Basophilic Mast Cell Research Articles

Single-cell molecular profiling provides a high-resolution map of basophil and mast cell development.

BackgroundBasophils and mast cells contribute to the development of allergic reactions. Whereas these mature effector cells are extensively studied, the differentiation trajectories from hematopoietic progenitors to basophils and mast cells are largely uncharted at the single‐cell level.MethodsWe performed multicolor flow cytometry, high‐coverage single‐cell RNA sequencing analyses, and cell fate assays to chart basophil and mast cell differentiation at single‐cell resolution in mouse.ResultsAnalysis of flow cytometry data reconstructed a detailed map of basophil and mast cell differentiation, including a bifurcation of progenitors into two specific trajectories. Molecular profiling and pseudotime ordering of the single cells revealed gene expression changes during differentiation. Cell fate assays showed that multicolor flow cytometry and transcriptional profiling successfully predict the bipotent phenotype of a previously uncharacterized population of peritoneal basophil‐mast cell progenitors.ConclusionsA combination of molecular and functional profiling of bone marrow and peritoneal cells provided a detailed road map of basophil and mast cell development. An interactive web resource was created to enable the wider research community to explore the expression dynamics for any gene of interest.

Temporal and Spatially Regulated Oncogenic KIT Expression and Loss of Dnmt3a Cooperate to Drive MPN Development: Role of PI3Kinase in Dnmt3a Loss Induced Hyperproliferation in Myeloid Cells

Recent studies, including studies from our group have shown that somatic mutations in DNA methyltransferase type 3A (DNMT3A) are frequently associated with systemic mastocytosis (SM), myelodysplastic syndrome and AML. Importantly, the presence of this mutation is associated with poor prognosis and overall patient survival. We have shown that approximately 12% of patients with SM possess DNMT3A mutations, which are commonly co-expressed with an activating mutation of KIT (KITD816V in humans and KITD814V in mice). However, it is unclear how DNMT3A mutations contribute to myeloid lineage derived mast cell growth and differentiation either alone or in conjunction with KITD816V. Further, the mechanisms by which loss of DNMT3A impairs various mast cell functions either alone or in cooperation with KITD816V are poorly understood. Utilizing mice conditionally deficient in Dnmt3a in the hematopoietic compartment, we show that loss of Dnmt3a in vivo results in increased numbers of mast cells relative to controls. In vitro, loss of Dnmt3a results in accelerated and enhanced differentiation of mast cells from its bone marrow (BM) precursors. Previous studies have shown that the level of expression of the transcription factor, Microthalamia (MITF) (high MITF expression drives mast cell commitment) and CEBPα (high C/EBPα drives basophil commitment) in Basophil-Mast cell common progenitor (BMPs) drives the fate of these cells towards either basophilic lineage or mast cell lineage. To assess why loss of Dnmt3a results in increased mast cell lineage commitment, we performed RNAseq analysis and found elevated expression of MITF and a profound repression in the expression of C/EBPα in BM precursors. Importantly, restoring the expression of Dnmt3a in Dnmt3a deficient BM cells, completely restored enhanced differentiation, along with the expression of MITF and C/EBPα. To our surprise, no appreciable differences in the expression of Gata-1, Gata-2 or PU.1 were noted in the absence of Dnmt3a relative to controls. These results demonstrate that early loss of C/EBPα in Dnmt3a null cells contributes to enhanced maturation of mast cells from their precursors. We next assessed the growth and survival potential of Dnmt3a mast cells and found a significant increase in SCF-mediated growth of these cells compared to controls. Biochemical analysis revealed greater PI3Kinase activation in Dnmt3a null cells, and these findings were supported by genome wide transcriptome analysis involving Ingenuity pathway analysis (IPA) and Gene set enrichment analysis (GSEA). Importantly, mast cells derived from Dnmt3a null BM lacking the expression of p85α regulatory subunit of PI3Kinase (Dnmt3a-/-:p85α-/-) or pharmacologic inhibition of PI3Kinase completely corrected SCF induced hyper proliferation in these cells to near normal levels. The increase in PI3Kinase activity in Dnmt3a null cells was associated with the loss of PTEN expression. To understand how loss of Dnmt3a and expression of an oncogenic form of KIT cooperate to drive MPN, we generated mice in which the expression of oncogenic KIT was temporally and spatially regulated in a cell type and tissue specific manner using the Mx-Cre system along with the expression of Dnmt3a. Consistent with earlier studies, we observed that KitD814V/+ mutation alone only resulted in the development of lymphoid neoplasms. In contrast, Dnmt3a-/- :KitD814V/+ mice manifested robust signs of MPN development including splenomegaly with increased numbers of neutrophils and monocytes in the peripheral blood associated with a significant reduction in lymphoid cells including B cells. Importantly, we noted a 5-fold increase in the fraction of LSKs and GMPs in the BM of Dnmt3a-/- :KitD814V/+ mice compared to controls and consequently an increase in the presence of mature Gr-1 and Mac-1 positive myeloid cells in the BM, spleen and in the PB of these mice relative to controls. The observed MPN was more aggressive in Dnmt3a-/- :KitD814V/+ mice compared to Dnmt3a+/- :KitD814V/+ mice and of stem cell origin as transplantation of BM cells from these mice recapitulated the development of MPN in recipient mice, similar to that observed in the original animals. The impact of PI3Kinase inhibition on mitigating oncogenic KIT and loss of Dnmt3a induced MPN and overall survival will be discussed. DisclosuresNo relevant conflicts of interest to declare.

