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Query: UNIPROT:P15088 (mast cell)
 14,925 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mast cells may be cultured from human peripheral blood in the presence of recombinant human stem cell factor (rhSCF). The characteristics of the cells in peripheral blood that give rise to mast cells are unknown. Because mast cell precursors in human marrow are CD34+, human peripheral blood mononuclear cells from patients with mastocytosis and normal controls were sorted on the basis of CD34 expression and the positive and negative cell populations were cultured in rhSCF, recombinant human interleukin-3 (rhIL-3), or both for 6 weeks. Cell cultures were examined every 2 weeks for total and mast cell number and cell differential using Wright Giemsa and acid toluidine blue stains and antibodies to mast cell tryptase and chymase, cell-associated histamine, and expression of CD34, c-kit, Fc epsilon RI, and Fc gamma RII using flow cytometric analysis. The ultrastructural anatomy of mast cells was examined by electron microscopy. Peripheral blood CD34+ cells cultured in rhSCF with or without rhIL-3 gave rise to cell cultures consisting of greater than 80% mast cells by 6 weeks. CD34+ cells cultured in rhIL-3 alone did not give rise to mast cells, whereas rhIL-3 plus rhSCF increased the final mast cell number eightfold when compared with cells cultured in rhSCF alone. Mast cells increased concomitantly with a decrease in large undifferentiated mononuclear cells. CD34- cells did not give rise to mast cells. Histamine content per cell at 6 weeks was approximately 5 pg. Electron microscopy of 4-week cultures showed immature mast cells containing predominantly tryptase-positive granules that were either homogeneous or contained lattice structures, partial scroll patterns, or central dense cores and mixtures of vesicles, fine granular material, and particles. The CD34+ population at day 0 expressed Kit (65%) and Fc gamma RII (95%), but not Fc epsilon RI, by fluorescence-activated cell sorter analysis. At 6 weeks, CD34+-derived mast cells exhibited Fc epsilon RI in addition to Kit and Fc gamma RII, and were negative for CD34 antigen. Patients with mastocytosis showed a higher number of mast cells per CD34+ cell cultured compared with normal controls. Thus, the mast cell precursor in human peripheral blood is CD34+/Fc epsilon RI- and gives rise to mast cells in the presence of rhSCF with or without rhIL-3, and the number of mast cells arising per CD34+ cell in culture is greater when the CD34+ cells are obtained from patients with mastocytosis compared with normal subjects.
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PMID:Mast cells cultured from the peripheral blood of normal donors and patients with mastocytosis originate from a CD34+/Fc epsilon RI- cell population. 752 30

The generation of murine mast cells is supported by several cytokines, and mast cell lines are frequently established in long-term cultures of normal murine marrow cells. In contrast, growth of human mast cells was initially dependent on coculture with murine fibroblasts. The growth factor produced by murine fibroblasts and required to observe differentiation of human mast cells is attributable in part to stem cell factor (SCF). However, other factors are likely involved. We have previously shown that the combination of SCF and interleukin-3 (IL-3) efficiently sustains proliferation and differentiation of colony-forming cells (CFCs) from pre-CFC enriched from human umbilical cord blood by CD34+ selection. With periodic medium changes and the addition of fresh growth factors, five consecutive cultures of different cord blood samples gave rise to differentiated cells and CFCs for more than 2 months. Although differentiated cells continued to be generated for more than 5 months, CFCs were no longer detectable by day 50 of culture. The cells have the morphology of immature mast cells, are Toluidine blue positive, are karyotypically normal, are CD33+, CD34-, CD45+, c-kit-, and c-fms-, and die in the absence of either SCF or IL-3. These cells do not form colonies in semisolid culture and are propagated in liquid culture stimulated with SCF and IL-3 at a seeding concentration of no less than 10(4) cells/mL. At refeedings, the cultures contain a high number (> 50%) of dead cells and have a doubling time ranging from 5 to 12 days. This suggests that subsets of the cell population die because of a requirement for a growth factor other than SCF or IL-3. These results indicate that the combination of cord blood progenitor and stem cells, plus a cocktail of growth factors including SCF and IL-3, is capable with high efficiency of giving rise in serum-deprived culture to human mast cells that behave like factor-dependent cell lines. These cells may represent a useful tool for studies of human mast cell differentiation and leukemia.
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PMID:Long-term generation of human mast cells in serum-free cultures of CD34+ cord blood cells stimulated with stem cell factor and interleukin-3. 752 46

