Protective Effect of Dermal Brimonidine Applications Against UV Radiation‐induced Skin Tumors,Epidermal Hyperplasia and Cell Proliferation in the Skin of Hairless Mice

Brimonidine at 0.18%, 1% and 2% concentrations applied topically in hairless mice significantly decreased tumor burden and incidences of erythema, flaking, wrinkling and skin thickening induced by UVR. The unbiased median week to tumor ≥1 mm was increased by the 1% and 2% concentrations. The tumor yield was reduced by all concentrations at week 40 for all tumor sizes but the ≥4 mm tumors with the 0.18% concentration. At week 52, the tumor yield was reduced for all tumor sizes and all brimonidine concentrations. The tumor incidence was reduced by all concentrations at week 40 for all tumor sizes, but the ≥4 mm tumor with the 0.18% concentration and at week 52 for all tumor sizes with the 1% and 2% concentrations and with the 0.18% concentration only for the ≥4 mm tumors. Reductions in ≥4 mm tumor incidences compared to the vehicle control group were 54%, 91% and 86% by week 52 for the 0.18%, 1% and 2% concentrations, respectively. Brimonidine at 2% applied 1 h before or just after UVB irradiation on hairless mice decreased epidermal hyperplasia by 23% and 32% and epithelial cell proliferation by 59% and 64%, respectively, similar to an epidermal growth factor receptor (EGFR) inhibitor.     

Effective Photoprotection of Human Skin against Infrared A Radiation by Topically Applied Antioxidants: Results from a Vehicle Controlled,Double‐Blind,Randomized Study

Infrared A radiation (IRA) from solar sunlight contributes to photoaging of human skin, e.g. by upregulating MMP‐1 expression in dermal fibroblasts, indicating the need for photoprotection of human skin against IRA. Up to now, however, there has been no controlled study to show that effective protection of human skin against IRA radiation is possible. Here, we have conducted a randomized, controlled, double‐blinded prospective study in 30 healthy volunteers to assess the capacity of an SPF 30 sunscreen versus the same sunscreen supplemented with an antioxidant cocktail containing grape seed extract, vitamin E, ubiquinone and vitamin C to protect human skin against IRA radiation‐induced MMP‐1 upregulation. As expected, exposure to IRA radiation significantly upregulated MMP‐1 expression, as compared to unirradiated skin, and this response was significantly reduced, if the SPF30 sunscreen plus the antioxidant cocktail had been applied prior to IRA radiation. In contrast, treatment of human skin with the SPF30 sunscreen alone did not provide significant protection. These results indicate that topically applied antioxidants effectively protect human skin against IRA radiation and that regular sunscreens need to be supplemented with specific antioxidants in order to achieve IRA photoprotection.     

Protective Effect of Magnesium-L-Ascorbyl-2 Phosphate Against Skin Damage Induced by UVB Irradiation

The protective effect of magnesium-L-ascorbyl-2-phosphate (MAP) on cutaneous photodamage such as lipid peroxidation and inflammation induced by ultraviolet B (UVB) exposure (290–320nm, max. 312 nm) was investigated using hairless mice. When MAP was administered intraperitoneally to mice at a dose of 100 mg of ascorbic acid (AS) per kg body weight base immediately before irradiation (15 kJ/m²), the expected increases in thiobarbituric acid reactive substance (TBARS) formation in skin and serum sialic acid, indices of lipid peroxidation and inflammatory reaction, respectively, were significantly reduced. However, the expected decrease in the level of cutaneous AS was unchanged. Similar results were observed for animals given 100 mg of AS-Na per kg body weight before UVB irradiation. When MAP was administered intracutaneously immediately before irradiation, the expected UVB-induced increases in TBARS and sialic acid were again significantly prevented. Ascorbic acid-Na had a less protective effect than intracutaneous MAP administration. The cutaneous AS level was significantly higher in the MAP-treated mice than in the controls, and the UVB-induced decrease in tissue AS was prevented by intracutaneous MAP administration. These results suggest that MAP protects against UVB irradiation-induced lipid peroxidation and inflammation in cutaneous tissue, regardless of the drug administration route. We found, in an in vitro experiment, that MAP was converted to AS as it crossed the epidermis, but that AS-Na did not pass through the epidermis. Furthermore, MAP was also converted to AS in serum. These results suggest that the protective effect of MAP on UVB-induced cutaneous damage is due to conversion of MAP to AS.     

