Phenotypic and functional characteristics of hematopoietic cell lineages in CD69-deficient mice

Abstract AIM/CD69 is the earliest leukocyte activation antigen and is expressed mainly by activated T, B, and natural killer (NK) cells. It is also constitutively expressed by platelets, by bone marrow myeloid precursors, and by small subsets of resident lymphocytes in the secondary lymphoid tissues. The engagement of CD69 by specific antibodies induces intracellular signals, including Ca++ flux, cytokine synthesis, and cell proliferation. To investigate the physiological relevance of CD69, we generated mice deficient in CD69 (CD69-/-) by gene targeting in embryonic stem cells. CD69 (-/-) mice showed largely normal hematopoietic cell development and normal T-cell subpopulations in thymus and periphery. Furthermore, studies of negative- and positive-thymocyte selection using a T-cell receptor transgenic model demonstrated that these processes were not altered in CD69 (-/-) mice. In addition, natural killer and cytotoxic T lymphocyte cells from CD69-deficient mice displayed cytotoxic activity similar to that of wild-type mice. Interestingly, B-cell development was affected in the absence of CD69. The B220hiIgMneg bone marrow pre-B cell compartment was augmented in CD69 (-/-) mice. In addition, the absence of CD69 led to a slight increase in immunoglobulin (Ig) G2a and IgM responses to immunization with T-dependent and T-independent antigens. Nevertheless, CD69-deficient lymphocytes had a normal proliferative response to different T-cell and B-cell stimuli. Together, these observations indicate that CD69 plays a role in B-cell development and suggest that the putative stimulatory activity of this molecule on bone marrow-derived cells may be replaced in vivo by other signal transducing receptors.

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Phenotypic and functional characteristics of hematopoietic cell lineages in CD69-deficient mice

Blood ◽

10.1182/blood.v95.7.2312.007k16_2312_2320 ◽

2000 ◽

Vol 95 (7) ◽

pp. 2312-2320 ◽

Cited By ~ 3

Author(s):

Pilar Lauzurica ◽

David Sancho ◽

Miguel Torres ◽

Beatriz Albella ◽

Mónica Marazuela ◽

...

Keyword(s):

Bone Marrow ◽

T Cell ◽

B Cell ◽

Natural Killer ◽

Embryonic Stem ◽

Cell Development ◽

B Cell Development ◽

Hematopoietic Cell ◽

Lymphoid Tissues ◽

Deficient Mice

AIM/CD69 is the earliest leukocyte activation antigen and is expressed mainly by activated T, B, and natural killer (NK) cells. It is also constitutively expressed by platelets, by bone marrow myeloid precursors, and by small subsets of resident lymphocytes in the secondary lymphoid tissues. The engagement of CD69 by specific antibodies induces intracellular signals, including Ca++ flux, cytokine synthesis, and cell proliferation. To investigate the physiological relevance of CD69, we generated mice deficient in CD69 (CD69-/-) by gene targeting in embryonic stem cells. CD69 (-/-) mice showed largely normal hematopoietic cell development and normal T-cell subpopulations in thymus and periphery. Furthermore, studies of negative- and positive-thymocyte selection using a T-cell receptor transgenic model demonstrated that these processes were not altered in CD69 (-/-) mice. In addition, natural killer and cytotoxic T lymphocyte cells from CD69-deficient mice displayed cytotoxic activity similar to that of wild-type mice. Interestingly, B-cell development was affected in the absence of CD69. The B220hiIgMneg bone marrow pre-B cell compartment was augmented in CD69 (-/-) mice. In addition, the absence of CD69 led to a slight increase in immunoglobulin (Ig) G2a and IgM responses to immunization with T-dependent and T-independent antigens. Nevertheless, CD69-deficient lymphocytes had a normal proliferative response to different T-cell and B-cell stimuli. Together, these observations indicate that CD69 plays a role in B-cell development and suggest that the putative stimulatory activity of this molecule on bone marrow-derived cells may be replaced in vivo by other signal transducing receptors.

