Journal of the Korean Association of Oral and Maxillofacial Surgeons

The Korean Association of Oral and Maxillofacial Surgeons (대한구강악안면외과학회)
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EFFECTS OF bFGF AND PDGF-BB ON OSTEOBLAST DIFFERENTIATION OF BONE MARROW-DERIVED MESENCHYMAL STEM CELL IN RAT

bFGF, PDGF-BB가 백서 골수기원 간엽 줄기세포의 조직골세포 분화에 미치는 영향에 관한 연구

  • Song, Gin-Ah (Department of Oral and Maxillofacial Surgery, College of Dentistry, Graduate School, Seoul National University) ;
  • Choi, Jin-Young (Department of Oral and Maxillofacial Surgery, College of Dentistry, Graduate School, Seoul National University)
  • 송진아 (서울대학교 치과대학 구강악안면외과학교실) ;
  • 최진영 (서울대학교 치과대학 구강악안면외과학교실)
  • Published : 2006.12.31
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Abstract

In this study we evaluate the effects of bFGF-BB and PDGF on in vitro proliferation, differentiation and mineralization of mesenchymal stem cells (MSCs) from rat. MSCs were prepared from the bone marrow of 6 or 7-week-old male rats with a technique previously described by Maniatopoulos et al. in 1988. Lineage differentiation to osteogenesis, chondrogenesis and adipogenesis were performed. At first, we characterized the cultured cell on passage 1, 3, 5, 7 with immunocytochemical staining using CD29, 44, 34, 45, ${\alpha}$-SMA and type I collagen. And to study the effects of bFGF and PDGF-BB on proliferation, differentiation and mineralization, we seeded the expanded cell at a density of 6 $6{\times}10^3\;cells/cm^2$ to 100-mm dish for evaluation of cell proliferation and MTT assay was carried out on day 2, 4, 7, 9. We also resuspended the cells with same density $(6{\times}10^3\;cells/cm^2)$ to 24 well plates for subculture. On the following day, the attached cells were exposed to 2.5ng/ml bFGF and/or 25ng/ml PDGF-BB daily during 5 days. The osteocalcin (OC) level was assessed and mineral contents were evaluated with alizarin red S staining on subculture day 2, 7, 14, 21. We identified the mesenchymal stem cell from the bone marrow derived cells of rat through their successful multi-differentiation and stable display of its phenotype. And bFGF and PDGF-BB showed the effect that inhibited osteoblastic differentiation and mineralization mildly in above concentration at in vitro culture. This study was supported by grant 04-2004-0120 from the Seoul National University Hospital Research Fund.

