Developmentally regulated cell surface expression and function of c-kit receptor during lymphocyte ontogeny in the embryo and adult mice

Development ◽  
1992 ◽  
Vol 115 (4) ◽  
pp. 1133-1147 ◽  
Author(s):  
R. Palacios ◽  
S. Nishikawa
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
B Lymphocyte ◽  
Fetal Liver ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Lymphocyte Development ◽  
Fetal Thymus ◽  
Kit Receptor ◽  
B Lymphocyte Development

We have used a c-kit-specific monoclonal antibody, immuno-fluorescence staining and flow fluorocytometry or microscopy analysis to assess the cell surface expression of the c-kit receptor on a panel of non-transformed clones representing different stages of T- and B-lymphocyte development, freshly isolated lymphoid cells from thymus, bone marrow and spleen of young adult C57BL/6 mice and cells from yolk sac, thymus and liver of developing C57BL/6 mouse embryos. Pro-T, Pro-B and Pre-B clones derived from thymus or liver of 14-day embryos are c-kit+. Starting at day 8 to 8.5 in yolk sac, day-10 in fetal liver, and day 11 to 12 in fetal thymus, there are many c-kit+ cells. The number of c-kit+ cells in liver and thymus increases up to day 15 and progressively decreases thereafter. Cell sorter purified c-kit+ day 14 fetal liver cells fully reconstitute the T and B cell compartments of immunodeficient Scid mice. Stromal cells or epithelial cells derived from fetal thymus or liver, which can support growth and differentiation of c-kit+ lymphocyte progenitor clones, synthesize mRNA for Steel Factor (SF), the ligand of c-kit. In the adult mouse, however, c-kit expression is restricted to very early stages of T- and B-lymphocyte development (multipotent progenitors, B-cell/myelocytic progenitors, Pro-T and Pro-B lymphocyte progenitors). Most cells at the Pre-T, Pre-B and later stages of development do not bear detectable c-kit. Using Cos-1 cells transfected with mouse SF-cDNA and an antagonistic c-kit receptor-specific antibody, we show that the c-kit/SF system contributes to the survival of lymphocyte progenitors and enhances the proliferative responses of these cells to other growth factors (i.e. IL2, IL3, IL4, IL7). However, the c-kit receptor/SF ligand pair is neither sufficient nor necessary for the differentiation of lymphocyte progenitors into mature T- or B-lymphocytes. Finally, in stromal cell lines from fetal liver and adult bone marrow and thymic epithelial cell lines the level of steady state SF-RNA transcripts is inversely correlated with that of IL-7-mRNA. Moreover, IL7 inhibits the synthesis of SF-mRNA in stromal cells and rIL6 abrogates this inhibitory effect of rIL7. Thus, the expression of SF in stromal cells is subjected to complex regulation by other cytokines produced by the same stromal cells or by neighboring cells in a given microenvironment.(ABSTRACT TRUNCATED AT 400 WORDS)

STAT5 Has Tumor Suppressor-Like Activity in a Murine Model of Myc-Initiated Acute B-Lymphoblastic Leukemia/Lymphoma

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 919-919 ◽  
Author(s):  
Zhengqi Wang ◽  
Geqiang Li ◽  
Zizhen Kang ◽  
Silvia T Bunting ◽  
William Tse ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cell ◽  
B Lymphocyte ◽  
Fetal Liver ◽  
Lymphoblastic Leukemia ◽  
Conditional Knockout ◽  
Lymphocyte Development ◽  
Wild Type ◽  
B Lymphocyte Development ◽  
B Lymphoid

