Characterization of Bone Marrow Laminins and Identification of 5-Containing Laminins as Adhesive Proteins for Multipotent Hematopoietic FDCP-Mix Cells

Blood ◽  
1999 ◽  
Vol 93 (8) ◽  
pp. 2533-2542 ◽  
Author(s):  
Yuchen Gu ◽  
Lydia Sorokin ◽  
Madeleine Durbeej ◽  
Tord Hjalt ◽  
Jan-Ingvar Jönsson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mouse Bone Marrow ◽  
Bone Marrow Cultures ◽  
Adhesive Interactions ◽  
Adhesive Proteins ◽  
Polypeptide Chains ◽  
Laminin 1

Laminins are extracellular matrix glycoproteins that influence the phenotype and functions of many types of cells. Laminins are heterotrimers composed of , β, and γ polypeptides. So far five , three β, and two γ polypeptide chains, and 11 variants of laminins have been proposed. Laminins interact in vitro with mature blood cells and malignant hematopoietic cells. Most studies have been performed with laminin-1 (1β1γ1), and its expression in bone marrow is unclear. Employing an antiserum reacting with most laminin isoforms, we found laminins widely expressed in mouse bone marrow. However, no laminin 1 chain but rather laminin 2, 4, and 5 polypeptides were found in bone marrow. Our data suggest presence of laminin-2 (2β1γ1), laminin-8 (4β1γ1), and laminin-10 (5β1γ1) in bone marrow. Northern blot analysis showed expression of laminin 1, 2, 4, and 5 chains in long-term bone marrow cultures, indicating upregulation of laminin 1 chain expression in vitro. Laminins containing 5 chain, in contrast to laminin-1, were strongly adhesive for multipotent hematopoietic FDCP-mix cells. Integrin 6 and β1 chains mediated this adhesion, as shown by antibody perturbation experiments. Our findings indicate that laminins other than laminin-1 are functional in adhesive interactions in bone marrow.

Characterization of Bone Marrow Laminins and Identification of 5-Containing Laminins as Adhesive Proteins for Multipotent Hematopoietic FDCP-Mix Cells

Blood ◽  
1999 ◽  
Vol 93 (8) ◽  
pp. 2533-2542 ◽  
Author(s):  
Yuchen Gu ◽  
Lydia Sorokin ◽  
Madeleine Durbeej ◽  
Tord Hjalt ◽  
Jan-Ingvar Jönsson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mouse Bone Marrow ◽  
Bone Marrow Cultures ◽  
Adhesive Interactions ◽  
Adhesive Proteins ◽  
Polypeptide Chains ◽  
Laminin 1

Abstract Laminins are extracellular matrix glycoproteins that influence the phenotype and functions of many types of cells. Laminins are heterotrimers composed of , β, and γ polypeptides. So far five , three β, and two γ polypeptide chains, and 11 variants of laminins have been proposed. Laminins interact in vitro with mature blood cells and malignant hematopoietic cells. Most studies have been performed with laminin-1 (1β1γ1), and its expression in bone marrow is unclear. Employing an antiserum reacting with most laminin isoforms, we found laminins widely expressed in mouse bone marrow. However, no laminin 1 chain but rather laminin 2, 4, and 5 polypeptides were found in bone marrow. Our data suggest presence of laminin-2 (2β1γ1), laminin-8 (4β1γ1), and laminin-10 (5β1γ1) in bone marrow. Northern blot analysis showed expression of laminin 1, 2, 4, and 5 chains in long-term bone marrow cultures, indicating upregulation of laminin 1 chain expression in vitro. Laminins containing 5 chain, in contrast to laminin-1, were strongly adhesive for multipotent hematopoietic FDCP-mix cells. Integrin 6 and β1 chains mediated this adhesion, as shown by antibody perturbation experiments. Our findings indicate that laminins other than laminin-1 are functional in adhesive interactions in bone marrow.

scholarly journals Early pre-B-cell transformation induced by the v-fms oncogene in long-term mouse bone marrow cultures.

1989 ◽  
Vol 9 (9) ◽  
pp. 3973-3981 ◽  
Author(s):  
G V Borzillo ◽  
C J Sherr
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Chain Gene ◽  
Mouse Bone Marrow ◽  
Feeder Layers ◽  
Bone Marrow Cultures ◽  
B Lymphoid ◽  
Marrow Cultures

Murine long-term bone marrow cultures that support B-lymphoid-cell development were infected with a helper-free retrovirus containing the v-fms oncogene. Infection of B-lymphoid cultures resulted in the rapid clonal outgrowth of early pre-B cells, which grew to high cell densities on stromal cell feeder layers, expressed v-fms-coded glycoproteins, and underwent immunoglobulin heavy-chain gene rearrangements. Late-passage cultures gave rise to factor-independent variants that proliferated in the absence of feeder layers, developed resistance to hydrocortisone, and became tumorigenic in syngeneic mice. The v-fms oncogene therefore recapitulates known effects of the v-abl and bcr-abl oncogenes on B-lineage cells. The ability of v-fms to induce transformation of early pre-B cells in vitro underscores the capacity of oncogenic mutants of the colony-stimulating factor-1 receptor to function outside the mononuclear phagocyte lineage.

