A new Fluorescence Lifetime Imaging Microscopy technique reveals skin penetration and differential cellular uptake of tecto-dendrimer nanoparticles in human reconstructed skin models

C. ZOSCHKE; A. BOREHAM; P. SCHILRREFF; R. BRODWOLF; E.L. ROMERO; M. SCHÄFER-KORTING; U. ALEXIEV

Conferencia; Gordon Research Conference on "Barrier Function of Mammalian Skin"; 2015

Dendritic nanoparticles possess many characteristics,such as high loading capacity for guest molecules and tunable end groups facilitating surface functionalizing, that have brought them into the focus as a new treatment option for therapy-resistant diseases.Nanoparticles can enhance poor skin penetration of topically applied agents [1], and tecto-dendrimer nanoparticles even induced selective toxicity to a melanoma cell line [2]. However the skin penetration and toxicity of the dendritic nanoparticle itself remains unknown. To further deepen our insight into skin penetration as obtained from dendritic core-multishell (CMS) nanocarriers [3] we investigated the skin penetration of fluorescently labelled (FITC) tecto-dendrimer (G5.G2.5) nanoparticles [2] into reconstructed human skin and an organotypic skin disease model (cutaneous squamous cell carcinoma (SCC) model).For visualization of the local distribution of tecto-dendrimers in the skin a new Fluorescence Lifetime Imaging Microscopy (FLIM) technique [4,5] was applied. This technique combines information from excited-state fluorescence decay curves of the fluorescing molecules with a sophisticated analysis method that uses multivariate analysis algorithm identifying unique fluorescence lifetime species.Analysis of sections from reconstructed human skin showed no penetration of tecto-dendrimer nanoparticles into the skin. In sections from the diseased reconstructed skin, however, a penetration of the tecto-dendrimer nanoparticles into the viable epidermis of the skin was observed. Thus, skin diseases specifically influence the penetration of the tecto-dendrimer nanoparticles in agreement with a less functional barrier. Moreover, regions of concentrated nanoparticle uptake were observed that appeared to correlate with the occurrence of SCC cells within the skin disease model. Therefore,monolayer cell experiments were performed to confirm the selective uptake of tecto-dendrimer nanoparticles into SCC cells.While the tecto-dendrimer nanoparticles remain in the membrane of normal keratinocytes, SCC cells take up the particles into the cytoplasm via an active process.Such differential uptake behaviour in the skin provides new avenues in targeted drug delivery by topical application.