The goal of the study was to investigate changes in expression of selected growth factors tentatively involved in regeneration of haematopoietic tissues (bone marrow and spleen) following cyclophosphamide (CY) damage in the mouse. The bone marrow (BM) and spleen were examined separately, since the regenerating pattern for haematopoietic progenitor cells (HPC) markedly differs in these two haematopoietically active organs after CY. Cytokines assumed to have a stimulatory effect on HPC – stem cell factor (SCF), fetal liver tyrosine kinase 3-ligand (flt3-ligand), thrombopoietin (TPO), stromal cell-derived factor 1 (SDF-1), oncostatin M (OSM) –, a suppressive effect on HPC proliferation – macrophage inflammatory protein-1α (MIP-1α), transforming growth factor-β1 (TGFβ1), tumour necrosis factor-α (TNFα) – and to be involved in migration of HPC (SCF, flt3-ligand, MIP1α, SDF-1) were examined at the level of mRNA expression by means of real-time RT-PCR. The expression of a particular cytokine appears to be similar in both BM and spleen of untreated mice. CY administration changed the expression pattern of the studied genes in BM and spleen. In BM, the levels of mRNAs for SCF and SDF-1 were increased and that for TGFβ1 decreased at time intervals at which HPC are known to proliferate intensively during BM regeneration. In contrast, stimulated proliferation of HPC in spleen was accompanied by increased expression of flt3-ligand and oncostatin M. Upon mobilization of HPC from BM into blood after CY, the expression of SCF, TPO, SDF-1 and TGFβ1 tends to decrease in BM. Accumulation of HPC in spleen is accompanied by increased mRNA for flt3-ligand and OSM. Our findings demonstrate that different cytokines may be involved in the proliferation and mobilization/homing of HPC during recovery after CY damage in BM and spleen.

Keywords:
Cyclophosphamide, Cytokines, Gene expression, Haematopoietic progenitor cells, Real-time RT-PCR
1.
McNiece IK, Langley KE, Zsebo KM: Recombinant human stem cell factor synergises with GM-CSF, G-CSF, IL-3 and epo to stimulate human progenitor cells of the myeloid and erythroid lineages. Exp Hematol 1991;19:226–231.
2.
Heinrich MC, Dooley DC, Freed AC, Band L, Hoatlin ME, Keeble WW, Peters ST, Silvey KV, Ey FS, Kabat D: Constitutive expression of steel factor gene by human stromal cells. Blood 1993;82:771–783.
3.
Toksoz D, Zsebo KM, Smith KA, Hu S, Brankow D, Suggs SV, Martin FH, Williams DA: Support of human hematopoiesis in long-term bone marrow cultures by murine stromal cells selectively expressing the membrane-bound and secreted forms of the human homolog of the steel gene product, stem cell factor. Proc Natl Acad Sci USA 1992;89:7350–7354.
4.
Okomura N, Tsuji K, Ebihara Y, Tanaka I, Sawai N, Koike K, Komiyama A, Nakahata T: Chemotactic and chemokinetic activities of stem cell factor on murine hematopoietic progenitor cells. Blood 1996;87:4100–4108.
5.
Kim CH, Broxmeyer HE: In vitro behavior of hematopoietic progenitor cells under the influence of chemoattractants: Stromal cell-derived factor-1, steel factor, and the bone marrow environment. Blood 1998;91:100–110.
6.
Kobayashi M, Laver JH, Kato T, Miyazaki H, Ogawa M: Thrombopoietin supports proliferation of human primitive hematopoietic cells in synergy with steel factor and/or interleukin-3. Blood 1996;88:429–436.
7.
Graham GJ, Wright EG, Hewick SD, Wolpe SD, Wilkie NM, Donaldson D, Lorimore S, Pragnell IB: Identification and characterization of an inhibitor of haemopoietic stem cell proliferation. Nature 1990;344:442–444.
8.
Jacobsen SE, Ruscetti FW, Ortiz M, Gooya JM, Keller JR: The growth response of Lin-Thy-1+ hematopoietic progenitors to cytokines is determined by the balance between synergy of multiple stimulators and negative cooperation of multiple inhibitors. Exp Hematol 1994;22:985–989.
9.
Jacobsen SE, Veiby OP, Myklebust J, Okkenhaug C, Lyman SD: Ability of flt3 ligand to stimulate the in vitro growth of primitive murine hematopoietic progenitors is potently and directly inhibited by transforming growth factor-beta and tumor necrosis factor-alpha. Blood 1996;87:5016–5026.
10.
Kohn FR, Landkamer GJ, Sladek NE: Effect of the aldehyde dehydrogenase inhibitor, cyanamide, on the ex vivo sensitivity of murine multipotent and committed hematopoietic progenitor cells to mafosfamide (ASTA Z 7557). Immunopharmacol Immunotoxicol 1987;9:163–176.
11.
Hatzfeld J, Li ML, Brown EL, Sookdeo H, Levesque JP, O’Toole T, Gurney C, Clark SC, Hatzfeld A: Release of early human hematopoietic progenitors from quiescence by antisense transforming growth factor beta 1 or Rb oligonucleotides. J Exp Med 1991;174:925–929.
