한국간담췌외과학회지 (Annals of Hepato-Biliary-Pancreatic Surgery)

  • 제13권4호
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  • Pages.198-204
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  • 2009
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  • 2508-5778(pISSN)
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  • 2508-5859(eISSN)
한국간담췌외과학회 (The Korean Association of Hepato-Biliary-Pancreatic Surgery)
권호 표지

Human Adipose Tissue-Derived Mesenchymal Stem Cells 배양 시 효율적인 Extracellular Matrix의 증명

The Extracellular Matrix Affected Proliferation and Cell Adhesion of Human Adipose Tissue Derived Mesenchymal Stem Cells in vitro

  • 민선옥 (연세대학교 의과대학 외과학교실) ;
  • 이상우 (연세대학교 의과대학 외과학교실) ;
  • 최새별 (고려대학교 의과대학 외과학교실) ;
  • 김경식 (연세대학교 의과대학 외과학교실)
  • Min, Seon-Ok (Department of Surgery, Yonsei University College of Medicine) ;
  • Lee, Sang-Woo (Department of Surgery, Yonsei University College of Medicine) ;
  • Choi, Sae-Byeol (Department of Surgery, Korea University College of Medicine) ;
  • Kim, Kyung-Sik (Department of Surgery, Yonsei University College of Medicine)
  • 발행 : 2009.12.31

⟨ 이전 논문 다음 논문 ⟩

초록

Purpose: Human mesenchymal stem cells (hMSCs) have the potency for self-renewal and differentiation into various kinds of cells. The hMSCs are obtained from the various tissues, including adipose tissue, bone marrow and cord blood. The extracellular matrix (ECM) is an important factor that affects cell adherence, growth, migration, apoptosis and differentiation both in vitro and vivo. The adipose-derived mesenchymal stem cells (AD-MSCs) have CD29 (integrin) on the cell surface, which is the receptor for fibronectin. The aim of this study is to validate the efficacy of ECM, and especially fibronectin, for cell expansion. Methods: The AD-MSCs were obtained from the abdominal fat of humans. These cells were seeded onto culture plates coated with fibronectin-Human (FN) and plates without ECM (control). The cells were incubated for 3 passages and the cellular morphology was simultaneously observed with microscopy. CCK-8 assay was performed to compare the proliferation ability in each condition at the same passage. Immunocytochemistry staining for integrin-beta1 was performed to observe the cell to cell interaction. Results: The hAD-MSCs in the FN-coated and non-coated plates exhibited cytoplasm staining for integrin-beta1. In all the cultures, extended fibroblastic-shaped cells that turned into rhomboid cells were most frequently observed. The cell growth rates for the non coated culture plate were lower than those for the FN coated plates. After 72 hour culture under the different coated concentrations of FN and the non coated condition (control), the control group had a lower growth rate. In the culture with a FN coated plate, a significant change was observed as compared with that of the control group. We observed an increase in cell proliferation, with a maximum of 140%, on the FN coated plate by performing CCK-8 assay. In comparison, integrin ${\beta}1$ on the cells was more expressed in the FN-coated plates than that in the non-coated plates. Conclusion: The cell morphology can be changed faster in the FN coated culture plates than that in the non coated culture plates. Because proliferation and adhesion with FN can enhance the expansion, the culture within a FN coated plate is needed to encourage hAD-MSCs to proliferate in vitro.

