Epidermal Cell Confluence and Implications for a Two-Step Mechanism of Wound Closure

In contrast to freshly isolated cells, some cultured keratinocytes have the ability to adhere and spread in protein-free media. Reported here are experiments testing the hypothesis that the social history of keratinocytes influences their ability to spread in defined media. The experiments indicate that confluent cells lack the ability to spread in defined media while subconfluent cells have this property. The inability of dissociated confluent cells to spread in protein-free media is referred to phenomenologically as a “confluent block.” The confluent block is acquired rapidly (1–3 days) and lost slowly (5–7 days). The ability of subconfluent cells to spread in the absence of media protein is sensitive to cycloheximide. Aortic endothelial cells and dermal fibroblasts do not demonstrate a confluent block. These observations are consonant with a two-step mechanism of epidermal wound repair: the first occurs immediately after wounding during which the cells require substratum-active proteins, and the second occurs 5–7 days later when the cells are able to synthesize their own substratum