Psoriasis is a common inflammatory skin disease with a multifactorial genetic basis. A dysregulated interplay between keratinocytes and infiltrating immune cells underlies the cutaneous inflammation in psoriasis. Keratinocytes are important producers of antimicrobial peptides such as hBD-2 and LL37 and cytokines such as TNF-alpha, which are essential elements in this process of cell-cell communication . Recently, miRNA-203 was identified as an important contributor to this dysfunctional cross talk . We have previously developed a new lipid-based nanosome (SECosome) that enables the effective delivery of siRNA into human skin . The aim of this project is to knockdown mRNA encoding hBD-2, LL37, TNF-alpha and miRNA-203 by tranfection of keratinocytes with SECosomes for the delivery of siRNAs and anti-miRNAs. Ultimately, we want to create a new therapy for psoriasis by intervening at genetic level by means of a topical therapy.
Materials and Methods
An optimized cytokine mix was used to induce a psoriatic phenotype starting from normal human keratinocytes. Complexes of siRNA or anti-miRNA and SECosomes were made and validated prior to transfection. 24h post-tranfection, qPCR analysis was performed to evaluate mRNA expression levels.
Transfection experiments with the complexes showed a stable knockdown efficiency of more than 80% of hBD-2, LL37, TNF-alpha and miR-203 mRNA.
In this in vitro work we prepared and characterized siRNA and anti-miRNA complexes with SECosomes against hBD-2, LL37, TNF-alpha and miR-203 respectively. These complexes efficiently knock-down the targeted genes with concomitant downregulation of the associated proteins. Hereafter we will test the therapeutic applicability of our complexes in xenografted psoriatic skin by topical application.
Geusens B, Van Gele M, Braat S, De Smedt SC, Stuart M, Prow T, Sanchez W, Roberts M, Sanders NN, J L: Flexible Nanosomes (SECosomes) Enable Efficient siRNA Delivery in Cultured Primary Skin Cells and in the Viable Epidermis of Ex Vivo Human Skin.