Summary
Granulocyte colony-stimulating factor (G-CSF) was linked to NHS-biotin to yield biotinylated G-CSF (b-G-CSF), which retained the ability to stimulate colony formation by normal bone marrow (BM) cells in methylcellulose. The use of streptivadin-phycoerythrin conjugate in conjunction with flow cytometry demonstrated that the binding of biotnylated G-CSF to its receptor is saturable, competitive, and specific. A 100-fold molar excess of unlabeled G-CSF almost completely inhibited the binding of the biotinylated G-CSF to the human leukemia cell line U937, which is known to posses the G-CSF receptor. G-CSF receptors were clearly detected by flow cytometry on adult human peripheral granulocytes and monocytes, but not on lymphocytes. Using this method, the expression of G-CSF receptors on hematopoietic progenitor cells in bone marrow and umbilical cord blood, detected as CD34-positive (CD34+) cells, were examined. A small but significant number of CD34+ cells were detected among the bone marrow mononuclear cells and umbilical-cord-blood mononuclear cells (4.28%±0.31%, 1.09%±0.20%, respectively). The percentage of CD34+ BM mononuclear cells was significantly higher than for cord blood mononuclear cells (P<0.01). These CD34+ cells were then analyzed by biotinylated G-CSF binding. CD34+ cells from bone marrow contained 25.8%±7.9% G-CSF receptor positive cells and those from cord blood possessed 29.2% ±7.0% of G-CSF receptor-positive cells. The difference was not statistically significant.
Similar content being viewed by others
References
Begley CD, Metcalf D, Nicola NA (1987) Primary human myeloid leukemia cells: comparative responsiveness to proliferative stimulation by GM-CSF or G-CSF and membrane expression of CSF receptors. Leukemia 1:1–8
Berenson RJ, Andrews RG, Bensinger WI, Kalamasz D, Knitter G (1988) Antigen CD34+ marrow cells engraft lethally irradiated baboons. J Clin Invest 81:951–955
Broxmeyer HE, Douglas GW, Hangoc G, Cooper S, Bard J, English D, Arny M, Thomas L, Boyse EA (1989) Human umbilical cord blood as a potential source of transplantable hematopoietic stem/progenitor cells. Proc Natl Acad Sci USA 86:3828–3832
Budel LM, Touw IP, Delwel R, Lowenberg B (1989) Granulocyte colony-stimulating factor receptors in human acute myelocytic leukemia. Blood 74:2668–2673
Civin IC, Strauss LC, Brovall C, Fackler MJ, Schwartz JF, Shaper JH (1984) Antigen analysis of hematopoiesis. III A hematopoietic progenitor, cell surface antigen defined by a monoclonal antibody raised against KG-1a cells. J Immunol 133:157–165
Civin CI, Banquerigo ML, Strauss LC, Loken MR (1987) Antigenic analysis of hematopoiesis. VI. Flow cytometric characterization of My-10-positive progenitor cells in normal human bone marrow. Exp Hematol 15:10–17
Clark SC, Kamen R (1987) The human hematopoietic colony-stimulating factors. Science 236:1229–1237
Cohen AM, Krisztina MZ, Inoue H, Hines D, Boone TC, Chazin VR, Tsai L, Ritch T, Souza LM (1987)In vivo stimulation of granulopoiesis by recombinant human granulocyte colony-stimulating factor. Proc Natl Acad Sci USA 84:2484–2488
Ema H, Suda T, Niura Y, Nakauchi H (1990) Colony formation of clone-sorted human hematopoietic progenitors. Blood 75:1941–1946
Ema H, Suda T, Nagayoshi K, Miura Y, Civin CI, Nakauchi H (1990) Target cells for granulocyte colon-stimulating factor, interleukin-3, and interleukin-5 in differentiation pathways of neutrophil and eosinophils. Blood 76:1956–1961
Gluckman E, Broxmeyer HA, Auerbach AD, Friedman HS, Douglas GW, Devergie A, Esperou H, Thierry D, Socie G, Lehn P, Cooper S, English D, Kurtzberg J, Bard J, Boyse E (1989) Hematopoietic reconstitution in a patient with Fanconi's anemia by means of umbilical-cord blood from HLA-identical sibling. N Engl J Med 321:1174–1178
Hoshino S, Oshimi K, Tsudo M, Miyasaka M, Teramura M, Masuda M, Motoji T, Mizoguchi H (1990) Flow cytometric analysis of expression of interleukin-2 receptor β chain (p70–75) on various leukemic cells. Blood 76:767–774
Ikebuchi K, Clark SC, Ihle JN, Souza LM, Ogawa M (1988) Granulocyte colony-stimulating factor enhances interleukin-3 dependent proliferation of multipotential hemopoietic progenitors. Proc Natl Acad Sci USA 85:3445–3449
Kondo S, Okamura S, Ohhara N, Niho Y (1990) Direct action of human granulocyte colony-stimulating factor on mature human neutrophils: Flow-cytometric analysis. Res Commun Chem Pathol Pharmacol 67:249–257