Although mast cells are hematopoietic cells, little is known about the origin of their precursors in vivo. In this study, the origin (donor v recipient genotype) of human mast cells (MCs) was analyzed in a patient who underwent allogeneic bone marrow transplantation (BMT). The patient presented with secondary acute myeloid leukemia (French-American-British classification, M2) arising from refractory anemia with excess of blast cells and bone marrow (BM) mastocytosis. Transplantation was performed in chemotherapy-induced complete remission. On days 88, 126, 198, and 494 after BMT, mast cells were enriched to homogeneity from bone marrow mononuclear cells (BM MNCs) by cell sorting for CD117+/CD34- cells. Purified mast cell populations were CD117(c-kit)+ (> 95%), CD34- (< 1%), CD3- (< 1%), CD14- (< 1%), and virtually free of contaminating cells as assessed by Giemsa staining. The genotype of MCs was analyzed after amplification by polymerase chain reaction (PCR) of a variable number tandem repeat (VNTR) region within intron 40 of the von Willebrand factor (vWF) gene. Unexpectedly, on days 88 and 126 after BMT, sorted MCs displayed recipient genotype as shown by vWF.VNTR-PCR. However, on days 198 and 494, PCR analysis showed a switch to donor genotype in isolated mast cells. Peripheral blood (PB) and BM MNC as well as highly enriched (sorted) CD3+ T cells (PB, BM), CD4+ helper T cells (PB), CD8+ T cells (PB), CD19+ B cells (PB), CD14+ monocytes (PB, BM), and CD34+ precursor cells (BM) showed donor genotype throughout the observation period. Together, these results provide evidence that human MCs developed in vivo from transplanted hematopoietic stem cells. Engraftment and in vivo differentiation of MCs from early hematopoietic progenitor cells may be a prolonged process.
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PMID:Origin of human mast cells: development from transplanted hematopoietic stem cells after allogeneic bone marrow transplantation. 794 67

In the present paper we have used a three-color immunofluorescence procedure combined with flow cytometry cell analysis and sorting for the identification and enumeration of human mast cells in both normal and pathological bone marrow samples. Our results show that bone marrow mast cells are clearly identifiable on the basis of their light-scatter properties and strong CD117 expression. These cells were negative for the CD34, CD38, and BB4 antigens. In addition, they were CD33+ and displayed a high reactivity for the anti-IgE monoclonal antibody. The identity of the CD117-strong+ cells (mast cells) was confirmed by both microscopic examination and flow cytometry analysis. The overall frequency of mast cells in the bone marrow samples analyzed in the present study was constantly lower than 1%. The lowest frequencies corresponded to normal human bone marrow samples (0.0080 +/- 0.0082%) and the highest to those patients suffering from indolent systemic mast cell disease (0.40 +/- 0.13%). In summary, our results show that the identification and enumeration of bone marrow mast cells can be achieved using multiparametric flow cytometry. Moreover, once identified, mast cells are suitable for being characterized from the phenotypic and the functional point of view, facilitating the comparison between normal and abnormal mast cells.
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PMID:Flow cytometric analysis of mast cells from normal and pathological human bone marrow samples: identification and enumeration. 890 39

