as SCF. Using both FL and SCF throughout the culture period enhanced total cell recovery fourfold above that of baseline control cultures on day 12 compared with > or =2.5-fold if either one was stopped on day 5. When both FL and SCF were used for 12 days, DC recovery was fivefold that of control cultures, whereas it was to three- to 3.5-fold when either one was stopped on day 5. A similar trend was noted for CD15+ cells, and, to a lesser extent, for CD14+ cells. Finally, using SCF and FL for 12 days, with IL-4 or IL-13 added from day 5 onwards, led to comparably enhanced cell yields relative to control cultures with approximately 60% DC. These data underline the need to use appropriate cytokine combinations and schedules to optimize generation of DCs from CD34+ progenitors. Associated with GM-CSF and TNF-alpha, IL-4 or IL-13 promotes differentiation and maturation of DCs over other myeloid cells. Under the same baseline conditions, FL appears to potentiate SCF throughout the culture period, inducing proliferation and development of DC as well as of other myeloid cells.

Antigens, CD34↗
PubMed1997

Differentiation of human dendritic cells from monocytes in vitro.

Since either macrophages (Mphi) or dendritic cells (DC) differentiate from monocytes (MO) depending on culture conditions, we investigated the relationship of the DC and Mphi differentiation pathways. Culturing MO-enriched blood mononuclear cells with Mphi colony-stimulating factor (M-CSF) or with granulocyte/Mphi (GM)-CSF induced Mphi with a different morphology and CD14/CD1a expression. In contrast, in cultures with GM-CSF and interleukin (IL)-4, cells rapidly became nonadherent and acquired DC morphology, ultrastructure, CD1a expression, and most DC markers; they lost membrane CD14 and CD64 and capacity of phagocytosis, displayed less CD68 than Mphi, but retained nonspecific esterase activity. These DC directly developed from MO without proliferation inasmuch as only day 0 FACS-sorted MO, but not small CD14- cells, differentiated into DC when cultured with GM-CSF and IL-4, or to Mphi with M-CSF While overall cell numbers declined, DC numbers plateaued from culture day 2 onwards, indicating that most had differentiasted by then. This differentiation was radioresistant and occurred without [3H]thymidine incorporation. Commitment to differentiate into DC with GM-CSF and IL-4 was irreversible by day 2, since discontinuing IL-4 at this point did not revert cells to Mphi. Alternatively, cells rapidly converted to DC when IL-4 was added from day 2 to cultures initiated with GM-CSF only. If cultures were initiated with M-CSF and switched to GM-CSF and IL-4 after 2 or 5 days, about half of the cells still converted to DC. Thus, the capacity of MO and even of Mphi to differentiate into DC was conserved for at least this period. The increased capacity to stimulate the mixed leukocyte reaction correlated with the relative number of CD1a+ cells at any time and under each condition tested, a confirmation that these cells functionally qualify as DC. Thus, MO and even Mphi can be directed to differentiate into DC depending on the cytokine microenvironment.

Antigens, CD1↗
PubMed1997

In vitro generation of human dendritic cells and cell therapy.

Dendritic cells (DC) are the most potent antigen-presenting cells: they, only, can prime naive T lymphocytes and even elicit generation of cytotoxic T lymphocytes to soluble antigens. Thus ex vivo antigen-pulsed DC represent a potentially powerful tool to elicit T-cell mediated responses against viral or tumor-associated antigens. Because isolation of DC as such from the blood is hampered by their scarcity, culture methods to generate them from different progenitors or precursors have been developed. Indeed, the possibility of obtaining relatively high numbers of DC from bone marrow, cord blood or adult blood CD34+ progenitors, or even blood monocytes, in cultures with different combinations of growth factors--mainly based on the use of GM-CSF, TNF-alpha and IL-4--has allowed the study of their ontogeny, the characterization of the different types of DC obtained under diverse conditions, and the assessment of whether they relate to a single pathway of differentiation. For example, the finding that monocytes and even macrophages can differentiate into DC depending on the cytokines used has to be reconciled with evidence that supports earlier branching off of the macrophage and DC lineages, and raises questions as to the identity of the latter lineage. Also, besides DC of myeloid origin, DC arise from lymphoid progenitors, and lymphoid DC display different properties than myeloid DC--at least in mice. From a practical point of view, there is a need to define the most appropriate cytokine combinations and schedules to optimize proliferation, differentiation and maturation of DC from different sources. In addition, because the capacity of DC to capture, process and present antigens varies according to their differentiation/maturation stage and origin, it appears necessary to define which type of DC to use for cell therapy in the setting of a given pathology for efficient and safe use.

Cell Differentiation↗
PubMed1996

The effect of in vitro human immunodeficiency virus infection on dendritic-cell differentiation and function.

