Stress In Human Skin Research Articles

Night Cream Containing Melatonin, Carnosine and Helichrysum italicum Extract Helps Reduce Skin Reactivity and Signs of Photodamage: Ex Vivo and Clinical Studies

IntroductionExtrinsic factors, such as solar radiation and urban pollution, cause damage that alters the structure, function and appearance of skin. The aim of this study was to determine the ability of a night cream containing melatonin, carnosine and Helichrysum italicum extract (referred to here as Night Cream) to reduce extrinsic skin damage, and to evaluate the efficacy of this Night Cream to reduce clinical signs of age and photodamage under normal conditions of use.MethodsRecovery from extrinsic damage was assessed by exposing human skin explants to ultraviolet (UV) A, infrared light, blue light or pollution and then treating the stress-exposed explants with Night Cream. Markers of oxidative stress were examined by immunohistochemistry. Anti-aging and calming properties were determined in four single-center, open-label trials involving 117 individuals. Subjects applied Night Cream to their face once nightly for up to 12 weeks. Improvements in clinical signs of age and photodamage, and reduction of lactic acid-induced stinging were evaluated by investigator assessment and subject self-assessment.ResultsNight Cream significantly reduced oxidative stress in human skin ex vivo. Clinically, hydration (+ 64.4%; p < 0.05) and transepidermal water loss (TEWL) values (− 10.0%; p < 0.05) were improved within 1 h of use. Wrinkle counts were reduced by up to 18.9% (p < 0.05), and brown and UV spot numbers by 5.5% (p < 0.05) and 13.2% (p < 0.05), respectively. Lactic acid-induced stinging was significantly reduced within 7 days of use, with 86.7% of subjects reporting that their skin felt calmer.ConclusionThese findings suggest that Night Cream reduces skin damage caused by environmental factors and that its nightly use can improve clinical signs of aging with additional skin calming benefits.

LB1586 Noninvasive imaging of UV-induced oxidative stress in human skin using ultra-weak photon emission

LB1586 Noninvasive imaging of UV-induced oxidative stress in human skin using ultra-weak photon emission

MMP-1 suppressing activity of high-molecular weight fraction isolated from the roots of Cynanchum wilfordii Hemsley

Cynanchum wilfordii Hemsley is a traditional herbal medicine that has been widely used in Asia. In the present study, we investigated the MMP-1 suppressing activity of a high-molecular weight fraction isolated from the roots of C. wilfordii Hemsley (HMF). In human skin tissue, UVB irradiation significantly increased matrix metalloproteinase-1 (MMP-1) expression, while HMF suppressed this effect. The inhibitory effect of HMF was confirmed in both primary human dermal fibroblasts (HDFs) and a human keratinocyte (HaCaT) model. An investigation of the underlying mechanisms responsible revealed that MKK4-JNK, MEK-ERK, and MKK3/6-p38 phosphorylation was downregulated by HMF treatment. HMF also induced a 30–40% increase in DPPH radical scavenging activity and recovered expression levels of the antioxidant enzyme superoxide dismutase. Taken together, these findings suggest that HMF elicits MMP-1 suppressing activity via inhibition of the signaling that drives UVB-induced oxidative stress in human skin.

The B6 -vitamer Pyridoxal is a Sensitizer of UVA-induced Genotoxic Stress in Human Primary Keratinocytes and Reconstructed Epidermis.

UVA-driven photooxidative stress in human skin may originate from excitation of specific endogenous chromophores acting as photosensitizers. Previously, we have demonstrated that 3-hydroxypyridine-derived chromophores including B6 -vitamers (pyridoxine, pyridoxamine and pyridoxal) are endogenous photosensitizers that enhance UVA-induced photooxidative stress in human skin cells. Here, we report that the B6 -vitamer pyridoxal is a sensitizer of genotoxic stress in human adult primary keratinocytes (HEKa) and reconstructed epidermis. Comparative array analysis indicated that exposure to the combined action of pyridoxal and UVA caused upregulation of heat shock (HSPA6, HSPA1A, HSPA1L, HSPA2), redox (GSTM3, EGR1, MT2A, HMOX1, SOD1) and genotoxic (GADD45A, DDIT3, CDKN1A) stress response gene expression. Together with potentiation of UVA-induced photooxidative stress and glutathione depletion, induction of HEKa cell death occurred only in response to the combined action of pyridoxal and UVA. In addition to activational phosphorylation indicative of genotoxic stress [p53 (Ser15) and γ-H2AX (Ser139)], comet analysis indicated the formation of Fpg-sensitive oxidative DNA lesions, observable only after combined exposure to pyridoxal and UVA. In human reconstructed epidermis, pyridoxal preincubation followed by UVA exposure caused genomic oxidative base damage, procaspase 3 cleavage and TUNEL positivity, consistent with UVA-driven photooxidative damage that may be relevant to human skin exposed to high concentrations of B6 -vitamers.

