Functional Studies on Subpopulations of B-Lymphocytes and Bone Marrow Cells

1982 ◽  
pp. 53-59
Author(s):  
A. Chayen ◽  
S. Marshall-Clarke ◽  
R. M. E. Parkhouse
Keyword(s):  
Bone Marrow ◽  
B Lymphocytes ◽  
Bone Marrow Cells ◽  
Functional Studies ◽  
Marrow Cells

Mesenchymal Stromal Cells Enhance G-CSF-Induced Mobilization Of Hematopoietic Stem- and Progenitor Cells

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2460-2460
Author(s):  
Evert-Jan F. M. de Kruijf ◽  
Ingmar van Hengel ◽  
Jorge M Perez-Galarza ◽  
Willem E. Fibbe ◽  
Melissa van Pel
Keyword(s):  
Bone Marrow ◽  
B Lymphocytes ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Peripheral Blood ◽  
Bone Marrow Cells ◽  
Calf Serum ◽  
Intraperitoneal Administration ◽  
Hematopoietic Stem ◽  
Marrow Cells

Abstract Hematopoietic stem- and progenitor cell (HSPC) mobilization is a property of most hematopoietic growth factors, such as Granulocyte Colony Stimulating Factor (G-CSF). Not all donors mobilize equally well and therefore the number of HSPC that are obtained following mobilization may be limited. Mesenchymal stromal cells (MSC) have the capacity to differentiate into cells of the mesodermal lineage and have immunomodulatory properties in vivo and in vitro. Here, we have investigated the effect of MSC co-administration on G-CSF-induced HSPC mobilization. MSC were obtained from bone marrow cells (bone marrow-derived) or bone fragments (bone-derived) and were expanded in alpha-MEM containing 10% fetal calf serum until sufficient cell numbers were obtained. Bone marrow or bone-derived MSC were administered intravenously for three days at a dose of 200 x103 cells per day to male C57BL/6 recipients that were simultaneously mobilized with G-CSF (10 μg per day intraperitoneally for 3 days) or PBS as a control. Co-injection of G-CSF and MSC lead to a 2-fold increase in HSPC mobilization compared to G-CSF alone (8,563 ± 3,309 vs. 4,268 ± 1,314 CFU-C per ml peripheral blood respectively; n=13, p<0.01). Administration of MSC alone did not induce HSPC mobilization (273 ± 229 CFU-C/ml blood; n=13). Furthermore, co-injection of splenocytes and G-CSF did not enhance HSPC mobilization, showing that the administration of exogeneous cells as such is not sufficient for enhancement of HSPC mobilization. It has been reported that G-CSF-induced HSPC mobilization is associated with a decrease in the number of osteal macrophages, B lymphocytes and erythroid progenitors. Administration of MSC alone induced a significant decrease in the frequency of osteal macrophages (7.9 ± 1.2 vs 6.2 ± 1.4% bone marrow cells for PBS vs. MSC respectively; n=8, p<0.05), but did not affect osteoblast numbers. Furthermore, the frequency of B lymphocytes was significantly decreased following MSC administration (29.9 ± 4.0 vs. 16.5 ± 4.9% bone marrow cells for PBS vs. MSC respectively; n=13, p<0.0001). No differences were observed in erythroid numbers following MSC administration. To investigate the mechanisms underlying these observations, the migratory capacity of luciferase transduced MSC was studied through bioluminescence imaging. Following intravenous injection, MSC were detected in the lungs, but not in other organs. In addition, no difference in MSC migration was observed between G-CSF and PBS treated mice. Moreover, intraperitoneal administration of G-CSF and MSC resulted in increased HSPC mobilization compared to G-CSF alone (10,178 ±3,039 vs. 5,158 ± 2,436 CFU-C per ml peripheral blood; n=5-12). Together, these data point to an endocrine effect of MSC on G-CSF-induced HSPC mobilization. No differences in IL-6, CXCL-12 or M-CSF levels in bone marrow extracellular fluid were observed. In conclusion, G-CSF-induced HSPC mobilization is enhanced by injection of MSC. We hypothesize that the MSC-induced partial depletion of B lymphocytes and osteal macrophages in the bone marrow are crucial factors involved in the enhancement of G-CSF-induced HSPC mobilization. Disclosures: No relevant conflicts of interest to declare.

