Transcriptional profiling of epidermal barrier formation in vitro.

BACKGROUND: Barrier function is integral to the health of epithelial tissues. Currently, there is a broad need to develop and improve our knowledge with regard to barrier function for reversal of mild skin irritation and dryness. However, there are few in vitro models that incorporate modulations of both lipids and epidermal differentiation programs for pre-clinical testing to aid in the understanding of barrier health. OBJECTIVE: We have generated a reconstituted epidermis on a decellularized dermis (DED) and characterized its barrier properties relative to human epidermis in order to determine its utility for modeling barrier formation and repair. METHODS: We followed the process of epidermal differentiation and barrier formation through immunocytochemistry and transcriptional profiling. We examined barrier functionality through measurements of surface pH, lipid composition, stratum corneum water content, and the ability to demonstrate topical dose-dependent exclusion of surfactant. RESULTS: Transcriptional profiling of the epidermal model during its formation reveals temporal patterns of gene expression associated with processes regulating barrier function. The profiling is supported by gradual formation and maturation of a stratum corneum and expression of appropriate markers of epidermis development. The model displays a functional barrier and a water gradient between the stratum corneum and viable layers, as determined by confocal Raman spectroscopy. The stratum corneum layer displays a normal acidic pH and an appropriate composition of barrier lipids. CONCLUSION: The epidermal model demonstrates its utility as an investigative tool for barrier health and provides a window into the transcriptional regulation of multiple aspects of barrier formation.

Auto-classification of acne lesions using multimodal imaging.

Differentiating inflammatory and non-inflammatory acne lesions and obtaining lesion counts is pivotal part of acne evaluation. Manual lesion counting has reliably demonstrated the clinical efficacy of anti-acne products for decades. However, maintaining assessment consistency within and across acne trials is an important consideration since lesion counting can be subjective to the individual evaluators, and the technique has not been rigorously standardized. VISIA-CR is a multi-spectral and multi-modal facial imaging system. It captures fluorescence images of Horn and Porphyrin, absorption images of Hemoglobin and Melanin, and skin texture and topography characterizing broad-spectrum polarized and non-polarized images. These images are analyzed for auto-classification of inflammatory and non-inflammatory acne lesion, measurement of erythema, and post-acne pigmentation changes. In this work the accuracy of this acne lesion auto-classification technique is demonstrated by comparing the auto-detected lesions counts with those counted by expert physicians. The accuracy is further substantiated by comparing and confirming the facial location and type of every auto-identified acne lesion with those identified by the physicians. Our results indicate a strong correlation between manual and auto-classified lesion counts (correlation coefficient >0.9) for both inflammatory and non inflammatory lesions This technology has the potential to eliminate the tedium of manual lesion counting, and provide an accurate, reproducible, and clinically relevant evaluation of acne lesions. As an aid to physicians it will allow development of a standardized technique for evaluating acne in clinical research, as well as accurately choosing treatment options for their patients according to the severity of a specific lesion type in clinical practice

Practical application of genomics to the development of a topical cosmetic anti-aging regimen.

The development of topical cosmetic anti-aging products is becoming increasingly sophisticated. This is demonstrated by the benefit agents selected and the scientific approaches used to identify them, treatment protocols that increasingly incorporate multi-product regimens, and the level of rigor in the clinical testing used to demonstrate efficacy. Consistent with these principles, a new cosmetic anti-aging regimen was recently developed. The key product ingredients were identified based on an understanding of the key mechanistic themes associated with aging at the genomic level coupled with appropriate in vitro testing. The products were designed to provide optimum benefits when used in combination in a regimen format. This cosmetic regimen was then tested for efficacy against the appearance of facial wrinkles in a 24-week clinical trial compared with 0.02% tretinoin, a recognized benchmark prescription treatment for facial wrinkling. The cosmetic regimen significantly improved wrinkle appearance after 8 weeks relative to tretinoin and was better tolerated. Wrinkle appearance benefits from the two treatments in cohorts of subjects who continued treatment through 24 weeks were also comparable.

Efficacy of anti-aging products for periorbital wrinkles as measured by 3-D imaging.

