Cyclosporine A is an immunosuppressive drug used not only to prevent organ or tissue rejection, but also in curing dermal diseases such as atopic dermatitis, pyoderma gangrenosum or pemphigus. Cyclosporine is metabolized by the cytochrome P450 isoform CYP3A4 mainly, but also works as an inhibitor of CYP3A4, CYP2C19 and CYP2D6. At the current state of our knowledge, we do not exactly know how cyclosporine A influences healthy dermal cells.

The aim of this study was to estimate how cyclosporine A acts on gene expression of cytochrome P450 isoforms in normal human dermal fibroblasts (NHDF) using microarray analysis.

Normal human fibroblasts cells (CC-2511, Lonza) were cultured in the FGM medium (Fibroblast Growth Medium; Lonza, Basel, Switzerland) on 25 cm² plates with Nunclon surface, equipped with bacteriological filters (Nunc, Wiesbaden, Germany). After the confluence state, there was added cyclosporine A (sigma Aldrich) at a concentration of 100 ng mL^-1. Next with the use of TRIZOL® the full cell RNA had been extracted accordingly to the producer protocol (Invitrogen Life Technologies, USA). Extracted RNA was purified and used to cDNA and cRNA synthesis and next was hybridized with the HG-U133A 2.0 microarrays accordingly to the protocol (Affymetrix Inc. California, USA). The array plate was scanned by the GeneArray Scanner 3000 7G (Affymetrix Inc.) obtaining fluorescence signal of 22277 mRNA. Statistical analysis was performed using t-test. The significance level was set at  p ≤ 0.05.

After comparing fluorescence signals for 91 mRNA isoforms of cytochrome P450 in NHDF culture with and without cyclosporine A were stated that 31 mRNA of CYP’s over-expressed in the presence of cyclosporine and 15 down-regulated. Between all of them the eight mRNA had statistically significant changes, but only CYP1B1 and CYP19A1 characterized highest expression.

We observed changes in the gene expression of CYP’s connected with steroids metabolism after an eight-hour cyclosporine A exposure in human dermal fibroblasts.

This work was supported by Medical University of Silesia grants KNW-1-013/N/3/0 and KNW-1-060/K/3/0.

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