scholarly journals THE LIFE-SPAN AND RECIRCULATION OF MARROW-DERIVED SMALL LYMPHOCYTES FROM THE RAT THORACIC DUCT

1972 ◽  
Vol 135 (2) ◽  
pp. 185-199 ◽  
Author(s):  
Jonathan C. Howard
Keyword(s):  
Bone Marrow ◽  
Life Span ◽  
B Lymphocytes ◽  
Thoracic Duct ◽  
Bone Marrow Cells ◽  
Thoracic Duct Lymph ◽  
Clear Cut ◽  
Duct Lymph ◽  
Order Of Magnitude ◽  
Marrow Cells

These experiments describe the preparation of pure marrow-derived lymphocyte suspensions from the thoracic duct of thymectomized, irradiated rats reconstituted with bone marrow cells. The majority of marrow-derived cells were small lymphocytes morphologically indistinguishable from small lymphocytes in thoracic duct lymph of normal donors. Marrow-derived small lymphocytes (B lymphocytes) were a predominantly long-lived population; the frequency of short-lived B lymphocytes in the thoracic duct was not significantly higher than the frequency of short-lived small lymphocytes in normal lymph. B lymphocytes transferred to normal recipients recirculated from blood to lymph. The first appearance of intravenously injected B lymphocytes in the thoracic duct was delayed relative to lymphocytes from normal donors and there was no clear cut modal recirculation time. Nevertheless their recirculation over a 48 hr period after transfusion was of the same order of magnitude as that of lymphocytes from normal donors.

scholarly journals Evidence for the clonal abortion theory of B-lymphocyte tolerance.

1975 ◽  
Vol 141 (4) ◽  
pp. 904-917 ◽  
Author(s):  
G J Nossal ◽  
B L Pike
Keyword(s):  
Bone Marrow ◽  
Tissue Culture ◽  
B Lymphocytes ◽  
B Lymphocyte ◽  
Cultured Cells ◽  
Bone Marrow Cells ◽  
Mouse Bone Marrow ◽  
Immune Competence ◽  
Self Recognition ◽  
Marrow Cells

This paper deals with the behavior of adult mouse bone marrow cells placed in tissue culture with or without antigen, and subsequently assessed for immune competence after adoptive transfer into lethally X-irradiated, syngeneic hosts. Attention was focussed on B lymphocytes through using hapten human gamma globulin (HGG) preparations as putative tolerogens in tissue culture, the T-cell-independent antigens DNP-POL and NIP-POL as challenge injections in adoptive hosts, and numbers of hapten-specific PFC in host spleens for the quantitation of immune competence. It was found that the capacity of bone marrow cells to mount an adoptive immune response rose by a factor of about fivefold over 3 days in tissue culture. This rise was completely abolished by the presence in the culture of hapten-HGG conjugates with about one mole of hapten per carrier molecule. The prevention of the emergence of immune competence amongst maturing B cells was termed clonal abortion tolerogenesis. Dose-response studies showed the lowest effective antigen concentration to be between 2.5 times 10- minus 10 and 2.5 times 10- minus 9 M, and a standard concentration of 2.5 times 10- minus 8 M was chosen as producing near maximal effects. The tolerance was antigen-specific and time-dependent, being maximal only when antigen was present continuously as the cultured cells was maturing. It did not depend on the presence of T lymphocytes in marrow, and was not of an "infectious" type. In contrast to tolerogenesis of mature B lymphocytes by high antigen concentrations, it could not be abolished by lipopolysaccharide. We speculate that clonal abortion may be a tolerance mechanism of great physiological significance for self-recognition, and discuss the results in the framework of other recent tolerance models, including those involving receptor blockade and suppressor T cells.

Effect of B lymphocytes from different organs on hematopoietic colony formation in the spleen by bone marrow cells

1992 ◽  
Vol 113 (2) ◽  
pp. 221-224
Author(s):  
V. M. Man'ko ◽  
O. A. Guseva ◽  
I. V. Miroshnichenko ◽  
A. A. Yarilin
Keyword(s):  
Bone Marrow ◽  
B Lymphocytes ◽  
Colony Formation ◽  
Bone Marrow Cells ◽  
Marrow Cells

scholarly journals Defective lymphopoiesis in bone marrow of motheaten (me/me) and viable motheaten (mev/mev) mutant mice. I. Analysis of development of prothymocytes, early B lineage cells, and terminal deoxynucleotidyl transferase-positive cells.

