scholarly journals Conversion of Normal Ly-1-Positive B-Lineage Cells into Ly-1-Positive Macrophages in Long-Term Bone Marrow Cultures

1990 ◽  
Vol 1 (2) ◽  
pp. 113-125 ◽  
Author(s):  
Shigeki Katoh ◽  
Akira Tominaga ◽  
Masahiro Migita ◽  
Akira Kudo ◽  
Kiyoshi Takatsu
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Cell Lines ◽  
Germ Line ◽  
Chain Gene ◽  
Nonspecific Esterase ◽  
Light Chain Gene ◽  
B Lineage ◽  
Region Segment

We obtained eight different cell lines in the long-term bone marrow culture system that showed a germ-line configuration of the joining (J) region segments of the Ig heavy-chain (IgH) genes. Their surface markers were CD45R+, Ly-1+, Lyb-2+, cIgM-, sIgM-, Ia-, Thy-1-, Mac-1-, and IL-2R (Tac)+. Use of very young mice and the presence of IL-5 were important for preferential promotion of the survival of B-lineage lymphocytes bearing the Ly-1 markers. When we treated two of them (J8 and J10) with 5-azacytidine for 24 h followed by co-culture with stromal cells and IL-.5, they became Ly-1+, sIgM+B cells, and Ly-1+, Mac-1+macrophagelike cells, respectively. After other early lymphoid lines (J1, J8, and J13) were maintained by co-culture with ST2 and IL-5 for more than a year, they showed a heterogeneous DNA rearrangement profile of the J region segment of the IgH gene, although only J13 rearranged theκ-light chain gene. Northern blot analysis revealed that these cell lines expressed Cμ-mRNA, andλ5-mRNA, consistent with normal pre-B cells. Intriguingly, J1, J8, and J13 expressed c-fmsmRNA constitutively. When J13 cells were co-cultured with ST2 and GM-CSF in place of ST2 and IL-5, they acquired Mac-1 expression and retained Ly-1 expression. They were morphologically macrophages, nonspecific-esterase-positive, and showed phagocytosis of latex beads. These results support evidence for a close relationship between the myeloid and Ly-1+B-cell pathways of differentiation, and indicate that our IL- 5-dependent clones are multipotential intermediates in differentiation from pro-B cells to B cells and macrophages.

scholarly journals Normal human B cells display ordered light chain gene rearrangements and deletions.

1982 ◽  
Vol 156 (4) ◽  
pp. 975-985 ◽  
Author(s):  
S J Korsmeyer ◽  
P A Hieter ◽  
S O Sharrow ◽  
C K Goldman ◽  
P Leder ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Lines ◽  
Light Chain ◽  
Developmental Process ◽  
Germ Line ◽  
Chain Gene ◽  
Gene Rearrangements ◽  
Lambda Light Chain ◽  
Light Chain Gene

Human kappa-producing B cell lines and leukemias retain their excluded lambda light chain genes in the germ line configuration, whereas transformed lambda-producing B cells uniformly rearrange or delete their kappa genes (12). Whether the unexpected lambda gene recombinations within malignant lambda-producing B cells reflect a normal developmental process or are secondary to transformation and specific chromosomal translocations was uncertain. To resolve this issue, we purified circulating lambda-bearing B cells from a normal individual to 97% purity by using a series of negative selection steps and a final positive selection on a cell sorter. Over 95% of the collective kappa genes in these lambda B cells were no longer in their germ line form, with the majority (60%) deleted and the remainder present but in a rearranged state. The chromosomal loss of the germ line kappa genes included the joining (J kappa) segments as well as the constant (C kappa) region, yet the particular variable (V kappa) gene family studied was spared. In addition, the incidence of kappa gene deletions was higher in long-term than in freshly transformed lambda B cell lines. This implies that the deletion of aberrantly rearranged kappa genes may occur as a second event. Such a mechanism would serve to eliminate aberrant transcripts and light chain fragments that might interfere with the synthesis and assembly of effective immunoglobulin molecules. Thus, despite the nearly equal usage of kappa and lambda light chain genes in man, there appears to be a sequential order to their expression during normal B cell ontogeny in which kappa gene rearrangements precede those of lambda.

Transformation of murine bone marrow cells with combined v-raf-v-myc oncogenes yields clonally related mature B cells and macrophages

1990 ◽  
Vol 10 (7) ◽  
pp. 3562-3568
Author(s):  
M Principato ◽  
J L Cleveland ◽  
U R Rapp ◽  
K L Holmes ◽  
J H Pierce ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Cell Lines ◽  
Bone Marrow Cells ◽  
Hematopoietic Differentiation ◽  
Developmental Relationships ◽  
Murine Bone Marrow ◽  
B Lineage ◽  
Marrow Cells

Murine bone marrow cells infected with replication-defective retroviruses containing v-raf alone or v-myc alone yielded transformed pre-B cell lines, while a retroviral construct containing both v-raf and v-myc oncogenes produced clonally related populations of mature B cells and mature macrophages. The genealogy of these transformants demonstrates that mature myeloid cells were derived from cells with apparent B-lineage commitment and functional immunoglobulin rearrangements. This system should facilitate studies of developmental relationships in hematopoietic differentiation and analysis of lineage determination.

