Segal Nadav, Andriani Frank, Pfeiffer Lawrence, Kamath Padmaja, Lin Ning, Satyamurthy Kapettu, Egles Christophe, Garlick Jonathan A
Department of Oral Biology and Pathology, School of Dental Medicine, SUNY at Stony Brook, Stony Brook, New York, United States.
Matrix Biol. 2008 Apr;27(3):163-70. doi: 10.1016/j.matbio.2007.09.002. Epub 2007 Sep 22.
Epithelial-mesenchymal interactions promote the morphogenesis and homeostasis of human skin. However, the role of the basement membrane (BM) during this process is not well-understood. To directly study how BM proteins influence epidermal differentiation, survival and growth, we developed novel 3D human skin equivalents (HSEs). These tissues were generated by growing keratinocytes at an air-liquid interface on polycarbonate membranes coated with individual matrix proteins (Type I Collagen, Type IV Collagen or fibronectin) that were placed on contracted Type I Collagen gels populated with dermal fibroblasts. We found that only keratinocytes grown on membranes coated with the BM protein Type IV Collagen showed optimal tissue architecture that was similar to control tissues grown on de-epidermalized dermis (AlloDerm) that contained intact BM. In contrast, tissues grown on proteins not found in BM, such as fibronectin and Type I Collagen, demonstrated aberrant tissue architecture that was linked to a significant elevation in apoptosis and lower levels of proliferation of basal keratinocytes. While all tissues demonstrated a normalized, linear pattern of deposition of laminin 5, tissues grown on Type IV Collagen showed elevated expression of alpha6 integrin, Type IV Collagen and Type VII Collagen, suggesting induction of BM organization. Keratinocyte differentiation (Keratin 1 and filaggrin) was not dependent on the presence of BM proteins. Thus, Type IV Collagen acts as a critical microenvironmental factor in the BM that is needed to sustain keratinocyte growth and survival and to optimize epithelial architecture.
上皮-间充质相互作用促进人类皮肤的形态发生和内稳态。然而，在此过程中基底膜（BM）的作用尚未得到充分理解。为了直接研究BM蛋白如何影响表皮分化、存活和生长，我们开发了新型的三维人类皮肤等效物（HSE）。这些组织是通过将角质形成细胞在气液界面上培养于涂有单个基质蛋白（I型胶原蛋白、IV型胶原蛋白或纤连蛋白）的聚碳酸酯膜上而生成的，这些膜放置在充满真皮成纤维细胞的收缩I型胶原凝胶上。我们发现，只有在涂有BM蛋白IV型胶原蛋白的膜上生长的角质形成细胞显示出最佳的组织结构，类似于在含有完整BM的去表皮真皮（AlloDerm）上生长的对照组织。相比之下，在BM中未发现的蛋白质（如纤连蛋白和I型胶原蛋白）上生长的组织表现出异常的组织结构，这与基底角质形成细胞凋亡显著增加和增殖水平降低有关。虽然所有组织都表现出层粘连蛋白5的正常化、线性沉积模式，但在IV型胶原蛋白上生长的组织显示出α6整合素、IV型胶原蛋白和VII型胶原蛋白的表达升高，提示BM组织的诱导。角质形成细胞分化（角蛋白1和丝聚蛋白）不依赖于BM蛋白的存在。因此，IV型胶原蛋白是BM中的关键微环境因子，是维持角质形成细胞生长和存活以及优化上皮结构所必需的。
