Characterization of a Three-Dimensional (3-D) Organotypic Skin Model using Keratinocytes and Mesenchymal Stem Cells Immortalized by hTERT

In this study, we compared primary keratinocytes (ATCC PCS-200-010) to hTERT immortalized keratinocytes (Ker-CT; ATCC CRL-4048), co-cultured with either primary fibroblasts (ATCC PCS-201-010), primary adipose-derived mesenchymal stem cells (MSCs; ATCC PCS-500-011), hTERT-immortalized fibroblasts (BJ5ta; ATCC CRL-4001), or hTERT-immortalized MSCs (hTERT-MSCs; ATCC SCRC-4000). We confirmed that both primary keratinocytes and Ker-CT are able to fully differentiate into skin equivalents in a 3-D culture model when co-cultured with primary fibroblasts, primary MSCs, BJ-5ta, or hTERT-MSCs. To confirm the functionality of the co-culture models, both the primary keratinocytes and the Ker-CT air-liquid interface (ALI) cocultures were subjected to a scratch assay. Re-epithelialization occurred in both cell lines, and interleukin 8 (IL-8) showed an increase in expression from day 0 to day 1 and 3, corresponding to migration of cells into the wound. The continuous nature of the Ker-CT cell line makes it an invaluable model for the research of keratinocyte biology, as it eliminates the issue of short life span and donor variation seen with primary cells.