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目的用携带人乳头瘤病毒11型(HPV11)基因组的角质形成细胞(HPV11.HaCaT细胞),在体外建立皮肤类似培养物三维模型,观察HPV11衣壳蛋白L1是否表达,为进一步建立体外病毒复制系统和药效筛选模型奠定基础。方法用人成纤维细胞和I型鼠尾胶原制备真皮类似物凝胶,接种HPV11.HaCaT细胞进行三维培养。同时以正常HaCaT细胞作对照。15天时取皮肤类似物进行冰冻切片,HE染色进行组织结构检查,免疫组化法检查角蛋白10、泛角蛋白以及HPV11L1蛋白表达。结果HE染色显示,HPV11.HaCaT细胞和正常HaCaT细胞的层数明显增多。免疫组化显示,两种细胞的皮肤类似物角蛋白10、泛角蛋白表达全部阳性。HPV11.HaCaT细胞表达L1蛋白阳性,而正常HaCaT表达L1蛋白阴性。结论三维培养方式可诱导HPV11.HaCaT细胞分化,皮肤类似培养物出现细胞分化标志,并且表达HPV11型衣壳蛋白L1。
OBJECTIVE: To establish a three-dimensional model of skin-like culture in vitro with keratinocytes carrying human papillomavirus type 11 (HPV11) genome (HPV11.HaCaT cells) and observe the expression of HPV11 capsid protein L1. To further establish an in vitro virus replication system And pharmacodynamic screening model to lay the foundation. Methods Human dermal analog gels were prepared from human fibroblasts and type I rat tail collagen and inoculated with HPV11.HaCaT cells for three-dimensional culture. Meanwhile, normal HaCaT cells were used as control. At 15 days, the skin analogs were frozen and stained with HE for histological examination. The expressions of keratin 10, pan-keratin and HPV11L1 protein were examined by immunohistochemistry. Results HE staining showed that the number of layers of HPV11.HaCaT cells and normal HaCaT cells increased significantly. Immunohistochemistry showed that the skin analogues keratin 10 and pan-keratin of both cells were all positive. HPV11.HaCaT cells express L1 protein positive, while normal HaCaT expression L1 protein negative. Conclusion Three-dimensional culture method can induce the differentiation of HPV11.HaCaT cells, the signs of cell differentiation in skin-like cultures, and the expression of HPV type 11 capsid protein L1.