Kondo S, Okamura S, Harada M, Niho Y (1991) Human granulocyte colony-stimulating factor receptors in acute myelogenous leukemia. Eur J Haematol 46:223–230
Leary AG, Ogawa M, Straus LC, Civin CI (1984) Single cell origin of multilineage colonies in culture. J Clin Invest 74:2193–2197
Metcalf D (1988) The molecular control of cell division, differentiation, commitment and maturation in haematopoietic cells. Nature 339:27–30
Nakahata T, Ogawa M (1982) Hemopoietic colony-forming cells in umbilical cord blood with extensive capability to generate mono- and multipotential hemopoietic progenitors. J Clin Invest 70:1324–1328
Newman W, Beall LW, Bertolini DR, Cone JL (1989) Modulation of TGF-β type 1 receptor: flow cytometric detection with biotinylated TGF-β. J Cell Physiol 141:170–180
Newman W, Beall LD, Levine MA, Cone JL, Randhawa ZI, Bertolini DR (1989) Biotinylated parathyroid hormone as a probe for the parathyroid hormone receptor: structure function analysis and detection of specific binding to cultured bone cells by flow cytometry. J Biol Chem 264:16359–16365
Nicola NA (1987) Why do hemopoietic growth factors receptors interact with each other? Immunol Today 8:134–140
Nicola NA, Metcalf D (1985) Binding of125I-labeled granulocyte colony-stimulating factor to normal murine hemopoietic cells. J Cell Physiol 124:313–321
Ohhara N, Okamura S, Hayashi S, Otsuka T, Niho Y (1988) Granulocyte-macrophage colony-formation in vitro using human non-phagocytic bone marrow cells. Res Exp Med 188:405–409
Okamura S, Crane F, Jamal N, Messner HA, Mak TW (1980) Single-cell immunofluorescence assay for terminal transferase: human leukemic and non-leukemic cells. Br J Cancer 41:159–167
Okamura S, Omori F, Haga K, Baba H, Sugimachi K, Niho Y (1990) Influence of bestatin on production of colony-stimulating factor from human peripheral mononuclear cells in vitro. Acta Oncol 29:795–797
Omori F, Okamura S, Takaku F, Niho Y (1989) Measurement of human G-CSF by enzyme-linked immunosorbent assay using monoclonal antibody. Res Exp Med 189:163–171
Park LS, Waldron PE, Friend D, Sassenfeld HM, Price V, Anderson D, Cosman D, Andrews RG, Bernstein ID, Urdal DL (1989) Interleukin-3, GM-CSF, and G-CSF receptor expression on cell lines and primary leukemic cells: receptor heterogeneity and relationship to growth factor responsiveness. Blood 74:56–65
Saeland S, Caux C, Favre C, Aubry JP, Mannoni P, Pebusque MJ, Gentilhomme O, Otsuka T, Yokota T, Arai K, Banchereau J, Vries JE (1988) Effects of recombinant human interleukin-3 on CD34-enriched normal hematopoietic progenitors and on myeloblastic leukemia cells. Blood 72:1580–1588.
Shimoda K, Okamura S, Omori F, Mizuno Y, Hara T, Aoki N, Ueda K, Niho Y (1991) Granulocyte clony-stimulating factor in cerebrospinal fluid from patients with meningitis. Blood 77:2214–2217
Sieff CA (1990) Hematopoietic growth factors. J Clin Invest 79:1459–1557
Souza LM, Boone TC, Gabrilove J, Lai PH, Zsebo KM, Murdock DC, Chazin VR, Bruszewski J, Lu H, Chen KK, Barendt J, Platzer E, Moore MAS, Meltelzmann R, Welte K (1986) Recombinant human granulocyte colony-stimulating factor: effects on normal and leukemic myeloid cells. Science 232:61–65
Strauss LC, Rowley SD, La Russa VF, Sharkis SJ, Stuart RK, Civin CI (1986) Antigen analysis of hematopoiesis. V. Characterization of My-10 antigen expression by normal lymphohematopoietic progenitor cells. Exp Hematol 14:878–886
Welte K, Bonilla MA, Gillio AP, Boone TC, Potter GK, Gabrilove JL, Moore MAS, O'Reilly RJ, Souza LM (1987) Recombinant human granulocyte colony-stimulating factor: effects on hematopoiesis in normal and cyclophosphamide-treated primates. J Exp Med 165:941–948
Wongnum AW, Lansdorp PM, Humphries RK, Krystal G (1990) Detection and isolation of the erythropoietin receptor using biotinylated erythropoietin. Blood 76:697–705
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Shimoda, K., Okamura, S., Harada, N. et al. Detection of the granulocyte colony-stimulating factor receptor using biotinylated granulocyte colony-stimulating factor: presence of granulocyte colony-stimulating factor receptor on CD34-positive hematopoietic progenitor cells. Res. Exp. Med. 192, 245–255 (1992). https://doi.org/10.1007/BF02576281
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02576281