Dawson R A, Goberdhan N J, Freedlander E, MacNeil S
Department of Medicine, Clinical Sciences Centre University of Sheffield, UK.
Burns. 1996 Mar;22(2):93-100. doi: 10.1016/0305-4179(95)00092-5.
The aim of this study was to investigate whether prior culture of cells on ECM proteins might positively influence the performance of keratinocytes when cells are transferred to a dermal in vitro wound bed model. Keratinocytes were cultured using a method for producing cultured epithelial autografts for severely burned patients (essentially using Green's medium, a mitogen-rich medium containing fetal calf serum, cholera toxin, EGF, insulin, transferrin and triiodothyronine). Cells were cultured either on irradiated 3T3 fibroblasts (as in the standard Rheinwald and Green technique) or, alternatively, on collagen I, collagen IV, matrigel, RGD, vitronectin or fibronectin. Under these conditions matrigel, collagen I and IV enhanced initial attachment, RGD, vitronectin, fibronectin and irradiated 3T3 fibroblasts did not. Proliferation of cells was positively influenced by matrigel, collagen I and IV and irradiated 3T3 fibroblasts; of these, however, only matrigel and 3T3 fibroblasts had sustained significant effects on keratinocyte proliferation over 4 days. Cells on fibronectin showed significantly reduced proliferation. An acellular non-viable dermis was then used to mimic the homograft allodermis onto which cultured epithelial autograft sheets are grafted clinically and cells cultured on the various ECM proteins for 96 h were transferred to this in vitro wound model. None of the substrates enhanced keratinocyte performance on this model. It was concluded that under these conditions some ECM proteins can significantly affect keratinocyte attachment and, to a lesser extent, proliferation but that the culture of keratinocytes on these ECM proteins does not appear to confer any lasting benefit to the attachment of these keratinocytes to an in vitro wound-bed model.
本研究的目的是调查当细胞被转移到真皮体外伤口床模型时，预先在细胞外基质（ECM）蛋白上培养细胞是否会对角质形成细胞的性能产生积极影响。角质形成细胞采用一种为严重烧伤患者生产培养上皮自体移植片的方法进行培养（基本上使用格林氏培养基，一种富含促有丝分裂原的培养基，含有胎牛血清、霍乱毒素、表皮生长因子、胰岛素、转铁蛋白和三碘甲状腺原氨酸）。细胞要么在经辐照的3T3成纤维细胞上培养（如标准的莱茵瓦尔德和格林技术），要么在I型胶原、IV型胶原、基质胶、RGD、玻连蛋白或纤连蛋白上培养。在这些条件下，基质胶、I型胶原和IV型胶原增强了初始附着，而RGD、玻连蛋白、纤连蛋白和经辐照的3T3成纤维细胞则没有。基质胶、I型胶原和IV型胶原以及经辐照的3T3成纤维细胞对细胞增殖有积极影响；然而，在这几种物质中，只有基质胶和3T3成纤维细胞在4天内对角质形成细胞增殖有持续的显著影响。在纤连蛋白上的细胞增殖明显减少。然后使用无细胞的无活力真皮来模拟临床移植培养上皮自体移植片的同种异体真皮，并将在各种ECM蛋白上培养96小时的细胞转移到这个体外伤口模型中。在这个模型中，没有一种底物能增强角质形成细胞的性能。得出的结论是，在这些条件下，一些ECM蛋白可以显著影响角质形成细胞的附着，并在较小程度上影响增殖，但在这些ECM蛋白上培养角质形成细胞似乎并不会给这些角质形成细胞附着到体外伤口床模型带来任何持久的益处。