The Protective Effect of Violaxanthin from Nannochloropsis oceanica against Ultraviolet B‐Induced Damage in Normal Human Dermal Fibroblasts

Skin photoaging, which is mainly induced by ultraviolet B (UVB) radiation, is prevented by the application of UV‐protective agents. The microalga Nannochloropsis oceanica (N. oceanica) has been primarily reported as a potential biofuel; however, in this study, we investigated whether N. oceanica extracts exerted photoprotective effects against UVB‐irradiated human dermal fibroblasts (HDFs) and which single component was responsible for the protective effect of the extracts. Two extracts—pigment and nonpigment—were prepared from N. oceanica biomass. WST‐1 assay and expression analysis of interleukin genes showed that the pigment extracts were not significantly cytotoxic to HDFs. Further experiments revealed that treatment with the pigment extract upregulated the expression of collagen genes and significantly blocked UVB‐induced damage such as decreased cell viability and increased ROS production. Next, to investigate the pigment composition of the extracts, HPLC analysis was conducted and violaxanthin was identified as the major pigment. The UVB photoprotective effect of the pigment extracts was confirmed in violaxanthin‐treated HDFs. In addition, violaxanthin significantly attenuated UVB‐induced G1 phase arrest, senescence‐associated β‐galactosidase activation, p16 and p21 upregulation, ERK phosphorylation and the downregulation of ECM molecules in HDFs. Therefore, we concluded that violaxanthin was a potential antiphotoaging agent.     

Photo‐co‐carcinogenesis of Topically Applied Retinyl Palmitate in SKH‐1 Hairless Mice

Cosmetic products that contain retinyl palmitate are popular as antiaging skin treatments; however, recent studies suggest a risk for enhanced skin tumor development with topical retinyl palmitate applications and exposure to solar ultraviolet radiation (UVR). In this study, we investigated the potential of retinyl palmitate to enhance UVR‐induced photo‐co‐carcinogenesis. Groups of 36 male and 36 female SKH‐1 hairless mice were exposed to simulated solar light (SSL) and treated with the control cream or creams containing retinyl palmitate, 5 days per week for 40 weeks. Other groups of mice were exposed to SSL and received no cream treatment or received cream treatments and were exposed to ultraviolet‐A or ultraviolet‐B. Mice were monitored for the development of skin tumors, and the incidences and multiplicities of squamous cell neoplasia were determined by histopathology. In both the absence and presence of SSL, mice administered the control cream developed skin tumors earlier and had higher incidences and multiplicities of skin squamous cell neoplasms than mice that received no cream treatment. Compared to the control cream groups, mice exposed to SSL and administered the retinyl palmitate creams demonstrated earlier onsets of skin tumors and had increased incidences and multiplicities of squamous cell skin neoplasms.     

Asiatic Acid Glucosamine Salt Alleviates Ultraviolet B-induced Photoaging of Human Dermal Fibroblasts and Nude Mouse Skin

Herbal extracts including asiatic acid (AA) have become popular candidates of anti-photoaging agents due to their anti-inflammatory and antioxidant properties and minimal side effect. Nevertheless, low bioavailability due to poor solubility limits their practical application. In this study, a highly bioavailable form of AA called AAGS (compounded by asiatic acid and glucosamine) was investigated for its anti-photoaging effect using both in vitro and in vivo models along with UVB irradiation. The results showed that AAGS alleviated UVB-induced cell proliferation inhibition by reducing G2 phase arrest and cell apoptosis rate as well as the gene expressions of P53, BAX, CASPASE 3 and CASPASE 9, but enhancing BCL-2 expression. It also reduced the production of reactive oxygen species along with increased gene expression of GPX-1 and downregulated the gene expression of IL-1β, IL-6, IL-8, IL-17 and TNF-α compared to nontreated cells. In vivo results demonstrated the antiphotodamaging effects by restoring skin thickness, collagen content and reducing MMPs expression, which are also supported by reduced MMPs gene expression and enhanced collagen I and TGF-β1 gene expression in vitro. Thus, AAGS may become a potential anti-photoaging agent for topical use due to its capability of self-assembling into a water gel.     