Resveratrol suppresses type 2 cytokine induction in basophilic mast cell and Th2 cell infiltration in PCA mice

Cytokines released from mast cells, may play roles to initiate and maintain allergic responses. The high amount of cytokines accumulation can excite the late‐phase allergic responses, and it happens in consistence with migration of other leukocytes including neutrophils, lymphocytes, eosinophils and macrophages. Therefore, this study was conducted to test the resveratrol effects on IgE‐mediated basophilic mast cell and passive cutaneous anaphylaxis (PCA) in mice <em style=”mso‐bidi‐font‐style: normal;”>via cytokine suppression. Basophilic mast cells were sensitized with anti‐dinitrophenyl‐immunoglobulin E (anti‐DNP‐Ig E) and then pre‐treated with 1‐25 uM resveratrol depending on treatments. The DNP‐human serum albumin antigen was subsequently stimulated and all cells were then sampled for analysis. Similar <em style=”mso‐bidi‐font‐style: normal;”>in vivo experiment was conducted due to oral administration of 10 mg/kg resveratrol to BALB/c mouse (n = 5). Resveratrol diminished the secretion of interleukin (IL)‐3, IL‐4, IL‐13 and tumor necrosis factor(TNF)‐α in anti‐DNP‐IgE‐sensitized cells. Oral administration of 10 mg/kg resveratrol also reduced serum IL‐3, IL‐4, IL‐13 and TNF‐α level, on the contrary high level of IL‐3, ‐4 and ‐13 and TNF‐a were determined when BALB/c were sensitized and then challenged to DNP‐HSA. All DNP‐HSA mouse had high Tyrosine kinase (Tyk) 2 and Signal Transducer and Activator of Transcription (STAT). Resveratrol alleviated T cell infiltration of ear and dorsal skin in mice. In summary, the results indicated that the resveratrol administration lightened Type 2 cytokine releases and then alleviated the cytokine‐mediated allergic responses.

Ectopic Expression of Lin28 Biases Myeloid Differentiation to Promote an Immature Mast Cell State and Is Implicated in Aggressive Mastocytosis