Mitochondrial DNA damage as a biomarker for ultraviolet radiation exposure and oxidative stress

The skin is regularly exposed to the harmful effects of sunlight, such as ultraviolet radiation (UVR), which leads to ageing effects as well as clinical precancerous lesions and skin cancer. The accumulation of mitochondrial DNA (mtDNA) damage has been strongly associated as an underlying cause of the general ageing process in tissues and mtDNA damage has been associated with cancer development in many tissues including human skin. This scenario is linked to the key roles of mitochondrial function and mtDNA both in terms of energy production and also oxidative stress production as well as a mediator of apoptosis. We and others have pioneered the use of mtDNA damage as a highly sensitive biomarker of UVR exposure and oxidative stress in human skin; furthermore, ageing-dependent mtDNA mutations can be accelerated by exposure to sunlight. In addition, this review will also highlight useful applications of mtDNA as a biomarker of UVR-induced oxidative stress including effects of antioxidants.

Investigating the role of melanin in UVA/UVB- and hydrogen peroxide-induced cellular and mitochondrial ROS production and mitochondrial DNA damage in human melanoma cells

Skin cancer incidence is dramatically increasing worldwide, with exposure to ultraviolet radiation (UVR) a predominant factor. The UVA component initiates oxidative stress in human skin, although its exact role in the initiation of skin cancer, particularly malignant melanoma, remains unclear and is controversial because there is evidence for a melanin-dependent mechanism in UVA-linked melanoma studies. Nonpigmented (CHL-1, A375), moderately pigmented (FM55, SKmel23), and highly pigmented (FM94, hyperpigmented FM55) human melanoma cell lines have been used to investigate UVA-induced production of reactive oxygen species using FACS analysis, at both the cellular (dihydrorhodamine-123) and the mitochondrial (MitoSOX) level, where most cellular stress is generated. For the first time, downstream mtDNA damage (utilizing a quantitative long-PCR assay) has been investigated. Using UVA, UVB, and H2O2 as cellular stressors, we have explored the dual roles of melanin as a photoprotector and photosensitizer. The presence of melanin has no influence over cellular oxidative stress generation, whereas, in contrast, melanin protects against mitochondrial superoxide generation and mtDNA damage (one-way ANOVA with post hoc Tukey's analysis, P<0.001). We show that if melanin binds directly to DNA, it acts as a direct photosensitizer of mtDNA damage during UVA irradiation (P<0.001), providing evidence for the dual roles of melanin.

Terpenoids isolated from Tunisian Nigella sativa L. essential oil with antioxidant activity and the ability to inhibit nitric oxide production

ABSTRACTFractionation of the essential oil from the seeds of Nigella sativa (Ranunculaceae) led to the isolation of four terpenoids: trans‐ (1), cis‐sabinene hydrate methyl ether (2), 1,2‐epoxy‐menth‐4‐ene (3) and 1,2‐epoxy‐menth‐4(8)‐ene (4). Structure elucidation was performed using NMR (1D and 2D NMR) experiments and mass spectral data. Compounds 3 and 4 are reported for the first time as natural products. Bioactivities of the compounds were assessed. The four compounds exhibit antioxidant activity in vitro, as assessed by the oxygen radical absorbance capacity test. Moreover, compounds 1, 2 and 4 displayed strong ability to inhibit oxidative stress in human skin WS‐1 fibroblasts cells, with IC50 values of 0.32, 0.005 and 0.43 µ m, respectively. In addition, the four tested compounds significantly inhibited nitric oxide release by lipopolysaccharide‐activated RAW 264.7 macrophages. The results show that the most effective compound was cis‐sabinene hydrate methyl ether (2). In addition, the cis‐configurated compound (2) exerted stronger activities in comparison with the trans‐configurated compound (1), suggesting geometric stereoselectivity for the biological activities. Copyright © 2011 John Wiley &amp; Sons, Ltd.