키워드

참고문헌

  1. Pittenger MF, Mackay AM, Beck SC, et al. Multilineage potential of adult human mesenchymal stem cells. Science 1999;284:143-147. https://doi.org/10.1126/science.284.5411.143
  2. Song GB, Ju Y, Soyama H. Growth and proliferation of bone marrow mesenchymal stem cells affected by type I collagen, fibronectin and bFGF. Materials Science & Engineering CB 2008;28:1467-1471. https://doi.org/10.1016/j.msec.2008.04.005
  3. Hutchings H, Ortega N, Plouët J. Extracellular matrix-bound vascular endothelial growth factor promotes endothelial cell adhesion, migration, and survival through integrin ligation. FASEB J 2003;17:1520-1522. https://doi.org/10.1096/fj.02-0691fje
  4. Goessler UR, Bugert P, Bieback K, et al. Integrin expression in stem cells from bone marrow and adipose tissue during chondrogenic differentiation. Int J Mol Med 2008;21:271-279.
  5. Hynes RO. Integrins: Bidirectional, allosteric signaling machines. Cell 2002;110:673-687. https://doi.org/10.1016/S0092-8674(02)00971-6
  6. Flaim CJ, Chien S, Bhatia SN. An extracellular matrix microarray for probing cellular differentiation. Nat Methods 2005; 2:119-125. https://doi.org/10.1038/nmeth736
  7. Taléns-Visconti R, Bonora A, Jover R, et al. Human mesenchymal stem cells from adipose tissue: Differentiation intohepatic lineage. Toxicology in vitro 2007;21:324-329. https://doi.org/10.1016/j.tiv.2006.08.009
  8. Kolf CM, Cho E, Tuan RS. Mesenchymal stromal cells. Biology of adult mesenchymal stem cells: regulation of niche, self- renewal and differentiation. Arthritis Res Ther 2007;9: 204. https://doi.org/10.1186/ar2116
  9. Gimble JM, Katz AJ, Bunnell BA. Adipose-derived stem cells for regenerative medicine. Circ Res 2007;100:1249-1260. https://doi.org/10.1161/01.RES.0000265074.83288.09
  10. Prockop DJ. Marrow stromal cells as stem cells for continual renewal of nonhematopoietic tissues and as potential vectors for gene therapy. J Cell Biochem Suppl 1998;30-31:284-285.
  11. Etheridge SL, Spencer GJ, Heath DJ, Genever PG. Expression profiling and functional analysis of Wnt signaling mechanisms in mesenchymal stem cells. Stem Cells 2004;22:849-860. https://doi.org/10.1634/stemcells.22-5-849
  12. Qian L, Saltzman WM. Improving the expansion and neuronal differentiation of mesenchymal stem cells through culture surface modification. Biomaterials 2004;25:1331-1337. https://doi.org/10.1016/j.biomaterials.2003.08.013
  13. Choi FJ, Kwon JY, Kim Ho, Kim SH, Choi YJ, Cho JA. The characterization of the mesenchymal stem cells derived from fat, cord blood, placenta tissues. Korean J HBP Surgery 2006; 10:1-6.
  14. Kern S, Eichler H, Stoeve J, Klüter H, Bieback K. Comparative analysis of mesenchymal stem cells from bone marrow, umbilical cord blood, or adipose tissue. Stem Cells 2006; 24:1294-1301. https://doi.org/10.1634/stemcells.2005-0342
  15. Deng W, Obrocka M, Fischer I, Prockop DJ. in vitro differentiation of human marrow stromal cells into early progenitors of neural cells by conditions that increase intracellular cyclic AMP. Biochem Biophys Res Commun 2001;282:148-152. https://doi.org/10.1006/bbrc.2001.4570
  16. Banfi A, Muraglia A, Dozin B, Mastrogiacomo M, Cancedda R, Quarto R. Proliferation kinetics and differentiation potential of ex vivo expanded human bone marrow stromal cells: implications for their use in cell therapy. Exp Hematol 2000; 28:707-715. https://doi.org/10.1016/S0301-472X(00)00160-0
  17. Digirolamo CM, Stokes D, Colter D, Phinney DG, Class R, Prockop DJ. Propagation and senescence of human marrow stromal cells in culture: a simple colony-forming assay identifies samples with the greatest potential to propagate and differentiate. Br J Haematol 1999;107:275-281. https://doi.org/10.1046/j.1365-2141.1999.01715.x
  18. Conget PA, Minguell JJ. Phenotypical and functional properties of human bone marrow mesenchymal progenitor cells. J Cell Physiol 1999;181:67-73. https://doi.org/10.1002/(SICI)1097-4652(199910)181:1<67::AID-JCP7>3.0.CO;2-C
  19. Hynes RO. Integrins: versatility, modulation, and signaling in cell adhesion. Cell 1992;69:11-25. https://doi.org/10.1016/0092-8674(92)90115-S
  20. Harnett EM, Alderman J, Wood T. The surface energy of various biomaterials coated with adhesion molecules used in cell culture. Colloids Surf B-Biointerfaces 2007;55:90-97. https://doi.org/10.1016/j.colsurfb.2006.11.021
  21. Salasznyk RM, Williams WA, Boskey A, Batorsky A, Plopper GE. Adhesion to vitronectin and collagen I promotes osteogenic differentiation of human mesenchymal stem cells. J Biomed Biotechnol 2004;2004:24-34. https://doi.org/10.1155/S1110724304306017