Dysregulated mast cell development leads to systemic mastocytosis, a clinically variable but often devastating family of hematologic disorders. Lin28 is a heterochronic gene and pluripotency factor implicated in many types of malignancy, and prior studies suggest that Lin28 expression can restore a fetal hematopoietic program in adult mice. However, the role of Lin28 in hematologic malignancy remains controversial. In our study, we induced expression of Lin28 in adult mice using a doxycycline-responsive transgenic system. Lin28 induction caused marked mast cell accumulation in target organs such as the skin and peritoneal cavity. In vitro assays revealed a skewing of myeloid commitment in Lin28-expressing hematopoietic progenitors, with increased levels of Lin28 in common myeloid progenitors and basophil-mast cell progenitors altering gene expression patterns to favor cell fate choices that enhance mast cell specification. In addition, Lin28-induced mast cells appeared phenotypically and functionally immature, and in vitro assays suggested a slowing of mast cell terminal differentiation in the context of Lin28 upregulation. Finally, interrogation of human mast cell leukemia samples revealed upregulation of LIN28 in abnormal mast cells from patients with aggressive systemic mastocytosis (ASM). This work identifies Lin28 as a novel regulator of innate immune function and a new protein of interest in mast cell disease. DisclosuresNo relevant conflicts of interest to declare.

Resveratrol Inhibits IgE-Mediated Basophilic Mast Cell Degranulation and Passive Cutaneous Anaphylaxis in Mice

AbstractResveratrol is a phytoalexin abundantly found in red grape skin and is effective in antitumor and antiinflammation associated with immune responses. This study investigated whether resveratrol suppressed immunoglobulin (Ig)E-mediated allergic responses and passive cutaneous anaphylaxis (PCA) in rat RBL-2H3 mast cells and in BALB/c mice. The release of β-hexosaminidase and histamine was enhanced in mast cells sensitized with anti-dinitrophenyl (DNP)-IgE and subsequently stimulated by DNP-human serum albumin (HSA), indicative of mast cell degranulation. When mast cells were pretreated with nontoxic resveratrol at 1–25 μmol/L, such induction was dose dependently diminished. Spleen tyrosine kinase (Syk) and phospholipase Cγ (PLCγ) of sensitized mast cells were activated by stimulation with DNP-HSA antigen, which was dampened by ≥5 μmol/L resveratrol. The phosphorylation of protein kinase C (PKC)μ and PKCθ was attenuated by administering resveratrol to DNP-HSA–exposed mast cells, whereas quiescent PKCζ/λ in sensitized cells was dose-dependently activated by resveratrol. Male BALB/c mice were sensitized for 24 h with DNP-IgE and orally administered with resveratrol 1 h before the DNP-HSA challenge. The histamine concentration was enhanced in sensitized mice challenged to DNP-HSA, which was reversed by administration of 10 mg/kg resveratrol. Additionally, it encumbered the tissue activation of Syk, PLCγ, and PKCμ in antigen-exposed mice. Resveratrol decreased IgE-mediated PCA and alleviated allergic edema of mouse ear and dorsal skin. Mast cell degranulation and allergic inflammation, accompanying the induction of monocyte chemotactic protein-1 and macrophage inflammatory protein-2, were inhibited by supplementing resveratrol to antigen-challenged mice. Resveratrol inhibited mast cell-derived, immediate-type allergic reactions, and these responses of resveratrol suggest possible therapeutic strategies in preventing allergic inflammatory diseases.

Dengue virus selectively induces human mast cell chemokine production.

Severe dengue virus infections usually occur in individuals who have preexisting anti-dengue virus antibodies. Mast cells are known to play an important role in host defense against several pathogens, but their role in viral infection has not yet been elucidated. The effects of dengue virus infection on the production of chemokines by human mast cells were examined. Elevated levels of secreted RANTES, MIP-1alpha, and MIP-1beta, but not IL-8 or ENA-78, were observed following infection of KU812 or HMC-1 human mast cell-basophil lines. In some cases a >200-fold increase in RANTES production was observed. Cord blood-derived cultured human mast cells treated with dengue virus in the presence of subneutralizing concentrations of dengue virus-specific antibody also demonstrated significantly (P < 0.05) increased RANTES production, under conditions which did not induce significant degranulation. Chemokine responses were not observed when mast cells were treated with UV-inactivated dengue virus in the presence or absence of human dengue virus-specific antibody. Neither antibody-enhanced dengue virus infection of the highly permissive U937 monocytic cell line nor adenovirus infection of mast cells induced a RANTES, MIP-1alpha, or MIP-1beta response, demonstrating a selective mast cell response to dengue virus. These results suggest a role for mast cells in the initiation of chemokine-dependent host responses to dengue virus infection.

