사람유래 지방줄기세포와 양수줄기세포의 분리 및 지방세포로의 분화에 따른 특성 비교
Characterization and differentiation into adipocytes of mesenchymal stem cells (MSCs) from human adipose tissue and amniotic fluid
2
조회
0
다운로드
3
인용
0
참고문헌
키워드
초록
목적: 본 연구자는 지방조직에서 분리한 세포와 산전진단검사를 하고 남은 양수 세포를 이용하여 중간엽 줄기세포의 분리방법을 확립하고, 지방유래 줄기세포와 양수유래 줄기세포의 특성과 지방세포로의 분화 후 그 특징을 평가하여 향후 줄기세포를 이용한 여러 분야의 학문에 이용할 수 있는 세포 치료제로써의 가능성을 확인하고자 하였다. 연구 방법: 지방 흡입술을 통해 획득한 사람의 지방에서 분리한 지방유래 줄기세포와 산전진단검사 후 남은 양수세포에서 분리한 양수유래 줄기세포를 유세포 분석 및 노화도 측정을 통해 동정하였고, 지방세포로 분화를 유도한 다음 면역형광세포 염색법과 역전사 중합효소 연쇄반응 (RT-PCR)으로 그 결과를 평가하였다. 결과: 지방과 양수에서 중간엽 줄기세포가 성공적으로 분리, 배양 및 증식함을 확인하였고, 지방유래 줄기세포와 양수유래 줄기세포 모두 노화가 지연되는 것을 β-galactosidase를 이용해 관찰하였다. 두 가지의 줄기세포를 지방세포 (adipocyte)로 분화한 후 분화 정도를 분석하였을 때, 발현 정도의 차이는 있었지만 지방세포 특이적 Oil red O 염색과 유전자 발현이 양성으로 확인되었다. 결론: 지방유래 줄기세포와 양수유래 줄기세포의 특정 세포로의 분화 능력을 평가함으로써, 향후 재생의학과 조직공학 및 성형의학에 이르는 세포 치료에 있어서 주요한 공급원이 될 수 있음을 확인함으로써, 세포 치료제로의 적용에 있어 중요한 기초자료가 될 것으로 사료된다.
Abstract
Objective: Mesenchymal stem cells (MSCs) are potentially very useful for regenerative and reparative medicine as well as therapeutic possibilities. The aim of this study is to examine the ability of ADSCs and AFSCs to be phenotypically and functionally differentiated into adipocyte and to determine the appropriate stem cell source and conditions for efficient adipocyte regeneration. Methods: Adipogenic differentiation was induced by culturing confluent ADSCs and AFSCs in adipogenic medium for 2~4 weeks. During the differentiation inducing period, we evaluated the successful adipogenesis by performing immunocytochemistry and RT-PCR to detect the lipid producibility and several adipogenic gene expressions including lipoprotein lipase (LPL), peroxisome proliferator-activated receptor γ2 (PPAR γ2) and adiponectin. Results: ADSCs and AFSCs are expanded easily in vitro and exhibited a fibroblast-like morphology as previously known in MSCs from bone marrow and a commercial source. Flow cytometric analysis showed that ADSCs and AFSCs expressed several CD marker antigens similar to those observed on bone marrow-derived MSCs. Adipogenic induction of ADSCs and AFSCs resulted in the extended cell morphology, intracellular staining of an established lipid dye Oil Red O, and expression of adipocyte-specific genes. Conclusion: Both ADSCs and AFSCs successfully differentiate in vitro into adipogenic cells in the presence of the lineage-specific induction factors although ADSC showed the greater capability. Therefore, the results suggest that ADSCs and AFSCs may be an excellent choice for many future tissue engineering strategies and cell-based therapies.
