Effective ex vivo expansion of hematopoietic stem cells using osteoblast-differentiated mesenchymal stem cells is CXCL12 dependent
Seiji Mishima
Central of Clinical Laboratory, Shimane University Hospital
Search for more papers by this authorAtsushi Nagai
Department of Laboratory Medicine, Shimane University Faculty of Medicine, Izumo, Japan
Search for more papers by this authorSk Abdullah
Department of Laboratory Medicine, Shimane University Faculty of Medicine, Izumo, Japan
Search for more papers by this authorChikashi Matsuda
Central of Clinical Laboratory, Shimane University Hospital
Search for more papers by this authorTakeshi Taketani
Division of Blood Transfusion, Shimane University Hospital
Search for more papers by this authorShunichi Kumakura
Department of Education for Rural Medicine, Shimane University Faculty of Medicine
Search for more papers by this authorHiroshi Shibata
Central of Clinical Laboratory, Shimane University Hospital
Search for more papers by this authorSeung U. Kim
Division of Neurology, UBC Hospital, University of British Columbia, Vancouver, Canada
Medical Research Institute, Chung-Ang University College of Medicine, Seoul, Korea
Search for more papers by this authorJunichi Masuda
Department of Laboratory Medicine, Shimane University Faculty of Medicine, Izumo, Japan
Search for more papers by this authorSeiji Mishima
Central of Clinical Laboratory, Shimane University Hospital
Search for more papers by this authorAtsushi Nagai
Department of Laboratory Medicine, Shimane University Faculty of Medicine, Izumo, Japan
Search for more papers by this authorSk Abdullah
Department of Laboratory Medicine, Shimane University Faculty of Medicine, Izumo, Japan
Search for more papers by this authorChikashi Matsuda
Central of Clinical Laboratory, Shimane University Hospital
Search for more papers by this authorTakeshi Taketani
Division of Blood Transfusion, Shimane University Hospital
Search for more papers by this authorShunichi Kumakura
Department of Education for Rural Medicine, Shimane University Faculty of Medicine
Search for more papers by this authorHiroshi Shibata
Central of Clinical Laboratory, Shimane University Hospital
Search for more papers by this authorSeung U. Kim
Division of Neurology, UBC Hospital, University of British Columbia, Vancouver, Canada
Medical Research Institute, Chung-Ang University College of Medicine, Seoul, Korea
Search for more papers by this authorJunichi Masuda
Department of Laboratory Medicine, Shimane University Faculty of Medicine, Izumo, Japan
Search for more papers by this authorAbstract
Effective ex vivo expansion of hematopoietic stem cells (HSCs) is a prerequisite for HSC transplantation. Growth and maintenance of HSC is dependent on cytokine and niche factors. We investigated whether mesenchymal stem cells (MSCs) or osteogenic cytokine-differentiated MSCs play a role in HSC expansion. We used the human HM3.B10 (B10) MSC cell line and the osteoblast-differentiated B10 (Ost-B10) as a feeder layer and examined ex vivo expansion of CD34+CD38− HSCs obtained from peripheral blood (PB) and cord blood (CB) with or without several growth cytokines. Both undifferentiated B10 and Ost-B10 cells exhibited similar effects on total HSC expansion; however, Ost-B10 demonstrated a higher potency in CD34+CD38− cell-specific proliferation in the presence of cytokines compared to undifferentiated B10 HSCs. Colony-forming cell assay and long-term culture initiating cell assay revealed that Ost-B10 displayed multipotent differentiation and enabled long-term ex vivo culture of HSCs. We next examined the relationship between HSC expansion and the presence of various chemokines. CXCL4 and CXCL12 expression were increased in Ost-B10 cells compared with the B10 cells. CD34+CD38− cells were significantly increased with CXCL12, but not CXCL4 treatment. siRNA inhibition of CXCL12 decreased CXCL12 secretion in both B10 and Ost-B10, whereas expansion of CD34+CD38− cells was decreased in Ost-B10 alone. These results demonstrated that ex vivo expansion of HSCs may be highly effective through osteoblast-differentiated MSCs acting as a feeder layer, and likely operates through the CXCL12 chemokines signaling pathway.