Lymphatic mast cell response and effect of compound 48/80 on popliteal lymph node reaction in rats following intracutaneous injection ofStaphylococcus aureus

To investigate the significance of mast cells in the popliteal lymph node during the development of an inflammatory response, rats were inoculated with 12 x 10(7) colony-forming units of Staphylococcus aureus in the hind foot pad. Numerical changes in mast cells were then measured in the corresponding popliteal lymph node. Six days after inoculation, despite the enlargement of the responding lymph node, a marked decrease in granulated mast cell number, relative to the contralateral node, was observed in the cortical and medullary compartments. Popliteal lymph nodes from rats treated with compound 48/80 and then inoculated with S. aureus showed a higher cortical and medullary hypertrophic response and a significant increase in degranulated/weakly basophilic mast cell number in the lymph node tissue. The findings suggest that (1) Staphylococcus aureus induces a reduction in granulated mast cell number in the cortical and medullary compartments of regional lymph nodes; (2) pretreatment with compound 48/80 appears to contribute to the lymphoid cell proliferation and the hypertrophic response of lymph nodes induced by S. aureus; and (3) granulated mast cells have a regulatory role on lymphoid cell proliferation.

Progressive inactivation of the expression of an erythroid transcriptional factor in GM- and G-CSF-dependent myeloid cell lines

The transcriptional binding protein NFE-1 (also called GF-1 and Ery-f1) is thought to play a necessary, but not sufficient, role in the regulation of differentiation-related gene expression in a subset of hematopoietic lineages (erythroid, megakaryocytic, and basophil-mast cell). In order to clarify the mechanism which underlies the lineage-specificity of the NFE-1 expression, as well as the relationship between the expression of this factor and growth factor responsiveness, we have evaluated the capacity of erythropoietin (Epo)-, granulomonocytic (GM)-colony stimulating factor (CSF)-, and granulocyte (G)-CSF-dependent subclones derived from the interleukin 3 (IL-3)-dependent cell line 32D, to express 1) NFE-1 mRNA, 2) NFE-1-related nuclear proteins, and 3) chloramphenicol acetyl transferase (CAT) activity when transfected with a CAT gene under the control of NFE-1 cognate sequences. NFE-1 mRNA was found to be expressed not only in cells with mast cell (IL-3-dependent 32D) and erythroid (Epo-dependent 32D Epo1) phenotypes, but also in cells with predominantly granulocyte/macrophage properties, such as the GM-CSF- (early myelomonocytic) and G-CSF- (myelocytic) dependent subclones of 32D. However, a gradient of expression, correlating with the lineage, the stage of differentiation, and the growth factor responsiveness of the cell lines, was found among the different subclones: Epo greater than or equal to IL-3 greater than GM-CSF greater than G-CSF. Binding experiments demonstrated NFE-1 activity in all cell lines except the G-CSF-dependent line. Function of the NFE-1 protein was assessed by the expression of the CAT gene linked to the SV40 promoter and a mutant (-175 T----C) HPFH gamma-globin promoter. High level CAT expression was seen only in the Epo1 cells although low level expression was also seen in the parent 32D. These results demonstrate that the specificity of the expression of NFE-1 for the erythroid--megakaryocytic--mast cell lineages is obtained by progressive inactivation of its expression in alternative lineages.

Splenic accumulation of interleukin-3-dependent hematopoietic cells in Friend erythroleukemia.

Interleukin-3-dependent hematopoietic stem cells commonly accumulate in spleens of mice infected with leukemia viruses. To study their origins, a molecularly tagged helper-free Friend spleen focus-forming virus was used to produce erythroleukemias. Uninfected interleukin-3-dependent basophil-mast cell progenitors coproliferated amidst the spleen focus-forming virus-infected leukemic cells. Splenic proliferation of normal stem cells is apparently a host response to leukemogenesis, and we propose that it may contribute to certain retroviral diseases.