Interaction of a functionalized complex of the flavonoid hesperetin with the AhR pathway and CYP1A1 expression: involvement in its protective effects against benzo[a]pyrene-induced oxidative stress in human skin

Skin cancer pathogenesis is partially associated to the oxidative stress conditions induced by environmentally carcinogens such as benzo[a]pyrene (BaP). The protective effects against BaP-induced oxidative stress of the flavonoid hesperetin as a complex with the 2-hydroxypropyl-β-cyclodextrin (HE/HP-β-CyD) have been evaluated using an ex vivo human skin model. Human healthy skin has been pre-treated with the functionalized complex HE/HP-β-CyD (0.5-50 μM) before BaP (5 μM) application simulating occupational and environmental exposure. Oxidative stress was evaluated in terms of 3-(4, 5-dimethylthiazol-2-yl)-2, 5-dipheyltetrazolium bromide reduction, protein peroxidation and reactive oxygen species (ROS) formation. Additionally, it has been investigated whether the potential protective effects of HE/HP-β-CyD may be correlated to the interaction with aryl hydrocarbon receptor (AhR) pathway. A significant protection by HE/HP-β-CyD against the BaP-induced increase in ROS and carbonyl compound production, as well as reduction in tissue viability, has been observed (p<0.001). Results obtained showed that HE/HP-β-CyD was also able to reduce BaP-induced AhR and CYP1A1 protein expression (p<0.001). Experimental evidences provided from this study suggest significant preventive properties of HE/HP-β-CyD in the toxicity caused by environmental carcinogens such as PAHs.

The malondialdehyde-derived fluorophore DHP-lysine is a potent sensitizer of UVA-induced photooxidative stress in human skin cells

Light-driven electron and energy transfer involving non-DNA skin chromophores as endogenous photosensitizers induces oxidative stress in UVA-exposed human skin, a process relevant to photoaging and photocarcinogenesis. Malondialdehyde is an electrophilic dicarbonyl-species derived from membrane lipid peroxidation. Here, we present experimental evidence suggesting that the malondialdehyde-derived protein epitope dihydropyridine (DHP)-lysine is a potent endogenous UVA-photosensitizer of human skin cells. Immunohistochemical analysis revealed the abundant occurrence of malondialdehyde-derived and DHP-lysine epitopes in human skin. Using the chemically protected dihydropyridine-derivative (2S)-Boc-2-amino-6-(3,5-diformyl-4-methyl-4H-pyridin-1-yl)-hexanoic acid-t-butylester as a model of peptide-bound DHP-lysine, photodynamic inhibition of proliferation and induction of cell death were observed in human skin Hs27 fibroblasts as well as primary and HaCaT keratinocytes exposed to the combined action of UVA and DHP-lysine. DHP-lysine photosensitization induced intracellular oxidative stress, p38 MAPkinase activation, and upregulation of heme oxygenase-1 expression. Consistent with UVA-driven ROS formation from DHP-lysine, formation of superoxide, hydrogen peroxide, and singlet oxygen was detected in chemical assays, but little protection was achieved using SOD or catalase during cellular photosensitization. In contrast, inclusion of NaN 3 completely abolished DHP-photosensitization. Taken together, these data demonstrate photodynamic activity of DHP-lysine and support the hypothesis that malondialdehyde-derived protein-epitopes may function as endogenous sensitizers of UVA-induced oxidative stress in human skin.

Exposure of human skin to benzo[a]pyrene: Role of CYP1A1 and aryl hydrocarbon receptor in oxidative stress generation

Exposure to benzo[a]pyrene (BaP) can induce inflammatory skin diseases and skin cancer, which are both associated to oxidative stress. BaP is known to bind with high specificity to the aryl hydrocarbon receptor (AhR), modifying the expression of CYP1A1, involved both in cancer and inflammation. While the current knowledge is based on murine skin and cell culture data, in this study human healthy skin has been treated with 5 μM BaP in conditions simulating occupational and environmental exposure. AhR and CYP1A1 expression was evaluated by Western blotting, which revealed their presence even in control untreated skin; both enzyme and receptor increased more than twofold after exposure to BaP. AhR expression level was lower than CYP1A1 in basal conditions and following induction. Oxidative stress was evaluated in terms of MTT reduction, protein peroxidation and reactive oxygen species (ROS) formation. A significant increase in ROS and carbonyl compound production, as well as reduced tissue viability have been determined by BaP. The results of this experiment indicate that BaP, an AhR agonist, can significantly increase receptor and CYP1A1 expression and induce oxidative stress in human skin, confirming the involvement of this pathway in the pathogenesis of tissue damage due to polycyclic aromatic hydrocarbons.

