Kobayashi M, Yakuwa T, Sasaki K, Sato K, Kikuchi A, Kamo I, Yokoyama Y, Sakuragawa N
Department of Regenerative Medicine, School of Allied Health Sciences, Kitasato University, Nishi-hashimoto, Kanagawa, Japan.
Cell Transplant. 2008;17(3):291-301. doi: 10.3727/096368908784153904.
Side population (SP) cells were isolated by FACS from a human amnion mesenchymal cell (AMC) layer soon after enzyme treatment. The yield of SP cells from AMC layer (AMC-SP cells) was about 0.1-0.2%. AMC-SP cells grew well with cell doublings of 40-80 days of culture. FACS profiles and immunocytostaining showed that AMC-SP cells were composed of two different cells immunologically: HLA I(-)/II(-) and HLA P/II(-). Oct-3/4 was detected in the nucleus of AMC-SP cells, when the culture was examined at the third, sixth, and 10th passages. RT-PCR showed that AMC-SP cells expressed the Oct-4, Sox-2, and Rex-1 genes. Immunocytochemistry revealed that all AMC-SP cells were vimentin+, CK19+, and nestin+. In addition, flow cytometry analysis showed that SP cells had high expression of CD13, CD29, CD44, CD46, CD49b, CD49c, CD49e, CD59, CD140a, and CD166 but low expression of CD 49d, and CD51. No evidence of expression was obtained for CD34, CD45, CD49a, CD56, CD90, CD105, CD106, CD117, CD133, CD271, or Flk-1. Upon appropriate differentiation protocols, AMC-SP cells differentiated to several cell lineages such as neuroectodermal, osteogenic, chondrogenic, and adipogenic cells. These results indicate that AMC-SP cells have multilineage potential to several cell lineages with unique immunological characteristics such as HLA I(-)/II(-) or HLA I+/II(-). AMC-SP cells should be of considerable value for regenerative medicine because they do not induce acute rejection after allotransplantation, they do not cause ethical issues, and there is no limit of supply.
酶处理后不久，通过荧光激活细胞分选术（FACS）从人羊膜间充质细胞（AMC）层中分离出侧群（SP）细胞。AMC层中SP细胞（AMC-SP细胞）的产量约为0.1%-0.2%。AMC-SP细胞生长良好，培养40-80天细胞可倍增。FACS分析图谱和免疫细胞化学显示，AMC-SP细胞在免疫学上由两种不同的细胞组成：HLA I(-)/II(-)和HLA I+/II(-)。在第3、6和10代培养时检查发现，AMC-SP细胞核中可检测到Oct-3/4。逆转录聚合酶链反应（RT-PCR）显示，AMC-SP细胞表达Oct-4、Sox-2和Rex-1基因。免疫细胞化学显示，所有AMC-SP细胞波形蛋白、细胞角蛋白19（CK19）和巢蛋白均呈阳性。此外，流式细胞术分析显示，SP细胞高表达CD13、CD29、CD44、CD46、CD49b、CD49c、CD49e、CD59、CD140a和CD166，但CD49d和CD51表达较低。未获得CD34、CD45、CD49a、CD56、CD90、CD105、CD106、CD117、CD133、CD271或Flk-1表达的证据。经过适当的分化方案，AMC-SP细胞可分化为几种细胞谱系，如神经外胚层、成骨、软骨生成和脂肪生成细胞。这些结果表明，AMC-SP细胞对几种细胞谱系具有多向分化潜能，具有独特的免疫学特征，如HLA I(-)/II(-)或HLA I+/II(-)。AMC-SP细胞在再生医学中应具有相当大的价值，因为它们在同种异体移植后不会引发急性排斥反应，不会引起伦理问题，且供应无限制。