Avian erythroblastosis virus transforms a novel mast cell-basophil precursor target in the Japanese quail

Hematopoietic cells of the Japanese quail were transformed by avian erythroblastosis virus in vivo and in vitro. In both circumstances, the infected hematopoietic tissues exhibited a dual oncogenic response of erythroid and mast cell-basophil elements. The erythroid transformants escaped the avian erythroblastosis virus block in differentiation and progressed to hemoglobinization. Resulting basophilic cells were morphologically, biochemically, and ultrastructurally identical to mast cell-basophils observed in other species. None of the virally transformed cells actively produced reverse transcriptase activity. Nonproducer cell lines synthesized viral RNA and both v-erbA and v-erbB proteins. These results indicate that the Japanese quail has a viral target cell different from that of the chicken. The implications of a single bipotential transformation target yielding both erythroid and mast cell-basophil colonies is discussed.

Basophil mast cell and eosinophil growth and differentiation factors in human allergic disease.

Clinical & Experimental AllergyVolume 19, Issue 3 p. 249-254 Basophil mast cell and eosinophil growth and differentiation factors in human allergic disease J. A. DENBURG, Corresponding Author J. A. DENBURG Department of Medicine, McMaster University, Ontario, CanadaDr Judah A. Denburg, Department of Medicine, McMaster University Medical Centre, Room 4H21—1200 Main Street West, Hamilton, Ontario L8N 3Z5, Canada.Search for more papers by this authorJ. DOLOVICH, J. DOLOVICH Department of Pediatrics, McMaster University, Ontario, CanadaSearch for more papers by this authorD. HARNISH, D. HARNISH Department of Pathology, McMaster University, Ontario, CanadaSearch for more papers by this author J. A. DENBURG, Corresponding Author J. A. DENBURG Department of Medicine, McMaster University, Ontario, CanadaDr Judah A. Denburg, Department of Medicine, McMaster University Medical Centre, Room 4H21—1200 Main Street West, Hamilton, Ontario L8N 3Z5, Canada.Search for more papers by this authorJ. DOLOVICH, J. DOLOVICH Department of Pediatrics, McMaster University, Ontario, CanadaSearch for more papers by this authorD. HARNISH, D. HARNISH Department of Pathology, McMaster University, Ontario, CanadaSearch for more papers by this author First published: May 1989 https://doi.org/10.1111/j.1365-2222.1989.tb02379.xCitations: 51AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume19, Issue3May 1989Pages 249-254 RelatedInformation

Basophil-mast cell response syndromes: a unified clinical approach.

Allergic antibodies, now known to be of the IgE class, have been associated with basophil and mast cell activation and thus with histamine release since these cells contain histamine. Common allergen induced diseases such as allergic rhinitis have been correlated with IgE antibody, basophils, mast cells and histamine release and partial control with pharmacologic use of histamine receptor blockade. This report now classifies all immediate-type reactions in reference to basophils and mast cells and activation of these cells. This approach appears particularly important in relation to those immediate-type reactions which simulate IgE mediated responses but in which no allergenic stimulus is present. The mechanisms of pharmacologic control of these basophil-mast cell response syndromes and potential research approaches are emphasized.

Human Nasal Polyp Epithelial Basophil/Mast Cell and Eosinophil Colony-stimulating Activity: The Effect is T-Cell-dependent

We have previously reported highly potent basophil/mast cell (BMC) and basophil/eosinophil (Eo) colony-stimulating activities (CSA) in conditioned medium derived from cultured human nasal polyp epithelial scrapings (NP-CM). We now have examined the involvement of peripheral blood T-cells in the NP-CM stimulation of colony-forming units (cfu) from the blood of atopic and nonatopic subjects. Because the number of BMC- and Eo-cfu was significantly higher in cultures of peripheral blood from subjects with out-of-season ragweed allergic rhinitis than from control subjects (23.8 +/- 4.1 versus 9.0 +/- 2.4, p less than 0.01), we asked whether the observed colony stimulation could be a T-cell-dependent effect. Indeed, peripheral blood target cells consisting of a reconstituted mixture of T-cells and T-cell-depleted peripheral blood mononuclear cells (non-T-cells) yielded a significantly higher number of colonies in the presence of NP-CM than the non-T-cells alone. NP-CM did not stimulate colony formation by isolated T-cells. These observations point to interactions among nasal epithelial growth and differentiation factors, blood-borne progenitors and T-cells in the local accumulation of basophils, mast cells, and eosinophils in nasal polyps.