Keywords

References

  1. Friedenstein AJ, Piatetzky S, II, Petrakova KV: Osteogenesis in transplants of bone marrow cells. J Embryol Exp Morphol 1966;16(3):381-90
  2. Ohgushi H, Goldberg VM, Caplan AI: Repair of bone defects with marrow cells and porous ceramic. Experiments in rats. Acta Orthop Scand 1989;60(3):334-9 https://doi.org/10.3109/17453678909149289
  3. Dennis JE, Haynesworth SE, Young RG, Caplan AI: Osteogenesis in marrow-derived mesenchymal cell porous ceramic composites transplanted subcutaneously: effect of fibronectin and laminin on cell retention and rate of osteogenic expression. Cell Transplant 1992;1(1):23-32 https://doi.org/10.1177/096368979200100106
  4. Perka C, Schultz O, Spitzer RS, Lindenhayn K, Burmester GR, Sittinger M: Segmental bone repair by tissue-engineered periosteal cell transplants with bioresorbable fleece and fibrin scaffolds in rabbits. Biomaterials 2000;21(11):1145-53 https://doi.org/10.1016/S0142-9612(99)00280-X
  5. Wakitani S, Yamamoto T: Response of the donor and recipient cells in mesenchymal cell transplantation to cartilage defect. Microsc Res Tech 2002;58(1):14-8 https://doi.org/10.1002/jemt.10111
  6. Young RG, Butler DL, Weber W, Caplan AI, Gordon SL, Fink DJ: Use of mesenchymal stem cells in a collagen matrix for Achilles tendon repair. J Orthop Res 1998;16(4):406-13 https://doi.org/10.1002/jor.1100160403
  7. Awad HA, Butler DL, Harris MT, et al.: In vitro characterization of mesenchymal stem cell-seeded collagen scaffolds for tendon repair: effects of initial seeding density on contraction kinetics. J Biomed Mater Res 2000;51(2):233-40 https://doi.org/10.1002/(SICI)1097-4636(200008)51:2<233::AID-JBM12>3.0.CO;2-B
  8. Pittenger MF, Mackay AM, Beck SC, et al.: Multilineage potential of adult human mesenchymal stem cells. Science 1999;284(5411):143-7 https://doi.org/10.1126/science.284.5411.143
  9. Phinney DG, Kopen G, Isaacson RL, Prockop DJ: Plastic adherent stromal cells from the bone marrow of commonly used strains of inbred mice: variations in yield, growth, and differentiation. J Cell Biochem 1999;72(4):570-85 https://doi.org/10.1002/(SICI)1097-4644(19990315)72:4<570::AID-JCB12>3.0.CO;2-W
  10. Jiang Y, Jahagirdar BN, Reinhardt RL, et al.: Pluripotency of mesenchymal stem cells derived from adult marrow. Nature 2002;418(6893):41-9 https://doi.org/10.1038/nature00870
  11. Tropel P, Noel D, Platet N, Legrand P, Benabid AL, Berger F: Isolation and characterisation of mesenchymal stem cells from adult mouse bone marrow. Exp Cell Res 2004;295(2):395-406 https://doi.org/10.1016/j.yexcr.2003.12.030
  12. Barry FP, Murphy JM: Mesenchymal stem cells: clinical applications and biological characterization. Int J Biochem Cell Biol 2004;36(4):568-84 https://doi.org/10.1016/j.biocel.2003.11.001
  13. Laurie SW, Kaban LB, Mulliken JB, Murray JE: Donor-site morbidity after harvesting rib and iliac bone. Plast Reconstr Surg 1984;73(6):933-8 https://doi.org/10.1097/00006534-198406000-00014
  14. Younger EM, Chapman MW: Morbidity at bone graft donor sites. J Orthop Trauma 1989;3(3):192-5 https://doi.org/10.1097/00005131-198909000-00002
  15. Stevenson S, Shaffer JW, Goldberg VM: The humoral response to vascular and nonvascular allografts of bone. Clin Orthop Relat Res 1996(326):86-95
  16. Friedlaender GE, Horowitz MC: Immune responses to osteochondral allografts: nature and significance. Orthopedics 1992;15(10):1171-5
  17. Jootar S, Angchaisuksiri P, Chiewsilp P, Sathapatayavongs B, Chuncharunee S, Tanprasert S: HIV infection after autologous bone marrow transplantation despite HIV-antibody and HIV-antigen screening. Bone Marrow Transplant 1993;12(2):167-8
  18. Carlson ER, Marx RE, Buck BE: The potential for HIV transmission through allogeneic bone. A review of risks and safety. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 1995;80(1):17-23 https://doi.org/10.1016/S1079-2104(95)80010-7
  19. Conrad EU, Gretch DR, Obermeyer KR, et al.: Transmission of the hepatitis-C virus by tissue transplantation. J Bone Joint Surg Am 1995;77(2):214-24 https://doi.org/10.2106/00004623-199502000-00007
  20. Hallfeldt KK, Stutzle H, Puhlmann M, Kessler S, Schweiberer L: Sterilization of partially demineralized bone matrix: the effects of different sterilization techniques on osteogenetic properties. J Surg Res 1995;59(5):614-20 https://doi.org/10.1006/jsre.1995.1213
  21. Spangrude GJ, Heimfeld S, Weissman IL: Purification and characterization of mouse hematopoietic stem cells. Science 1988;241(4861):58-62 https://doi.org/10.1126/science.2898810
  22. Vacanti V, Kong E, Suzuki G, Sato K, Canty JM, Lee T: Phenotypic changes of adult porcine mesenchymal stem cells induced by prolonged passaging in culture. J Cell Physiol 2005;205(2):194-201 https://doi.org/10.1002/jcp.20376
  23. Niemeyer P, Krause U, Fellenberg J, et al.: Evaluation of mineralized collagen and alpha-tricalcium phosphate as scaffolds for tissue engineering of bone using human mesenchymal stem cells. Cells Tissues Organs 2004;177(2):68-78 https://doi.org/10.1159/000079182
  24. Sharma B, Elisseeff JH: Engineering structurally organized cartilage and bone tissues. Ann Biomed Eng 2004;32(1):148-59 https://doi.org/10.1023/B:ABME.0000007799.60142.78
