Comparison of therapeutic antibiotic treatments on tissue-engineered human skin substitutes.

For regenerative medicine to gain clinical acceptance, the effects of commonly used treatment regimens on bioengineered organs must be considered. The antibiotics mafenide acetate (mafenide) and neomycin plus polymyxin (neo/poly) are routinely used to irrigate postoperative skin grafts on contaminated wounds. The effects of these clinically used antibiotics were investigated using tissue-engineered human skin substitutes generated with primary human keratinocytes or the near-diploid human keratinocyte cell line, Near-diploid Immortal Keratinocytes. Following topical or dermal treatment, the skin substitutes were assayed for viability, tissue morphology, glycogen content, and the expression of active caspase 3. Mafenide, but not neo/poly, induced morphological and biochemical changes in tissue-engineered skin substitutes. Keratinocytes in all histological layers of mafenide-treated skin substitutes exhibited ballooning degeneration and glycogen depletion. Mafenide-treatment also triggered separation of basal keratinocytes from the underlying dermis. None of the antibiotic treatments induced apoptosis, as measured by active caspase 3 immunostaining. The results demonstrate that mafenide, but not neo/poly, is detrimental to the viability and structural integrity of tissue-engineered human skin substitutes. These findings highlight the need to identify treatment regimens that are compatible with and hence enable the therapeutic efficacy of first-generation bioengineered organs such as skin.