Dual Role of Stratum Corneum Carotenes/Lycopene Against the Development of Chemotherapy-induced PPE in Patients With Cancer.

Palmar-plantar erythrodysaesthesia (PPE) is a common side effect of chemotherapy treatment in patients with cancer. The exact pathophysiologic mechanisms of the development of PPE remain unclear. Here, we report two important physiological functions of carotenoids without hydroxyl groups (α-carotene, ß-carotene, γ-carotene, ξ-carotene, lycopene, phytoene, phytofluene and their isomers) in the stratum corneum (SC) of glabrous skin: The powerful antioxidant protection of the integrity of the SC components against the destructive action of free radicals and maintaining the skin barrier function by the creation of an orthorhombic organization of intercellular lipids within lamellae using carotenoids as a skeleton. The dual protective role of carotenoids without hydroxyl groups is important for both healthy skin and, in the authors' opinion, for the skin of chemotherapy-treated patients against the development of PPE, as the chemotherapy-induced reduction of the carotenoid concentration in the stratum corneum considerably weakens the skin resistance to cytotoxic and other adverse reactions.

Atopic Dermatitis: Molecular Alterations between Lesional and Non-Lesional Skin Determined Noninvasively by In Vivo Confocal Raman Microspectroscopy.

Atopic dermatitis (AD)/atopic eczema is a chronic relapsing inflammatory skin disease affecting nearly 14% of the adult population. An important pathogenetic pillar in AD is the disrupted skin barrier function (SBF). The atopic stratum corneum (SC) has been examined using several methods, including Raman microspectroscopy, yet so far, there is no depth-dependent analysis over the entire SC thickness. Therefore, we recruited 21 AD patients (9 female, 12 male) and compared the lesional (LAS) with non-lesional atopic skin (nLAS) in vivo with confocal Raman microspectroscopy. Our results demonstrated decreased total intercellular lipid and carotenoid concentrations, as well as a shift towards decreased orthorhombic lateral lipid organisation in LAS. Further, we observed a lower concentration of natural moisturising factor (NMF) and a trend towards increased strongly bound and decreased weakly bound water in LAS. Finally, LAS showed an altered secondary and tertiary keratin structure, demonstrating a more folded keratin state than nLAS. The obtained results are discussed in comparison with healthy skin and yield detailed insights into the atopic SC structure. LAS clearly shows molecular alterations at certain SC depths compared with nLAS which imply a reduced SBF. A thorough understanding of these alterations provides useful information on the aetiology of AD and for the development/control of targeted topical therapies.

Altered structure indicating reduced barrier function of lesional compared to non-lesional psoriatic skin-A non-invasive in vivo study of the human stratum corneum with confocal Raman micro-spectroscopy.

Psoriasis, one of the most common skin diseases affecting roughly 2%-3% of the world population, is associated with a reduced skin barrier function (SBF) that might play an important role in its pathophysiology. The SBF is provided primarily by the stratum corneum (SC) of the skin. Previous studies have revealed a higher trans-epidermal water loss, lower hydration, abnormal concentration and composition of intercellular lipids, as well as alterations in secondary keratin structure in the psoriatic SC. We compared on molecular level lesional psoriatic skin (LPS) with non-lesional psoriatic skin (nLPS) from 19 patients non-invasively in vivo, using confocal Raman micro-spectroscopy. By analysing the corresponding Raman spectra, we determined SBF-defining parameters of the SC depth-dependently. Our results revealed a lower total lipid concentration, a shift of lamellar lipid organisation towards more gauche-conformers and an increase of the less dense hexagonal lateral packing of the intercellular lipids in LPS. Furthermore, we observed lower natural moisturising factor concentration, lower total water as well as a strong tendency towards less strongly bound and more weakly bound water molecules in LPS. Finally, we detected a less stable secondary keratin structure with increased ß-sheets, in contrast to the tertiary structure, showing a higher degree of folded keratin in LPS. These findings clearly suggest structural differences indicating a reduced SBF in LPS, and are discussed in juxtaposition to preceding outcomes for psoriatic and healthy skin. Understanding the alterations of the psoriatic SC provides insights into the exact pathophysiology of psoriasis and paves the way for optimal future treatments.