Abstract Abstract 919 Signal transducer and activator of transcription 5 (STAT5) is a critical regulator of normal and leukemic lympho-myeloid hematopoiesis through activation downstream of early-acting cytokines, their receptors, and janus kinases (JAKs). Despite upstream activating mutations driving JAK-STAT phosphorylation in precursor-B acute lymphoblastic leukemia (B-ALL), activated JAK-STAT is absent from the aggressive “double hit” lymphomas expressing myc and bcl-2. Using C57BL/6 background transgenic mouse models for myc and bcl-2, we set out to determine whether endogenous STAT5 functions in guarding against B-ALL induced by combined myc/bcl-2 or myc alone. We first determined whether constitutive expression of bcl-2 driven from the H2K promoter and Moloney murine leukemia virus enhancer in C57BL/6 background STAT5-deficient hematopoietic cells could bypass blocks in B-lymphocyte development. Transgenic H2K/bcl-2 expression in hypomorphic STAT5abDN/DN mice, which are leaky and still produce some mature B-lymphocytes, largely rescued peripheral B-lymphocyte survival to near normal levels but could only rescue about 10% of the multilineage hematopoietic stem cell (HSC) competitive repopulating defect. Complete deletion of the entire STAT5ab locus resulted in the expected severe block of B-cell development at the pre-pro-B-cell stage following transplantation of STAT5ab null/null fetal liver cells into irradiated wild type or common γC−/− recipients. Peripheral B-lymphocyte development could not be restored by transgenic bcl-2 alone in the absence of STAT5. However, transgenic myc driven from an immunoglobulin H chain enhancer (Emu/myc) combined with H2K/bcl-2 induced B-ALL peripheral counts as high as 1.1 × 105 B-cells/ul and reduced latency (a median survival of 44 days) compared to wild-type control (a median survival of 91 days) in either lethally-irradiated (P<0.001; N range from 8–14 mice/group) or sub-lethally-irradiated cohorts of fetal liver transplanted mice (P=0.007; N range 10–20 mice/group). B-ALL in mice with or without STAT5 was a mix of Pro-B and Pre-B ALL (IgM-CD43+B220+CD19+/−CD4+/−) and morphologically similar in the spleen and bone marrow. Multi-parameter flow cytometry analysis of bone marrow cells from STAT5ab null/null fetal liver transplanted mice (N=4) showed that deletion of STAT5 significantly reduced by 11.5-fold (P=0.004) the fraction of long-term repopulating HSC (CD150+CD48-) c-Kit+Lin-Sca-1+ (KLS). In an independent adult Mx1-Cre conditional knockout of STAT5 by pI:pC treatment, lymphomas induced by Myc alone were also accelerated (P=0.05; N range 14–15 mice/group) with STAT5 maintained deleted in sorted B-cells. These mice also had reduced CD150+CD48- KLS cells (5.6-fold; N=4; P=0.006). Interestingly, several phenotypes recently reported as associated with increased HSC cycling and lymphoid-biased differentiation were observed. The mean fluorescence intensity of slamf1 (CD150) was reduced 2.2-fold (P<0.001; N=4) in conditional knockout mice and the B-lymphoid biased CD48+CD150+ or CD48-CD150- KLS cells representing short-term HSC/multipotent progenitors were not significantly reduced. Microarray analyses of the KLS fraction provided evidence that STAT5 promotes HSC maintenance and myeloid potential (limiting lymphoid commitment, cycling) in the KLS compartment. The deletion of STAT5 reduced expression of HSC self-renewal and quiescence promoting genes and increased immunoglobulin and B-lymphoid transcripts. Combined with the pre-pro-B-cell block, loss of STAT5 promotes accumulation of B-lineage committed progenitors as potential ALL initiating cells. The effects of bcl-2 and myc hits on STAT5 null/null hematopoietic cells are currently being further characterized with respect to B-cell developmental blocks and molecular heterogeneity. B-ALL has a high relapse rate and is driven by clonally diverse tumor propagating populations. Our work may have important implications for ALL drug therapy. In conclusion, we demonstrate that STAT5, considered primarily as functioning like an oncogene in hematologic malignancies upon persistent activation, can play a tumor suppressor-like role in subsets of B-ALL. These data add to an emerging understanding that endogenous STAT5 can suppress some cancers and transcriptionally regulate several cell cycle inhibitors. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Flow cytometric analysis of human bone marrow. II. Normal B lymphocyte development