UVB‐Induced Inflammatory Cytokine Release,DNA Damage and Apoptosis of Human Oral Compared with Skin Tissue Equivalents

People can get oral cancers from UV (290–400 nm) exposures. Besides high outdoor UV exposures, high indoor UV exposures to oral tissues can occur when consumers use UV‐emitting tanning devices to either tan or whiten their teeth. We compared the carcinogenic risks of skin to oral tissue cells after UVB (290–320 nm) exposures using commercially available 3D‐engineered models for human skin (EpiDerm?), gingival (EpiGing?) and oral (EpiOral?) tissues. To compare the relative carcinogenic risks, we investigated the release of cytokines, initial DNA damage in the form of cyclobutane pyrimidine dimers (CPDs), repair of CPDs and apoptotic cell numbers. We measured cytokine release using cytometric beads with flow cytometry and previously developed a fluorescent immunohistochemical assay to quantify simultaneously CPD repair rates and apoptotic cell numbers. We found that interleukin‐8 (IL‐8) release and the initial CPDs are significantly higher, whereas the CPD repair rates and apoptotic cell numbers are significantly lower for oral compared with skin tissue cells. Thus, the increased release of the inflammatory cytokine IL‐8 along with inefficient CPD repair and decreased death rates for oral compared with skin tissue cells suggests that mutations are accumulating in the surviving population of oral cells increasing people's risks for getting oral cancers.     

The Dynamics of Laser-Induced Changes in Human Skin Autofluorescence—Experimental Measurements and Theoretical Modeling

To study the temporal dynamics of human skin autofluorescence photobleaching, we measured the autofluorescence spectral changes of skin in vivo during continuous exposure to 442 nm (He-Cd) laser light. Integral intensities were calculated for various spectral wavelength bands and plotted as a function of time. Mathematical analysis of the time function revealed a double-exponential photobleaching process: I(t) = a exp(-t/τ¹,) + b exp(-t/τ²) + c, in which t¹, and t² differed by an order of magnitude. A hypothesis for the mechanism of the double-exponential photobleaching dynamics was proposed and evaluated using Monte Carlo modeling of light propagation in the skin and autofluorescence escape from skin. By combining the fluorophore microdistributions, Monte Carlo simulation results and the variation in fluorescence decrease parameters (a, b, c, τ¹τ²) with increasing exposure intensities a biophysical explanation for the double-exponential photobleaching function was elucidated. The fast decrease term corresponds to laser-induced photobleaching in the stratum corneum, while the slow decrease term represents fluorophore changes in the dermis. The measured autofluorescence photobleaching dynamics can be used to determine the fractional contributions of different skin layers to the total autofluorescence signal measured in vivo.     

Protective Effect of Baicalin Against TLR4‐mediated UVA‐induced Skin Inflammation

UVA irradiation is known to cause photoaging via production of reactive oxygen species (ROS) and activation of inflammatory processes. Previously, we have demonstrated that baicalin, a plant‐derived flavonoid possessing both antioxidant and anti‐inflammatory activity, protects mouse keratinocytes against damage from UVB irradiation. However, the role of baicalin in vivo has not been well studied, particularly in the setting of UVA irradiation. To explore the protective effects and mechanisms of baicalin treatment in mice after UVA irradiation, mice were exposed to acute and chronic doses of UVA irradiation with or without baicalin or vehicle. Skin samples were collected for histological staining, RNA isolation, flow cytometry and protein extraction. Our results demonstrate the protective effect of baicalin against UVA‐induced oxidative damage and inflammation in mouse skin. These effects are likely mediated via the TLR4 pathway, which may serve as a target for photochemoprevention against skin inflammation.     