Allergic rhinitis nasal mucosal conditioned medium stimulates growth and differentiation of basophil/mast cell and eosinophil progenitors from atopic blood

In previous studies of human nasal polyps and nasal mucosa from subjects with allergic rhinitis, we have demonstrated hematopoietic progenitors, measured as colony-forming units (CFU) in culture, and colony-stimulating activities for basophil/mast cells (BMC) and basophil/ eosinophils (Eo). In the present study, conditioned medium (CM) from short-term cultures of nasal mucosal scrapings of subjects with allergic rhinitis are compared with those from normal subjects for colony-stimulating activities, with BMC and Eo CFU in methylcellulose assays of peripheral blood from atopic (N = 6) or nonatopic (N = 6) individuals. CM derived from cultures of nasal mucosal scrapings from subjects with allergic rhinitis stimulated a significantly greater number of BMC/Eo CFU from the peripheral blood of atopic (out-of-season and ragweed-allergic) subjects than from peripheral blood of nonatopic subjects in a series of matched-control experiments (20.6 ± 4.6 versus 1.9 ± 0.8; p < 0.002); in contrast, nasal mucosal CM from normal control subjects did not stimulate an increase in these CFU in peripheral blood from atopic compared to nonatopic subjects. These studies further support the hypothesis that in situ hematopoietic mechanisms are operative in allergic inflammatory reactions and suggest the presence of cells in affected (allergic) epithelium that are “activated” to produce large amounts of BMC and Eo growth and differentiation factors.

Contribution of basophil/mast cell and eosinophil growth and differentiation to the allergic tissue inflammatory response.

The mechanisms underlying allergic tissue basophil/mast cell (BMC) or eosinophil (Eo) accumulation are unclear, especially since chemotaxis or IgE levels do not offer a sufficient explanation. We have found that a formaldehyde-blockable, steroid-responsive nasal metachromatic cell (NMC) population predominates in epithelium and correlates well with symptoms and signs in patients with allergic rhinitis. Circulating BMC and Eo progenitors (colony-forming cells in culture; CFU-c) are increased in atopic patients, inversely related to NMC counts, and fall as NMC numbers rise during seasonal allergen (ragweed pollen) stimulation. The metachromatic cell progeny of these CFU-c are also formaldehyde-blockable in their staining reaction and thus may correspond to NMC. Human nasal polyps yield BMC CFU-c. Nasal polyp epithelial scrapings or mononuclear cells, T lymphocytes or keratinocytes in vitro all produce potent BMC or Eo colony-stimulating activities (CSA) as well as an interleukin-3-like activity, each of which is partially separable from the others. Nasal epithelial cells cultured from scrapings of atopic, as opposed to nonatopic, patients also produce BMC or Eo CSA with an enhanced effect of the former on atopic peripheral blood CFU-c growth. These studies support the hypothesis that BMC and Eo accumulate in allergic inflammation as a result of in situ growth and differentiation of progenitors stimulated by soluble hemopoietic factors derived from mucosal cell populations.

ヒト好塩基球-肥満細胞系から放出されるproteoglycanの抗凝固活性に関する研究

ヒト好塩基球-肥満細胞系から放出されるproteoglycanの抗凝固活性に関する研究

ヒト好塩基球-肥満細胞系から放出されるproteoglycanの抗凝固活性に関する研究

ヒト好塩基球-肥満細胞系から放出されるproteoglycanの抗凝固活性に関する研究

Murine basophil-mast differentiation: toward optimal conditions for selective growth and maturation of basophil-mast or allied cells.

