Fundación Inbiomed, San Sebastián, Spain.
J Tissue Eng Regen Med. 2013 Jun;7(6):479-90. doi: 10.1002/term.548. Epub 2012 Jan 31.
Full thickness wounds require a dermal component to achieve functional permanent skin restoration. Currently available tissue-engineered skin substitutes lack a subcutaneous fat layer that would functionally contribute some of the mechanical and thermoregulatory properties of normal skin. To generate a trilayer engineered skin equivalent, we included bone marrow mesenchymal (BM-MSC) or adipose tissue-derived (ASC) stromal cells in a human plasma hydrogel exposed to adipogenic clues for three weeks. Approximately half of the cells differentiated under these conditions into mature adipocytes that survived for two years in culture with minimal medium change. In vitro generation of bona fide fully differentiated adipocytes was assessed by leptin secretion and ultrastructurally demonstrated through semithin to ultrathin sectioning and lipid staining with osmium tetroxide. Furthermore, presence of BM-MSCs or ASCs within the subcutaneous layer contributed to the epidermal differentiation program, with more proliferating basal cells depositing basal membrane proteins and differentiating into mature keratinocytes that were able to generate a pluristratified epithelium. In conclusion, we engineered a fully differentiated human skin trilayer that could present multiple applications such as use for in vitro drug absorption tests and regenerative therapies.
全层创面需要真皮成分才能实现功能性永久性皮肤修复。目前可用的组织工程皮肤替代物缺乏皮下脂肪层，而该脂肪层将在功能上为正常皮肤的一些机械和温度调节特性做出贡献。为了生成三层工程皮肤等效物，我们将骨髓间充质（BM-MSC）或脂肪组织来源（ASC）基质细胞包含在人血浆水凝胶中，并暴露于成脂线索中 3 周。在此条件下，大约有一半的细胞分化为成熟脂肪细胞，在更换少量培养基的情况下在培养中存活了两年。通过瘦素分泌和半薄到超薄切片以及用锇四氧化物进行脂质染色在体外评估真正完全分化的脂肪细胞的生成。此外，BM-MSCs 或 ASC 存在于皮下层有助于表皮分化程序，更多增殖的基底细胞沉积基底膜蛋白并分化为成熟的角蛋白细胞，这些细胞能够生成多细胞层上皮。总之，我们构建了一种完全分化的人类皮肤三层结构，可用于多种应用，例如用于体外药物吸收测试和再生疗法。