Spectroscopic biofeedback on cutaneous carotenoids: A powerful tool for primary prevention in advanced age.

Antioxidants exhibit a powerful defense mechanism against aging and chronic disease. The human skin reflects the overall antioxidant status of the body. The cutaneous carotenoid concentration is a biomarker for individual nutritional intake of antioxidants, as it correlates with the overall antioxidant status. The cutaneous carotenoid concentrations of 44 adults were measured using a multiple spatially resolved reflection spectroscopy. During the first phase of the study, measurements of carotenoid concentrations were performed without revealing the antioxidant status, followed by an intervention phase during which the volunteers were informed about their individual values by biofeedback. During the third phase, biofeedback was combined with an additional intake of fruit juices. Across time points, participants showed increasing levels of carotenoid status. Thus, biofeedback leads to an improvement of the carotenoid value of the skin. Providing a biofeedback measurement to monitor the individual antioxidative status may be an easy and cost-effective way of primary prevention.

Tattoo Pigments Are Localized Intracellularly in the Epidermis and Dermis of Fresh and Old Tattoos: In vivo Study Using Two-Photon Excited Fluorescence Lifetime Imaging.

BACKGROUND: The knowledge about the location and kinetics of tattoo pigments in human skin after application and during the recovery is restricted due to the limitation of in vivo methods for visualizing pigments. Here, the localization and distribution of tattoo ink pigments in freshly and old tattooed human skin during the regeneration of the epidermis and dermis were investigated in vivo. METHODS: Two-photon excited fluorescence lifetime imaging (TPE-FLIM) was used to identify tattoo ink pigments in human skin in vivo down to the reticular dermis. One subject with a freshly applied tattoo and 10 subjects with tattoos applied over 3 years ago were investigated in the epidermal and dermal layers in vivo. One histological slide of tattooed skin was used to localize skin-resident tattoo pigment using light microscopy. RESULTS: The carbon black particles deposited around the incision have still been visible 84 days after tattoo application, showing delayed recovery of the epidermis. The TPE-FLIM parameters of carbon black tattoo ink pigments were found to be different to all skin components except for melanin. Distinction from melanin in the skin was based on higher fluorescence intensity and agglomerate size. Using TPE-FLIM in vivo tattoo pigment was found in 75% of tattoos applied up to 9 years ago in the epidermis within keratinocytes, dendritic cells, and basal cells and in the dermis within the macrophages, mast cells, and fibroblasts. Loading of highly fluorescent carbon black particles enables in vivo imaging of dendritic cells in the epidermis and fibroblasts in the dermis, which cannot be visualized in native conditions. The collagen I structures showed a higher directionality similar to scar tissue resulting in a greater firmness and decreased elasticity of the tattooed skin. CONCLUSIONS: Here, we show the kinetics and location of carbon black tattoo ink pigment immediately after application for the first time in vivo in human skin. Carbon black particles are located exclusively intracellularly in the skin of fresh and old tattoos. They are found within macrophages, mast cells, and fibroblasts in the dermis and within keratinocytes, dendritic cells, and basal cells in the continuously renewed epidermis even in 9-year-old tattoos in skin showing no inflammation.

Quantitative determination of concentration profiles of skin components and topically applied oils by tailored multivariate curve resolution-alternating least squares using in vivo confocal Raman micro-spectroscopy.

The main components of the stratum corneum (SC), water, lipids, and proteins, are non-homogeneously distributed throughout the depth. The quantitative determination of their concentration profiles and penetration depth of topically applied substances are urgent topics of dermatological and cosmetic research. Confocal Raman micro-spectroscopy has distinct advantages when determining semi-quantitative concentrations of SC components and topically applied substances non-invasively and in vivo. In this work, we applied a tailored multivariate curve resolution-alternating least squares (tMCR-ALS) method to analyze Raman spectra of the SC in the 2000-4000 cm-1 region for quantitatively determining the concentrations of water, lipids, proteins, and topically applied oils using substance-related spectral loadings which were allowed to change depth-dependently from the SC's surface toward its bottom. tMCR-ALS makes matching of depth-dependent signal attenuation, that is, the normalization on keratin, unnecessary and requires only a few additional experiments for calibration - Raman spectra of the pure materials and their densities.