1986 ◽  
Vol 164 (4) ◽  
pp. 1129-1144 ◽  
Author(s):  
D L Greiner ◽  
I Goldschneider ◽  
K L Komschlies ◽  
E S Medlock ◽  
F J Bollum ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Lymphocytes ◽  
Bone Marrow Cells ◽  
Precursor Cells ◽  
T And B Lymphocytes ◽  
Normal Numbers ◽  
B Lineage ◽  
Marrow Cells ◽  
B Lineage Cells

This study identifies defects in the early stages of lymphopoiesis that may contribute to the abnormalities in the development and/or function of peripheral T and B lymphocytes in mice homozygous for the motheaten (me/me) and viable motheaten (mev/mev) mutations. The results indicate that in me/me and mev/mev mice prothymocytes in bone marrow are present in essentially normal numbers, as determined by intrathymic injection, but apparently lack the ability to home effectively to the thymus, as determined by intravenous transfer; early B lineage cells in bone marrow, identified by the B220 antigen, are markedly depleted, including immature B cells (sIg+), pre-B cells (cIg+, sIg-), and pro-B cells (B220+, cIg-, sIg-); TdT+ bone marrow cells, especially a subset that expresses the B220 B lineage antigen, are markedly depleted by two weeks of age; normal numbers of TdT+ thymocytes are present during the first 3 wk of postnatal life, but rapidly decrease thereafter. The results further indicate that neither the defective thymus homing capacity of prothymocytes nor the deficiency of TdT+ bone marrow cells is due to autoantibodies. The possible relationship of the defective development of lymphoid precursor cells to the premature onset of thymic involution and to the abnormalities of peripheral T and B lymphocytes in me/me and mev/mev mice is discussed; as are the results of in vitro studies (presented in a companion paper), which suggest that a primary defect in the stromal microenvironment of the bone marrow is responsible for the abnormal development of the lymphoid precursor cells.

Dysfunctional Double-Strand DNA Break Repair In Primary t-AML/t-MDS Myeloblasts.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3366-3366
Author(s):  
Meagan A Jacoby ◽  
Rigoberto de Jesus ◽  
Matthew J. Walter
Keyword(s):  
Bone Marrow ◽  
Dna Repair ◽  
Array Cgh ◽  
Bone Marrow Cells ◽  
Dsb Repair ◽  
Functional Studies ◽  
Primary Bone ◽  
Nuclear Foci ◽  
Repair Genes ◽  
Marrow Cells

