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目的探讨ES源表皮干细胞(ESC)与高分子材料构建人工皮肤InspectskinⅠ的方法,为化妆品皮肤毒性检验替代模型奠定基础。方法采用嵌入式培养法,先将3-羟基丁酸-co-3-羟基戊酸共聚物(PHBV)支架与原代小鼠成纤维细胞共培养5 d构建真皮,再于表面放置ESC细胞,气-液界面培养7~10 d,形态学和β1整合素、CK10、CK19免疫组化观察。结果成纤维细胞可在真皮支架内粘附,ESC细胞可在支架表面增殖分化形成含基底层、棘层和角化层的复层表皮结构,免疫组化表明基底层以上细胞中表达CK10明显增多,基底层有些细胞仍呈β1整合素和CK19阳性。结论InspectskinⅠ替代皮肤模型是由ESC细胞分化形成的角化复层表皮结构和PHBV真皮结构共同构成的全层皮肤模型。
Objective To explore the method of constructing ESCs Inspectskin Ⅰ by ES-derived epidermal stem cells (ESCs) and macromolecule materials, and lay a foundation for a surrogate model of cosmetic dermal toxicity test. METHODS: Embryonic epithelial cells were co-cultured with primary mouse fibroblasts with 3-hydroxybutyrate-co-3-hydroxyvaleric acid (PHBV) The air-liquid interface was cultured for 7-10 days, and the morphology and β1 integrin, CK10 and CK19 were observed by immunohistochemistry. Results Fibroblasts could adhere in dermis scaffolds. ESC cells proliferated and differentiated into scaffolds with basal, stratum spinosum and keratinized layers. Immunohistochemistry showed that the expression of CK10 was significantly increased in basal lamina propria , Some basal cells still showed β1 integrin and CK19 positive. Conclusion The Inspectskin Ⅰ alternative skin model is a full-thickness skin model composed of keratinized epidermal structure and PHBV dermis formed by ESC cell differentiation.