Supporting Information
Table S1. Primer pairs to analyze expression of growth factors, morphogens, and adhesion molecules.
Table S2. Primers and TaqMan probe to detect CC chemokines.
Table S3. Primers and TaqMan probe to detect CXC chemokines.
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References
- 1 Kessinger A, Armitage JO, Landmark JD, Weisenburger DD. Reconstitution of human hematopoietic function with autologous cryopreserved circulating stem cells. Exp Hematol 1986; 14: 192–6.
- 2 Korbling M, Dorken B, Ho AD, Pezzutto A, Hunstein W, Fliedner TM. Autologous transplantation of blood-derived hemopoietic stem cells after myeloablative therapy in a patient with Burkitt’s lymphoma. Blood 1986; 67: 529–32.
- 3 Anderlini P, Donato M, Chan KW, Huh YO, Gee AP, Lauppe MJ, Champlin RE, Korbling M. Allogeneic blood progenitor cell collection in normal donors after mobilization with filgrastim: the m.D. Anderson cancer center experience. Transfusion 1999; 39: 555–60.
- 4 Osawa M, Hanada K, Hamada H, Nakauchi H. Long-term lymphohematopoietic reconstitution by a single cd34-low/negative hematopoietic stem cell. Science 1996; 273: 242–5.
- 5 Heike T, Nakahata T. Ex vivo expansion of hematopoietic stem cells by cytokines. Biochim Biophys Acta 2002; 1592: 313–21.
- 6 Mourcin F, Grenier N, Mayol JF, Lataillade JJ, Sotto JJ, Herodin F, Drouet M. Mesenchymal stem cells support expansion of in vitro irradiated CD34(+) cells in the presence of SCF, FLT3 ligand, TPO and IL3: potential application to autologous cell therapy in accidentally irradiated victims. Radiat Res 2005; 164: 1–9.
- 7 Piacibello W, Sanavio F, Severino A, et al. Engraftment in nonobese diabetic severe combined immunodeficient mice of human CD34(+) cord blood cells after ex vivo expansion: evidence for the amplification and self-renewal of repopulating stem cells. Blood 1999; 93: 3736–49.
- 8 McNiece IK, Almeida-Porada G, Shpall EJ, Zanjani E. Ex vivo expanded cord blood cells provide rapid engraftment in fetal sheep but lack long-term engrafting potential. Exp Hematol 2002; 30: 612–16.
- 9 Zhang J, Niu C, Ye L, et al. Identification of the haematopoietic stem cell niche and control of the niche size. Nature 2003; 425: 836–41.
- 10 Arai F, Hirao A, Ohmura M, Sato H, Matsuoka S, Takubo K, Ito K, Koh GY, Suda T. Tie2/angiopoietin-1 signaling regulates hematopoietic stem cell quiescence in the bone marrow niche. Cell 2004; 118: 149–61.
- 11 Uchida N, Fleming WH, Alpern EJ, Weissman IL. Heterogeneity of hematopoietic stem cells. Curr Opin Immunol 1993; 5: 177–84.
- 12 Brandt JE, Galy AH, Luens KM, et al. Bone marrow repopulation by human marrow stem cells after long-term expansion culture on a porcine endothelial cell line. Exp Hematol 1998; 26: 950–61.
- 13 Kawada H, Ando K, Tsuji T, et al. Rapid ex vivo expansion of human umbilical cord hematopoietic progenitors using a novel culture system. Exp Hematol 1999; 27: 904–15.
- 14 Yildirim S, Boehmler AM, Kanz L, Mohle R. Expansion of cord blood CD34+ hematopoietic progenitor cells in coculture with autologous umbilical vein endothelial cells (HUVEC) is superior to cytokine-supplemented liquid culture. Bone Marrow Transplant 2005; 36: 71–9.
- 15 Calvi LM, Adams GB, Weibrecht KW, et al. Osteoblastic cells regulate the haematopoietic stem cell niche. Nature 2003; 425: 841–6.
- 16 Nagai A, Kim WK, Lee HJ, Jeong HS, Kim KS, Hong SH, Park IH, Kim SU. Multilineage potential of stable human mesenchymal stem cell line derived from fetal marrow. PLoS ONE 2007; 2: e1272.