Label-free imaging of M1 and M2 macrophage phenotypes in the human dermis in vivo using two-photon excited FLIM.

Macrophages (ΜΦs) are important immune effector cells that promote (M1 ΜΦs) or inhibit (M2 ΜΦs) inflammation and are involved in numerous physiological and pathogenic immune responses. Their precise role and relevance, however, are not fully understood for lack of noninvasive quantification methods. Here, we show that two-photon excited fluorescence lifetime imaging (TPE-FLIM), a label-free noninvasive method, can visualize ΜΦs in the human dermis in vivo. We demonstrate in vitro that human dermal ΜΦs exhibit specific TPE-FLIM properties that distinguish them from the main components of the extracellular matrix and other dermal cells. We visualized ΜΦs, their phenotypes and phagocytosis in the skin of healthy individuals in vivo using TPE-FLIM. Additionally, machine learning identified M1 and M2 MФs with a sensitivity of 0.88±0.04 and 0.82±0.03 and a specificity of 0.89±0.03 and 0.90±0.03, respectively. In clinical research, TPE-FLIM can advance the understanding of the role of MФs in health and disease.

Penetration Depth of Propylene Glycol, Sodium Fluorescein and Nile Red into the Skin Using Non-Invasive Two-Photon Excited FLIM.

The stratum corneum (SC) forms a strong barrier against topical drug delivery. Therefore, understanding the penetration depth and pathways into the SC is important for the efficiency of drug delivery and cosmetic safety. In this study, TPT-FLIM (two-photon tomography combined with fluorescence lifetime imaging) was applied as a non-invasive optical method for the visualization of skin structure and components to study penetration depths of exemplary substances, like hydrophilic propylene glycol (PG), sodium fluorescein (NaFl) and lipophilic Nile red (NR) into porcine ear skin ex vivo. Non-fluorescent PG was detected indirectly based on the pH-dependent increase in the fluorescence lifetime of SC components. The pH similarity between PG and viable epidermis limited the detection of PG. NaFl reached the viable epidermis, which was also proved by laser scanning microscopy. Tape stripping and confocal Raman micro-spectroscopy were performed additionally to study NaFl, which revealed penetration depths of ≈5 and ≈8 µm, respectively. Lastly, NR did not permeate the SC. We concluded that the amplitude-weighted mean fluorescence lifetime is the most appropriate FLIM parameter to build up penetration profiles. This work is anticipated to provide a non-invasive TPT-FLIM method for studying the penetration of topically applied drugs and cosmetics into the skin.

Carotenoids in Human SkinIn Vivo: Antioxidant and Photo-Protectant Role against External and Internal Stressors.

The antioxidant system of the human body plays a crucial role in maintaining redox homeostasis and has an important protective function. Carotenoids have pronounced antioxidant properties in the neutralization of free radicals. In human skin, carotenoids have a high concentration in the stratum corneum (SC)-the horny outermost layer of the epidermis, where they accumulate within lipid lamellae. Resonance Raman spectroscopy and diffuse reflectance spectroscopy are optical methods that are used to non-invasively determine the carotenoid concentration in the human SC in vivo. It was shown by electron paramagnetic resonance spectroscopy that carotenoids support the entire antioxidant status of the human SC in vivo by neutralizing free radicals and thus, counteracting the development of oxidative stress. This review is devoted to assembling the kinetics of the carotenoids in the human SC in vivo using non-invasive optical and spectroscopic methods. Factors contributing to the changes of the carotenoid concentration in the human SC and their influence on the antioxidant status of the SC in vivo are summarized. The effect of chemotherapy on the carotenoid concentration of the SC in cancer patients is presented. A potential antioxidant-based pathomechanism of chemotherapy-induced hand-foot syndrome and a method to reduce its frequency and severity are discussed.

