CLONAL PROLIFERATION OF MURINE LYMPHOHEMATOPOIETIC PROGENITORS IN CULTURE

被引:144
作者
HIRAYAMA, F
SHIH, JP
AWGULEWITSCH, A
WARR, GW
CLARK, SC
OGAWA, M
机构
[1] MED UNIV S CAROLINA,DEPT MED,CHARLESTON,SC 29403
[2] MED UNIV S CAROLINA,DEPT BIOCHEM & MOLEC BIOL,CHARLESTON,SC 29403
[3] RALPH H JOHNSON VET AFFAIRS MED CTR,CHARLESTON,SC 29403
[4] GENET INST,CAMBRIDGE,MA 02140
关键词
HEMATOPOIETIC STEM CELL; GROWTH FACTOR; LYMPHOCYTE-B;
D O I
10.1073/pnas.89.13.5907
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
We have used a two-step clonal culture system to unequivocally demonstrate that individual primitive lymphohemopoietic progenitor cells have the capacity for differentiation along either the myeloid or the B-lymphoid lineage. Highly enriched murine marrow cells were plated individually in culture by micromanipulation in the presence of pokeweed mitogen-stimulated spleen cell conditioned medium, erythropoietin, steel factor (SF), and interleukin (IL) 7. Forty-five percent of the single cells formed primary colonies expressing multiple hemopoietic lineages. When aliquots from individual colonies were replated in secondary methyl cellulose culture containing SF and IL-7, 41% of the primary colonies gave rise to lymphocyte colonies. Cells of the lymphocyte colonies were blast-like and B220+, sIg-, Mac-1-, Gr-1-, Ly-1-, L3T4-, Ly-2-, and CD3-. Thirty to 70% of the cells were Thy-1+. Mu-chain mRNA was detected in most of the cells by in situ hybridization with an antisense RNA probe. When lymphocyte colonies derived from a single cell were pooled and individually injected into scid mice, donor-type IgM was measurable in the serum of mice and spleens contained donor-type B cells. We then carried out initial screening of growth factors to identify growth factors that might replace pokeweed mitogen-stimulated spleen cell conditioned medium in the primary culture. Combinations of two factors that included SF plus IL-6, IL-11, or granulocyte colony-stimulating factor were all effective in the primary culture in the maintenance of the B-lymphoid potential. Interestingly, IL-3 could neither replace nor act synergistically with SF to support the lymphoid potential of the primary cultures. Our observations demonstrate that many primitive progenitors previously believed to be myeloid-committed also possess B-lymphoid potential. This culture system should prove valuable for elucidation of the mechanisms regulating early stages of lymphohemopoiesis.
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页码:5907 / 5911
页数:5
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