  25. Richmon JD, Sage AB, Shelton E, Schumacher BL, Sah RL, Watson D: Effect of growth factors on cell proliferation, matrix deposition, and morphology of human nasal septal chondrocytes cultured in monolayer. Laryngoscope 2005;115(9):1553-60 https://doi.org/10.1097/01.MLG.0000175541.31131.A5
  26. Salgado AJ, Coutinho OP, Reis RL: Bone tissue engineering: state of the art and future trends. Macromol Biosci 2004;4(8):743-65 https://doi.org/10.1002/mabi.200400026
  27. Van de Putte KA, Urist MR: Osteogenesis in the interior of intramuscular implants of decalcified bone matrix. Clin Orthop Relat Res 1965;43:257-70 https://doi.org/10.1097/00003086-196500430-00026
  28. Marx RE, Carlson ER, Eichstaedt RM, Schimmele SR, Strauss JE, Georgeff KR: Platelet-rich plasma: Growth factor enhancement for bone grafts. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 1998;85(6):638-46 https://doi.org/10.1016/S1079-2104(98)90029-4
  29. Nash TJ, Howlett CR, Martin C, Steele J, Johnson KA, Hicklin DJ: Effect of platelet-derived growth factor on tibial osteotomies in rabbits. Bone 1994;15(2):203-8 https://doi.org/10.1016/8756-3282(94)90709-9
  30. Radomsky ML, Aufdemorte TB, Swain LD, Fox WC, Spiro RC, Poser JW: Novel formulation of fibroblast growth factor-2 in a hyaluronan gel accelerates fracture healing in nonhuman primates. J Orthop Res 1999;17(4):607-14 https://doi.org/10.1002/jor.1100170422
  31. Jingushi S, Heydemann A, Kana SK, Macey LR, Bolander ME: Acidic fibroblast growth factor (aFGF) injection stimulates cartilage enlargement and inhibits cartilage gene expression in rat fracture healing. J Orthop Res 1990;8(3):364-71 https://doi.org/10.1002/jor.1100080308
  32. Wang JS, Aspenberg P: Basic fibroblast growth factor enhances bone-graft incorporation: dose and time dependence in rats. J Orthop Res 1996;14(2):316-23 https://doi.org/10.1002/jor.1100140222
  33. Chaudhary LR, Hofmeister AM, Hruska KA: Differential growth factor control of bone formation through osteoprogenitor differentiation. Bone 2004;34(3):402-11 https://doi.org/10.1016/j.bone.2003.11.014
  34. Maniatopoulos C, Sodek J, Melcher AH: Bone formation in vitro by stromal cells obtained from bone marrow of young adult rats. Cell Tissue Res 1988;254(2):317-30
  35. Chen SJ, Bradley ME, Lee TC: Chemical hypoxia triggers apoptosis of cultured neonatal rat cardiac myocytes: modulation by calciumregulated proteases and protein kinases. Mol Cell Biochem 1998;178(1-2):141-9 https://doi.org/10.1023/A:1006886421795
  36. Deschaseaux F, Gindraux F, Saadi R, Obert L, Chalmers D, Herve P: Direct selection of human bone marrow mesenchymal stem cells using an anti-CD49a antibody reveals their CD45med,low phenotype. Br J Haematol 2003;122(3):506-17 https://doi.org/10.1046/j.1365-2141.2003.04469.x
  37. Zhang Y, Lin HK, Frimberger D, Epstein RB, Kropp BP: Growth of bone marrow stromal cells on small intestinal submucosa: an alternative cell source for tissue engineered bladder. BJU Int 2005;96(7):1120-5 https://doi.org/10.1111/j.1464-410X.2005.05741.x
  38. Spector M: Musculoskeletal connective tissue cells with muscle: expression of muscle actin in and contraction of fibroblasts, chondrocytes, and osteoblasts. Wound Repair Regen 2001;9(1):11-8 https://doi.org/10.1046/j.1524-475x.2001.00011.x
  39. Andreassen TT, Oxlund H: Local anabolic effects of growth hormone on intact bone and healing fractures in rats. Calcif Tissue Int 2003;73(3):258-64 https://doi.org/10.1007/s00223-002-2074-6
  40. Kimoto T, Hosokawa R, Kubo T, Maeda M, Sano A, Akagawa Y: Continuous administration of basic fibroblast growth factor (FGF-2) accelerates bone induction on rat calvaria--an application of a new drug delivery system. J Dent Res 1998;77(12):1965-9 https://doi.org/10.1177/00220345980770120301
  41. Bak B, Jorgensen PH, Andreassen TT: Dose response of growth hormone on fracture healing in the rat. Acta Orthop Scand 1990;61(1):54-7 https://doi.org/10.3109/17453679008993067
  42. Takita H, Tsuruga E, Ono I, Kuboki Y: Enhancement by bFGF of osteogenesis induced by rhBMP-2 in rats. Eur J Oral Sci 1997;105(6):588-92 https://doi.org/10.1111/j.1600-0722.1997.tb00222.x
  43. Unda FJ, Martin A, Hernandez C, et al.: FGFs-1 and -2, and TGF beta 1 as inductive signals modulating in vitro odontoblast differentiation Evidence that fibroblast growth factors 1 and 4 participate in regulation of cardiogenesis Skeletal growth factors. Adv Dent Res 2001;15(4):34-7 https://doi.org/10.1177/08959374010150010801
  44. Zhang L, Leeman E, Carnes DC, Graves DT: Human osteoblasts synthesize and respond to platelet-derived growth factor. Am J Physiol 1991;261(2 Pt 1):C348-54 https://doi.org/10.1152/ajpcell.1991.261.2.C348
  45. Hanada K, Dennis JE, Caplan AI: Stimulatory effects of basic fibroblast growth factor and bone morphogenetic protein-2 on osteogenic differentiation of rat bone marrow-derived mesenchymal stem cells. J Bone Miner Res 1997;12(10):1606-14 https://doi.org/10.1359/jbmr.1997.12.10.1606
  46. Hakki SS, Nohutcu RM, Hakki EE, Berry JE, Akkaya MS, Somerman MJ: Dexamethasone and basic-fibroblast growth factor regulate markers of mineralization in cementoblasts in vitro. J Periodontol 2005;76(9):1550-8 https://doi.org/10.1902/jop.2005.76.9.1550