ES细胞源性表皮干细胞与胶原海绵构建组织工程皮肤

Reconstructed Tissue-engineered Skin with ES Cell-derived Epidermal Stem Cells and Collagen Sponge

【目的】以胚胎干细胞(ES细胞)源性表皮干细胞为种子细胞与胶原海绵构建组织工程皮肤,探讨其对皮肤缺损修复的作用。【方法】胎鼠皮肤成纤维细胞与胶原海绵构建类真皮,植入小鼠全层皮肤缺损创面,以生物膜为载体,把羊膜诱导后带有核标记的ES细胞源性表皮干细胞覆盖在类真皮上,术后1～8周连续取材,HE染色,β1整合素、CK15、CK19、CK10和CEA免疫荧光双标和免疫组化观察。【结果】植入后3周,创面完全长合,较厚新生皮完全覆盖创面,基底层细胞增生,形成许多大小不一的细胞柱伸向真皮层,新生表皮中可见核标记的细胞呈β1整合素、CK15、CK19阳性,真皮中的管腔样结构呈核荧光和β1整合素、CEA免疫组化双标阳性,4～8周新生表皮基底层细胞呈CK19、CK10阳性,新生表皮下可见毛囊样、皮脂腺样结构。【结论】ES细胞源性表皮干细胞为种子细胞与胶原海绵构建的组织工程皮肤可以修复缺损皮肤,在其下真皮层有分化为毛囊样、皮脂腺样和汗腺样的结构,但是是否来源于ES细胞源性表皮干细胞仍需进一步证实。

[Objective] Using tissue-engineered skin reconstructed with ES cell-derived epidermal stem cells and collagen sponge to restore the full thickness skin defects. [Methods] The dermal scaffolds were reconstructed by collagen sponge compounded with mice fetus skin fibroblasts. The scaffolds were transplanted into mice full thickness skin defects. ES cell-derived epidermal stem cells, labeled with Hoechst 33342, carried by a layer of biomembrane, were transplanted into the dermal scaffolds. The differentiation tissue of the tissue-engineered skin was sampled each week. The sections were observed with HE staining, immunohistochemical and di-labeled immunofluorescence methods to test β1 integrin, CK15, CK19, CK10, and CEA. [Results] The full thickness skin defects, covered with tissue-engineered skin, were healed within 3 weeks. The newborn skin was thicker than normal skin, The basal layer of newborn skin was increased. The labeled cells in the epidermis formed much bulky cellular poles towards dermis and expressed β1 integrin, CK15 and CK19. The labeled tubular or follicular structure expressed β1 integrin and CEA in dermis within 2 weeks. The basal cells of keratinized stratified squamous epithelium expressed CK19 and CK10. There were sebaceous glands-like and hair follicles-like structures in newborn skin after 4 weeks. [Conclusion] Reconstructed tissue-engineered skin with ES cell-derived epidermal stem cells and collagen sponge can restore the mice full thickness skin defect. There are epidermis, sweat glandslike, sebaceous glands-like and hair follicles-like structures in newborn skin.