Using ultra-weak photon emission to determine the effect of oligomeric proanthocyanidins on oxidative stress of human skin

Exposure of skin to ultraviolet (UV) radiation triggers oxidative stress in skin tissue that can lead to erythema, early skin aging or even cancer. It is suggested that oligomeric proanthocyanidins (OPCs), phytonutrients that belong to the polyphenol family have an anti-oxidant/anti-inflammatory activity on the skin. Measuring ultra-weak photon emission (UPE) is a non-invasive, fairly-sensitive and convenient technique for continuously monitoring oxidative stress. The present study was undertaken to confirm anti-oxidant activity of the specific OPCs cream formulation in human skin by measuring UPE of skin. In the present study 25 healthy female subjects participated. As a baseline measurement of skin, UPE was recorded from the dorsal surface of the subjects’ hands before (spontaneous UPE) and after exposure to UV (UV-induced UPE). The effects of the OPCs cream on spontaneous and UV-induced UPE were measured using a fractionated UV exposure protocol. UV exposure resulted in an increase in UPE from both hands. Repeat UV exposure also resulted in a long-term increase of spontaneous UPE. This is likely due to depletion of anti-oxidant capacity of skin resulting in sensitization of skin to UV. It was assessed by measuring spontaneous UPE at 80 min after each UV exposure. Application of the OPCs cream immediately after UV exposure resulted in a significant (approx. 30%) decrease in UV-induced UPE. Topical OPCs cream application also reduced sensitization of skin to UV following repeated UV exposure (i.e., reduced long-term increase in spontaneous UPE). This study indicates that the specific OPCs cream formulation significantly decreases UV-induced oxidative stress in human skin based on UPE measurement. It therefore suggests that regular use of this OPCs cream might protect skin from harmful effects of UV.

Topical formulation containing natural oligomeric proanthocyanidins (OPCS) reduces UV-induced oxidative stress in skin as assessed by ultra-weak photon emission (UPE)

Background OPCs are natural phytonutrients of the flavanol family. The dietary supplement ingredient MASQUELIER's ® OPCs has been demonstrated to exhibit antioxidant, anti-inflammatory and vasculo-protective properties in humans, which suggested potential benefits as a topical formulation for skin health. Exposure of skin to UV triggers oxidative stress in skin tissue that can lead to erythema, skin aging or even cancer. Our previous studies have shown that a specific topical formulation (cream containing MASQUELIER's ® OPCs) can significantly reduce UV-induced erythema in skin of human subjects and in animals, indicating it has antioxidant activity in skin. Objective This study was undertaken to further establish antioxidant activity of the specific OPCs cream formulation in human skin by measuring ultra-weak photon emission (UPE) of skin. UPE measurement in skin is a non-invasive, fairly-sensitive and convenient technique for continuously monitoring oxidative stress (Sauermann, 1999). Methods The study was an open-label study in 25 healthy volunteers. Measurement of skin UPE was performed on the dorsal surface of the subjects’ hands before (spontaneous UPE) and after exposure to UV (UV-induced UPE). The effects of the OPCs cream on spontaneous and UV-induced UPE were measured using a fractionated UV exposure protocol. Results Within each subject, spontaneous UPE of the right and left hand were found to be significantly correlated ( r =0.9; p =0.000). UV-induced UPE of the right and left hand were also significantly correlated ( r =0.87; p =0.000). UV exposure resulted in a long-term (>80 min) increase of spontaneous UPE. This is likely due to depletion of antioxidant capacity of skin, which is quantifiable by the fractionated UV-exposure protocol. Application of the OPCs cream immediately after UV exposure resulted in a significant (approx. 30%) decrease in UV-induced UPE. Topical OPCs application also reduced UV-induced long-term increase in spontaneous UPE, indicating prevention depletion of antioxidant capacity of skin by UV. Conclusions This study indicates that the specific OPCs cream formulation significantly decreases UV-induced oxidative stress in human skin based on UPE measurement, thereby suggesting that regular use of this OPCs cream might protect skin from harmful effects of UV.