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1316-1324 ◽  
Author(s):  
MR Loken ◽  
VO Shah ◽  
KL Dattilio ◽  
CI Civin
Keyword(s):  
Bone Marrow ◽  
Cell Surface ◽  
B Lymphocyte ◽  
Human Bone ◽  
Lymphocyte Development ◽  
Hla Dq ◽  
B Lymphocyte Development ◽  
B Lineage ◽  
B Lymphoid ◽  
B Lineage Cells

Abstract A panel of B lymphoid-reactive monoclonal antibodies was used to analyze the phenotypic changes that accompany B lymphocyte development in normal human bone marrow. The B lymphoid cells were identified using light scattering and the expression of CD19 on a flow cytometer. Quantitative three-color immunofluorescence was then used to correlate other cell surface antigens on these cells identified as B lymphoid in normal marrow. CD10 and CD20 identified almost exclusive populations and provided a convenient means of discriminating between the less and more mature B lineage cells. The CD10+ cells could be further subdivided using CD34. The population of CD19+, CD10+, CD34+ cells comprised only 0.6% of marrow cells, but these contained the majority of terminal deoxynucleotidyl transferase (TdT+) cells. In the assessment of class II antigens, HLA-DR was expressed on all B lineage cells whereas HLA-DP preceded HLA-DQ in appearance during the developmental process. Among the later antigens expressed on B lineage cells, cell surface IgM, CD20, and HLA-DQ were expressed at essentially the same time. Cell surface CD10 was lost at the time when CD21 and CD22 were acquired on the cell surface. These data illustrate that multiparameter flow cytometry can be used to define a continuous progression of stages of B lymphocyte development based on cell surface antigen expression even though these cells represent a minor fraction of normal marrow cells.

Two Pathways of B-Lymphocyte Development in Mouse Bone Marrow and the Roles of Surrogate L Chain in this Development

1994 ◽  
Vol 137 (1) ◽  
pp. 185-201 ◽  
Author(s):  
Antonius Rolink ◽  
Hajime Karasuyama ◽  
Dirk Haasner ◽  
Ulf Grawunder ◽  
Inga-Lill Martensson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Lymphocyte ◽  
Lymphocyte Development ◽  
Mouse Bone Marrow ◽  
B Lymphocyte Development

scholarly journals Delay of Early B-Lymphocyte Development by Gamma 2b Immunoglobulin Transgene: Effect on Differentiation-Specific Molecules

1990 ◽  
Vol 1 (2) ◽  
pp. 105-112 ◽  
Author(s):  
Kathleen A. Denis ◽  
Scott Provost ◽  
Owen N. Witte ◽  
Ralph L. Brinster ◽  
Ursula Storb
Keyword(s):  
Bone Marrow ◽  
Gene Rearrangement ◽  
B Lymphocyte ◽  
Bone Marrow Cells ◽  
Leukemia Virus ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Lymphocyte Development ◽  
B Cell Differentiation ◽  
B Lymphocyte Development

Mice transgenic forγ2b Ig heavy chain were examined for alterations in B-cell differentiation and endogenous Ig gene rearrangement and expression. Fresh bone marrow from these mice was markedly reduced in BP-1+cells and there were small reductions in B220+and sIg+cells. A-MuLV (Abelson murine leukemia virus) transformants from these bone marrow cells showed little alteration in Ig gene rearrangement and expression when compared to controls, however. Isolation of the B-lymphoid compartment from these mice in vitro using LBMC (lymphoid bone marrow cultures) enabled more detailed characterization of the effects of the transgene. LBMC derived fromγ2b transgenic mice had similar growth kinetics, but a 4-5-week delay in the expression of endogenous mu Ig in comparison to control cultures. Nucleic acids derived from these early cultures prior to endogenous mu Ig expression showed reduced Ig JHrearrangements, some sterile mu transcription, low levels of BP-1 expression, and virtually undetectable TdT (terminal deoxynucleotidyl transferase) expression. Thus, thisγ2b transgene appears able to affect early B-lymphocyte development.