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A Novel Molecular Complex Expressed on Immature B Cells: A Possible Role in T Cell-Independent B Cell Development

Developmental Immunology ◽

10.1155/1996/21973 ◽

1996 ◽

Vol 5 (1) ◽

pp. 67-78 ◽

Cited By ~ 5

Author(s):

Philip J. Griebel ◽

Paolo Ghia ◽

Ulf Grawunder ◽

Giorgio Ferrari

Keyword(s):

Bone Marrow ◽

Cell Death ◽

T Cell ◽

B Cells ◽

B Cell ◽

Cell Line ◽

Molecular Complex ◽

Cell Development ◽

B Cell Development ◽

Lymphoid Tissues

To identify surface molecules that may play a role in regulating ileal Peyer's patch (PP) B cell growth, we generated monoclonal antibodies (mAbs) and then selected them for a unique reactivity with ileal PP B cells. Flow cytometric analysis identified a mAb (SIC4.8R) that labeled 97% of ileal and 50–60% of jejunal PP sIgM+B cells. SIC4.8R also labeled a subpopulation of cortical thymocytes but few B or T cells in other lymphoid tissues, including bone marrow. Immunohistochemistry revealed intense SIC4.8R staining of B cells in the cortex of ileal PP follicles. SIC4.8R also labeled bovine PP B cells, a murine pro-B cell line, and pre-B cells in human bone marrow. Protein chemistry revealed that a structurally similar molecular complex was expressed on sheep ileal PP B cells and thymocytes and murine pro-B cells. Addition of soluble SIC4.8R to cultured ileal PP B cells reduced apoptotic cell death, elevated proliferative responses, partially inhibited anti-Ig-induced cell death, and induced IL-4 responsiveness. In contrast, soluble SIC4.8R had an antiproliferative effect on a mouse pro-B cell line. Finally, SIC4.8R labeling declined following the stimulation of ileal PP B cells with CD40 ligand. In conclusion, the present investigation determined that SIC4.8R identified a novel molecular complex that is expressed at several stages of T cell-independent B cell development in a variety of mammalian species. This observation confirmed that PP B cells are developmentally distinct from other B cell populations in sheep and suggested that the bone marrow may not be a site of B lymphopoiesis in young lambs.

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Altered Erythrocytes and a Leaky Block in B-Cell Development in CD24/HSA-Deficient Mice

Blood ◽

10.1182/blood.v89.3.1058 ◽

1997 ◽

Vol 89 (3) ◽

pp. 1058-1067 ◽

Cited By ~ 88

Author(s):

P.J. Nielsen ◽

B. Lorenz ◽

A.M. Müller ◽

R.H. Wenger ◽

F. Brombacher ◽

...

Keyword(s):

T Lymphocytes ◽

B Cell ◽

Embryonic Stem ◽

Cell Development ◽

B Cell Development ◽

Surface Glycoprotein ◽

Malarial Parasite ◽

Heat Stable Antigen ◽

Deficient Mice

Abstract The heat stable antigen (HSA, or murine CD24) is a glycosyl phosphatidylinositol-linked surface glycoprotein expressed on immature cells of most, if not all, major hematopoietic lineages, as well as in developing neural and epithelial cells. It has been widely used to stage the maturation of B and T lymphocytes because it is strongly induced and then repressed again during their maturation. Terminally differentiated lymphocytes, as well as most myeloid lineages, are negative for HSA. Erythrocytes are an exception in that they maintain high levels of HSA expression. HSA on naive B cells has been shown to mediate cell-cell adhesion, while HSA on antigen-presenting cells has been shown to mediate a costimulatory signal important for activating T lymphocytes during an immune response. Here, we characterize mice that lack a functional HSA gene, constructed by homologous recombination in embryonic stem cells. While T-cell and myeloid development appears normal, these mice show a leaky block in B-cell development with a reduction in late pre-B and immature B-cell populations in the bone marrow. Nevertheless, peripheral B-cell numbers are normal and no impairment of immune function could be detected in these mice in a variety of immunization and infection models. We also observed that erythrocytes are altered in HSA-deficient mice. They show a higher tendency to aggregate and are more susceptible to hypotonic lysis in vitro. In vivo, the mean half-life of HSA-deficient erythrocytes was reduced. When infected with the malarial parasite Plasmodium chabaudi chabaudi, the levels of parasite-bearing erythrocytes in HSA-deficient mice were also significantly elevated, but the mice were able to clear the infection with kinetics similar to wild-type mice and were immune to a second challenge. Thus, apart from alterations in erythrocytes and a mild block in B-cell development, the regulated expression of HSA appears to be dispensable for the maturation and functioning of those cell lineages that normally express it.