- 17 Alakel N, Jing D, Muller K, Bornhauser M, Ehninger G, Ordemann R. Direct contact with mesenchymal stromal cells affects migratory behavior and gene expression profile of CD133+ hematopoietic stem cells during ex vivo expansion. Exp Hematol 2009; 37: 504–13.
- 18 Pelus LM, Fukuda S. Chemokine-mobilized adult stem cells; defining a better hematopoietic graft. Leukemia 2008; 22: 466–73.
- 19 Blank U, Karlsson G, Karlsson S. Signaling pathways governing stem-cell fate. Blood 2008; 111: 492–503.
- 20 Yoshihara H, Arai F, Hosokawa K, et al. Thrombopoietin/MPL signaling regulates hematopoietic stem cell quiescence and interaction with the osteoblastic niche. Cell Stem Cell 2007; 1: 685–97.
- 21 Noort WA, Kruisselbrink AB, In’t Anker PS, et al. Mesenchymal stem cells promote engraftment of human umbilical cord blood-derived CD34(+) cells in nod/scid mice. Exp Hematol 2002; 30: 870–8.
- 22 Feugier P, Jo DY, Shieh JH, MacKenzie KL, Rafii S, Crystal RG, Moore MA. Ex vivo expansion of stem and progenitor cells in co-culture of mobilized peripheral blood CD34+ cells on human endothelium transfected with adenovectors expressing thrombopoietin, c-kit ligand, and Flt-3 ligand. J Hematother Stem Cell Res 2002; 11: 127–38.
- 23 Gan OI, Murdoch B, Larochelle A, Dick JE. Differential maintenance of primitive human scid-repopulating cells, clonogenic progenitors, and long-term culture-initiating cells after incubation on human bone marrow stromal cells. Blood 1997; 90: 641–50.
- 24 Shimakura Y, Kawada H, Ando K, Sato T, Nakamura Y, Tsuji T, Kato S, Hotta T. Murine stromal cell line hess-5 maintains reconstituting ability of ex vivo-generated hematopoietic stem cells from human bone marrow and cytokine-mobilized peripheral blood. Stem Cells 2000; 18: 183–9.
- 25 Ara T, Tokoyoda K, Sugiyama T, Egawa T, Kawabata K, Nagasawa T. Long-term hematopoietic stem cells require stromal cell-derived factor-1 for colonizing bone marrow during ontogeny. Immunity 2003; 19: 257–67.
- 26 Nagasawa T, Hirota S, Tachibana K, Takakura N, Nishikawa S, Kitamura Y, Yoshida N, Kikutani H, Kishimoto T. Defects of b-cell lymphopoiesis and bone-marrow myelopoiesis in mice lacking the CXC chemokine PBSF/SDF-1. Nature 1996; 382: 635–8.
- 27 Broxmeyer HE, Cooper S, Hangoc G, Kim CH. Stromal cell-derived factor-1/cxcl12 selectively counteracts inhibitory effects of myelosuppressive chemokines on hematopoietic progenitor cell proliferation in vitro. Stem Cells Dev 2005; 14: 199–203.
- 28 Patel VP, Kreider BL, Li Y, et al. Molecular and functional characterization of two novel human C-C chemokines as inhibitors of two distinct classes of myeloid progenitors. J Exp Med 1997; 185: 1163–72.
- 29 Kortesidis A, Zannettino A, Isenmann S, Shi S, Lapidot T, Gronthos S. Stromal-derived factor-1 promotes the growth, survival, and development of human bone marrow stromal stem cells. Blood 2005; 105: 3793–801.
- 30 Strieter RM, Polverini PJ, Arenberg DA, Kunkel SL. The role of CXC chemokines as regulators of angiogenesis. Shock 1995; 4: 155–60.
- 31 Zhang J, Lu SH, Liu YJ, Feng Y, Han ZC. Platelet factor 4 enhances the adhesion of normal and leukemic hematopoietic stem/progenitor cells to endothelial cells. Leuk Res 2004; 28: 631–8.
- 32 Sugiyama T, Kohara H, Noda M, Nagasawa T. Maintenance of the hematopoietic stem cell pool by CXCL12-CXCR4 chemokine signaling in bone marrow stromal cell niches. Immunity 2006; 25: 977–88.
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