Protective effects of a cream containing Dead Sea minerals against UVB‐induced stress in human skin

Dead Sea (DS) mud and water are known for their unique composition of minerals, and for their therapeutic properties on psoriasis and other inflammatory skin diseases. Their mode of action, however, remains poorly known. To analyse the ability of Dermud, a leave-on skin preparation containing DS mud and other ingredients like DS water, zinc oxide, aloe-vera extract, pro-vitamin B5 and vitamin E, to antagonize biological effects induced by UVB irradiation in skin when topically applied in organ cultures. We have used human skin organ cultures as a model to assess the biological effects of UVB irradiation and of Dermud cream topical application. Skin pieces were analysed for mitochondrial activity by MTT assay, for apoptosis by caspase 3 assay, for cytokine secretion by solid phase ELISA, for overall antioxidant capacity by ferric reducing antioxidant power and Oxygen radical absorbance capacity assays (epidermis) or by cyclic voltammetry (external medium), and for uric acid (UA) content by HPLC. We report that UVB irradiation decreases cell viability, total antioxidant capacity and UA contents in the epidermis of skin organ cultures, while increasing the levels of apoptosis in cells and their cytokine secretion. Topical application of Dermud decreased all these effects significantly. Our results clearly show that Dermud has protective, anti-oxidant and anti-inflammatory properties that can antagonize biological effects of UVB irradiation in skin. It may therefore be able to reduce skin photodamage and photoaging, and more generally to reduce oxidative stress and inflammation in skin pathologies.

Dermal contributions to UVA‐induced oxidative stress in skin

When the skin is exposed to solar irradiation, UVA photons interact with skin tissues and induce excessive reactive oxygen species, resulting in oxidative stress. We have shown in a previous study that in vivo chemiluminescence's measurement can be used to evaluate the overall level of UVA-induced oxidative stress in human skin. However, the origin of the observed chemiluminescence signals remains unclear. UVA-induced chemiluminescence measurements were conducted: (a) in vitro on collagen solutions and solid collagen sheet preparations, (b) ex vivo on human and mouse skin biopsies, and (c) in vivo on human skin of various constitutive pigmentation levels. Fluorescence was measured on collagen in vitro as well as on skin for the in vivo experiments. We found in the in vitro experiments that UVA-induced chemiluminescence increases with the presence of collagen cross-links. When dermal sides were exposed to UVA irradiation, both mouse and human skin biopsies demonstrated significantly higher chemiluminescence levels than when epidermal sides were exposed to UVA. The amount of collagen cross-links decreases slightly following UVA exposure, as shown both by in vivo fluorescence and by UVA-induced chemiluminescence. Finally, there was less measurable UVA-induced chemiluminescence in dark skin compared with light pigmented skin in vivo. The dermis is very sensitive to UVA photons. Dermal cross-links are potential UVA sensitizers. The oxidative stress induced by UVA and measured by chemiluminescence may largely be attributed to the breakdown of dermal collagen cross-links.

Ferulic Acid Stabilizes a Solution of Vitamins C and E and Doubles its Photoprotection of Skin

Ferulic acid is a potent ubiquitous plant antioxidant. Its incorporation into a topical solution of 15%l-ascorbic acid and 1%alpha-tocopherol improved chemical stability of the vitamins (C+E) and doubled photoprotection to solar-simulated irradiation of skin from 4-fold to approximately 8-fold as measured by both erythema and sunburn cell formation. Inhibition of apoptosis was associated with reduced induction of caspase-3 and caspase-7. This antioxidant formulation efficiently reduced thymine dimer formation. This combination of pure natural low molecular weight antioxidants provides meaningful synergistic protection against oxidative stress in skin and should be useful for protection against photoaging and skin cancer.