Bone Marrow B Lymphocyte Development in c-abl-Deficient Mice

1995 ◽  
Vol 165 (1) ◽  
pp. 44-54 ◽  
Author(s):  
Jeff D. Hardin ◽  
Sharon Boast ◽  
Pamela L. Schwartzberg ◽  
Grace Lee ◽  
Fred W. Alt ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Lymphocyte ◽  
Lymphocyte Development ◽  
B Lymphocyte Development ◽  
Deficient Mice

scholarly journals Flow cytometric analysis of human bone marrow. II. Normal B lymphocyte development

Blood ◽  
1987 ◽  
Vol 70 (5) ◽  
pp. 1316-1324 ◽  
Author(s):  
MR Loken ◽  
VO Shah ◽  
KL Dattilio ◽  
CI Civin
Keyword(s):  
Bone Marrow ◽  
Cell Surface ◽  
B Lymphocyte ◽  
Human Bone ◽  
Lymphocyte Development ◽  
Hla Dq ◽  
B Lymphocyte Development ◽  
B Lineage ◽  
B Lymphoid ◽  
B Lineage Cells

A panel of B lymphoid-reactive monoclonal antibodies was used to analyze the phenotypic changes that accompany B lymphocyte development in normal human bone marrow. The B lymphoid cells were identified using light scattering and the expression of CD19 on a flow cytometer. Quantitative three-color immunofluorescence was then used to correlate other cell surface antigens on these cells identified as B lymphoid in normal marrow. CD10 and CD20 identified almost exclusive populations and provided a convenient means of discriminating between the less and more mature B lineage cells. The CD10+ cells could be further subdivided using CD34. The population of CD19+, CD10+, CD34+ cells comprised only 0.6% of marrow cells, but these contained the majority of terminal deoxynucleotidyl transferase (TdT+) cells. In the assessment of class II antigens, HLA-DR was expressed on all B lineage cells whereas HLA-DP preceded HLA-DQ in appearance during the developmental process. Among the later antigens expressed on B lineage cells, cell surface IgM, CD20, and HLA-DQ were expressed at essentially the same time. Cell surface CD10 was lost at the time when CD21 and CD22 were acquired on the cell surface. These data illustrate that multiparameter flow cytometry can be used to define a continuous progression of stages of B lymphocyte development based on cell surface antigen expression even though these cells represent a minor fraction of normal marrow cells.

The characterization of distinct populations of murine skeletal cells that have different roles in B lymphopoiesis

Blood ◽  
2021 ◽  
Author(s):  
Alanna Claire Green ◽  
Gavin Tjin ◽  
Samuel C Lee ◽  
Alistair M Chalk ◽  
Lenny Straszkowski ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Leptin Receptor ◽  
B Lymphocyte ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
B Lymphopoiesis ◽  
Functional Studies

Hematopoiesis is extrinsically controlled by cells of the bone marrow microenvironment, including skeletal lineage cells. The identification and subsequent studies of distinct subpopulations of maturing skeletal cells is currently limited due to a lack of methods to isolate these cells. We found that murine Lineage-CD31-Sca-1-CD51+ cells can be divided into four subpopulations using flow cytometry, based on their expression of the platelet derived growth factor receptors ⍺ and β (PDGFR⍺ and PDGFRβ). The use of different skeletal lineage reporters confirmed the skeletal origin of the four populations. Multiplex immunohistochemistry studies revealed that all four populations were localized near the growth plate and trabecular bone and were rarely found near cortical bone regions or in central bone marrow. Functional studies revealed differences in their abundance, colony-forming unit-fibroblast capacity and potential to differentiate into mineralized osteoblasts or adipocytes in vitro. Furthermore, the four populations had distinct gene expression profiles and differential cell surface expression of leptin receptor (LEPR) and vascular cell adhesion molecule 1 (VCAM-1). Interestingly, we discovered that one of these four different skeletal populations showed the highest expression of genes involved in the extrinsic regulation of B lymphopoiesis. This cell population varied in abundance between distinct hematopoietically active skeletal sites, and significant differences in the proportions of B lymphocyte precursors were also observed in these distinct skeletal sites. It also supported pre-B lymphopoiesis in culture. Our method to isolate four distinct maturing skeletal populations will assist in elucidating the roles of distinct skeletal niche cells in regulating hematopoiesis and bone.

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