以HaCaT细胞为种子细胞构建新型人组织工程皮肤

Construction of Human Tissue Engineering Skin by Seed Cell HaCaT

目的:评估以人永生化角朊细胞HaCaT为种子细胞构建新型人组织工程皮肤的可行性。方法:对HaCaT进行无血清培养(DK-SFM)及传代培养;取处于对数生长期的细胞进行接种;利用物理化学方法,采用SD大鼠网状层真皮制备(Acelluar dermal matrix,ADM);应用MTT比色法作生长曲线观察HaCaT细胞长满真皮支架所需时间。HE染色法观察脱细胞前后真皮支架和细胞单层及初步形成的细胞多层。结果:脱细胞真皮的细胞成分消失,组织疏松,为理想的组织皮肤支架;将HaCaT细胞按2×10~5/cm^2密度接种到ADM上,细胞长满支架的时间为5天。

Objective： To evaluate the feasibility of constructing human tissue engineering skin by seed cell human immortal keratinocyte （HaCaT）. Methods： Human immortal skin keratinocytes （HaCaT） were cultured and subcultured in defined keratinocyte-serum free medium（DK-SFM）. The HaCaT keratinocytes in log phase growth period were trypsinized and suspended for seed cell on ADM.The xenogeneic ADM was made from the reticular layer of rat（SD） dermis and prepared by physical and chemical methods; HaCaT reached a conflunent monolayer after seeding, which was detected by MTT method; contrast of effect before and after removel of cells or monolayer or even mutilayer on ADM with HE staining. Results. Cytocomponent of Acelluar dermal matrix （ADM） disappeared,and its tissue loosed, which is an ideal skeleton of tissue skin. The time of monolayer formation was 5 days if HaCaT cells were seeded as 2xl0^5/cm^2. Conclusions ： Human tissue engineering skin can be constructed by seed cell HaCaT on the skeleton of ADM in SD rats.