• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of pnasPNASInfo for AuthorsSubscriptionsAboutThis Article
Proc Natl Acad Sci U S A. Feb 28, 1995; 92(5): 1570–1574.

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.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.2M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Uckun FM. Regulation of human B-cell ontogeny. Blood. 1990 Nov 15;76(10):1908–1923. [PubMed]
  • Rolink A, Melchers F. Molecular and cellular origins of B lymphocyte diversity. Cell. 1991 Sep 20;66(6):1081–1094. [PubMed]
  • Cooper MD. Current concepts. B lymphocytes. Normal development and function. N Engl J Med. 1987 Dec 3;317(23):1452–1456. [PubMed]
  • Kincade PW, Lee G, Pietrangeli CE, Hayashi S, Gimble JM. Cells and molecules that regulate B lymphopoiesis in bone marrow. Annu Rev Immunol. 1989;7:111–143. [PubMed]
  • Kincade PW. B lymphopoiesis: global factors, local control. Proc Natl Acad Sci U S A. 1994 Apr 12;91(8):2888–2889. [PMC free article] [PubMed]
  • Dorshkind K. Regulation of hemopoiesis by bone marrow stromal cells and their products. Annu Rev Immunol. 1990;8:111–137. [PubMed]
  • Whitlock CA, Witte ON. Long-term culture of B lymphocytes and their precursors from murine bone marrow. Proc Natl Acad Sci U S A. 1982 Jun;79(11):3608–3612. [PMC free article] [PubMed]
  • Scherle PA, Dorshkind K, Witte ON. Clonal lymphoid progenitor cell lines expressing the BCR/ABL oncogene retain full differentiative function. Proc Natl Acad Sci U S A. 1990 Mar;87(5):1908–1912. [PMC free article] [PubMed]
  • Faust EA, Saffran DC, Toksoz D, Williams DA, Witte ON. Distinctive growth requirements and gene expression patterns distinguish progenitor B cells from pre-B cells. J Exp Med. 1993 Apr 1;177(4):915–923. [PMC free article] [PubMed]
  • Dorshkind K, Denis KA, Witte ON. Lymphoid bone marrow cultures can reconstitute heterogeneous B and T cell-dependent responses in severe combined immunodeficient mice. J Immunol. 1986 Dec 1;137(11):3457–3463. [PubMed]
  • Wolf ML, Buckley JA, Goldfarb A, Law CL, LeBien TW. Development of a bone marrow culture for maintenance and growth of normal human B cell precursors. J Immunol. 1991 Nov 15;147(10):3324–3330. [PubMed]
  • McGinnes K, Letarte M, Paige CJ. B-lineage colonies from normal, human bone marrow are initiated by B cells and their progenitors. Blood. 1991 Mar 1;77(5):961–970. [PubMed]
  • McGinnes K, Quesniaux V, Hitzler J, Paige C. Human B-lymphopoiesis is supported by bone marrow-derived stromal cells. Exp Hematol. 1991 May;19(4):294–303. [PubMed]
  • Moreau I, Duvert V, Banchereau J, Saeland S. Culture of human fetal B-cell precursors on bone marrow stroma maintains highly proliferative CD20dim cells. Blood. 1993 Mar 1;81(5):1170–1178. [PubMed]
  • Moreau I, Duvert V, Caux C, Galmiche MC, Charbord P, Banchereau J, Saeland S. Myofibroblastic stromal cells isolated from human bone marrow induce the proliferation of both early myeloid and B-lymphoid cells. Blood. 1993 Oct 15;82(8):2396–2405. [PubMed]
  • Saeland S, Duvert V, Pandrau D, Caux C, Durand I, Wrighton N, Wideman J, Lee F, Banchereau J. Interleukin-7 induces the proliferation of normal human B-cell precursors. Blood. 1991 Nov 1;78(9):2229–2238. [PubMed]
  • Villablanca JG, Anderson JM, Moseley M, Law CL, Elstrom RL, LeBien TW. Differentiation of normal human pre-B cells in vitro. J Exp Med. 1990 Jul 1;172(1):325–334. [PMC free article] [PubMed]
  • Ryan DH, Nuccie BL, Ritterman I, Liesveld JL, Abboud CN. Cytokine regulation of early human lymphopoiesis. J Immunol. 1994 Jun 1;152(11):5250–5258. [PubMed]
  • Cicuttini FM, Martin M, Salvaris E, Ashman L, Begley CG, Novotny J, Maher D, Boyd AW. Support of human cord blood progenitor cells on human stromal cell lines transformed by SV40 large T antigen under the influence of an inducible (metallothionein) promoter. Blood. 1992 Jul 1;80(1):102–112. [PubMed]
  • Thalmeier K, Meissner P, Reisbach G, Falk M, Brechtel A, Dörmer P. Establishment of two permanent human bone marrow stromal cell lines with long-term post irradiation feeder capacity. Blood. 1994 Apr 1;83(7):1799–1807. [PubMed]
  • Sutherland HJ, Eaves CJ, Lansdorp PM, Thacker JD, Hogge DE. Differential regulation of primitive human hematopoietic cells in long-term cultures maintained on genetically engineered murine stromal cells. Blood. 1991 Aug 1;78(3):666–672. [PubMed]
  • Issaad C, Croisille L, Katz A, Vainchenker W, Coulombel L. A murine stromal cell line allows the proliferation of very primitive human CD34++/CD38- progenitor cells in long-term cultures and semisolid assays. Blood. 1993 Jun 1;81(11):2916–2924. [PubMed]