In Vitro Cytotoxic Protective Effect of Alginate-Encapsulated Capsaicin Might Improve Skin Side Effects Associated with the Topical Application of Capsaicin

Chronic neuropathic pain, particularly peripheral pain, is a cause of great concern for diabetic patients. Current treatments include numerous agents such as capsaicinoids, a known deterrent of neuropathic pain despite the inconvenience associated with local side effects. In this context, the current work aims to elucidate the potential mechanisms involved in cytotoxicity by capsaicin and proposes an efficient formulation of capsaicin in alginate microcapsules, which significantly reduces side effects from capsaicin topical administration. For this, human dermal fibroblast cells were treated with alginate-microencapsulated capsaicin extracts and screened for potential cytotoxic effects produced by the treatment. Cell viability and morphology were examined, as well as oxidative stress status and anti-inflammatory potential. Our results show that the alginate encapsulated formulation of capsaicin exerted lower cytotoxic effects on human dermal fibroblasts as measured by cell viability and reactive oxygen species (ROS) production. Furthermore, the expression profiles of inflammatory cytokines were significantly altered by the treatment as compared with the control culture.     

The Assessment of Meloxicam Phototoxicity in Human Normal Skin Cells: In Vitro Studies on Dermal Fibroblasts and Epidermal Melanocytes

Meloxicam (MLX), which belongs to the oxicam nonsteroidal anti-inflammatory drug derivatives, is an inhibitor of the cyclooxygenase-2 (COX-2) enzyme. Cutaneous adverse effects caused by interaction between UVA radiation and exogenous factors can manifest as phototoxic reactions. Phototoxicity may be a reason for the accumulation of genetic and molecular changes in long-lived cells with low proliferation potential, leading to tumor development. There are several potentially phototoxic drugs, the active component of which is meloxicam. The research aimed to evaluate the influence of MLX and UVAR on skin cells—fibroblasts and melanocytes homeostasis. The obtained results indicated that co-treatment with MLX and UVAR inhibited skin cell proliferation, proportionally to the drug concentration. The observation was confirmed by cytometric analysis of the cell number and viability. The phototoxic effect of MLX was revealed in morphological changes. It was stated that MLX with UVAR lowered the mitochondrial transmembrane potential and changed the cell cycle profile. Additionally, MLX and UVAR caused the disruption of redox homeostasis by lowering the intracellular level of reduced thiols. The presented study revealed that the phototoxic activity of MLX is associated with oxidative stress induction and disruptions in cell homeostasis. The differences in the phototoxic effects of MLX at the cellular level may be related to the different content of melanin pigments.     

Screening and Research on Skin Barrier Damage Protective Efficacy of Different Mannosylerythritol Lipids

Mannosylerythritol lipids (MELs) may prevent skin barrier damage, although their protective mechanisms and active monomeric constituents remain unclear. Here, three MELs were extracted from Candida antarctica cultures containing fermented olive oil then purified using silica gel-based column chromatography and semipreparative HPLC. All three compounds (MEL-A, MEL-B, MEL-C) were well separated and stable, and reliable materials were used for NMR and HRESIMS chemical structure determinations and for assessing MELs’ protective effects against skin damage. Notably, MEL-B and MEL-C effectively protected HaCaT cells from UVB-induced damage by upregulating the contents of filaggrin (FLG) and transglutaminase-1 (TGM1), as determined via ELISA. Moreover, MEL-B treatment (20 μg/mL) of UVB-irradiated HaCaT cells led to the upregulation of both the expression of mRNA genes and the key proteins FLG, LOR, and TGM1, which are known to be decreased in damaged skin cells. Additionally, histopathological analysis results revealed a markedly reduced intracellular vacuolation and cell damage, reflecting improved skin function after MEL-B treatment. Furthermore, immunofluorescence results revealed that MEL-B protected EpiKutis^® three-dimensional cultured human skin cells from sodium dodecyl sulfate-induced damage by up-regulating FLG, LOR, and TGM1 expression. Accordingly, MELs’ protection against skin barrier damage depended on MEL-B monomeric constituent activities, thus highlighting their promise as beneficial ingredients for use in skin-care products.     

