INTERACTIVE EFFECTS OF 2,3,7,8-TETRACHLORODIBENZO-P-DIOXIN AND RETINOIDS ON PROLIFERATION AND DIFFERENTIATION IN CULTURED HUMAN KERATINOCYTES - QUANTIFICATION OF CROSS-LINKED ENVELOPE FORMATION

Dioxins are potent inducers of chloracne in humans. This skin aberrati on can be interpreted as an altered differentiation pattern of acinar sebaceous base cells and a change in the rate of terminal differentiat ion of the keratinocytes. We measured this rate induced by 2,3,7,8-tet rachlorodibenzo-p-dioxin (TCDD) in primary cultures of human keratinoc ytes. As parameters for differentiation, we quantified the S-35-methio nine incorporation into cross-linked envelopes (revealing the total CL E biomass), as well as the number of microscopically visible CLEs. It was shown that TCDD is a very potent inducer of both CLE biomass and n umber with a half-maximal effect concentration (EC(50)) Of 1.4 nM. CLE biomass was maximally increased 10-fold and the number of cells in cu lture producing a CLE was increased from 15% in control cultures to ma ximally 75% of the cells in TCDD-treated cultures. Both effects were C a2+-dependent and increased with elevated cell density, being optimal in post-confluent cultures. Retinoic acid dose-dependently decreased t he effect of 10(-8) M TCDD, 10(-6) M having a nearly complete antagoni stic action. This interaction of retinoic acid with TCDD-induced diffe rentiation was non-competitive. Retinol was equally potent as an antag onist of the TCDD-induced elevation of CLE formation as compared with retinoic acid. Retinyl palmitate and etretinate were not very effectiv e as TCDD antagonists. Supplementation of hydrocortisone suppressed th e TCDD-induced keratinocyte differentiation. It was concluded that CLE biomass quantification provides a reliable and sensitive parameter fo r keratinocyte differentiation. In this in vitro system it is shown th at TCDD strongly induces a switch from proliferation to terminal diffe rentiation and that this effect can be antagonized effectively by reti noic acid and retinol.