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IL-7 Effects on Thymocyte Progenitors.

Blood ◽

10.1182/blood.v106.11.3318.3318 ◽

2005 ◽

Vol 106 (11) ◽

pp. 3318-3318

Author(s):

Nahed El Kassar ◽

Baishakhi Choudhury ◽

Francis Flomerfelt ◽

Philip J. Lucas ◽

Veena Kapoor ◽

...

Keyword(s):

Bone Marrow ◽

T Cell ◽

B Cell ◽

Notch Signaling ◽

Stromal Cells ◽

T Cell Development ◽

Fold Increase ◽

Cell Development ◽

B Cell Development ◽

Over Expression

Abstract IL-7 is a non-redundant cytokine in T cell development. We studied the role of IL-7 in early T-cell development using a model of transgenic (Tg) mice with the murine IL-7 gene under control of the lck proximal promoter. At high IL-7 over-expression (x39 fold increase at day 1 in total thymic tissue), we observed a disruption of TCRαβ development along with increased B cell development in the thymus (7- to 13-fold increase) (El Kassar, Blood, 2004). In order to further explore abnormal T and B cell thymic development in these mice, we first confirmed that they both arise in parallel and were non-cell autonomous, by in vivo injection of neutralizing anti-IL-7 MAb and mixed bone marrow chimera experiments. Using a six color flow cytometry analysis, we found a dramatic decrease of the early thymocyte progenitors (ETPs, lin−CD44+CD25−c-kithiIL-7R−/lo) in the adult Tg mice (x4.7 fold decrease). Lin−CD44+CD25−c-kit+ thymocytes were sorted and cultured on OP9 and OP9 delta-like1 (OP9-DL1) stromal cells (kindly provided by Pr Zuniga Pflucker). At day 14, we observed an important decrease of T cell development (54% vs. 1% of DP cells) and an increase of NK cells (x5 fold increase) in the Tg-derived DN1 cell culture. DN2 (Lin−CD44+CD25−c-kit+) Tg thymocytes showed the same, but less dramatic abnormalities. While DN1 progenitors developed effectively into B220+CD19+ cells on OP9 stromal cells, no B cell development was observed on OP-DL stromal cells from DN1-Tg derived progenitors or by addition of increasingly high doses of IL-7 (x10, x40, x160) to normal B6-derived DN1 progenitors. Instead, a block of T-cell development was observed with increased IL-7. We hypothesized a down regulation of Notch signaling by IL-7 over-expression and analyzed by FACS Notch expression in the DN thymocytes. By staining the intra-cellular part of Notch cleaved after Notch 1/Notch ligand activation, Tg-derived DN2 cells showed decreased Notch signaling. More importantly, HES expression was decreased in the DN2, DN3 and DN4 fractions by semi-quantitative PCR. Sorted Pro/Pre B cells from Tg thymi showed TCR Dβ1-Jβ1 rearrangement indicating their T specific origin, in opposition to Pro/Pre B cells sorted from the bone marrow of the same mice. We suggest that more than one immature progenitor seeds the thymus from the bone marrow. While ETPs had T and NK proliferative capacity, another thymic progenitor with B potential may be responsible for thymic B cell development in normal and IL-7 Tg mice. Finally, IL-7 over-expression may induce a decreased Notch signaling in thymic progenitors, inducing a switch of T vs. B lineage development.

