Development and Reproduction (한국발생생물학회지:발생과생식)

The Korean Society of Developmental Biology (한국발생생물학회)
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Comparison of Neural Cell Differentiation of Human Adipose Mesenchymal Stem Cells Derived from Young and Old Ages

연령별 지방 중간엽 유래 줄기세포의 신경세포로의 분화 능력 비교

  • 조정윤 (알앤엘바이오 중앙연구소) ;
  • 강성근 (알앤엘바이오 중앙연구소) ;
  • 최인수 (알앤엘바이오 중앙연구소) ;
  • 라정찬 (알앤엘바이오 중앙연구소)
  • Published : 2009.12.31
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Abstract

Recently, adipose mesenchymal stem cells (AdMSC) that are similar to bone marrow MSC and blood derived MSC are thought to be another source for stem cell therapy. However, the diseases that can be applied for stem cells therapy are age-dependent degenerative diseases. Accordingly, the present study investigated the growth and differentiation potential to neural cells of human AdMSC (hAdMSC) obtained from aged thirty, forty and fifty. The growth of cells and cell viability were measured by passage and neural differentiation of hAdMSC was induced in neural differentiation condition for 10 days. Our results demonstrated that cell number, viability and morphology were not different from hAdMSC by age and passage. Immunofluorescence analysis of neural cell marker (TuJ1, NSE, Sox2, GFAP or MAP2) demonstrated no significant differences in neural cell differentiation by age and passage. As the number of passage was increased, the mRNA level of MAP2 and Sox2 was decreased in hAdMSC from age of 50 compared to hAdMSC from age of 30. In conclusion, the present study demonstrated that ability of neural cell differentiation of hAdMSC was maintained with ages, suggesting that autologous stem cells from aged people can be applied for stem cell therapy with age-dependent neural disease with the same stem cell quality and ability as stem cell derived from young age.

최근 골수와 혈액으로 유래된 중간엽 줄기세포와 비슷한 능력을 가지는 것으로 알려진 지방 유래 중간엽줄기세포가 새로운 세포 치료제로 떠오르고 있다. 하지만 줄기세포를 이용하여 치료하려는 질병은 나이가 들어감에 따라 발병하는 퇴행성 질환들이 대부분인데, 노화가 진행됨에 따라 줄기세포의 능력이 차이가 있다고 알려져 있다. 이에 본 연구에서는 노화가 일어남에 따라 발생되는 신경성 질환을 자가 유래 지방 중간엽 줄기세포를 이용하여 치료함에 있어서 노화가 진행됨에 따라 얻어진 지방줄기세포가 세포학적으로 변화는 없는지에 대해 줄기세포 성장능, 생존율과 신경세포로의 분화유도 능력을 비교하였다. 30대, 40대, 50대에서 사람 지방 유래 줄기세포를 분리 배양하여 연령별 계대에 따른 세포수와 생존율을 측정하고, 줄기세포 성장능력을 비교 분석하였고, 지방 줄기세포를 신경세포 배양 조건 하에서 10일 동안 배양하여 신경 분화능력을 연령별로 비교하였다. 실험결과, 세포수와 생존율, 세포 모양이 연령과 계대별에 의해 차이가 없다는 것을 확인하였다. 신경 분화 후 면역형광염색법을 통해 분석한 결과, 연령에 따른 신경 분화능력의 차이가 관찰되지 않았다. 분자 유전적학으로 신경세포 마커의 발현을 mRNA 수준에서 분석한 결과, 연령별 간의 차이가 몇 개의 유전자 발현을 제외하고는 차이가 발견되지 못했다. 하지만 계대가 진행될수록 50대군의 줄기세포에서 MAP2와 Sox2의 mRNA 발현이 30대군의 줄기세포에 비해 상대적으로 낮게 발현됨이 확인되었다. 결론적으로 자가 지방 중간엽 줄기세포의 신경세포 분화능력이 연령에 상관없이 차이가 없음이 관찰되었으며, 이는 나이 든 사람으로부터 얻어진 지방 줄기세포도 젊은 사람에서 얻어진 세포와 마찬가지 능력으로 자가 세포 치료제로 사용될 수 있다는 점을 말해주고 있다.