Early pre-B-cell transformation induced by the v-fms oncogene in long-term mouse bone marrow cultures

1989 ◽  
Vol 9 (9) ◽  
pp. 3973-3981
Author(s):  
G V Borzillo ◽  
C J Sherr
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Chain Gene ◽  
Mouse Bone Marrow ◽  
Feeder Layers ◽  
Bone Marrow Cultures ◽  
B Lymphoid ◽  
Marrow Cultures

Murine long-term bone marrow cultures that support B-lymphoid-cell development were infected with a helper-free retrovirus containing the v-fms oncogene. Infection of B-lymphoid cultures resulted in the rapid clonal outgrowth of early pre-B cells, which grew to high cell densities on stromal cell feeder layers, expressed v-fms-coded glycoproteins, and underwent immunoglobulin heavy-chain gene rearrangements. Late-passage cultures gave rise to factor-independent variants that proliferated in the absence of feeder layers, developed resistance to hydrocortisone, and became tumorigenic in syngeneic mice. The v-fms oncogene therefore recapitulates known effects of the v-abl and bcr-abl oncogenes on B-lineage cells. The ability of v-fms to induce transformation of early pre-B cells in vitro underscores the capacity of oncogenic mutants of the colony-stimulating factor-1 receptor to function outside the mononuclear phagocyte lineage.

scholarly journals Stromal cells in myeloid and lymphoid long-term bone marrow cultures can support multiple hemopoietic lineages and modulate their production of hemopoietic growth factors

Blood ◽  
1986 ◽  
Vol 68 (6) ◽  
pp. 1348-1354 ◽  
Author(s):  
A Johnson ◽  
K Dorshkind
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Culture Conditions ◽  
B Lymphopoiesis ◽  
Bone Marrow Cultures ◽  
Factor Secretion ◽  
Marrow Cultures

Abstract Hemopoiesis in long-term bone marrow cultures (LTBMC) is dependent on adherent stromal cells that form an in vitro hemopoietic microenvironment. Myeloid bone marrow cultures (MBMC) are optimal for myelopoiesis, while lymphoid bone marrow cultures (LBMC) only support B lymphopoiesis. The experiments reported here have made a comparative analysis of the two cultures to determine whether the stromal cells that establish in vitro are restricted to the support of myelopoiesis or lymphopoiesis, respectively, and to examine how the different culture conditions affect stromal cell physiology. In order to facilitate this analysis, purified populations of MBMC and LBMC stroma were prepared by treating the LTBMC with the antibiotic mycophenolic acid; this results in the elimination of hemopoietic cells while retaining purified populations of functional stroma. Stromal cell cultures prepared and maintained under MBMC conditions secreted myeloid growth factors that stimulated the growth of granulocyte-macrophage colonies, while no such activity was detected from purified LBMC stromal cultures. However, this was not due to the inability of LBMC stroma to mediate this function. Transfer of LBMC stromal cultures to MBMC conditions resulted in an induction of myeloid growth factor secretion. When seeded under these conditions with stromal cell- depleted populations of hemopoietic cells, obtained by passing marrow through nylon wool columns, the LBMC stromal cells could support long- term myelopoiesis. Conversely, transfer of MBMC stroma to LBMC conditions resulted in a cessation of myeloid growth factor secretion; on seeding these cultures with nylon wool-passed marrow, B lymphopoiesis, but not myelopoiesis, initiated. These findings indicate that the stroma in the different LTBMC are not restricted in their hemopoietic support capacity but are sensitive to culture conditions in a manner that may affect the type of microenvironment formed.

Identification and characterization of undifferentiated mast cells in mouse bone marrow

Blood ◽  
2005 ◽  
Vol 105 (11) ◽  
pp. 4282-4289 ◽  
Author(s):  
Maria Célia Jamur ◽  
Ana Cristina G. Grodzki ◽  
Elsa H. Berenstein ◽  
Majed M. Hamawy ◽  
Reuben P. Siraganian ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mast Cell ◽  
Immunoglobulin E ◽  
Mouse Bone Marrow ◽  
Immunomagnetic Isolation ◽  
Polymerase Chain ◽  
Identification And Characterization ◽  
Mast Cell Population

Abstract Sequential immunomagnetic isolation with 2 monoclonal antibodies was used to purify and characterize an undifferentiated mast cell in adult mouse bone marrow that had not been previously recognized. This cell represents 0.02% of the cells in the bone marrow, is CD34+, CD13+, and c-kit+, and does not express FcϵRI. However, by polymerase chain reaction (PCR) the cell contains message for the α and β subunits of FcϵRI, mast cell–specific proteases, and carboxypeptidase A. Morphologically, this cell has a large nucleus, little cytoplasm, few cytoplasmic organelles, and no cytoplasmic granules. In vitro, in the presence of interleukin-3 (IL-3) and stem cell factor (SCF) these cells differentiate only into a granulated mast cell that now expresses CD13, c-kit, mast cell–specific gangliosides, FcϵRI, and binds immunoglobulin E (IgE). When injected into lethally irradiated mice, these cells are able to reconstitute the mast cell population in the spleen.