Current Views on Noninvasive in vivo Determination of Physiological Parameters of the Stratum Corneum Using Confocal Raman Microspectroscopy.

Confocal Raman microspectroscopy is widely used in dermatology and cosmetology for analysis of the concentration of skin components (lipids, natural moisturizing factor molecules, water) and the penetration depth of cosmetic/medical formulations in the human stratum corneum (SC) in vivo. In recent years, it was shown that confocal Raman microspectroscopy can also be used for noninvasive in vivo depth-dependent determination of the physiological parameters of the SC, such as lamellar and lateral organization of intercellular lipids (ICLs), folding properties of keratin, water mobility, and hydrogen bonding states. The results showed that the strongest skin barrier function, which is primarily manifested by the orthorhombic organization of ICLs, is provided at ≈20-40% SC depth, which is related to the maximal bonding state of water with surrounding components in the SC. The secondary and tertiary structures of keratin determine water binding in the SC, which is depth-dependent. This paper shows the technical possibility and advantage of confocal Raman microspectroscopy in noninvasive investigation of the skin and summarizes recent results on in vivo investigation of the human SC.

Safety and efficacy of combined essential oils for the skin barrier properties: In vitro, ex vivo and clinical studies.

OBJECTIVE: To evaluate the safety and the synergistic effects of tea tree, lavender, eucalyptus and tangerine essential oils in combination on the skin using in vitro, ex vivo and clinical studies. METHODS: The phototoxicity was predicted using 3T3 neutral red uptake phototoxicity test (OECD TG 432). Skin penetration was evaluated by confocal Raman microspectroscopy using direct application of essential oils to pig ears. For the clinical studies, 40 participants were enrolled and randomized in three groups: (1) lavender, eucalyptus and tangerine, (2) the same essential oils plus melaleuca and (3) placebo group. The skin was evaluated by noninvasive techniques before and after a 90-day period of topical use. RESULTS: The essential oils were non-phototoxic, but the tangerine oil showed dose-dependent cytotoxicity (IC50: 33.1 µg/ml), presenting 35% of penetration in the viable epidermis. On the contrary, 17.7 µg/ml in combination was applied per day in the clinical study and the penetration rate for the combinations (10%, 1.77 µg/ml achieving the viable epidermis) guaranteed the safety, since in the clinical study, the application of the four essential oils improved skin barrier and morphologic skin characteristics, as well as increased skin hydration and decreased sebum levels, with no unwanted effects reported. CONCLUSIONS: All essential oils studied were considered non-cytotoxic or non-phototoxic separately except tangerine, which present a dose-dependent cytotoxicity. Finally, the essential oils in combination in an appropriate amount were safe and effective in the improvement of the hydrolipidic balance and morphological properties of the skin.

OBJECTIF: évaluer la sécurité d'emploi et les effets synergiques des associations d'huiles essentielles d'arbre à thé, de lavande, d'eucalyptus et de mandarine sur la peau à l'aide d'études in vitro, ex vivo et cliniques. MÉTHODES: la phototoxicité a été prédite avec le test de phototoxicité de fixation du rouge neutre 3T3 (OCDE TG 432). La pénétration cutanée a été évaluée par microspectroscopie confocale de Raman grâce à l'application directe d'huiles essentielles sur les oreilles de cochons. Pour les études cliniques, 40 participants ont été inclus et randomisés dans trois groupes : (1) lavande, eucalyptus et mandarine, (2) les mêmes huiles essentielles plus melaleuca et (3) un groupe placebo. La peau a été évaluée par des techniques non invasives avant et après une période d'utilisation topique de 90 jours. RÉSULTATS: les huiles essentielles se sont avérées non phototoxiques, mais l'huile de mandarine a montré une cytotoxicité dose-dépendante (CI 50 : 33,1 µg/ml), représentant 35 % de pénétration dans l'épiderme viable. À l'inverse, dans l'étude clinique, une quantité de 17,7 µg/ml par jour en association a été appliquée, et le taux de pénétration des associations (10 %, soit 1,77 µg/ml atteignant l'épiderme viable) a garanti la sécurité d'emploi, puisque dans l'étude clinique, l'application des quatre huiles essentielles a amélioré la barrière cutanée et les caractéristiques morphologiques de la peau, et a entraîné une augmentation de l'hydratation cutanée et une diminution des taux de sébum, sans signalement d'effets indésirables. CONCLUSIONS: chacune des huiles essentielles étudiées a été considérée comme non cytotoxique ou non phototoxique, à l'exception de la mandarine, qui présente une cytotoxicité dose-dépendante. Enfin, l'association d'huiles essentielles en quantité appropriée a démontré sa sécurité d'emploi et son efficacité dans l'amélioration de l'équilibre hydrolipidique et des propriétés morphologiques de la peau.