Keywords

References

  1. Ashijan PH, Elbarbary AS, Edmonds B, DeUgarte D, Zhu M, Zuk PA, Lorenz HP, Benhaim P, Hedrick MH (2003) In vitro differentiation of human processed lipoaspirate cells into early neural progenitors. Plast Reconstr Surg 111:1922-1231. https://doi.org/10.1097/01.PRS.0000055043.62589.05
  2. Black IB, Woodbury D (2001) Adult rat and human bone marrow stromal cells differentiate into neurons. Blood Cells Mol Dis 27:632-636. https://doi.org/10.1006/bcmd.2001.0423
  3. Bobis S, Jarocha D, Majka M. (2006) Mesenchymal stem cell: characteristic and clinical applications. Folia Histochem Cytobiol 44:215-230.
  4. Bonab MM, Alimoghaddam K, Talebian F, Ghaffari SH, Ghavamzadeh A, Nikbin B (2006) Aging mesenchymal stem cell in vitro. BMC Cell Biol 7:14-21. https://doi.org/10.1186/1471-2121-7-14
  5. Chhabra P, Mirmira RG, Brayman KL (2009) Regenerative medicine and tissue engineering: contribution of stem cells in organ transplantation. Curr Opin Organ Transplant 14:46-50. https://doi.org/10.1097/MOT.0b013e328322f989
  6. Colter DC, Class R, DiGirolamo CM, Prockop DJ (2000) Rapid expansion of recyciling stem cells in culture of plastic-adherent cells from human bone marrow. Pro Natl Acad Sci USA 97:3213-3218. https://doi.org/10.1073/pnas.97.7.3213
  7. Deng YB, Yuan QT, Liu XG, Liu XL, Liu Y, Liu ZG, Zhang C (2005) Functional recovery after rhesus monkey spinal cord injury by transplantiation of bone marrow mesenchymal stem cell-derived neurons. Chin Med J 118, 18:1533-1541.
  8. Fibble WE, Noort WA (2003) Mesenchymal stem cells hematopoietic stem cell transplantation. Ann NY Acad Sci 996:235-244. https://doi.org/10.1111/j.1749-6632.2003.tb03252.x
  9. Gordon D, Scolding NJ (2009) Human mesenchymal stem cell culture for neural transplantation. Methods Mol Biol 549:103-118. https://doi.org/10.1007/978-1-60327-931-4_8
  10. Gimble J, Guilak F (2003) Adipose-derived adult stem cells: isolation, characterization, and differentiation potential. Cytotherapy 5:362-369. https://doi.org/10.1080/14653240310003026
  11. Honczarenko M, Le Y, Swierkowski M, Ghiran I, Glodek AM, Silberstein LE (2006) Human bone mrow stromal cells express a distinct set of biologically functional chemokin receptors. Stem Cells 24:1030-1041. https://doi.org/10.1634/stemcells.2005-0319
  12. Izadpanah R, Kaushal D, Kriedt C, Tsien F, Patel B, Dufour J, Bunnell BA (2008) Long term in vitro expansion alters the biology of adult mesenchymal stem cells. Cancer Res 68:4229-4238. https://doi.org/10.1158/0008-5472.CAN-07-5272
  13. Lin TM, Tsai JL, Lin SD, Lai CS, Chang CC (2005) Accelerated growth and prolonged lifespan of adipose tissue-derived human mesenchymal stem cells in a medium using reduced calcium and antioxidants. Stem Cells Dev 14:92-102. https://doi.org/10.1089/scd.2005.14.92
  14. Miki T, Lehmann T, Cai H, Stolz DB, Strom SC (2005) Stem cell characteristics of amniotic epithelial cells. Stem Cells 23:1549-1559. https://doi.org/10.1634/stemcells.2004-0357