Rational Design and Synthesis of Efficient Sunscreens To Boost the Solar Protection Factor

Skin cancer incidence has been increasing in the last decades, but most of the commercial formulations used as sunscreens are designed to protect only against solar erythema. Many of the active components present in sunscreens show critical weaknesses, such as low stability and toxicity. Thus, the development of more efficient components is an urgent health necessity and an attractive industrial target. We have rationally designed core moieties with increased photoprotective capacities and a new energy dissipation mechanism. Using these scaffolds, we have synthesized a series of compounds with tunable properties suitable for their use in sunscreens, and enhanced properties in terms of stability, light energy dissipation, and toxicity. Moreover, some representative compounds were included in final sunscreen formulations and a relevant solar protection factor boost was measured.     

Three-dimensional QSAR using the k-nearest neighbor method and its interpretation

In this paper we report a novel three-dimensional QSAR approach, kNN-MFA, developed based on principles of the k-nearest neighbor method combined with various variable selection procedures. The kNN-MFA approach was used to generate models for three different data sets and predict the activity of test molecules through each of these models. The three data sets used were the standard steroid benchmark, an antiinflammatory and an anticancerous data set. The study resulted in kNN-MFA models having better statistical parameters than the reported CoMFA models for all the three data sets. It was also found that stochastic methods generate better models resulting in more accurate predictions as compared to stepwise forward selection procedures. Thus, kNN-MFA method represents a good alternative to CoMFA-like methods.     

Protective Effect of Djulis (Chenopodium formosanum) Extract against UV- and AGEs-Induced Skin Aging via Alleviating Oxidative Stress and Collagen Degradation

Skin aging is a complex process involving photoaging and glycation stress, which share some fundamental pathways and have common mediators. They can cause skin damage and collagen degradation by inducing oxidative stress and the accumulation of reactive oxygen species (ROS). Chenopodium formosanum (CF), also known as Djulis, is a traditional cereal in Taiwan. This study investigated the protection mechanisms of CF extract against ultraviolet (UV) radiation and advanced glycation end products (AGEs)-induced stress. The results indicated that CF extract had strong antioxidant and free radical scavenging effects. It could reduce UV-induced intracellular ROS generation and initiate the antioxidant defense system by activating the nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) signaling pathway in human skin fibroblasts. CF extract modulated mitogen-activated protein kinase (MAPK) and transformed growth factor-beta (TGF-β) signaling pathways to alleviate oxidative stress-induced skin aging. Moreover, the results revealed that CF extract not only promoted collagen synthesis but also improved aging-induced collagen degradation. CF extract attenuated AGEs-induced ROS production and the upregulation of receptor for AGEs (RAGE). The overall results suggest that CF extract provides an effective anti-aging strategy by preventing skin damage from oxidative stress and collagen loss with potent antioxidant, anti-photoaging, and antiglycation activities.     

Peptide-derived from Seahorse Exerts a Protective Effect against Cholinergic Neuronal Death in in vitro Model of Alzheimer's Disease

Hippocampus trimaculatus are highly valued and the most heavily traded seahorse species for traditional medicine purposes in many Asian countries. One of the main interesting features in H.trimaculatus is their richness in protein which can be hydrolyzed into bioactive peptides. In the previous study, H. trimaculatus was hydrolyzed using commercial enzymes, the peptides responsible for neuroprotective activity was identified as HTP-1. In the present study, in vitro co-culture system of neuronal (PC12) cells and Aβ₄₂ oligomer-stimulated murine microglia cells (BV2 cells) were used to test neuroprotective effects of HTP-1. The co-culture system showed that HTP-1 protected PC12 cells from BV2 neurotoxic responses. In addition, the activation of PI3K/Akt signaling pathway by HTP-1 were confirmed. The PI3K/Akt activation was found to be mediated through transforming growth factor-β (TGF-β) induction by HTP-1. Furthermore, this signaling pathway was found to up-regulate the expression of pro survival protein expression in PC12 cells. Collectively, HTP-1 has potent protective effect against neuronal cells death in in vitro model of Alzheimer's disease.     

