Bio-derived hydroxystearic acid ameliorates skin age spots and conspicuous pores.

INTRODUCTION: We report on the preparation and efficacy of 10-hydroxystearic acid (HSA) that improves facial age spots and conspicuous pores. METHODS: The hydration of oleic acid into HSA was catalyzed by the oleate hydratase from Escherichia coli. Following treatment with HSA, collagen type I and type III was assessed in primary human dermal fibroblasts together with collagen type III, p53 protein levels and sunburn cells (SBC) after UVB irradiation (1 J cm-2 ) by immunohistochemistry on human ex vivo skin. UVB-induced expression of matrix metalloprotease-1 (MMP-1) was determined from full thickness skin by RT-qPCR. Modification of the fibroblast secretome by HSA was studied by mass-spectrometry-based proteomics. In a full-face, double blind, vehicle-controlled trial HSA was assessed for its effects on conspicuous facial pore size and degree of pigmentation of age spots in Caucasian women over an 8-week period. RESULTS: HSA was obtained in enantiomeric pure, high yield (≥80%). Collagen type I and type III levels were dose-dependently increased (96% and 244%; P < 0.01) in vitro and collagen type III in ex vivo skin by +57% (P < 0.01) by HSA. HSA also inhibited UVB-induced MMP-1 gene expression (83%; P < 0.01) and mitigated SBC induction (-34% vs. vehicle control) and reduced significantly UV-induced p53 up-regulation (-46% vs. vehicle control; P < 0.01) in irradiated skin. HSA modified the fibroblast secretome with significant increases in proteins associated with the WNT pathway that could reduce melanogenesis and proteins that could modify dermal fibroblast activity and keratinocyte differentiation to account for the alleviation of conspicuous pores. Docking studies in silico and EC50 determination in reporter gene assays (EC50 5.5 × 10-6  M) identified HSA as a peroxisomal proliferator activated receptor-α (PPARα) agonist. Clinically, HSA showed a statistically significant decrease of surface and volume of skin pores (P < 0.05) after 8 weeks of application and age spots became significantly less pigmented than the surrounding skin (contrast, P < 0.05) after 4 weeks. CONCLUSION: HSA acts as a PPARα agonist to reduce the signs of age spots and conspicuous pores by significantly modulating the expression of p53, SBC, MMP-1 and collagen together with major changes in secreted proteins that modify keratinocyte, melanocyte and fibroblast cell behavior.

INTRODUCTION: voici notre rapport sur la préparation et l'efficacité de l'acide 10-hydroxystéarique (AHS) qui atténue les taches de vieillesse faciale et améliore l'apparence des pores. MÉTHODES: l'hydratation de l'acide oléique en AHS a été catalysée par l'hydratase d'oléate à partir de l'Escherichia coli. Après un traitement par AHS, les collagènes de type I et de type III ont été analysés dans des fibroblastes dermiques humains primaires, ainsi que le taux de collagène de type III et de protéine p53, et les cellules provenant de coups de soleil (sunburn cells, SBC) après irradiation par UVB (1 J cm−2 ) par immunohistochimie sur de la peau humaine ex vivo. L'expression de la matrice métalloprotéase-1 (MMP-1) induite par les UVB a été déterminée à partir d'un échantillon de pleine épaisseur de peau par RT-qPCR. La modification du sécrétome des fibroblastes par l'AHS a été étudiée par analyse protéomique basée sur une spectrométrie de masse. Dans une étude du visage entier, en double aveugle, contrôlée par excipient, l'AHS a été évaluée pour ses effets sur la taille des pores apparents du visage et sur le degré de pigmentation de taches de vieillesse chez des femmes de race blanche sur une période de 8 semaines. RÉSULTATS: l'AHS a été obtenu à un haut rendement, énantiomérique pur (≥80 %). Les taux de collagènes de type I et de type III ont augmenté in vitro en fonction de la dose (96 % et 244 %; P < 0.01) et le collagène de type III dans de la peau ex vivo de +57 % (P < 0.01) lors d'un traitement par AHS. L'AHS a également inhibé l'expression génique MMP-1 induite par les UVB (83%; P < 0.01) et a atténuée l'induction des SBC (−34 % par rapport à l'excipient), et a réduit significativement la régulation à la hausse du p53 induite par les UV (−46% par rapport à l'excipient; P < 0.01) sur de la peau irradiée. L'AHS a modifié le sécrétome des fibroblastes avec des augmentations significatives des protéines associées à la voie WNT qui pouvaient réduire la mélanogenèse et des protéines qui pouvaient modifier l'activité des fibroblastes dermiques et la différenciation des kératinocytes pour une atténuation des pores apparents. Des études de docking in silico et la détermination de l'EC50 dans les dosages des gènes rapporteurs (EC50 5.5 × 9 10−6 M) ont identifié l'AHS comme un agoniste du récepteur-α activé par les proliférateurs de peroxysomes (peroxisomal proliferator activated receptor-α, PPARα). Cliniquement, l'AHS a permis une diminution statistiquement significative de la surface et du volume des pores de la peau (P < 0.05) après 8 semaines d'application, et les taches de vieillesse sont devenues significativement moins pigmentées par rapport à la peau environnante (contraste, P < 0,05) après 4 semaines. CONCLUSION: l'AHS agit comme un agoniste du PPARα pour réduire les signes de taches de vieillesse et l'apparence des pores par une modulation significative de l'expression de la protéine p53, des SBC, de la MMP-1 et du collagène avec des changements majeurs dans les protéines sécrétées qui modifient le comportement cellulaire des kératinocytes, des mélanocytes et des fibroblastes.