Role of Phagocyte Oxidase in UVA-Induced Oxidative Stress and Apoptosis in Keratinocytes

Chronic exposure to ultraviolet radiation including ultraviolet A (315-400 nm) (UVA) may cause photocarcinogenesis and photoaging. The UVA-induced production of reactive oxygen species (ROS) and the resultant oxidative stress exposure play an important role in these biological processes. Here we have investigated the role of phagocyte oxidase (PHOX, gp91phox) in the production of ROS, redox status change, and apoptosis after UVA exposure by using gp91phox-deficient (gp91phox-/-) primary keratinocytes. UVA radiation resulted in increased ROS production and oxidation of reduced glutathione (GSH) to its oxidized form (GSSG). The presence of diphenylene iodonium (DPI) inhibited ROS production by UVA. In comparison with wild-type cells, gp91phox-/- cells produced slightly less ROS and GSH oxidation. UVA radiation induced apoptosis in wild-type keratinocytes as detected by phosphatidylserine (PS) translocation, caspase activation, and DNA fragmentation. As compared with wild-type cells, UVA induced less PS translocation in gp91phox-deficient cells. No difference, however, was observed in caspase activation and DNA fragmentation after UVA exposure in wild-type and gp91phox-/- cells. These findings suggest that gp91phox plays a limited role in the UVA-induced ROS production, oxidative stress, and therefore the PS translocation, but has no effect on UVA-induced caspase activation and DNA fragmentation during apoptosis.

Effects of oral vitamin E and β-carotene supplementation on ultraviolet radiation–induced oxidative stress in human skin

Ultraviolet radiation (UVR) generates reactive oxygen species in skin that can play a role in skin damage, but reports about the photoprotective properties of oral antioxidant supplements are conflicting. We examined the ability of 2 lipid-soluble antioxidants, vitamin E and beta-carotene, to reduce markers of oxidative stress and erythema in human skin exposed to UVR. Sixteen healthy subjects took either alpha-tocopherol (n = 8; 400 IU/d) or beta-carotene (n = 8; 15 mg/d) for 8 wk. Biopsy samples before and after supplementation were taken from unexposed skin and skin 6 h after 120 mJ/cm(2) UVR. The effects of supplements on markers of oxidative stress in skin and the minimal erythema dose to UVR were assessed. Supplementary vitamin E was bioavailable, the plasma concentration increased from 14.0 +/- 0.66 (x +/- SEM) to 18.2 +/- 0.64 mug/mL (P < 0.01), and the skin concentration increased from 0.55 +/- 0.09 to 1.6 +/- 0.19 ng/mg protein (P < 0.01). Supplementary beta-carotene increased plasma concentrations from 1 +/- 0.3 to 2.25 +/- 0.3 mug/mL (P < 0.05), but skin concentrations were undetectable. Before vitamin E supplementation, UVR increased the skin malondialdehyde concentration from 0.42 +/- 0.07 to 1.24 +/- 0.16 nmol/mg protein (P < 0.01), whereas oxidized or total glutathione increased from 9.98 +/- 0.4% to 12.0 +/- 1.0% (P < 0.05). Vitamin E supplementation significantly decreased the skin malondialdehyde concentration, but neither vitamin E nor beta-carotene significantly influenced other measures of oxidation in basal or UVR-exposed skin. Vitamin E or beta-carotene supplementation had no effect on skin sensitivity to UVR. Although vitamin E supplements significantly reduced the skin malondialdehyde concentration, neither supplement affected other measures of UVR-induced oxidative stress in human skin, which suggested no photoprotection of supplementation.

Topical Activity of Ascorbic Acid: From in vitro Optimization to in vivo Efficacy

