人羊水干细胞分离方法及其生物学特性研究

Isolation and Biological Characterization of Human Amniotic Fluid-derived Stem Cells

目的建立体外培养人羊水来源CD117阳性干细胞的方法,初步探讨CD117阳性干细胞的特性。方法通过孕中期羊膜腔穿刺获得86例羊水标本。采用CD117磁珠分选表达CD117抗原的羊水干细胞。对经过反复冻存的CD117阳性干细胞进行核型分析。分别经成脂诱导和成骨诱导分化,再分别使用油红O染色、茜素红染色。结果从61例标本中成功分离培养出CD117阳性细胞,经核型分析显示其染色体核型正常。CD117阳性细胞经成骨和成脂诱导后,茜素红和油红O染色阳性,说明可以向脂肪细胞和成骨细胞分化。结论基于本研究,孕中期羊水CD117阳性细胞的分选成功率为70.93%。CD117阳性细胞在体外培养中保持形态和遗传学的稳定。在特殊培养条件下,人源羊水CD117阳性干细胞在体外,可以诱导分化成为脂肪细胞和成骨细胞,可能应用于细胞移植和再生医学。

Objective To establish in vitro culture procedure for human amniotic fluid-derived CD117 positive stem cells,and to identify the characteristics of CD117 positive stem cells.Methods 86 amniotic fluid samples（10 mL of each） were obtained by second-trimester amniocentesis.Isolation of amniotic fluid-derived stem cells expressing CD117 antigen was performed via magnetic cell sorting using the CD117 MicroBead Kit.The karyotype of CD117 positive stem cells was analysed throughrepeated freezing.Adipogenic differentiation of these CD117 positive stem cells was displayed by Oil Red O staining.Osteogeneic differentiation of these CD117 positive stem cells was confirmed by Alizarin Red staining.Results The CD117 positive stem cells were successfully isolated and cultured from 61 samples,with all showing normal karyotype.Product analysis of specific staining confirmed that under specific culture mediums,these cells could be successfully induced to differentiate into adipocytes and osteocytes.Conclusion Based on this study,we estimate that isolating CD117 positive stem cells from second-trimester amniotic fluid obtained by amniocentesis has a success rate of 70.93%.These cells maintain morphological and genetic stability in vitro.Human amniotic fluid-derived CD117 positive stem cells have the ability to differentiate in vitro into adipocytes and osteocytes under specific cuLture mediums and may be applied in cell transplantation and regenerative medicine.