Quantitative analysis of the proliferation of epidermal cells using a human skin organ culture system and the effect of DbcAMP using markers of proliferation (BrdU, Ki-67, PCNA)

Abstract The process of re-epithelialization of a wound in the epidermis comprises the following steps: proliferation of epidermal basal cells, migration of epidermal cells to the wound surface, and cell differentiation. In the present study, we evaluated the proliferation of epidermal basal cells, an important process in wound healing, in the wound margin using a human skin organ culture system and immunohistochemical labeling with bromodeoxyuridine (BrdU), Ki-67, and proliferating cell nuclear antigen (PCNA), as markers of cell proliferation. Dibutyryl cyclic adenosine monophosphate (DbcAMP) is a derivative of cAMP and has been shown to modulate human keratinocyte proliferation. The proliferation of keratinocytes was promoted by DbcAMP and particularly strong effects in terms of BrdU labeling index, Ki-67-positive ratio, and PCNA-positive ratio, were seen at 10^–5 M. The skin organ culture system presented here uses adult preputial skin and is a simple technique that uses easily available materials. In addition to identifying S phase cells using BrdU as an index of cell proliferation, the immunohistochemical method for evaluating the expression of Ki-67 and PCNA is very simple. Accordingly, the method described here seems to be useful for evaluating cell dynamics in wound healing.