Recent investigations revealed that basophil-mast cells were related to the hemopoietic system. Strikingly, murine bone marrow showed a singular paucity in cells with basophil-mast features; moreover in clonogenic assays (methylcellulose, agarose) bone marrow was found to be manifestly poor in basophil-mast progenitor cells. Our work brought to light several new facts concerning the culture and differentiation of this cell type: 1 degree pure and mixed mast clones can be derived in large numbers from bone marrow, provided progenitors are cultured in collagen matrix. Up to 1,382 hemopoietic clones were analysed in situ after staining: 30% contained mast cells (34 per 10(5) cells), thus the basophil-mast lineage was one of the most frequent. We concluded that other cloning media were noticeably nonoptimal for the growth and/or maturation of mast cells. We suggested that collagen and the molecular edifices derived from it, both found in variable amounts in the natural mast environments, should play essential roles in mast phenotype expression. 2 Degrees cholera toxin (CT) selectively eradicated nonmast progenies: mast progenitors and mast progenies were resistant. In this way, pure and rapidly expanding mast cell clones were obtained at a frequency never reported before. CT possibly acts both directly, as a stimulator of mast cell proliferation, or indirectly on marrow subpopulations which repress basophil-mast cell growth and maturation. In vitro culture conditions, specifically designed for basophil-mast lineage, should prove of interest in the search for an unifying hypothesis concerning the multiple forms of mast cells found in various tissues.

Natural cytotoxic (NC) cell activity in basophilic cells: release of NC-specific cytotoxic factor by IgE receptor triggering.

A murine interleukin 3 (IL 3)-dependent basophilic mast cell line, PT-18 (A17), and a rat basophilic leukemic cell line, RBL-2H3, were shown to be capable of selective natural cytotoxic (NC) but not natural killer (NK) cell activity. The basophilic cell types could also be augmented in their NC activity by bridging of their surface IgE receptors. IgE-mediated triggering of the basophilic cells was accomplished by coating the cells with IgE and exposing the IgE-bound cells to specific antigen or to anti-IgE monoclonal antibody. Another method of triggering was by direct binding of basophilic cells to anti-IgE receptor monoclonal antibody. Basophilic cells, triggered by these methods, not only displayed increased NC activity but also released a soluble factor capable of selectively lysing NC tumor targets, WEHI-164, but not three of the NK-sensitive targets, YAC-1, RLM1, and RBL-5. Normal C3H/HeJ mouse embryonic fibroblasts were also not lysed. Dose response and time course of the cytotoxic factor release from triggered RBL-2H3 cells were similar to those of tritiated serotonin release. As with serotonin or histamine release, the NC-specific cytotoxic factor (NCCF) was not released in the absence of extracellular calcium. Therefore, NCCF appears to be released along with other mediators during the triggering of basophilic cells by bridging of IgE receptors. The m.w. of the native form of this factor, determined by a gel filtration method, was about 43,000.

Basophil/Mast Cell Precursors in Human Peripheral Blood

Semisolid (methylcellulose) hemopoietic cultures revealed the presence of histamine-containing granulocyte colonies derived from precursors (CFU-C) present in human peripheral blood. Light microscopy and histochemical studies of cells in individual histamine-containing colonies demonstrated homogeneous populations of metachromatic basophil/mast cells (BMC) at various stages of maturation. By inverted microscopy, pure BMC colonies were more often found to have the overall appearance of the previously described "eosinophil" (type II), rather than "neutrophil-macrophage" (type I), colony type. Histamine-positive colonies constituted 58% (50/86) of all (type I and type II) granulocyte colonies in repeated cultures from a patient with systemic mastocytosis (SM), and 19% (13/67) of colonies in cultures from 8 patients with chronic myeloid leukemia (CML); this was in contrast to 8% (12/153) of colonies in cultures from 4 patients with urticaria pigmentosa (UP) and 6 normal controls (p less than 0.0001). Calculated frequency of BMC CFU-C was approximately 1 per 2 X 10(6) in normal and 1 per 2 X 10(5) nucleated cells in SM peripheral blood. Taking colony size into account, histamine content per cell in histamine-positive type II colonies in SM cultures was 1.1 +/- 0.19 pg, compared to 0.29 +/- 0.08 pg in CML and less than or equal to 0.10 in normals and UP. Electron microscopy (EM) of individual colonies revealed electron-dense granules with ultrastructural features of BMC in histamine-positive, but not histamine-negative, colonies. Use of these methods may help to further clarify the nature of BMC precursors and the regulation of their proliferation in bone marrow disorders and allergic states.