The primary role of protooncogene c-kit in mast cell differentiation is supported by the development of mast cells from CD34+/CD117+(c-kit) myeloid precursors. Growth factor independence, neoplastic transformation and differentiation of mast cells were found in association with c-kit activating mutations in both murine and human mastocytoma and mast cell diseases. We have identified a novel c-kit mutation (D816Y) in peripheral blood mononuclear cells from a patient with AML (M2), massive presence of mast cells in bone marrow and rapid progression of the disease. The mutation, a G-->T transversion at nt 2467 of the c-kit gene resulting in Asp816-->Tyr substitution, corresponds to the D814Y and D817Y mutations identified and characterized in the murine P815 mastocytoma and the rat RBL-2H3 mast cell leukemia cell lines. The absence of SCF transcripts that we found by RTPCR in the patient's blasts indicates that, also in humans, this activating mutation leads to SCF independent growth. The expression of the mutant allele on Kit signaling may be further enhanced by trisomy of chromosome 4 (carrying the c-kit gene) in the patient's blasts. From these findings it is concluded that mast cells could be generated from a leukemic CD34/CD117-positive clone, that combines the antigenic expression of mast cell precursor to the growth and differentiation factor-independence which was derived by the c-kit D816Y mutation.
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PMID:In vivo differentiation of mast cells from acute myeloid leukemia blasts carrying a novel activating ligand-independent C-kit mutation. 971 3

A subset of patients with systemic mastocytosis (SM) develop acute myeloid leukaemia (AML). However, little is known about the biology of such leukaemias and their relationship to the mast cell (MC) lineage. We report on two female patients who suffered from SM and AML. According to FAB criteria, the leukaemias were classified as AML-M4 (patient 1) and AML-MO (patient 2). The coexistence of the two distinct neoplasms (AML and SM) was demonstrable by immunostaining of serial bone marrow (BM) sections with monoclonal antibodies (mAb). In particular, the MC infiltrates were found to react with mAb against MC-tryptase and MC growth factor receptor c-kit (CD117), but not with mAb to CD15 or CD34. In contrast, the AML blasts were immunoreactive for CD15 (patient 1) or CD34 (patient 2), but did not express tryptase. The c-kit point mutation Asp-->Val at codon 816, considered to play a role in the transformation of MC progenitors, was detected in patient 1 in a BM cell fraction containing 4% MC. However, no c-kit mutation was found in pure AML blasts (<1% MC). These findings argue against an evolution of the AML clone from neoplastic MC or MC-committed progenitors.
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PMID:Systemic mastocytosis associated with acute myeloid leukaemia: report of two cases and detection of the c-kit mutation Asp-816 to Val. 985 25

The pattern of expression of several protein kinase C (PKC) isoforms (alpha, betaI, delta, epsilon, eta, and zeta) during the course of hematopoietic development was investigated using primary human CD34(+) hematopoietic cells and stable cell lines subcloned from the growth factor-dependent 32D murine hematopoietic cell line. Each 32D cell clone shows the phenotype and growth factor dependence characteristics of the corresponding hematopoietic lineage. Clear-cut differences were noticed between erythroid and nonerythroid lineages. (1) The functional inhibition of PKC-epsilon in primary human CD34(+) hematopoietic cells resulted in a twofold increase in the number of erythroid colonies. (2) Erythroid 32D Epo1 cells showed a lower level of bulk PKC catalytic activity, lacked the expression of epsilon and eta PKC isoforms, and showed a weak or absent upregulation of the remaining isoforms, except betaI, upon readdition of Epo to growth factor-starved cells. (3) 32D, 32D GM1, and 32D G1 cell lines with mast cell, granulo-macrophagic, and granulocytic phenotype, respectively, expressed all the PKC isoforms investigated, but showed distinct responses to growth factor readdition. (4) 32D Epo 1.1, a clone selected for interleukin-3 (IL-3) responsiveness from 32D Epo1, expressed the epsilon isoform only when cultured with IL-3. On the other hand, when cultured in Epo, 32D Epo1.1 cells lacked the expression of both epsilon and eta PKC isoforms, similarly to 32D Epo1. (5) All 32D cell lines expressed the mRNA for PKC-epsilon, indicating that the downmodulation of the epsilon isoform occurred at a posttranscriptional level. In conclusion, the PKC isoform expression during hematopoiesis appears to be lineage-specific and, at least partially, related to the growth factor response.
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PMID:Lineage-restricted expression of protein kinase C isoforms in hematopoiesis. 994 60