scholarly journals Stromal cells in myeloid and lymphoid long-term bone marrow cultures can support multiple hemopoietic lineages and modulate their production of hemopoietic growth factors

Blood ◽  
1986 ◽  
Vol 68 (6) ◽  
pp. 1348-1354 ◽  
Author(s):  
A Johnson ◽  
K Dorshkind
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Culture Conditions ◽  
B Lymphopoiesis ◽  
Bone Marrow Cultures ◽  
Factor Secretion ◽  
Marrow Cultures

Hemopoiesis in long-term bone marrow cultures (LTBMC) is dependent on adherent stromal cells that form an in vitro hemopoietic microenvironment. Myeloid bone marrow cultures (MBMC) are optimal for myelopoiesis, while lymphoid bone marrow cultures (LBMC) only support B lymphopoiesis. The experiments reported here have made a comparative analysis of the two cultures to determine whether the stromal cells that establish in vitro are restricted to the support of myelopoiesis or lymphopoiesis, respectively, and to examine how the different culture conditions affect stromal cell physiology. In order to facilitate this analysis, purified populations of MBMC and LBMC stroma were prepared by treating the LTBMC with the antibiotic mycophenolic acid; this results in the elimination of hemopoietic cells while retaining purified populations of functional stroma. Stromal cell cultures prepared and maintained under MBMC conditions secreted myeloid growth factors that stimulated the growth of granulocyte-macrophage colonies, while no such activity was detected from purified LBMC stromal cultures. However, this was not due to the inability of LBMC stroma to mediate this function. Transfer of LBMC stromal cultures to MBMC conditions resulted in an induction of myeloid growth factor secretion. When seeded under these conditions with stromal cell- depleted populations of hemopoietic cells, obtained by passing marrow through nylon wool columns, the LBMC stromal cells could support long- term myelopoiesis. Conversely, transfer of MBMC stroma to LBMC conditions resulted in a cessation of myeloid growth factor secretion; on seeding these cultures with nylon wool-passed marrow, B lymphopoiesis, but not myelopoiesis, initiated. These findings indicate that the stroma in the different LTBMC are not restricted in their hemopoietic support capacity but are sensitive to culture conditions in a manner that may affect the type of microenvironment formed.

scholarly journals Hematopoietic regulatory factors produced in long-term murine bone marrow cultures and the effect of in vitro irradiation

Blood ◽  
1984 ◽  
Vol 64 (2) ◽  
pp. 516-525 ◽  
Author(s):  
RJ Gualtieri ◽  
RK Shadduck ◽  
DG Baker ◽  
PJ Quesenberry
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Factor Vii ◽  
Murine Bone Marrow ◽  
L Cell ◽  
Bone Marrow Cultures ◽  
Macrophage Colony ◽  
Marrow Cultures

The nature of hematopoietic regulatory factors elaborated by the adherent (stromal) cells of long-term murine bone marrow cultures and the effect of in vitro stromal irradiation (XRT) on the production of these factors was investigated. Using an in situ stromal assay employing a double layer of semisolid agar, it was possible to demonstrate stromal elaboration of at least two colony-stimulating activities, ie, granulocyte/macrophage colony-stimulating activity (G/M- CSA) and megakaryocyte colony-stimulating activity (Meg-CSA). Exposure of the stroma to XRT resulted in dose-dependent elevations of both activities that correlated inversely with total myeloid cell mass as determined by concurrent reductions in total supernatant cell recoveries from irradiated cultures. Mixture experiments that combined control and irradiated stroma revealed that the hematopoietically active control stroma could block detection of XRT-related G/M-CSA elevations. These data implicate a local negative feedback mechanism in the regulation of hematopoiesis. Antiserum directed against purified L cell colony-stimulating factor (CSF) reduced granulocyte/macrophage colony formation in the target layer but did not effect the increased Meg-CSA. While a radioimmunoassay for L-cell type CSF was unable to detect significant differences in concentrated media from control and irradiated cultures, bioassays of these media revealed XRT-related G/M- CSA elevations. These results indicate that the G/M-CSA elaborated in these cultures is immunologically distinct from the Meg-CSA produced, and although distinct from L cell CSF, the G/M-CSA is crossreactive with the L cell CSF antiserum. Morphologic, histochemical, and factor VII antigen immunofluorescent studies were performed on the stromal cell population responsible for production of these stimulatory activities. In addition to “fat” cells, the stromal cells remaining after XRT were composed of two predominant cell populations. These included a major population of acid phosphatase and nonspecific esterase-positive macrophage-like cells and a minor population of factor VII antigen negative epithelioid cells.

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