CAJ | 학술논문

背景：人羊膜上皮细胞具有多系分化能力，是再生医学中重要的细胞来源。目前的研究多集中于对其分化能力的考察，而体外培养过程中羊膜上皮细胞的生物学特征如何变化尚不清楚。 　　目的：分析体外培养对人羊膜上皮细胞生长、表型及向心肌样细胞分化的能力等生物学特性的影响，探讨原代人羊膜上皮细胞干性标志物SSEA-4的表达水平与人羊膜上皮细胞生物学特性变化之间的关联性。 　　方法：使用统一分离方法获得原代羊膜上皮细胞并进行体外培养。利用CCK-8、流式细胞仪及real-time PCR等手段检测不同培养阶段人羊膜上皮细胞的增殖、表型以及向心肌样细胞分化的能力。 　　结果与结论：不同胎儿样本来源的原代人羊膜上皮细胞的SSEA-4表达在26.7%-97%，存在很大的个体差异。并且，随着传代次数的增加，人羊膜上皮细胞的SSEA-4表达水平显著降低，其下降程度与原代SSEA-4的表达水平无关。另外，培养后人羊膜上皮细胞的心肌分化潜能也存在很大个体差异，且其差异与原代人羊膜上皮细胞的 SSEA-4表达水平的高低无关。结果提示，不同胎儿样本来源的原代人羊膜上皮细胞的 SSEA-4表达水平受到个体差异的影响，需要建立更准确的临床样本筛选指标来稳定获得原代高表达 SSEA-4的胎儿样本，以实现对人羊膜上皮细胞的质量监控。另外，体外培养过程中 SSEA-4的表达水平受到培养条件的影响，需要继续优化培养条件以维持其高表达。此外，人羊膜上皮细胞向心肌样细胞分化的能力受到样本个体差异以及培养条件的影响，在今后还需要进一步研究。
배경：인양막상피세포구유다계분화능력，시재생의학중중요적세포래원。목전적연구다집중우대기분화능력적고찰，이체외배양과정중양막상피세포적생물학특정여하변화상불청초。 　　목적：분석체외배양대인양막상피세포생장、표형급향심기양세포분화적능력등생물학특성적영향，탐토원대인양막상피세포간성표지물SSEA-4적표체수평여인양막상피세포생물학특성변화지간적관련성。 　　방법：사용통일분리방법획득원대양막상피세포병진행체외배양。이용CCK-8、류식세포의급real-time PCR등수단검측불동배양계단인양막상피세포적증식、표형이급향심기양세포분화적능력。 　　결과여결론：불동태인양본래원적원대인양막상피세포적SSEA-4표체재26.7%-97%，존재흔대적개체차이。병차，수착전대차수적증가，인양막상피세포적SSEA-4표체수평현저강저，기하강정도여원대SSEA-4적표체수평무관。령외，배양후인양막상피세포적심기분화잠능야존재흔대개체차이，차기차이여원대인양막상피세포적 SSEA-4표체수평적고저무관。결과제시，불동태인양본래원적원대인양막상피세포적 SSEA-4표체수평수도개체차이적영향，수요건립경준학적림상양본사선지표래은정획득원대고표체 SSEA-4적태인양본，이실현대인양막상피세포적질량감공。령외，체외배양과정중 SSEA-4적표체수평수도배양조건적영향，수요계속우화배양조건이유지기고표체。차외，인양막상피세포향심기양세포분화적능력수도양본개체차이이급배양조건적영향，재금후환수요진일보연구。
BACKGROUND:Human amniotic epithelial cells are an important source of cells in regenerative medicine as its multipotentation, but new studies mainly focused on differentiation features and there were little research oneffect of culture in vitro on biological property of amniotic epithelial cells. OBJECTIVE:To analyze the effects of in vitro culture on growth, cellphenotype and differentiation capacity of human amniotic epithelial cells into cardiomyocyte-like cells, and explore the correlation of primarily cultured human amniotic epithelial cells marker SSEA-4 expression level and the change of biological characteristics of human amniotic epithelial cells. METHODS:Primarily cultured human amniotic epithelial cells were obtained from amniotic tissues by using the same separation protocol. Human amniotic epithelial cells were cultured in vitro. The proliferation, cellphenotype and the differentiation capacity of human amniotic epithelial cells into cardiomyocyte-like cells were evaluated by means of cellcounting kit-8, flow cytometry and real-time PCR. RESULTS AND CONCLUSION:The SSEA-4 positive cells in primarily cultured human amniotic epithelial cells from different fetal tissues were between 26.7%-97%, which indicated that there was great individual difference among amniotic tissue samples. Moreover, with passage, the SSEA-4 expression in human amniotic epithelial cells decreased significantly, which did not correlate with the SSEA-4 expression in primarily cultured human amniotic epithelial cells. Results indicated that there was great individual difference in SSEA-4 expression level in primarily cultured human amniotic epithelial cells from different amniotic tissue samples. Thus, it is necessary to set up clinical screening indexes to get samples with higher SSEA-4 expression stably and to control the quality of human amniotic epithelial cells. In addition, during culture period, SSEA-4 expression level was affected by culture conditions. The culture conditions of human amniotic epithelial cells should be optimized to maintain SSEA-4 expression at a high level. In addition, the differentiation capacity of human amniotic epithelial cells into cardiomyocyte-like cells was also affected by individual difference among different samples and culture conditions, which wil be further studied in the future.