  • Baum CM, Weissman IL, Tsukamoto AS, Buckle AM, Peault B. Isolation of a candidate human hematopoietic stem-cell population. Proc Natl Acad Sci U S A. 1992 Apr 1;89(7):2804–2808. [PMC free article] [PubMed]
  • Collins LS, Dorshkind K. A stromal cell line from myeloid long-term bone marrow cultures can support myelopoiesis and B lymphopoiesis. J Immunol. 1987 Feb 15;138(4):1082–1087. [PubMed]
  • Broxmeyer HE, Douglas GW, Hangoc G, Cooper S, Bard J, English D, Arny M, Thomas L, Boyse EA. Human umbilical cord blood as a potential source of transplantable hematopoietic stem/progenitor cells. Proc Natl Acad Sci U S A. 1989 May;86(10):3828–3832. [PMC free article] [PubMed]
  • Hows JM, Bradley BA, Marsh JC, Luft T, Coutinho L, Testa NG, Dexter TM. Growth of human umbilical-cord blood in longterm haemopoietic cultures. Lancet. 1992 Jul 11;340(8811):73–76. [PubMed]
  • Lu L, Xiao M, Shen RN, Grigsby S, Broxmeyer HE. Enrichment, characterization, and responsiveness of single primitive CD34 human umbilical cord blood hematopoietic progenitors with high proliferative and replating potential. Blood. 1993 Jan 1;81(1):41–48. [PubMed]
  • Moritz T, Keller DC, Williams DA. Human cord blood cells as targets for gene transfer: potential use in genetic therapies of severe combined immunodeficiency disease. J Exp Med. 1993 Aug 1;178(2):529–536. [PMC free article] [PubMed]
  • Billips LG, Petitte D, Dorshkind K, Narayanan R, Chiu CP, Landreth KS. Differential roles of stromal cells, interleukin-7, and kit-ligand in the regulation of B lymphopoiesis. Blood. 1992 Mar 1;79(5):1185–1192. [PubMed]
  • Henderson AJ, Johnson A, Dorshkind K. Functional characterization of two stromal cell lines that support B lymphopoiesis. J Immunol. 1990 Jul 15;145(2):423–428. [PubMed]
  • Noguchi M, Nakamura Y, Russell SM, Ziegler SF, Tsang M, Cao X, Leonard WJ. Interleukin-2 receptor gamma chain: a functional component of the interleukin-7 receptor. Science. 1993 Dec 17;262(5141):1877–1880. [PubMed]
  • Kondo M, Takeshita T, Higuchi M, Nakamura M, Sudo T, Nishikawa S, Sugamura K. Functional participation of the IL-2 receptor gamma chain in IL-7 receptor complexes. Science. 1994 Mar 11;263(5152):1453–1454. [PubMed]
  • Peschon JJ, Morrissey PJ, Grabstein KH, Ramsdell FJ, Maraskovsky E, Gliniak BC, Park LS, Ziegler SF, Williams DE, Ware CB, et al. Early lymphocyte expansion is severely impaired in interleukin 7 receptor-deficient mice. J Exp Med. 1994 Nov 1;180(5):1955–1960. [PMC free article] [PubMed]
  • Hardy RR, Carmack CE, Shinton SA, Kemp JD, Hayakawa K. Resolution and characterization of pro-B and pre-pro-B cell stages in normal mouse bone marrow. J Exp Med. 1991 May 1;173(5):1213–1225. [PMC free article] [PubMed]
  • Gibson LF, Piktel D, Landreth KS. Insulin-like growth factor-1 potentiates expansion of interleukin-7-dependent pro-B cells. Blood. 1993 Nov 15;82(10):3005–3011. [PubMed]
  • Landreth KS, Narayanan R, Dorshkind K. Insulin-like growth factor-I regulates pro-B cell differentiation. Blood. 1992 Sep 1;80(5):1207–1212. [PubMed]
  • Samal B, Sun Y, Stearns G, Xie C, Suggs S, McNiece I. Cloning and characterization of the cDNA encoding a novel human pre-B-cell colony-enhancing factor. Mol Cell Biol. 1994 Feb;14(2):1431–1437. [PMC free article] [PubMed]
  • Nagasawa T, Kikutani H, Kishimoto T. Molecular cloning and structure of a pre-B-cell growth-stimulating factor. Proc Natl Acad Sci U S A. 1994 Mar 15;91(6):2305–2309. [PMC free article] [PubMed]
  • Hannum C, Culpepper J, Campbell D, McClanahan T, Zurawski S, Bazan JF, Kastelein R, Hudak S, Wagner J, Mattson J, et al. Ligand for FLT3/FLK2 receptor tyrosine kinase regulates growth of haematopoietic stem cells and is encoded by variant RNAs. Nature. 1994 Apr 14;368(6472):643–648. [PubMed]
  • Matthews W, Jordan CT, Wiegand GW, Pardoll D, Lemischka IR. A receptor tyrosine kinase specific to hematopoietic stem and progenitor cell-enriched populations. Cell. 1991 Jun 28;65(7):1143–1152. [PubMed]
  • Rosnet O, Marchetto S, deLapeyriere O, Birnbaum D. Murine Flt3, a gene encoding a novel tyrosine kinase receptor of the PDGFR/CSF1R family. Oncogene. 1991 Sep;6(9):1641–1650. [PubMed]
  • Small D, Levenstein M, Kim E, Carow C, Amin S, Rockwell P, Witte L, Burrow C, Ratajczak MZ, Gewirtz AM, et al. STK-1, the human homolog of Flk-2/Flt-3, is selectively expressed in CD34+ human bone marrow cells and is involved in the proliferation of early progenitor/stem cells. Proc Natl Acad Sci U S A. 1994 Jan 18;91(2):459–463. [PMC free article] [PubMed]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences


Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...


  • PubMed
    PubMed citations for these articles
  • Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...