Abstract Abstract 3366 The gross chromosomal aberrations in treatment-related acute myeloid leukemia/myelodysplastic syndrome (t-AML/t-MDS) cells suggest that disease initiation and progression may result from the inability of cells to appropriately respond to double-strand DNA breaks (DSBs) induced by prior exposure to radiation, alkylator, or topoisomerase II inhibitor therapy. We hypothesize that dysregulation of DSB repair by homology-directed repair (HDR) or nonhomologous end joining (NHEJ) contributes to the development of t-AML/t-MDS. Dysregulation of DSB repair in t-AML/t-MDS may result from inherited variants or acquired mutations in HDR/NHEJ pathway genes. To directly test this possibility, we used next-generation sequencing technology and an array CGH platform to identify inherited and somatic genetic variants, including small indels and copy number alterations, in 21 canonical HDR and 9 NHEJ DNA repair genes, as well as a subset of 7 DNA damage response genes using tumor DNA and paired normal DNA obtained from 30 t-AML/t-MDS patients. All the data has been acquired and the analysis is ongoing. Because dysregulation of DNA repair pathways can result from epigenetic changes or post-translational modifications in DNA repair genes and would not be detected using sequencing and array CGH, we are also performing functional studies to interrogate DSB repair using primary bone marrow cells from 16 of these t-AML/t-MDS patients and CD34+ cells from 5 normal healthy controls. We performed a flow-based assay for DSB formation and repair by measuring the phosphorylated form of the variant histone H2AX (pH2AX), which is rapidly phosphorylated upon DSB formation, in myeloblasts (CD45 dim, low side scatter) and lymphocytes (a surrogate for normal cells) from leukemic bone marrow. Baseline measurements of unmanipulated primary bone marrow cells and a time course to measure the kinetics of DSB repair after gamma irradiation are used to assess a cell's basal DSB burden and the response to acute damage. We have validated this assay in a defined genetic system using isogenic cells deficient or not in BRCA2, a gene central to the HDR pathway, and were able to demonstrate that cells lacking BRCA2 have elevated pH2AX levels at 4–24 hours post DSB induction compared to controls (p=0.01). In addition, we have evaluated the ability of the cells to form nuclear foci of pH2AX and RAD51 (a protein central to the repair of DSB by HDR) by quantifying the relative numbers of immunofluorescent nuclear foci upon irradiation. We have performed these functional assays for 8 t-AML/t-MDS samples and 3 normal donors. We identified one t-AML/t-MDS sample whose myeloblasts show 2–3 fold greater pH2AX at baseline, compared to control CD34+ cells (p=0.006). Interestingly, the clearance of pH2AX is significantly faster in this sample compared to controls (p=0.001) despite showing elevated baseline pH2AX levels. This patient had a translocation involving chromosome 11q23. One additional patient in our cohort with an 11q23 translocation also showed similar increased baseline pH2AX levels. In addition, we identified abnormalities in a third t-AML/t-MDS sample that trends towards impaired maximal pH2AX induction (p=0.052). Furthermore, myeloblasts from a fourth t-AML/t-MDS sample, a known heterozygous BRCA2 mutation carrier, show a trend towards delayed pH2AX clearance (p=0.07). As predicted, bone marrow cells from this BRCA2 carrier also have 3.5 fold less RAD51 nuclear foci formation compared to control cells (8.6% vs 30%). Collectively, these results show that primary t-AML bone marrow cells can be used to assess the functional integrity of DSB response, and suggest that a large proportion of samples (4/8) may contain genetic alterations in DNA DSB response and/or repair genes. Functional studies of an additional 8 samples are ongoing. Next generation sequencing and array CGH data for the 37 HDR/NHEJ genes has been completed and will be correlated with the functional studies for each sample. Identification of abnormal DSB repair in leukemic patients may not only elucidate the molecular pathogenesis of the disease, but may provide rationale for testing agents to achieve selective killing of leukemic cells, as defects in one DNA repair pathway may increase reliance on another, making these cancer cells particularly susceptible to killing by inhibitors targeting the extant pathway(s). Disclosures: No relevant conflicts of interest to declare.

scholarly journals Lectin Histochemistry Reveals the Appearance of M-cells in Peyer's Patches of scid Mice After Syngeneic Normal Bone Marrow Transplantation

1998 ◽  
Vol 46 (2) ◽  
pp. 143-148 ◽  
Author(s):  
Ram Sharma ◽  
Udo Schumacher ◽  
Elizabeth Adam
Keyword(s):  
Bone Marrow ◽  
B Lymphocytes ◽  
Bone Marrow Cells ◽  
Scid Mice ◽  
Peyer's Patches ◽  
Peyer’S Patches ◽  
M Cells ◽  
Peyer’S Patch ◽  
Peyer's Patch ◽  
Marrow Cells

Peyer's patches in the intestinal mucosa are characterized by the presence of several lymphatic follicles and interfollicular T-cell regions. Luminal antigens are transported across the intestinal epithelium to stimulate the Peyer's patch pre-B-cells in the follicles that proliferate and migrate to distant sites. Evidence suggests that antigen priming of B-lymphocytes is responsible for the number and location of Peyer's patches during postnatal life, but little is known about the histogenesis of Peyer's patches and their overlying membranous (M) cells. To examine whether T- and B-lymphocytes in Peyer's patches have an influence on M-cell generation, we studied the development of Peyer's patches and M-cells in severe combined immunodeficient (scid) mice reconstituted with bone marrow cells from normal syngeneic mice. Our experiments demonstrate that the donor bone marrow cells in the host scid mice repopulate to form single (primary) follicles and aggregates of lymphoid nodules, the Peyer's patches. Use of the lectins Anguilla anguilla (AAA) and Ulex europaeus I (UEA-I) as positive markers of mouse Peyer's patch M-cells revealed that M-cells develop in the dome epithelium overlying the single primary follicles and Peyer's patches of reconstituted scid mice. This experimental system therefore allows the study of the histogenesis of Peyer's patches and M-cells.

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