CD1a+ dendritic cells (DC) differentiate from a major population of nonadherent CD13(hi)lin- cells that appear when human cord blood CD34+ hematopoietic progenitor cells are cultured with stem-cell factor, granulocyte/macrophage (MA) colony-stimulating factor, and tumor necrosis factor-alpha (TNF-alpha) for 5 days. CD13hilin- cells, which also comprise MA and granulocyte precursors, are CD4+ and can thus be targets of human immunodeficiency virus (HIV). Low replication was noted when these day 5 cells were infected with lymphotropic HIV-1LA1 (p24: < or = 4 ng/mL on day 8 postinfection [PI]), while high virus production occurred with MA-tropic HIV-1Ba-L, HIV-1Ada, or HIV-1-m-n. (p24: 50 to > or = 1,000 ng/mL). Strong cytopathicity (CPE) was then observed in nonadherent cells as in adherent MA. However, FACS analysis on day 7 PI showed that HIV did not affect differentiation of DC that survived CPE: apart from CD4 downmodulation related to HIV production, overall expression of CD40, CD80, and CD86 costimulatory molecules, and of HLA-DR, was unchanged relative to controls. At that time, the capacity of DC from HIV-infected cultures to stimulate the mixed leukocyte reaction was only altered less than 10-fold. Immunocytochemistry on day 7 PI showed that most HIV-infected cells were included in syncytia that were stained by anti-CD1a, anti-S100, and anti-CD14 antibodies, indicating that syncytia consisted of DC and cells of the MA lineage. Polymerase chain reaction analysis of FACS-sorted CD1a+ cells confirmed that they harbored then HIV DNA. Viral DNA was also detected in CD1a+ DC from noninfected cultures that had been exposed to HIV only after sorting. Therefore, we examined whether in infected cultures DC precursors were infected at the onset or if virus spread later from other infected cells to differentiated DC. This was answered by showing that, 24 hours postexposure to HIV, viral DNA was preferentially detected in day 5 sorted CD13hilin- versus CD13hilin- cells, and that it was found in the CD1a+ progeny of CD13(hi)lin- cells 48 hours later. In addition, HIV replication did not affect myeloid clonogenic progenitors in day 0 to day 7 PI cultures, although viral DNA was detected in colony-forming unit-granulocyte/macrophage (CFU-GM)/CFU-M colonies derived from day 3 and 7 PI cultures. Thus, precursors of DC and their progeny are susceptible to HIV in vitro, but, apart from CPE, the effect of virus production on DC differentiation or function is limited.

Antigens, CD↗
PubMed1996

Macrophage inflammatory protein-1alpha is induced by human immunodeficiency virus infection of monocyte-derived macrophages.

Disparate findings have been reported as to whether human immunodeficiency virus (HIV) affects cytokine production by macrophages (MA). We investigated production of different cytokines and of macrophage inflammatory protein (MIP)-1alpha by HIV-1Ba-L- or HIV-1Ada-infected blood-derived MA. Relative to controls, only MIP-1alpha levels increased twofold to > 10-fold in supernatants 2 to 3 weeks postinfection (PI), at the time of maximum virus production; levels of the other chemokines (RANTES, interleukin (IL)-8) and cytokines (IL-1alpha, IL-3, IL-6, granulocyte-macrophage colony-stimulating factor (GM-CSF), G-CSF, tumor necrosis factor (TNF)-alpha, transforming growth factor (TGF)-beta1) investigated were not affected. MIP-1alpha mRNA signal assessed by reverse transcriptase-polymerase chain reaction (RT-PCR) was, however, only occasionally greater in cells from infected cultures relative to controls. MIP-1alpha levels in supernatants remained in the same range as in control cultures when more than 10 mmol/L Zidovudine was added 24 hours PI, which indicates involvement of virus replication in the effect. Anti-MIP-1alpha antibody labeling identified a 10% to 25% subset of MA, strongly expressing HLA-DR and CD4, and also stained by anti-IL-6 and anti-TNF-alpha antibodies. Two weeks PI, dual staining showed that the majority of the 5% to 20% cells that were p24+ belonged to the MIP-1alpha+ population, which may define a MA subset capable to better sustain HIV replication. MIP-1alpha induced by HIV replication in MA might play a role in the pathophysiology of HIV infection; in impaired hematopoiesis; or as a CD4+ and CD8+ lymphocyte chemoattractant, by recruiting either or both HIV-susceptible and cytotoxic T lymphocytes to virus replication sites.

Antiviral Agents↗
PubMed1996

Human dendritic cell differentiation pathway from CD34+ hematopoietic precursor cells.