The effect of photodamage on the female Caucasian facial stratum corneum corneome using mass spectrometry-based proteomics.

BACKGROUND: The effect of photodamage on facial stratum corneum (SC) is still poorly understood. OBJECTIVE: To describe the SC proteome from tape strippings of Caucasian SC from photoexposed cheek and photoprotected post-auricular (PA) site, a global analysis of photodamage on the skin will be developed leading to a better understanding of keratinocyte signalling pathways and identification of new molecular targets for the treatment of photoaged skin. METHODS: Female Caucasian subjects had nine consecutive tape strippings taken from their cheeks and PA site. Proteins were extracted and the trypsin-digested peptides were analysed by nanochromatography coupled to a high-resolution mass spectrometer. Data-dependent acquisition allowed protein identification that was processed by Paragon algorithm of Protein Pilot software. RESULTS: Changes in the levels of epidermal differentiation proteins were apparent indicating poor epidermal differentiation and SC maturation (keratins, cornified envelope (CE) proteins) on photoexposed cheeks. Differences in protease-anti-protease balance were observed for corneodesmolysis (favouring desquamation) and filaggrinolysis (favouring reduced filaggrin processing). 12R-LOX, a CE maturation enzyme, was reduced in photodamaged skin but not transglutaminases. Changes in signal keratinocyte transduction pathway markers were demonstrated especially by reduced levels of downstream signalling markers such as calreticulin (unfolded protein response; UPR) and increased level of stratifin (target of rapamycin; mTOR). Evidence for impaired proteostasis was apparent by reduced levels of a key proteasomal subunit (subunit beta type-6). Finally, key antioxidant proteins were upregulated except catalase. CONCLUSION: Clear examples of poor keratinocyte differentiation and associated metabolic and signalling pathways together with reduced SC maturation were identified in photodamaged facial SC. Corneocyte immaturity was evident with changes in CE proteins. Particularly, the reduction in 12R-LOX is a novel finding in photodamaged skin and supports the lack of SC maturation. Moreover, filaggrinolysis was reduced, whereas corneodesmolysis was enhanced. From our results, we propose that there is a poor cross-talk between the keratinocyte endoplasmic reticulum UPR, proteasome network and autophagy machinery that possibly leads to impaired keratinocyte proteostasis. Superimposed on these aberrations is an apparently enhanced mTOR pathway that also contributes to reduced SC formation and maturation. Our results clearly indicate a corneocyte scaffold disorder in photodamaged cheek SC.