참고문헌 (26)
- Zuk PA, "Multilineage cells from human adipose tissue: implications for cell-based therapies", Tissue Eng, 7, pp.211~228 (2001)
- Tsai MS, "Isolation of human multipotent mesenchymal stem cells from second-trimester amniotic fluid using a novel two-stage culture protocol", Hum Reprod, 19, pp.1450~1456 (2004)
- Zuk PA, "Human Adipose Tissue Is a Source of Multipotent Stem Cells", Mol Biol Cell, 13, pp.4279~4295 (2002)
- Hauner H, "Glucocorticoids and insulin promote the differentiation of human adipocyte precursor cells into fat cells", J Clin Endocrinol Metabol, 64, pp.832~835 (1987)
- Pittenger MF, "Multilineage potential of adult human mesenchymal stem cells", Science, 284, pp.143~147 (1999)
- Bruder SP, "Bone regeneration by implantation of purified, culture-expanded human mesenchymal stem cells", J Orthop Res, 16, pp.155~162 (1998)
- Lin TM, "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, pp.92~102 (2005)
- Rodriguez LV, "Clonogenic multipotent stem cells in human adipose tissue differentiate into functional smooth muscle cells", Proc Natl Acad Sci U S A, 103, pp.12167~12172 (2006)
- Delo DM, "Amniotic fluid and placental stem cells", Methods Enzymology, 419, pp.426~438 (2006)
- Kaviani A, "The amniotic fluid as a source of cells for fetal tissue engineering", J Pediatr Surg, 36, pp.1662~1665 (2001)
- Kaviani A, "Fetal tissue engineering from amniotic fluid", J Am Coll Surg, 196, pp.592~597 (2003)
- Mizuno H, "Mesengenic potential and future clinical perspective of human processed lipoaspirate cells", J Nippon Med Sch, 70, pp.300~306 (2003)
- Katz AJ, "Emerging approaches to the tissue engineering of fat", Clin Plast Surg, 26, pp.587~603 (1999)
- Katz AJ, "Cell surface and transcriptional characterization of human adipose-derived adherent stromal (hADAS) cells", Stem cells, 23, pp.412~423 (2005)
- Varma MJ, "Phenotypical and functional characterization of freshly isolated adipose tissue-derived stem cells", Stem Cells Dev, 16, pp.91~104 (2007)
- Yoshimura K, "Characterization of freshly isolated and cultured cells derived from the fatty and fluid portions of liposuction aspirates", J Cell Physiol, 208, pp.64~76 (2006)
- Gimble JM, "Adipose-derived stem cells for regenerative medicine", Circ Res, 100, pp.1249~1260 (2007)
- In't Anker PS, "Amniotic fluid as a novel source of mesenchymal stem cells for therapeutic transplantation", Blood, 102, pp.1548~1549 (2003)
- Prusa, AR, "Neurogenic cells in human amniotic fluid", Am J Obstet Gynecol, 191, pp.309~314 (2004)
- Prusa AR, "Oct-4-expressing cells in human amniotic fluid:a new source for stem cell research?", Hum Reprod, 18, pp.1489~1493 (2003)
- Delo DM, "Amniniotic fluid and placental stem cells", Methods Enzymol, 419, pp.426~438 (2006)
- De Ugarte DA, "Differential expression of stem cell mobilization-associated molecules on multi-lineage cells from adipose tissue and bone marrow", Immunol Lett, 89, pp.267~270 (2003)
- De Coppi P, "Isolation of amniotic stem cell lines with potential for therapy", Nat Biotechnol, 25, pp.100~106 (2007)
- Kern S, "Comparative analysis of mesenchymal stem cells from bone marrow, umbilical cord blood, or adipose tissue", Stem Cells, 24, pp.1294~1301 (2006)
- Gimble JM, "Adipose-derived adult stem cells: isolation, characterization, and differentiation potential", Cytotherapy, 5, pp.362~369 (2003)
- Gronthos S, "Characterization of surface protein expression on human adipose tissue-derived stromal cells", J Cell Physiol, 189, pp.54~63 (2001)