Microdialysis on Ex Vivo Porcine Ear Skin Can Validly Study Dermal Penetration including the Fraction of Transfollicular Penetration-Demonstrated on Caffeine Nanocrystals.

Common ex vivo methods for penetration investigations often fail to monitor transfollicular penetration appropriately. In the present investigation, the validity of dermal microdialysis on the ex vivo porcine ear skin to investigate penetration kinetics, including transfollicular penetration, was studied. In setup A, a caffeine nanocrystal formulation was compared to a non-particular caffeine gel formulation. In setup B, two caffeine nanocrystal formulations of different sizes (200 nm, 700 nm) were compared to each other. Microdialysis samples were collected for 46 h. After sampling, the skin layers were separated, homogenized, and caffeine was quantified in all samples. In setup A the area under the curve (AUC) after crystal gel formulation application was 12 times higher than after non-particular formulation application. Setup B showed an increased AUC of 42% in the microdialysis data when the 700 nm caffeine crystals were applied compared to the 200 nm crystals. The microdialysis data was supported by the separation, homogenization and extraction data. Microdialysis performed on ex vivo porcine ear skin is a novel experimental setup. It is of high interest for further investigations since it is able to also capture the impact of follicular and transfollicular penetration kinetics as no other ex vivo setup can.

Retaining Skin Barrier Function Properties of the Stratum Corneum with Components of the Natural Moisturizing Factor-A Randomized, Placebo-Controlled Double-Blind In Vivo Study.

The influence of a topically applied formulation containing components of natural moisturizing factor (NMF) on barrier-related parameters of the stratum corneum (SC) was investigated in vivo using confocal Raman microspectroscopy in a randomized, placebo-controlled double-blind study on 12 volunteers for 14 days. This method allowed for the elucidation of subtle differences between the verum and the placebo even though the components of the verum naturally occur in the SC. This differentiation is not possible non-invasively by conventional methods. In this study, we found that the applied verum and placebo formulations disrupted the equilibrium of water, NMF and lipids in the SC. The adverse effects of the formulation could be mitigated by incorporating it into a simplified supplementation of NMF molecules. As a long-term effect, the amount of strongly bound water increases at 30-40% SC depth (p < 0.05) and the amount of weakly bound water decreases at 30-40% SC depth (p < 0.05) for the verum. This supplement was also unexpectedly able to prevent intercellular lipids (ICL) disorganization in selected depths. In the long term, the verum treatment limited the lateral disorganization of the ICL to the upper 20% SC depth. Further research is required to elucidate the interplay of these factors in the SC, to better understand their contribution to the equilibrium and barrier function of the skin. This understanding of the interaction of these naturally occurring components could help in the future to develop and optimize topical treatments for diseases like psoriasis, atopic dermatitis, ichthyosis where the skin barrier is disrupted.

Fiber-based SORS-SERDS system and chemometrics for the diagnostics and therapy monitoring of psoriasis inflammatory disease in vivo.

