Cellular metabolism and pore lifetime of human skin following microprojection array mediation.

Skin-targeting microscale medical devices are becoming popular for therapeutic delivery and diagnosis. We used cryo-SEM, fluorescence lifetime imaging microscopy (FLIM), autofluorescence imaging microscopy and inflammatory response to study the puncturing and recovery of human skin ex vivo and in vivo after discretised puncturing by a microneedle array (Nanopatch®). Pores induced by the microprojections were found to close by ~25% in diameter within the first 30 min, and almost completely close by ~6 h. FLIM images of ex vivo viable epidermis showed a stable fluorescence lifetime for unpatched areas of ~1000 ps up to 24 h. Only the cells in the immediate puncture zones (in direct contact with projections) showed a reduction in the observed fluorescence lifetimes to between ~518-583 ps. The ratio of free-bound NAD(P)H (α1/α2) in unaffected areas of the viable epidermis was ~2.5-3.0, whereas the ratio at puncture holes was almost double at ~4.2-4.6. An exploratory pilot in vivo study also suggested similar closure rate with histamine administration to the forearms of human volunteers after Nanopatch® treatment, although a prolonged inflammation was observed with Tissue Viability Imaging. Overall, this work shows that the pores created by the microneedle-type medical device, Nanopatch®, are transient, with the skin recovering rapidly within 1-2 days in the epidermis after application.

ZnO:SBA-15 Nanocomposites for Potential Use in Sunscreen: Preparation, Properties, Human Skin Penetration and Toxicity.

AIM: We evaluated the effects of the incorporation of zinc oxide (ZnO) nanoparticles in a mesoporous matrix, aiming to improve the textural, structural and morphological properties and verify their safety so that they can be applied in sunscreen cosmetics. MATERIALS AND METHODS: ZnO nano-particles were incorporated into an ordered mesoporous silica matrix known as Santa Barbara Amorphous-15 (SBA-15), using post-synthesis methodology. The resulting nanocomposites were characterized using X-ray diffraction, small angle X-ray scattering, N2 adsorption-desorption isotherms, Fourier transform infrared spectroscopy, scanning electron microscopy and predicted in vitro sun protector factor (SPF) estimation. Effectiveness and safety were evaluated by antimicrobial activity, in vitro cell toxicity and non-invasive multi-photon tomography with fluorescence lifetime imaging. RESULTS: The structure of the nanocomposites was similar to that of SBA-15, with little perturbation caused by ZnO incorporation. Nanocomposites had an increased in vitro SPF, reduced cytotoxic activity and favourable antimicrobial properties compared to ZnO. ZnO:SBA-15 nanocomposites exhibited no measurable toxicity when applied to human skin in vivo. CONCLUSION: Due to their suitable physicochemical properties and improved safety compared to bare ZnO nanoparticles, the ZnO:SBA-15 nanocomposites show promise for use in cosmetic applications.

Support for the Safe Use of Zinc Oxide Nanoparticle Sunscreens: Lack of Skin Penetration or Cellular Toxicity after Repeated Application in Volunteers.

Zinc oxide is a widely used broad-spectrum sunscreen, but concerns have been raised about the safety of its nanoparticle (NP) form. We studied the safety of repeated application of agglomerated zinc oxide (ZnO) NPs applied to human volunteers over 5 days by assessing the skin penetration of intact ZnO-NPs and zinc ions and measuring local skin toxicity. Multiphoton tomography with fluorescence lifetime imaging microscopy was used to directly visualize ZnO-NP skin penetration and viable epidermal metabolic changes in human volunteers. The fate of ZnO-NPs was also characterized in excised human skin in vitro. ZnO-NPs accumulated on the skin surface and within the skin furrows but did not enter or cause cellular toxicity in the viable epidermis. Zinc ion concentrations in the viable epidermis of excised human skin were slightly elevated. In conclusion, repeated application of ZnO-NPs to the skin, as used in global sunscreen products, appears to be safe, with no evidence of ZnO-NP penetration into the viable epidermis nor toxicity in the underlying viable epidermis. It was associated with the release and penetration of zinc ions into the skin, but this did not appear to cause local toxicity.

Space- and time-resolved investigation on diffusion kinetics of human skin following macromolecule delivery by microneedle arrays.

Microscale medical devices are being developed for targeted skin delivery of vaccines and the extraction of biomarkers, with the potential to revolutionise healthcare in both developing and developed countries. The effective clinical development of these devices is dependent on understanding the macro-molecular diffusion properties of skin. We hypothesised that diffusion varied according to specific skin layers. Using three different molecular weights of rhodamine dextran (RD) (MW of 70, 500 and 2000 kDa) relevant to the vaccine and therapeutic scales, we deposited molecules to a range of depths (0-300 µm) in ex vivo human skin using the Nanopatch device. We observed significant dissipation of RD as diffusion with 70 and 500 kDa within the 30 min timeframe, which varied with MW and skin layer. Using multiphoton microscopy, image analysis and a Fick's law analysis with 2D cartesian and axisymmetric cylindrical coordinates, we reported experimental trends of epidermal and dermal diffusivity values ranging from 1-8 µm2 s-1 to 1-20 µm2 s-1 respectively, with a significant decrease in the dermal-epidermal junction of 0.7-3 µm2 s-1. In breaching the stratum corneum (SC) and dermal-epidermal junction barriers, we have demonstrated practical application, delivery and targeting of macromolecules to both epidermal and dermal antigen presenting cells, providing a sound knowledge base for future development of skin-targeting clinical technologies in humans.

Development and Evaluation of Lipid Nanoparticles Containing Natural Botanical Oil for Sun Protection: Characterization and in vitro and in vivo Human Skin Permeation and Toxicity.

The use of sunscreen products is widely promoted by schools, government agencies, and health-related organizations to minimize sunburn and skin damage. In this study, we developed stable solid lipid nanoparticles (SLNs) containing the chemical UV filter octyl methoxycinnamate (OMC). In parallel, we produced similar stable SLNs in which 20% of the OMC content was replaced by the botanical urucum oil. When these SLNs were applied to the skin of human volunteers, no changes in fluorescence lifetimes or redox ratios of the endogenous skin fluorophores were seen, suggesting that the formulations did not induce toxic responses in the skin. Ex vivo (skin diffusion) tests showed no significant penetration. In vitro studies showed that when 20% of the OMC was replaced by urucum oil, there was no reduction in skin protection factor (SPF), suggesting that a decrease in the amount of chemical filter may be a viable alternative for an effective sunscreen, in combination with an antioxidant-rich vegetable oil, such as urucum. There is a strong trend towards increasing safety of sun protection products through reduction in the use of chemical UV filters. This work supports this approach by producing formulations with lower concentrations of OMC, while maintaining the SPF. Further investigations of SPF in vivo are needed to assess the suitability of these formulations for human use.