  15. Minguell JJ, Erices A, Conget P (2001) Mesenchymal stem cells. Exp Biol Med 226:507-520.
  16. Mizuno H (2009) Adipose-derived stem cells for tissue repair and regeneration: ten years of research and a literature review. J Nippon Med Sch 76:56-66. https://doi.org/10.1272/jnms.76.56
  17. Nakagami H, Morishita R, Maeda K, Kikuchi Y, Ogihara T, Kaneda Y (2006) Adipose tissue-derived stromal cells as a novel option for regenerative cell therapy. J Atherosceler Thrombo 13:77-81. https://doi.org/10.5551/jat.13.77
  18. Pittenger MF, Mackay AM, Beck SC, Jaiswal RK, Douglas R, Mosca JD, Moorman MA, Simonetti DW, Craig S, Marshak DR (1999) Multilineage potential of adult human mesenchymal stem cells. Science 284:143-147. https://doi.org/10.1126/science.284.5411.143
  19. Safford KM, Rice HE (2005) Stem cell therapy for neurologic disoders: therapeutic potential of adipose-derived stem cells. Curr Drug Targets 6:57-62. https://doi.org/10.2174/1389450053345028
  20. Sanchez-Ramos J, Song S, Cardozo-Pelaez F, Hazzi C, Stedeford T, Willing A, Freeman TB, Saporta S, Janssen W, Patel N, Cooper DR, Sanberg PR (2000) Adult bone marrow stromal cells differnetiatied into neural cells in vitro. Exp Neurol 164:247-256. https://doi.org/10.1006/exnr.2000.7389
  21. Shi YY, Nacamuli RP, Salim A, Longaker MT (2005) The osteogenic potential of adipose-derived mesenchymal cells is maintained with aging. Plast Reconstr Surg 116: 1686-1696. https://doi.org/10.1097/01.prs.0000185606.03222.a9
  22. Stenderup K, Justesen J, Clausen C, Kassen M (2003) Aging is associated with decreased maximal life span and accelerated senescence of bone marrow stromal cells. Bone 33:919-926. https://doi.org/10.1016/j.bone.2003.07.005
  23. Wagner W, Horn P, Castoldi M, Diehlmann A, Bork S, Saffrich R, Benes V, Blake J, Pfister S, Eckstein V, Ho AD (2008) Replicative senescence of mesenchymal stem cells: a continuous and organized process. PLoS One 3:2213-2225. https://doi.org/10.1371/journal.pone.0002213
  24. Williams BA, Keating A (2008) Cell therapy for age-related disorders: Myocardial infarction and stroke-a mini review. Gerontology 54:300-311. https://doi.org/10.1159/000156223
  25. Yuan XD, Yan X, Yang H, Gao ZQ, Li KH, Han Q, Lu S, Chen XW, Jiang H, Zhao CH (2009) In vitro study of directional inducible differentiation of inner ear hair cells from human adipose-derived mesenchymal stem cells. Zhonghua Er Bi Yan Hou Tou Jing Wai Ke Za Zhi 44:323-328.
  26. Zuk PA, Zhu M, Mizuno H, Huang J, Futrell JW, Katz AJ, Benhaim P, Lorenz HP, Hedrick MH (2001) Multilineage cells from human adipose tissue: implications for cellbased therapies. Tissue Eng 7:211-228. https://doi.org/10.1089/107632701300062859
  27. Zuk PA, Zhu M, Ashjian P, De Ugarte DA, Huang JI, Mizuno H, Alfonso ZC, Fraser JK, Benhaim P, Hedrick MH (2002). Human adipose tissue is a source of multipotent stem cells. Mol Biol Cell 13:4279-4295. https://doi.org/10.1091/mbc.E02-02-0105
  28. 라정찬 (2008) 계대 배양된 사람 지방 유래 줄기세포의 안정성 및 하지 허혈동물 모델에서 혈관 신생에 관한 연구. 제주대학교, p 99.