12.
Lataillade JJ, Clay D, Dupuy C, Rigal S, Jasmin C, Bourin P, Le Bousse-Kerdiles MC: Chemokine SDF-1 enhances circulating CD34(+) cell proliferation in synergy with cytokines: Possible role in progenitor survival. Blood 2000;95:756–768.
13.
Brown TJ, Liu J, Brashem-Stein C, Shoyab M: Regulation of granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor expression by oncostatin M. Blood 1993;82:33–37.
14.
Wallace PM, MacMaster JF, Rillema JR, Peng J, Burstein SA, Shoyab M: Thrombocytopoietic properties of oncostatin M. Blood 1995;86:1310–1315.
15.
Clegg CH, Rulffes JT, Wallace PM, Haugen HS: Regulation of an extrathymic T-cell development pathway by oncostatin M. Nature 1996;384:261–263.
16.
Brasel K, McKenna HJ, Morrissey PJ, Charrier K, Morris AE, Lee CC, Williams DE, Lyman SD: Hematologic effects of flt3 ligand in vivo in mice. Blood 1996;88:2004–2012.
17.
Liu F, Poursine-Laurent J, Link DC: The granulocyte colony-stimulating factor receptor is required for the mobilization of murine hematopoietic progenitors into peripheral blood by cyclophosphamide or interleukin-8 but not flt-3 ligand. Blood 1997;90:2522–2528.
18.
Kaushansky K, Lok S, Holly RD, Broudy VC, Lin N, Bailey MC, Forstrom JW, Buddle MM, Oort PJ, Hagen FS, Roth GJ, Papayannopoulou T, Foster DC: Promotion of megakaryocyte progenitor expansion and differentiation by the c-Mpl ligand thrombopoietin. Nature 1994;369:568–571.
19.
Šefc L, Pšenák O, Sýkora V, Nečas E: SDF-1 and its receptor CXCR-4 mRNA in murine hematopoietic tissues during increased migration of progenitors between bone marrow and spleen. Exp Hematol 2001;29(suppl 1):79.
20.
Ponomaryov T, Peled A, Petit I, Taichman RS, Habler L, Sandbank J, Arenzana-Seisdedos F, Magerus A, Caruz A, Fujii N, Nagler A, Lahav M, Szyper-Kravitz M, Zipori D, Lapidot T: Induction of the chemokine stromal-derived factor-1 following DNA damage improves human stem cell function. J Clin Invest 2000;106:1331–1339.
21.
Ghaffar A, Sigel MM, Huggins EM Jr: Resistance of helper T lymphocytes to cyclophosphamide. Int J Immunopharmacol 1985;7:449–454.
22.
Peterson VM, Adamovicz JJ, Elliott TB, Moore MM, Madonna GS, Jackson WE 3rd, Ledney GD, Gause WC: Gene expression of hematoregulatory cytokines is elevated endogenously after sublethal gamma irradiation and is differentially enhanced by therapeutic administration of biologic response modifiers. J Immunol 1994;153:2321–2330.
23.
Orazi A, Gordon MS, John K, Sledge G Jr, Neiman RS, Hoffman R: In vivo effects of recombinant human stem cell factor treatment. A morphologic and immunohistochemical study of bone marrow biopsies. Am J Clin Pathol 1995;103:177–184.
24.
Šefc L, Pšenák O, Sýkora V, Šulc K, Nečas E: Response of hematopoiesis to cyclophosphamide follows highly specific patterns in bone marrow and spleen. J Hematother Stem Cell Res, in press.
25.
Nikcevich AD, Young MR, Ellis NK, Newby M, Wepsic HT: Stimulation of hematopoiesis in untreated and cyclophosphamide treated mice by the inhibition of prostaglandin synthesis. J Immunopharmacol 1986;8:299–313.
26.
Rowley SD, Brashem-Stein C, Andrews R, Bernstein ID: Hematopoietic precursors resistant to treatment with 4-hydroperoxycyclophosphamide: Requirement for an interaction with marrow stroma in addition to hematopoietic growth factors for maximal generation of colony-forming activity. Blood 1993;82:60–65.
27.
Lyman SD, James L, Bos TV, de Vries P, Brasel K, Gliniak B, Hollingsworth LT, Picha KS, McKenna HJ, Splett RR: Molecular cloning of a ligand for the flt3/flk-2 tyrosine kinase receptor: A proliferative factor for primitive hematopoietic cells. Cell 1993;75:1157–1167.
28.
Minehata K, Sekine K, Hara T, Donovan P, Hirabayashi Y, Inoue T, Miyajima A: A role for oncostatin M in the bone marrow hematopoietic microenvironment. Exp Hematol 2001;29 (suppl 1):101.
29.
Byfield JE, Calabro-Jones PM: Resistance of human T-CFUcs to activated cyclophosphamide: A feature common with critical marrow stem cells? Br J Cancer 1985;52:403–405.
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