Human mast cells are derived from CD34(+) hematopoietic cells present in cord blood, bone marrow, and peripheral blood. However, little is known about the properties of the CD34(+) cells. We demonstrated here that mast cell progenitors that have distinct phenotypes from other hematopoietic cell types are present in cord blood by culturing single, sorted CD34(+) cells in 96-well plates or unsorted cells in methylcellulose. The CD34(+) mast cell-committed progenitors often expressed CD38 and often lacked HLA-DR, whereas CD34(+) erythroid progenitors often expressed both CD38 and HLA-DR and CD34(+) granulocyte-macrophage progenitors often had CD33 and sometimes expressed CD38. We then cultured single cord blood-derived CD34(+)CD38(+) cells under conditions optimal for mast cells and three types of myeloid cells, ie, basophils, eosinophils, and macrophages. Of 1,200 CD34(+)CD38(+) cells, we were able to detect 13 pure mast cell colonies and 52 pure colonies consisting of either one of these three myeloid cell types. We found 17 colonies consisting of two of the three myeloid cell types, whereas only one colony consisted of mast cells and another cell type. These results indicate that human mast cells develop from progenitors that have unique phenotypes and that committed mast cell progenitors develop from multipotent hematopoietic cells through a pathway distinct from myeloid lineages including basophils, which have many similarities to mast cells.
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PMID:Characterization of mast cell-committed progenitors present in human umbilical cord blood. 1023 86

The effects of thrombopoietin (TPO) and/or stem cell factor (SCF) on the development of human mast cells from CD34(+) bone marrow (BM) cells were investigated using a serum-deprived liquid culture system. Mast cells were identified by measurement of intracellular histamine content, immunocytochemical staining, and flow cytometric analysis. Whereas SCF alone generated only a small number of tryptase+ cells, the addition of TPO to the culture containing SCF resulted in an apparent production of mast cells from 3 weeks until at least 15 weeks. Some of the cells reacted with an antichymase monoclonal antibody as well. Based on the effects of growth factor(s) on a later phase of the mast cell growth, TPO may stimulate an early stage of mast cell development in combination with SCF, whereas subsequent growth seems to be supported by SCF alone. Single-cell culture studies indicated that the CD34(+)CD38(-)c-kit+ cells and CD34(+)CD38(+)c-kit+ cells were responsible for the SCF + TPO-dependent mast cell production. Two-step culture assays clearly showed that mast cells originated from multilineage colony-forming cells that had potential to differentiate into neutrophil/mast cell lineages, neutrophil/macrophage/mast cell lineages, or neutrophil/macrophage/mast cell/erythroid lineages. These results suggest that TPO plays an important role in the development of human mast cells from CD34(+) BM cells in concert with SCF, and provide direct evidence of the differentiation into the mast cell lineage of human multipotential BM-derived progenitors.
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PMID:Thrombopoietin augments stem cell factor-dependent growth of human mast cells from bone marrow multipotential hematopoietic progenitors. 1033 77

In the present study, we attempted to clarify the effects of interleukin-6 (IL-6) on the growth and properties of human mast cells using cultured mast cells selectively generated by stem cell factor (SCF) from CD34(+) cord blood cells. The addition of IL-6 to cultures containing mast cells resulted in a substantial reduction of the number of progenies grown by SCF in the liquid culture. This IL-6-mediated inhibition of mast cell growth may be due in part to the suppression at the precursor level, according to the results of a clonal cell culture assay. Moreover, a flow cytometric analysis showed that the cultured mast cells grown in the presence of SCF+IL-6 had decreased c-kit expression. The exposure of cultured mast cells to SCF+IL-6 also caused substantial increases in the cell size, frequency of chymase-positive cells, and intracellular histamine level compared with the values obtained with SCF alone. The flow cytometric analysis showed low but significant levels of expression of IL-6 receptor (IL-6R) and gp130 on the cultured mast cells grown with SCF. The addition of either anti-IL-6R antibody or anti-gp130 antibody abrogated the biological functions of IL-6. Although IL-4 exerted an effect similar to that of IL-6 on the cultured mast cells under stimulation with SCF, the results of comparative experiments suggest that the two cytokines use different regulatory mechanisms. Taken together, the present findings suggest that IL-6 modulates SCF-dependent human mast cell development directly via an IL-6R-gp130 system.
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PMID:Interleukin-6 directly modulates stem cell factor-dependent development of human mast cells derived from CD34(+) cord blood cells. 1039 17


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