Psoriasis is considered a widespread dermatological disease that can strongly affect the quality of life. Currently, the treatment is continued until the skin surface appears clinically healed. However, lesions appearing normal may contain modifications in deeper layers. To terminate the treatment too early can highly increase the risk of relapses. Therefore, techniques are needed for a better knowledge of the treatment process, especially to detect the lesion modifications in deeper layers. In this study, we developed a fiber-based SORS-SERDS system in combination with machine learning algorithms to non-invasively determine the treatment efficiency of psoriasis. The system was designed to acquire Raman spectra from three different depths into the skin, which provide rich information about the skin modifications in deeper layers. This way, it is expected to prevent the occurrence of relapses in case of a too short treatment. The method was verified with a study of 24 patients upon their two visits: the data is acquired at the beginning of a standard treatment (visit 1) and four months afterwards (visit 2). A mean sensitivity of ≥85% was achieved to distinguish psoriasis from normal skin at visit 1. At visit 2, where the patients were healed according to the clinical appearance, the mean sensitivity was ≈65%.

Release of the model drug SR101 from polyurethane nanocapsules in porcine hair follicles triggered by LED-derived low dose UVA light.

Hair follicles (HFs) are important drug delivery targets for the therapy of miscellaneous skin diseases and for skin antisepsis. Furthermore, HFs significantly contribute to drug delivery of topically applied substances. Nanoparticulate systems are excellently suited for follicular drug delivery as they entail the opportunity of directed drug transport into HFs. Moreover, they involve the possibility of an intrafollicular drug release initiated by extrinsic or intrinsic trigger mechanisms. In this study, we present a novel preclinical model for an anatomically and temporally targeted intrafollicular drug release. In vitro release kinetics of the model drug sulforhodamine 101 (SR101) from newly synthesized ultraviolet A (UVA)-responsive polyurethane nanocapsules (NCs) were investigated by fluorescence spectroscopy. Low power density UVA radiation provided by a UVA light emitting diode (LED) induced a drug release of over 50% after 2 min. We further utilized confocal laser scanning microscopy (CLSM) to investigate follicular penetration as well as intrafollicular drug release on an ex vivo porcine ear skin model. UVA-responsive degradation of the NCs at a mean follicular penetration depth of 509 ± 104 µm ensured liberation of SR101 in the right place and at the right time. Thus, for the first time a UVA-triggered drug release from NCs within HFs was demonstrated in the present study. Cytotoxicity tests revealed that NCs synthesized with isophorone diisocyanate show sufficient biocompatibility after UVA-induced cleavage. A considerable and controllable release of various water-soluble therapeutics could be reached by means of the presented system without risking any radiation-related tissue damage. Therefore, the implementation of the presented system into clinical routine, e.g. for preoperative antisepsis of HFs, appears very promising.

Noninvasive measurement of the 308 nm LED-based UVB protection factor of sunscreens.

The current method for determining the sun protection factor (SPF) requires erythema formation. Noninvasive alternatives have recently been suggested by several groups. Our group previously developed a functional sensor based on diffuse reflectance measurements with one UVB LED, which was previously evaluated on pig ear skin. Here we present the results of a systematic in vivo study using 12 sunscreens on 10 volunteers (skin types [ST] I-III). The relationship of the UVB-LED reflectance of unprotected skin and melanin index was determined for each ST. The spatial variation of the reflectance signal of different positions was analyzed and seems to be mainly influenced by sample inhomogeneity except for high-protection factors (PFs) where signal levels are close to detection noise. Despite the low-signal levels, a correlation of the measured LED-based UVB PF with SPF reference values from test institutes with R2 = 0.57 is obtained, suggesting a strong relationship of SPF and LED-based UVB-PF. Measured PFs tend to be lower for increasing skin pigmentation. The sensor design seems to be suitable for investigations where a fast measurement of relative changes of PFs, such as due to inhomogeneous application, bathing and sweating, is of interest.

Electrohydrodynamic spray applicator for homogenous application and reduced overspray of sunscreen.