We present here a new cosmetic formula system containing 3% ascorbic acid based on an optimized oil-in-water (O/W) emulsion. This formulation demonstrated a good long-term stability of the active ingredient and also of the emulsion itself. It could be deduced from in vitro release studies that this O/W emulsion enabled a better release of the hydrophilic active agent than an alternative W/O emulsion. By measuring the ultraweak photon emission, which is a well-established parameter for the oxidative stress in the skin, the high in vivo antioxidant capacity of 3% ascorbic acid was demonstrated after 1 week of product application. This placebo-controlled study also proved that ascorbic acid in an O/W cream reduced oxidative stress in human skin significantly better than the derivative sodium ascorbyl-2-phosphate, a more stable vitamin C replacement commonly used in cosmetic formulations. With increasing age, the number of papillae in the epidermal-dermal junction zone in human skin are reduced. This implies a possible consequence of reduced mechanical resistance of the skin and impaired supply of the epidermis with nutrients. In a 1-month placebo-controlled study on 25 human volunteers, a significant increase in the number of dermal papillae after application of the 3% ascorbic acid cream was demonstrated, using a confocal laser scanning microscope. Fine lines and wrinkles are a characteristic sign of aged and especially photo-aged skin. Application of 3% ascorbic acid in a 12-week placebo-controlled usage study indicated a significant reduction of facial wrinkles. Altogether, 3% ascorbic acid in a cosmetic O/W emulsion has been shown to be appropriately stable and to enable a good release of the active agent in vitro as a precondition for a high efficacy in vivo. Application in vivo resulted in a significant reduction of oxidative stress in the skin, an improvement of the epidermal-dermal microstructure and a reduction of fine lines and wrinkles in aged skin. These results were received within a relatively short period of time of product application.

A chemiluminescence study of UVA-induced oxidative stress in human skin in vivo.

Oxidative stress is defined as an imbalance between pro-oxidants and antioxidants in favor of pro-oxidants. Photon emission (also called chemiluminescence) has been widely used to study oxidative stress in biological systems in vitro. In vivo chemiluminescence has been proposed as a non-invasive method to assess oxidative stress in the skin. UVA (320-400 nm part of the ultraviolet radiation) exposure is generally accepted as a source of oxidative stress in the skin. In this study, UVA-induced oxidative stress was studied by using an in vivo chemiluminescence detection method. First, the dose response and the fluence rate response of the UVA-induced oxidative stress in human skin were investigated by examining the decay kinetics of the chemiluminescence signal following UVA exposure. A kinetic model was proposed to help differentiate these two responses. We found that the initial burst of the chemiluminescence signal depended on the UVA fluence rate, whereas the decay of the signal following exposure can be related to the UVA dose involved. Second, a significant reduction of UVA-induced chemiluminescence signal was observed after tape-stripping, indicating that stratum corneum is a major source of UVA-induced oxidative stress in the skin. Furthermore, the oxygen dependence of UVA-induced chemiluminescence signal was also confirmed by application of a pressure cuff, implying that some of the oxidative stress occurs in the deeper layers of the skin. Finally, topical application of vitamin C before exposure significantly reduced the UVA-induced chemiluminescence signal. We thus conclude that chemiluminescence is an effective method to assess the oxidative stress induced by UVA in human skin in vivo.

UVR-induced oxidative stress in human skin in vivo: effects of oral vitamin C supplementation.

Previous studies of cultured skin cells and murine skin in vivo have indicated that UVR-induced damage involves the generation of reactive oxygen species and depletion of endogenous antioxidant systems. In order to explore the relevance of this to UVR-induced damage to human skin, we have undertaken a detailed examination of the time-course of changes in markers of oxidative stress in human skin following exposure to physiological amounts of UVR in vivo. In addition, we have examined the skin bioavailability of a common nutritional antioxidant, vitamin C, and have assessed the effects of supplementation on markers of oxidative stress. Our hypothesis was that acute exposure of human skin to UVR in vivo would lead to oxidation of cellular biomolecules that could be prevented by prior vitamin C treatment. A UVR-challenge of 120 mJ/cm2 of broadband UVB (peak 310 nm, range 270-400 nm) was applied to buttock skin of 8 healthy volunteers. This caused a rapid and significant rise in activity of skin catalase at 1 h and an increase in the oxidized/total glutathione ratio at 6 h post-UVR. AP-1 DNA binding also peaked at 1-6 h post-UVR, then declined rapidly to baseline levels. No significant changes were seen in skin malonaldehyde content. Oral vitamin C supplements (500 mg/day) were taken by 12 volunteers for 8 weeks resulting in significant rises in plasma and skin vitamin C content. Supplementation had no effect on the UVR-induced erythemal response. The skin malonaldehyde content was reduced by vitamin C supplementation, but surprisingly, reductions in the skin content of total glutathione and protein thiols were also seen. We speculate that this apparently paradoxical effect could be due to regulation of total reductant capacity by skin cells, such that vitamin C may have been replacing other reductants in these cells. No evidence was obtained for an effect of the supplementary vitamin C on the mild oxidative stress seen in human skin following UVR exposure.