LIPID SUPPLEMENTED MEDIUM INDUCES LAMELLAR BODIES AND PRECURSORS OF BARRIER LIPIDS IN CULTURED ANALOGS OF HUMAN SKIN

Barrier function of cultured skin substitutes (CSS) is required for th eir effective use in clinical treatment of skin wounds, and for percut aneous absorption in vitro. Arachidonic, palmitic, oleic, and linoleic free fatty acids, in conjunction with the antioxidant alpha-tocophero l acetate . (lipid supplements, ''LS''), were added to nutrient media of CSS to provide precursors of epidermal barrier lipids. CSS were com posed of human keratinocytes (HK), fibroblasts (HF), and collagen-glyc osaminoglycan substrates, and were incubated for 14 d submerged or lif ted to the air-liquid interface in media based on MCDB 153 +/- LS. Dup licate samples (30 cm2) were harvested and the epidermal analogue was analyzed for total protein, total DNA, total lipid, lipid fractions in cluding acylglucosylceramide (AGC), and presence of lamellar bodies. S ignificant increases (p < 0.05) were detected between CSS incubated in + LS medium for total lipid, total DNA, ceramide, glucosylceramide, t riglycerides, and diglycerides. AGC and lamellar bodies were detected only in epithelia of CSS incubated in + LS medium. These data show tha t free fatty acids, vitamin E, and lifting of CSS promote increased ep ithelial morphogenesis compared to CSS cultured submerged without lipi d supplements. Presence of lamellar bodies and AGC suggests enhanced p roduction in vitro of barrier-associated epidermal lipids.