BACKGROUND: The recommended amount of sunscreen by hand application (2 mg/cm2 ) is in reality not achieved, which decreases the homogeneity and thereby the effective sun protection factor (SPF). MATERIALS AND METHODS: The homogeneity of sunscreen applied by a newly developed spray applicator using an electrostatically charged aerosol, for which a hand rubbing of the formulation is not necessary, is evaluated. In vivo experiments were performed on the volar forearms of human volunteers using the spray applicator compared to the standardized hand application according to ISO 24444. RESULTS: The distribution homogeneity was assessed qualitatively using in vivo laser scanning microscopy and quantitatively by absorption spectroscopy after tape stripping and by the standard deviation of multiple spatially displaced reflectance measurements for non-invasive SPF determination below the minimal erythemal dose, which showed a significantly higher homogeneity by 20.9% after spray application compared to hand application. CONCLUSION: Non-invasive SPF determination of multiple spatially displaced reflectance measurements was proven to be a suitable method for the non-invasive determination of the sunscreen distribution homogeneity. Electrostatically charged spray application increased the sunscreen distribution homogeneity on the skin and can reduce the amount of overspray.

In vivo sun protection factor and UVA protection factor determination using (hybrid) diffuse reflectance spectroscopy and a multi-lambda-LED light source.

The sun protection factor (SPF) values are currently determined using an invasive procedure, in which the volunteers are irradiated with ultraviolet (UV) light. Non-invasive approaches based on hybrid diffuse reflectance spectroscopy (HDRS) have shown a good correlation with conventional SPF testing. Here, we present a novel compact and adjustable DRS test system. The in vivo measurements were performed using a multi-lambda-LED light source and an 84-channel imaging spectrograph with a fiber optic probe for detection. A transmission spectrum was calculated based on the reflectance measured with sunscreen and the reflectance measured without sunscreen. The preexposure in vitro spectrum was fitted to the in vivo spectrum. Each of the 11 test products was investigated on 10 volunteers. The SPF and UVA-PF values obtained by this new approach were compared with in vivo SPF results determined by certified test institutes. A correlation coefficient R2 = 0.86 for SPF, and R2 = 0.92 for UVA-PF were calculated. Having examined various approaches to apply the HDRS principle, the method we present was found to produce valid and reproducible results, suggesting that the multi-lambda-LED device is suitable for in-vivo SPF testing based on the HDRS principle as well as for in-vivo UVA-PF measurements.

The impact of skin massage frequency on the intrafollicular transport of silica nanoparticles: Validation of the ratchet effect on an ex vivo porcine skin model.

The human hair follicle (HF) represents a promising drug delivery target as an anatomical entity by itself, but also as a gateway enabling dermal or systemic bioavailability of active cosmetic and pharmaceutical ingredients. Due to its morphological characteristics, the HF provides a mechanically driven transport process of nanoparticles (NPs) when external forces are applied. This mechanism was presented as the so-called ratchet effect within the framework of an in silico study published recently. To investigate the influence of massage frequency on the penetration depth of NPs, and, by this, to validate the results obtained in silico, we implemented a corresponding application protocol on an ex vivo porcine skin model. In this connection, we compared three different skin massage frequencies (4.2 Hz, 50 Hz, 100 Hz) for the topical application of cyanine 5-labeled silica NPs (Cy5-SNPs). To elucidate the interplay of frequency and particle size, we furthermore applied Cy5-SNPs of three different diameters (300 nm, 676 nm, 1000 nm). Confocal laser scanning microscopy was utilized to investigate the follicular penetration depth of Cy5-SNPs on cryohistological slices. By this, we could demonstrate that the massage frequency and the follicular penetration depth exhibit an inverse relation pattern. Thus, the highest follicular penetration depth was observed within the 4.2 Hz group, while the lowest follicular penetration depth was found within the 100 Hz group for each Cy5-SNP size category. Additionally, we found that 676 nm Cy5-SNPs penetrated significantly deeper into HFs than 300 nm Cy5-SNPs and 1000 nm Cy5-SNPs, respectively. Summarizing, our results show that a low massage frequency including a dominant radial direction component leads to deeper follicular penetration depths of NPs than automated 3D-oscillation massage at 50 Hz or 100 Hz. Thus, our findings are in line with recent in silico results. Regarding translational purposes, our results are of high interest, since a massage executed at 250BPM (4.2 Hz) is within a realizable range for manual application, e.g. for the implementation into clinical routines or the domestic use of drugs or cosmetics. Furthermore, the application of different massage frequencies offers the opportunity of patho-specific targeting as different anatomical parts of the HF can be reached.