Synthesis and Utilization of Nitroalkyne Equivalents in Batch and Continuous Flow

We report a method for overcoming the low stability of nitroalkynes through the development of nitrated vinyl silyltriflate equivalents. Because of their instability, nitroalkynes have only rarely been utilized in synthesis. The reactivity of these silyltriflates, which are prepared in situ, is exemplified by dipolar cycloaddition reactions with nitrones to give highly substituted 4‐nitro‐4‐isoxazolines in high yields. This approach has proven general for several different alkyl and aryl substituted alkynes. In order to minimize the accumulation of potentially hazardous reaction intermediates, we have also developed a continuous flow variant of this method that is capable of carrying out the entire reaction sequence in a good yield and a short residence time.     

Noninvasive Assessment of UV-induced Skin Damage: Comparison of Optical Measurements to Histology and MMP Expression

Acute exposure to UV radiation (UVR) causes visible skin damage such as erythema and results in local and systemic immunosuppression while chronic exposure can result in photocarcinogenesis. These deleterious effects can be quantified by histology and by bioassays of key biological markers, including matrix metalloproteinases (MMPs), or tryptophan moieties. We now report our results in quantifying UV skin damage with noninvasive optical methods based on reflectance and fluorescence spectroscopy and compare these noninvasive measurements to histopathology and MMP-13 expression. A solar simulator with spectral output nearly identical to that of solar radiation was developed and used in our experiments. SKH1 hairless mice were exposed to solar-simulated UVR at a total dose of 21 MED delivered over 10 weeks. Changes in oxygenated and deoxygenated hemoglobin were measured by diffuse reflectance spectroscopy, and tryptophan changes were monitored via a fluorescence monitor. Our results show that there is an increase in erythema, skin fluorescence, sunburn cells and MMP-13 after a series of suberythemal doses of UV irradiation on a hairless mouse animal model. Increased skin fluorescence is observed with increasing UV exposure. The levels of MMP-13 increase as the cumulative UV dose increases but their increase does not correspond to noninvasively measured changes.     

Hydrolytic Methods for the Quantification of Fructose Equivalents in Herbaceous Biomass

A low, but significant, fraction of the carbohydrate portion of herbaceous biomass may be composed of fructose/fructosyl-containing components (“fructose equivalents”); such carbohydrates include sucrose, fructooligosaccharides, and fructans. Standard methods used for the quantification of structural-carbohydrate-derived neutral monosaccharide equivalents in biomass are not particularly well suited for the quantification of fructose equivalents due to the inherent instability of fructose in conditions commonly used for hemicellulose/cellulose hydrolysis (>80% degradation of fructose standards treated at 4% sulfuric acid, 121°C, 1 h). Alternative time, temperature, and acid concentration combinations for fructan hydrolysis were considered using model fructans (inulin, β-2,1, and levan, β-2,6) and a grass seed straw (tall fescue, Festuca arundinacea) as representative feedstocks. The instability of fructose, relative to glucose and xylose, at higher acid/temperature combinations is demonstrated, all rates of fructose degradation being acid and temperature dependent. Fructans are shown to be completely hydrolyzed at acid concentrations well below that used for the structural carbohydrates, as low as 0.2%, at 121°C for 1 h. Lower temperatures are also shown to be effective, with corresponding adjustments in acid concentration and time. Thus, fructans can be effectively hydrolyzed under conditions where fructose degradation is maintained below 10%. Hydrolysis of the β-2,1 fructans at temperatures ≥50°C, at all conditions consistent with complete hydrolysis, appears to generate difructose dianhydrides. These same compounds were not detected upon hydrolysis of levan, sucrose, or straw components. It is suggested that fructan hydrolysis conditions be chosen such that hydrolysis goes to completion; fructose degradation is minimized, and difructose dianhydride production is accounted for.     

Three-dimensional paper microfluidic devices assembled using the principles of origami

We report a method, based on the principles of origami (paper folding), for fabricating three-dimensional (3-D) paper microfluidic devices. The entire 3-D device is fabricated on a single sheet of flat paper in a single photolithographic step. It is assembled by simply folding the paper by hand. Following analysis, the device can be unfolded to reveal each layer. The applicability of the device to chemical analysis is demonstrated by colorimetric and fluorescence assays using multilayer microfluidic networks.