In view of increasing numbers of dermatological disorders, transdermal drug delivery along with
in vitro
research is becoming increasingly popular. Herefore, qualified
in vitro
skin models are required. The objective of this study was the optimization and validation of a modified lactate dehydrogenase (LDH) release assay during the establishment of an
in vitro
viable human skin model, employable for a variety of skin associated disorders. Firstly, the most suitable LDH isoform for the study was determined. Subsequently, a stability study was conducted to investigate the best storage conditions of the LDH enzyme. Finally, the test system was validated in terms of linear range, range limits and system suitability. The results indicate LDH-5 as most suitable isoform due to its predominance in skin. The stability samples stored at −20 °C in the presence of polyethylene glycol (PEG) as cryoprotector displayed the targeted recovery of 100% ± 15 % until the end of the four-week study in contrast to other investigated conditions. A six-point calibration without PEG and a seven-point calibration with PEG including evaluation of system suitability and quantification limits were established with both correlation coefficients r
2
above 0.99 and all deviations below 15%. Concluding from those results, this method can be considered valid and useful for its employment in viability determination of viable
in vitro
skin models.
Transdermal drug delivery provides several advantages over conventional drug administration, such as the avoidance of first-pass metabolism and better patient compliance. In vitro research can abbreviate and facilitate the pharmaceutical development considerably compared to in vivo research as drug screening and clinical studies can be reduced. These advantages led to the development of corresponding skin models. Viable skin models are more useful than non-viable ones, due to the influence of skin metabolism on the results. While most in vitro studies concentrate on evaluating human-based models, the current study is designed for the investigation of both human and animal diseases. So far, there is little information available in the literature about viable animal skin cultures which are in fact intended for application in the veterinary and not the human field. Hence, the current study aims to fill the gap. For the in vitro viable skin model, specimens of human, porcine and canine skin were cultured over two weeks under serum-free conditions. To evaluate the influence of medium supplementation on skin viability, two different supplement mixtures were compared with basic medium. The skin specimens were maintained at a viability-level >50% until the end of the study. From the tested supplements, the addition of bovine pituitary extract and epidermal growth factor increased skin viability whereas hydrocortisone and insulin induced a decrease. This in vitro viable skin model may be a useful tool for the investigation of skin diseases, especially for the veterinary field.
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