以正常人皮肤成纤维细胞滋养层培养人角质形成细胞的研究

目的探索和建立人皮肤成纤维细胞(HDF)滋养层培养人角质形成细胞的技术方法。方法在HDF滋养层制备中，对丝裂霉素C的较适浓度、同一株不同代数的HDF滋养层、滋养层的细胞密度对正常人表皮角质形成细胞（NHEK）生长的影响及NHEK的最低接种密度进行了初步的探索。结果①终浓度为10μg/mL的丝裂霉素C是制备HDF滋养层的较适浓度。②同一株1～20代的HDF滋养层对不同来源的NHEK均有较好的促生长增殖作用，1～20代HDF的培养代数与9例NHEK生长达80％融合时间(平均为11.1d)之间相关无显著性(r=-0.27,P>0.05)。③用HDF滋养层培养NHEK可减少原代培养NHEK的接种量，用NHEK最小接种密度是0.1×10

Human Keratinocytes Grown in Mitomycin C-treated Human Dermal Fibroblast Feeder Layer

Objective To establish a method for the cultivation of keratinocytes in human dermal fibroblast (HDF) feeder layer.Methods The seeding efficacy, growth and confluence of normal human epidermal keratinocytes (NHEK) in mitomycin C-treated HDF feeder layers from various passages of same HDF line were observed.Results ① The concentration of mitomycin C suitable for HDF feeder layer was 10 μ g/mL.② The first to 20th passages of the same HDF line were preferable to establish different allogenic keratinocyte cultures.There was no correlation between the number of passages and periods needed for NHEK to reach 80％ confluence (r=-0.27,P >0.05).③ HDF feeder layer could improve the efficient growth of primary keratinocyte culture plated at very low seeding densities (0.1× 104 cells/cm2).Conclusions Different allogenic keratinocytes can grow in HDF feeder layers from various passages of the same HDF line.HDF feeder layer is able to improve the seeding efficacy of human keratinocytes, which may be the reason of primary keratinocyte culture plated at very low seeding densities.