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T and B lymphocyte abnormalities in bone marrow biopsies of common variable immunodeficiency

Blood ◽

10.1182/blood-2010-11-321695 ◽

2011 ◽

Vol 118 (2) ◽

pp. 309-318 ◽

Cited By ~ 66

Author(s):

Manuella L. Gomes Ochtrop ◽

Sigune Goldacker ◽

Annette M. May ◽

Marta Rizzi ◽

Ruth Draeger ◽

...

Keyword(s):

Bone Marrow ◽

T Cell ◽

B Cell ◽

Common Variable Immunodeficiency ◽

Plasma Cells ◽

Cell Development ◽

B Cell Development ◽

Bone Marrow Biopsies ◽

Cell Counts ◽

Common Variable

Abstract In common variable immunodeficiency (CVID) defects in early stages of B-cell development, bone marrow (BM) plasma cells and T lymphocytes have not been studied systematically. Here we report the first morphologic and flow cytometric study of B- and T-cell populations in CVID BM biopsies and aspirates. Whereas the hematopoietic compartment showed no major lineage abnormalities, analysis of the lymphoid compartment exhibited major pathologic alterations. In 94% of the patients, BM plasma cells were either absent or significantly reduced and correlated with serum immunoglobulin G levels. Biopsies from CVID patients had significantly more diffuse and nodular CD3+ T lymphocyte infiltrates than biopsies from controls. These infiltrates correlated with autoimmune cytopenia but not with other clinical symptoms or with disease duration and peripheral B-cell counts. Nodular T-cell infiltrates correlated significantly with circulating CD4+CD45R0+ memory T cells, elevated soluble IL2-receptor and neopterin serum levels indicating an activated T-cell compartment in most patients. Nine of 25 patients had a partial block in B-cell development at the pre-B-I to pre-B-II stage. Because the developmental block correlates with lower transitional and mature B-cell counts in the periphery, we propose that these patients might form a new subgroup of CVID patients.

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Differential Regulation of B Cell Development, Activation, and Death by the Src Homology 2 Domain–Containing 5′ Inositol Phosphatase (Ship)

Journal of Experimental Medicine ◽

10.1084/jem.191.9.1545 ◽

2000 ◽

Vol 191 (9) ◽

pp. 1545-1554 ◽

Cited By ~ 96

Author(s):

Anne Brauweiler ◽

Idan Tamir ◽

Joseph Dal Porto ◽

Robert J. Benschop ◽

Cheryl D. Helgason ◽

...

Keyword(s):

Bone Marrow ◽

B Cells ◽

B Cell ◽

Cell Development ◽

B Cell Development ◽

Inositol Phosphatase ◽

Src Homology 2 ◽

Src Homology ◽

Src Homology 2 Domain ◽

Deficient Mice

Although the Src homology 2 domain–containing 5′ inositol phosphatase (SHIP) is a well-known mediator of inhibitory signals after B cell antigen receptor (BCR) coaggregation with the low affinity Fc receptor, it is not known whether SHIP functions to inhibit signals after stimulation through the BCR alone. Here, we show using gene-ablated mice that SHIP is a crucial regulator of BCR-mediated signaling, B cell activation, and B cell development. We demonstrate a critical role for SHIP in termination of phosphatidylinositol 3,4,5-triphosphate (PI[3,4,5]P3) signals that follow BCR aggregation. Consistent with enhanced PI(3,4,5)P3 signaling, we find that splenic B cells from SHIP-deficient mice display enhanced sensitivity to BCR-mediated induction of the activation markers CD86 and CD69. We further demonstrate that SHIP regulates the rate of B cell development in the bone marrow and spleen, as B cell precursors from SHIP-deficient mice progress more rapidly through the immature and transitional developmental stages. Finally, we observe that SHIP-deficient B cells have increased resistance to BCR-mediated cell death. These results demonstrate a central role for SHIP in regulation of BCR signaling and B cell biology, from signal driven development in the bone marrow and spleen, to activation and death in the periphery.