BACKGROUND: The periorbital area is a key wrinkle-prone region, where the first signs of aging usually appear. AIMS: To demonstrate the ability of new anti-aging moisturizing products to improve overall smoothness and wrinkle depth appearance in the periorbital region via the Fast Optical in vivo Topometry of Human Skin (FOITS). METHODS: Two double-blind, randomized, controlled, split-face studies (n = 42, Study 1; n = 35, Study 2) were conducted in women 30-70 years old with moderate to distinct periorbital wrinkles. Subjects applied 0.5 g of individual products to half their face twice daily for 4 weeks. Four test products containing niacinamide, the peptides Pal-KT and Pal-KTTKS, and carnosine were used and included a daytime SPF 30 lotion also containing antioxidants, a night cream, an eye cream also containing caffeine, and a wrinkle treatment containing retinyl propionate. The wrinkle treatment was only tested in Study 2. The FOITS technique was used to measure changes in periorbital R(a) (mean roughness) and R(z) (average maximum roughness) at 2 and 4 weeks. RESULTS: In Study 1, the daytime SPF 30 lotion, night cream, and eye cream significantly improved crow's feet smoothness after 4 weeks relative to no treatment. After 4 weeks, the daytime SPF 30 lotion and night cream, but not the eye cream, were significantly better than no treatment at improving R(z). In Study 2, the night cream, eye cream, and wrinkle treatment, but not the daytime SPF 30 lotion, significantly improved both R(a) and R(z) after 4 weeks. To increase power and precision of estimates, a meta-analysis was performed; the pooled data showed all three products were significantly better than no treatment at improving R(a) and R(z) after 4 weeks. CONCLUSIONS: Four weeks of treatment with these products was shown to improve the smoothness of periorbital skin and to reduce the apparent depth of larger wrinkles.

Practical applications of genomics research for treatment of aging skin.

Skin aging integrates the impact of extrinsic skin insults (e.g., ultraviolet [UV] light, etc.) with chronological, genetically programmed decreases in cellular function. A genomic study of aged skin highlighted the mechanistic importance of skin barrier function, exfoliation, control of reactive oxygen species and maintenance of extracellular matrix to the aging process. A set of topical products designed to address these mechanistic themes was developed and clinically tested. The individual products improved skin barrier function, hydration and skin turnover, as well as the smoothness and depth of periorbital wrinkles. Treatment with a regimen of these products improved the appearance of facial wrinkles after eight weeks. Changes in treated subjects' stratum corneum protein biomarker levels were consistent with the mechanistic pathways identified in the genomic work. Thus, leveraging a genomic understanding of skin aging led to the development of a clinically efficacious, aesthetically pleasing cosmetic regimen that improved the appearance of aged skin.

Isolation and expression of a Malassezia globosa lipase gene, LIP1.

Dandruff and seborrheic dermatitis (D/SD) are common hyperproliferative scalp disorders with a similar etiology. Both result, in part, from metabolic activity of Malassezia globosa and Malassezia restricta, commensal basidiomycete yeasts commonly found on human scalps. Current hypotheses about the mechanism of D/SD include Malassezia-induced fatty acid metabolism, particularly lipase-mediated breakdown of sebaceous lipids and release of irritating free fatty acids. We report that lipase activity was detected in four species of Malassezia, including M. globosa. We isolated lipase activity by washing M. globosa cells. The isolated lipase was active against diolein, but not triolein. In contrast, intact cells showed lipase activity against both substrates, suggesting the presence of at least another lipase. The diglyceride-hydrolyzing lipase was purified from the extract, and much of its sequence was determined by peptide sequencing. The corresponding lipase gene (LIP1) was cloned and sequenced. Confirmation that LIP1 encoded a functional lipase was obtained using a covalent lipase inhibitor. LIP1 was differentially expressed in vitro. Expression was detected on three out of five human scalps, as indicated by reverse transcription-PCR. This is the first step in a molecular description of lipid metabolism on the scalp, ultimately leading toward a test of its role in D/SD etiology.

Three etiologic facets of dandruff and seborrheic dermatitis: Malassezia fungi, sebaceous lipids, and individual sensitivity.

Application of new molecular and biochemical tools has greatly increased our understanding of the organisms, mechanisms, and treatments of dandruff and seborrheic dermatitis. Dandruff results from at least three etiologic factors: Malassezia fungi, sebaceous secretions, and individual sensitivity. While Malassezia (formerly P. ovale) has long been a suspected cause, implicated by its presence on skin and lipophylic nature, lack of correlation between Malassezia number and the presence and severity of dandruff has remained perplexing. We have previously identified the Malassezia species correlating to dandruff and seborrheic dermatitis. In this report, we show that dandruff is mediated by Malassezia metabolites, specifically irritating free fatty acids released from sebaceous triglycerides. Investigation of the toxic Malassezia free fatty acid metabolites (represented by oleic acid) reveals the component of individual susceptibility. Malassezia metabolism results in increased levels of scalp free fatty acids. Of the three etiologic factors implicated in dandruff, Malassezia, sebaceous triglycerides, and individual susceptibility, Malassezia are the easiest to control. Pyrithione zinc kills Malassezia and all other fungi, and is highly effective against the Malassezia species actually found on scalp. Reduction in fungi reduces free fatty acids, thereby reducing scalp flaking and itch.