The most effective antigen-presenting cells for T lymphocytes are dendritic cells (DCs), the differentiation pathway of which, however, is incompletely characterized. We examined here how DCs differentiated from human cord blood CD34+ progenitor cells cultured with granulocyte-macrophage colony-stimulating factor, tumor necrosis factor-alpha, and stem cell factor. After 5 days, 2 of 3 nonadherent cells were CD13hiHLA-DRhiCD4+, half of them were also CD14+, and < or = 10% were CD1a+. When day-5 sorted CD13hiCD1a- and CD13lo cells were further cultured, CD1a+ cells appeared in the already CD13hi population, whereas CD13hi cells, a minority of which rapidly became CD1a+, emerged from the CD13lo population. By day 12, still 66% of bulk cells in suspension were CD13hi, most of which displayed high forward and side scatters of large granular cells. Half of CD13hi cells were CD1a+. All CD13hi cells expressed to the same extent DR, CD4, costimulatory and adhesion molecules, and various amounts of CD14. CD1a+ cells stimulated allogeneic lymphocytes more than CD13hiCD1a- cells and, although they were CD14+, both cell types were nonspecific esterase-negative nonphagocytic cells and were stronger mixed leukocyte reaction stimulators than were their macrophage counterparts. Eventually, the percentage of CD1a+ cells decreased. However, typical CD1a+ DCs still emerged in culture of sorted day-12 CD13hiCD1a- cells, and adding interleukin-4 to bulk cultures at that time led to the persistence of the CD1a+ population while diminishing CD14 expression. Thus, this system results first in the differentiation of CD13hi precursors that strongly express DR and CD4, from which more mature CD1a+ DCs continuously differentiate all along the culture period.

Antigens, CD↗
PubMed1995

Susceptibility of human bone marrow stromal cells to human immunodeficiency virus (HIV).

It is not known whether impaired hematopoiesis noted during human immunodeficiency virus (HIV) infection results from infection of stem/progenitor cells or of cells of the bone marrow microenvironment. Normal adherent primary stromal layers were exposed to HIV to determine which of this mixture of endothelial cells, fibroblasts, and macrophages are susceptible to the virus. Viral p24 in supernatants was noted with monocytotropic HIV-1Ada, HIV-1Ba-L, and HIV-1JR-FL but not with lymphotropic HIV-1LAI nor HIV-1MN strain, and only stromal macrophages expressed the viral antigens. Coculture of the layers with PHA-activated normal lymphocytes failed to rescue lymphotropic virus. No p24 was produced when macrophage-depleted stromal cells were exposed to either HIV-1Ba-L or HIV-1LAI; proviral DNA was then amplified by PCR in cells exposed to either virus, though coculture with lymphocytes rescued only HIV-1Ba-L. Altogether, these data indicate that macrophages are the major targets of HIV in cultured stromal layers. As virus replication in macrophages did not affect the profile of major cytokines involved in regulating hematopoiesis, HIV infection could alter hematopoiesis by other as yet unspecified mechanisms.

Base Sequence↗
PubMed1994

The MTCP-1/c6.1B gene encodes for a cytoplasmic 8 kD protein overexpressed in T cell leukemia bearing a t(X;14) translocation.

The t(X;14)(q28;q11.2) translocation is associated with mature T-cell proliferations. Recently this translocation has been shown to implicate the MTCP-1/c6.1B gene on chromosome Xq28, leading to aberrant or overexpressed MTCP-1 transcripts. The potential coding role of this gene was made uncertain by the lack of a long open reading frame in its major transcripts. However, a short 204 bases open reading frame is potentially coding for a 68 amino-acid protein. Here, we show that this open reading frame sequence and the deduced product are well conserved in mouse. A 8 kD protein (p8), which corresponds to the predicted molecular weight was revealed in transient transfectants and in cell lines by Western blotting, using a rabbit antiserum. This product was absent in lymphoblastoid cell lines with deletions of the MTCP-1/c6.1B locus. A dramatic overexpression of p8 was found in leukemic cells from a patient with a t(X;14). This small protein was localized in the cytoplasm by immunofluorescence. In conclusion, MTCP-1 encodes for a cytoplasmic 8 kD product. Its potential role in leukemogenesis is supported by its overexpression in leukemia with t(X;14), but its function remains unknown.

Amino Acid Sequence↗
PubMed1993

MTCP-1: a novel gene on the human chromosome Xq28 translocated to the T cell receptor alpha/delta locus in mature T cell proliferations.

T-cell lymphoproliferative diseases are often associated with recurrent chromosomal translocations involving T cell receptor genes (TCR) and genes that are thought to play a role in the pathogenesis of these diseases. Whereas numerous such genes have already been identified in acute T cell leukemias, no candidate gene has yet been identified to play a role in the heterogeneous group of T cell proliferations with a mature phenotype. We here report the molecular cloning of two examples of the rare but recurrent t(X;14) translocation. The first translocation was associated with a benign clonal proliferation in an ataxia telangiectasia patient and the second with a T cell prolymphocytic leukemia. Both translocations implicated the TCR alpha/delta locus and a common breakpoint region on chromosome Xq28. A previously unidentified gene, abnormally transcribed in both T cell proliferations, was characterized in the immediate proximity of the breakpoints. This Xq28 gene has no homology with known sequences, uses a complex alternative splicing pattern and demonstrates two short open reading frames. This gene, named MTCP-1 (Mature T Cell Proliferation-1) is the first candidate gene potentially involved in the leukemogenic process of mature T cell proliferations.

Alternative Splicing↗