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T and B cell development in pituitary deficient insulin-like growth factor-II transgenic dwarf mice

Journal of Endocrinology ◽

10.1677/joe.0.1550165 ◽

1997 ◽

Vol 155 (1) ◽

pp. 165-170 ◽

Cited By ~ 7

Author(s):

R Kooijman ◽

SC van Buul-Offers ◽

LE Scholtens ◽

RG Reijnen-Gresnigt ◽

BJ Zegers

Keyword(s):

Bone Marrow ◽

T Cell ◽

B Cells ◽

B Cell ◽

Bone Marrow Cells ◽

T Cell Development ◽

Cell Development ◽

B Cell Development ◽

Igf I ◽

Dwarf Mice

Treatment of mice with IGF-I stimulates T and B cell development. We showed that overexpression of IGF-II in transgenic FVB/N mice only stimulated T cell development. In the present study, we further addressed the in vivo effects of IGF-II in the absence of IGF-I to get more insight into the potential abilities of IGF-II to influence T and B cell development. To this end, we studied lymphocyte development in IGF-II transgenic Snell dwarf mice that are prolactin, GH and thyroid-stimulating hormone deficient and as a consequence show low serum IGF-I levels. We showed that T cell development was stimulated to the same extent as in IGF-II transgenic FVB/N mice. Furthermore, IGF-II increased the number of nucleated bone marrow cells and the number of immature B cells without having an effect on the number of mature B cells in spleen and bone marrow. Our data show that IGF-II has preferential effects on T cell development compared with B development, and that these preferential effects also occur in the absence of measurable IGF-I levels.

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Impaired bone marrow B-cell development in mice with a bronchiolitis obliterans model of cGVHD

Blood Advances ◽

10.1182/bloodadvances.2017014977 ◽

2018 ◽

Vol 2 (18) ◽

pp. 2307-2319 ◽

Cited By ~ 1

Author(s):

Oleg V. Kolupaev ◽

Trisha A. Dant ◽

Hemamalini Bommiasamy ◽

Danny W. Bruce ◽

Kenneth A. Fowler ◽

...

Keyword(s):

Bone Marrow ◽

T Cells ◽

T Cell ◽

B Cell ◽

Bronchiolitis Obliterans ◽

Critical Role ◽

Cell Development ◽

B Cell Development ◽

Allogeneic Bone ◽

B Cell Precursors

Abstract Chronic graft-versus-host disease (cGVHD) causes significant morbidity and mortality in patients after allogeneic bone marrow (BM) or stem cell transplantation (allo-SCT). Recent work has indicated that both T and B lymphocytes play an important role in the pathophysiology of cGVHD. Previously, our group showed a critical role for the germinal center response in the function of B cells using a bronchiolitis obliterans (BO) model of cGVHD. Here, we demonstrated for the first time that cGVHD is associated with severe defects in the generation of BM B lymphoid and uncommitted common lymphoid progenitor cells. We found an increase in the number of donor CD4+ T cells in the BM of mice with cGVHD that was negatively correlated with B-cell development and the frequency of osteoblasts and Prrx-1–expressing perivascular stromal cells, which are present in the B-cell niche. Use of anti-DR3 monoclonal antibodies to enhance the number of donor regulatory T cells (Tregs) in the donor T-cell inoculum ameliorated the pathology associated with BO in this model. This correlated with an increased number of endosteal osteoblastic cells and significantly improved the generation of B-cell precursors in the BM after allo-SCT. Our work indicates that donor Tregs play a critical role in preserving the generation of B-cell precursors in the BM after allo-SCT. Approaches to enhance the number and/or function of donor Tregs that do not enhance conventional T-cell activity may be important to decrease the incidence and severity of cGVHD in part through normal B-cell lymphopoiesis.

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