Long-term culture system for selective growth of human B-cell progenitors.
We describe a simple reproducible system for enrichment and long-term culture of human B-cell progenitors. Enriched CD34+ cord blood mononuclear cells are seeded onto a murine stromal cell line to establish a biphasic culture system. These cultures are characterized by transient growth of myeloid cells followed by outgrowth of cells highly enriched for early B-cell progenitors. Cultures consisting of > 90% early B-lineage cells [expressing CD10, CD19, CD38, and CD45 but lacking CD20, CD22, CD23, and surface IgM] are maintained for > 12 weeks without growth factor addition. Cells remain predominantly germ line at the immunoglobulin locus and express only low levels of cytoplasmic mu chain, terminal deoxynucleotidyltransferase, and recombination-activating gene 1 product. They are unresponsive to the pre-B-cell growth factors interleukin 7 or stem cell factor, or both, suggesting that growth support is provided by a cross-reactive murine stromal cell factor. Cultured B-cell progenitors are generated in large numbers ( > 10(8) cells from a typical cord blood specimen) suitable for use in biochemical analysis and gene-transfer studies. This system should be useful for study of normal and abnormal early human B-lymphopoiesis.
Rawlings
D J
DJ
Department of Microbiology and Molecular Genetics, University of California, Los Angeles 90024-1662.
Quan
S G
SG
Kato
R M
RM
Witte
O N
ON
D015703:Antigens, CD / Q000032:analysis; D001402:B-Lymphocytes / Q000166:cytology*; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D005312:Fetal Blood / Q000166:cytology; D015322:Gene Rearrangement, B-Lymphocyte; D005803:Genes, Immunoglobulin; D016298:Hematopoietic Cell Growth Factors / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D016130:Immunophenotyping; D066298:In Vitro Techniques; D015851:Interleukin-7 / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019009:Proto-Oncogene Proteins c-kit; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism; D016184:Receptors, Colony-Stimulating Factor / Q000378:metabolism; D018123:Receptors, Interleukin / Q000378:metabolism; D020395:Receptors, Interleukin-7; D019089:Stem Cell Factor; D013997:Time Factors
D015703:Antigens, CD; D016298:Hematopoietic Cell Growth Factors; D015851:Interleukin-7; D011518:Proto-Oncogene Proteins; D016184:Receptors, Colony-Stimulating Factor; D018123:Receptors, Interleukin; D020395:Receptors, Interleukin-7; D019089:Stem Cell Factor; D019009:Proto-Oncogene Proteins c-kit; D020794:Receptor Protein-Tyrosine Kinases