旋转细胞培养系统对老年人表皮干细胞增殖和干性维持的影响  

Effect of rotating cell culture system on proliferation and stemness maintenance of epidermal stem cells in the elderly

作　　者：张敏 董溪溪 黄萌 王朝祥 张鲁岳 曹均凯 Zhang Min;Dong Xixi;Huang Meng;Wang Chaoxiang;Zhang Luyue;Cao Junkai(Medical School of Chinese PLA,Beijing 100853,China;First Medical Center,Chinese PLA General Hospital,Beijing 100853,China;Fifth Medical Center of Chinese PLA General Hospital,Beijing 100071,China)

机构地区：[1]解放军医学院,北京市100853 [2]中国人民解放军总医院第一医学中心,北京市100853 [3]中国人民解放军总医院第五医学中心,北京市100071

出　　处：《中国组织工程研究》2025年第13期2698-2705,共8页Chinese Journal of Tissue Engineering Research

基　　金：国家重点研发计划资助(2020YFC2008900),项目负责人:曹均凯。

摘　　要：背景:由于人口老龄化和老年疾病的增加,对老年人表皮干细胞的研究和利用变得越来越重要。但由于老年人表皮干细胞的增殖和干性能力较弱,限制了其在生物医学和临床研究的应用。目的:探讨旋转细胞培养系统培养的老年人表皮干细胞的增殖和干性维持能力。方法:采用传统平面培养方法获取儿童(7-12岁)和老年人(60岁以上)表皮干细胞,经细胞形态学和细胞免疫荧光鉴定后,通过流式细胞术、CCK-8、细胞克隆形成实验分析儿童和老年人表皮干细胞增殖能力的差异;采用旋转细胞培养系统和3D TableTrix®微载体培养儿童和老年人表皮干细胞,通过活/死染色、细胞倍增效率、细胞倍增时间以及实时定量聚合酶链反应检测细胞增殖情况以及干性标志物的表达。结果与结论:①在平面培养条件下,儿童组表皮干细胞较老年组具有更强的增殖能力;②与平面培养相比,经旋转细胞培养系统动态培养的老年人表皮干细胞的细胞倍增效率更高(P<0.01),细胞倍增时间更短(P<0.01);③旋转细胞培养系统动态培养促使老年人表皮干细胞自组装形成基于微载体支架的三维细胞球,具有和儿童组表皮干细胞相似的增殖情况;④动态培养后的老年人表皮干细胞在干性标记物基因(ITGA6和ITGB1)、基底细胞标记物基因(K5)和分化标记物基因(K10和K1)表达上可达到和儿童组相近的表达水平。结果表明:旋转细胞培养系统动态培养不仅可以提高老年人表皮干细胞的培养效率,促进细胞增殖和自组装成三维细胞球,而且可维持老年人表皮干细胞一定的干性,未发生完全终末分化。BACKGROUND:Due to the aging population and the increase in elderly diseases,research and utilization of epidermal stem cells in the elderly have become increasingly important.However,due to the weak proliferation and drying ability of epidermal stem cells in the elderly,their application in biomedical and clinical research is limited.OBJECTIVE:To investigate the ability of rotating cell culture system to promote the proliferation and maintenance of epidermal stem cells in elderly individuals.METHODS:Epidermal stem cells were obtained from children(7-12 years old)and the elderly(over 60 years old)using traditional planar culture methods.After identification by cell morphology and cell immunofluorescence,the differences in proliferation ability of epidermal stem cells between the children and the elderly were analyzed by flow cytometry,CCK-8 assay,and cell clone formation assay.Elderly epidermal stem cells were cultured using a rotating cell culture system and 3D TableTrix®microcarriers.The cell proliferation and expression of stemness markers were detected by Live/Dead staining,cell doubling efficiency,cell doubling time,and real-time quantitative polymerase chain reaction.RESULTS AND CONCLUSION:(1)Under planar culture conditions,the children group had stronger proliferative ability than the elderly group.(2)Compared with planar culture,the dynamic cultivation of elderly epidermal stem cells using a rotating cell culture system had a higher population doubling(P<0.01)and a shorter population doubling time(P<0.01).(3)The dynamic cultivation of rotating cell culture system promoted the self-assembly of elderly epidermal stem cells to form 3D human epidermal stem cell spheres based on microcarrier scaffolds,which had similar epidermal stem cell proliferation as the children group.(4)After dynamic culture,the expression of stemness marker genes(ITGA6 and ITGB1),basal cell marker genes(K5)and differentiation marker genes(K10 and K1)of elderly epidermal stem cells reached the same level as that of children.The results

关 键 词：人表皮干细胞 老年人 旋转细胞培养系统 微载体 三维动态培养 干性 增殖 

分 类 号：R459.9[医药卫生—治疗学] R318[医药卫生—临床医学] R787