Serial Cultivation of Epidermal Keratinocytes from Psoriatic Plaques

Abstract

Using the combined techniques of Rheinwald and Green, and Liu and Karasek, large quantities of proliferative keratinocytes from involved (PP) or uninvolved (PN) skin of psoriatic patients and from normal skin of non-psoriatic donors (NN) can be obtained. Primary cultures, initiated from a 6-mm punch biopsy, are grown on a 3T3 feeder layer seeded on a collagen surface and fed with Dulbecco's Modified Eagle's Medium containing 20% fetal calf serum, hydrocortisone, epidermal growth factor, and cholera toxin. To test the hypothesis that the mechanism(s) responsible for the abnormal proliferation of psoriatic keratinocytes may be located within the cells themselves, primary cultures are passaged onto collagen surfaces without a feeder layer and maintained with medium plus serum, but no additional supplements, and the growth profiles of the 3 cell populations compared. No difference in morphology among these cells is observed in either primary or passaged cultures. In primary cultures, PP keratinocytes, especially those isolated from active lesions, seem to initiate growth at a slower rate than do PN and NN keratinocytes. The difference in the growth rate, as determined by cell number, DNA content, and mitotic activity, is insignificant among passaged PP, PN, and NN cells. Waves observed in the mitotic index and the metabolic activity of the passaged keratinocytes from all 3 sources suggest that the cells are synchronized during subculturing. The cells have high metabolic and mitotic activities during the first week after subculturing, indicative of an initial population of actively dividing cells. We have not found the characteristic feature of hyperproliferation seen in psoriatic keratinocytes in situ, in the cultured cells; however, it is too early to reach the definite conclusion that the mechanism(s) responsible for psoriasis does not exist in the keratinocyte itself. These 3 cell types may respond differently to agents that either enhance or inhibit cell growth and, by using the culture system outlined in this report, we may study these factors and their potential role in psoriasis.