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 Lipopolysaccharide-unresponsive mutant pre-B-cell lines blocked in NF-kappa B activation.

1990 ◽  
Vol 10 (1) ◽  
pp. 422-425 ◽  
Author(s):  
M Briskin ◽  
M Damore ◽  
R Law ◽  
G Lee ◽  
P W Kincade ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Lines ◽  
Chain Gene ◽  
Nf Kappa B ◽  
Light Chain Gene ◽  
Immunoglobulin Kappa ◽  
Variant Cell ◽  
Late Step ◽  
Mutant Lines

NF-kappa B activation is a crucial late step in the induction of immunoglobulin kappa light-chain gene expression in pre-B cells by lipopolysaccharide (LPS). We have analyzed NF-kappa B activation in three independent mutant lines of 70Z/3 pre-B cells which are unresponsive to LPS. All three variant cell lines failed to activate NF-kappa B when induced with LPS or the phorbol ester 12-O-tetradecanoylphorbol 13-acetate. However, all three cell lines contained functional NF-kappa B, as revealed by detergent treatment of cytoplasmic extracts. Moreover, cycloheximide induced limited activation of NF-kappa B comparable to that in wild-type 70Z/3 pre-B cells in two of the three variant lines. These results indicate that the mutations blocking kappa gene induction in these variant 70Z/3 pre-B-cell lines affect NF-kappa B activation.

Lipopolysaccharide-unresponsive mutant pre-B-cell lines blocked in NF-kappa B activation

1990 ◽  
Vol 10 (1) ◽  
pp. 422-425
Author(s):  
M Briskin ◽  
M Damore ◽  
R Law ◽  
G Lee ◽  
P W Kincade ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Lines ◽  
Chain Gene ◽  
Nf Kappa B ◽  
Light Chain Gene ◽  
Immunoglobulin Kappa ◽  
Variant Cell ◽  
Late Step ◽  
Mutant Lines

NF-kappa B activation is a crucial late step in the induction of immunoglobulin kappa light-chain gene expression in pre-B cells by lipopolysaccharide (LPS). We have analyzed NF-kappa B activation in three independent mutant lines of 70Z/3 pre-B cells which are unresponsive to LPS. All three variant cell lines failed to activate NF-kappa B when induced with LPS or the phorbol ester 12-O-tetradecanoylphorbol 13-acetate. However, all three cell lines contained functional NF-kappa B, as revealed by detergent treatment of cytoplasmic extracts. Moreover, cycloheximide induced limited activation of NF-kappa B comparable to that in wild-type 70Z/3 pre-B cells in two of the three variant lines. These results indicate that the mutations blocking kappa gene induction in these variant 70Z/3 pre-B-cell lines affect NF-kappa B activation.

scholarly journals Independent regulation of IgM, IgD, and Ia antigen expression in cultured immature B lymphocytes.

1986 ◽  
Vol 163 (4) ◽  
pp. 938-951 ◽  
Author(s):  
J R Dasch ◽  
P P Jones
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Bone Marrow Cells ◽  
Antigen Expression ◽  
Adult Mice ◽  
Immature B Cells ◽  
Similar Phenotype ◽  
B Lineage ◽  
Set Up

Long-term cultured bone marrow cells were characterized with respect to a number of B and pre-B cell markers. Cells expressing ThB, B-220, and IgM were found within cultures set up according to the procedure of Whitlock and Witte. This culture system was modified by placing sorted pre-B cells (ThB+, IgM-) from bone marrow in culture with previously-established bone marrow adherent layers. These cultures commenced growth without the lag associated with the Whitlock cultures. These cultured nonadherent cells show a high frequency of IgM+ cells, but do not express either IgD or Ia, and we refer to them as immature B cells. Cells with a similar phenotype (IgM+, Ia-, IgD-) are found within the spleens of young but not adult mice. The phorbol ester PMA induces expression of IgD on the cultured immature B cells, but has no effect on Ia expression. This suggests that the processing of H chain RNA transcripts may be affected by protein kinase C. These results demonstrate that the appearance of IgM, IgD, and Ia are independently controlled in long-term cultured B-lineage cells.

scholarly journals Transformation of murine bone marrow cells with combined v-raf-v-myc oncogenes yields clonally related mature B cells and macrophages.

1990 ◽  
Vol 10 (7) ◽  
pp. 3562-3568 ◽  
Author(s):  
M Principato ◽  
J L Cleveland ◽  
U R Rapp ◽  
K L Holmes ◽  
J H Pierce ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Cell Lines ◽  
Bone Marrow Cells ◽  
Hematopoietic Differentiation ◽  
Developmental Relationships ◽  
Murine Bone Marrow ◽  
B Lineage ◽  
Marrow Cells

Murine bone marrow cells infected with replication-defective retroviruses containing v-raf alone or v-myc alone yielded transformed pre-B cell lines, while a retroviral construct containing both v-raf and v-myc oncogenes produced clonally related populations of mature B cells and mature macrophages. The genealogy of these transformants demonstrates that mature myeloid cells were derived from cells with apparent B-lineage commitment and functional immunoglobulin rearrangements. This system should facilitate studies of developmental relationships in hematopoietic differentiation and analysis of lineage determination.

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