The effect of long-term cigarette smoking on selected skin barrier proteins and lipids.

The negative impact of cigarette smoking on the skin includes accelerated aging, pigmentation disorders, and impaired wound healing, but its effect on the skin barrier is not completely understood. Here, we studied the changes in selected epidermal proteins and lipids between smokers (45-66 years, smoking > 10 years, > 10 cigarettes per day) and non-smokers. Volar forearm epidermal and stratum corneum samples, obtained by suction blister and tape stripping, respectively, showed increased thickness in smokers. In the epidermis of smokers, we observed a significant upregulation of filaggrin, loricrin, and a trend of increased involucrin but no differences were found in the case of transglutaminase 1 and kallikrein-related peptidase 7, on the gene and protein levels. No significant changes were observed in the major skin barrier lipids, except for increased cholesterol sulfate in smokers. Liquid chromatography coupled with mass spectrometry revealed shorter acyl chains in ceramides, and an increased proportion of sphingosine and 6-hydroxysphingosine ceramides (with C4 trans-double bond) over dihydrosphingosine and phytosphingosine ceramides in smokers, suggesting altered desaturase 1 activity. Smokers had more ordered lipid chains found by infrared spectroscopy. In conclusion, cigarette smoking perturbs the homeostasis of the barrier proteins and lipids even at a site not directly exposed to smoke.

Common Cosmetic Compounds Can Reduce Air Pollution-Induced Oxidative Stress and Pro-Inflammatory Response in the Skin.

INTRODUCTION: Constantly increasing air pollution (AP) poses a concern affecting not only our health but also our skin. A typical manifestation of the skin damage induced by AP is its premature aging, irritation, skin barrier impairment, pigmentation disorders, and development or exacerbation of various skin diseases. For these reasons, it is crucial to protect the skin from the negative effects of AP. In this study, we evaluated the ability of some compounds commonly used in dermatological or cosmetic preparations with various biological activities to reduce AP-induced skin damage. METHODS: We established a new experimental model using porcine skin explants exposed to cigarette smoke (CS) in which we determined the level of reactive oxygen species (ROS) in the stratum corneum, skin barrier lipids peroxidation, and gene expression of the pro-inflammatory cytokine interleukin 6 in the epidermis. Then, we tested several polysaccharides and their derivatives such as sodium hyaluronate (SH) of different molecular weight (MW, 1.6 MDa, 300 kDa, 15 kDa, 5 kDa), yeast glucomannan, schizophyllan, and carboxymethyl ß-glucan, then vitamin C derivative sodium ascorbyl phosphate, niacinamide, and D-panthenol for their ability to prevent CS-induced skin damage. For the evaluation and comparison of their mechanism of action, film-forming effect was determined by TEWL and gloss measurements and the antioxidant properties were assessed by DPPH assay. RESULTS: In the skin samples exposed to CS, we observed significant negative changes such as the presence of large amount of ROS in the stratum corneum, high level of skin barrier lipids peroxidation and upregulated IL6 gene expression. Pretreatment of the skin samples with all the tested substances significantly prevented CS-induced skin damage. The most effective were high MW SH probably due to its best film-forming effect and sodium ascorbyl phosphate with the best antioxidant properties. CONCLUSION: AP leads to a significant skin damage which can be effectively prevented using some conventional cosmetic and dermatological ingredients with various mechanisms of action.

Seasonal variations in the skin parameters of Caucasian women from Central Europe.

BACKGROUND: The human skin is greatly affected by external factors such as UV radiation (UVR), ambient temperature (T), and air humidity. These factors oscillate during the year giving rise to the seasonal variations in the skin properties. The aim of this study was to evaluate the effect of seasons, environmental T, relative and absolute humidity on the skin parameters of Caucasian women, perform a literature review and discuss the possible factors lying behind the found changes. MATERIALS AND METHODS: We measured stratum corneum (SC) hydration, transepidermal water loss (TEWL), sebum level, erythema index, and elasticity parameters R2 and R7 on the forehead and the cheek of Caucasian women from the Czech Republic throughout the year. We also performed a non-systematic literature review focused on the seasonal variations in these skin parameters. RESULTS: We confirmed a well-documented low SC hydration and sebum production in winter. In spring, we found the lowest TEWL (on the forehead) and the highest SC hydration but also the highest erythema index and the lowest elasticity presumably indicating skin photodamage. For most of the skin parameters, the seasonal variations probably arise due to a complex action of different factors as we extensively discussed. CONCLUSION: The data about the seasonal variations in the skin parameters are still highly inconsistent and further studies are needed for better understanding of the normal skin changes throughout the year.

A comprehensive in vivo study of Caucasian facial skin parameters on 442 women.

Generally, skin properties are highly specific for each individual depending on various factors such as genetic predisposition, age, gender, body region, health and lifestyle. In this study, we measured various skin parameters on forehead, temple and cheek of 442 Caucasian women between 23 and 63 years, and evaluated differences between these facial regions and also the relationship between skin parameters and age of the volunteers. We measured transepidermal water loss (TEWL), stratum corneum hydration, skin gloss, melanin level, individual typology angle (ITA), erythema, sebum level and elasticity (R7). We observed significant negative relationship between age and TEWL, elasticity and skin lightness represented by ITA. Sebum, melanin and erythema levels increased up to the age of 50, when menopause usually takes place, and then decreased again. Evaluating the skin parameters on the forehead, temple and cheek area, we observed the biggest differences between the cheek and the forehead. The cheek possessed the worst skin parameters, such as the highest TEWL and erythema values and the lowest hydration.

The synthesis of a new unsaturated derivative of chondroitin sulfate with increased antioxidant properties.

Chondroitin sulfate (CS) was regio-specifically modified to an unsaturated derivative (ΔCS) with a double bond in positions 4 and 5 of N-acetyl-d-galactosamine. The structure of ΔCS was elucidated in detail by two dimensional nuclear magnetic resonance, ultraviolet spectroscopy and mass spectrometry. The introduction of a nucleophilic CC double bond into a polymer backbone had no influence on biocompatibility of CS, which was demonstrated by MTT live-dead assay and enzymatic degradation in vitro. On the other hand the chemical modification significantly enhanced the reactivity of ΔCS towards numerous oxidizing agents, which might be promising for a variety of biomedical and cosmetic applications.

Comparison of suction blistering and tape stripping for analysis of epidermal genes, proteins and lipids.

Analysis of epidermal genes, proteins and lipids is important in the research and diagnosis of skin diseases. Although punch biopsy is the first-choice technique for the skin sampling, it is unnecessarily invasive for obtaining a sample just for the epidermal analysis. Here we compare two less invasive methods, suction blistering (SB) and tape stripping (TS), for the analysis of selected epidermal genes (quantitative real-time reverse transcription PCR, qRT-PCR), proteins (western blotting, WB), and lipids in ten healthy volunteers. TS provided significantly less material than SB and no viable epidermal layers could be obtained according to the reflectance confocal microscopy. Consistently, only the SC protein filaggrin and housekeeping GAPDH together with FLG and RPL13A mRNA were detected by TS. In the SB samples, WB and qRT-PCR could easily detect all the selected proteins (claudin-1, occludin, filaggrin, laminin and GAPDH) and genes (CLDN1, OCLN, FLG, LAMA3 and RPL13A), respectively. A single SB sample further provided enough of material for immunohistochemistry and lipid analyses, which was not feasible with the TS samples. Immunohistochemistry of the SB samples showed intact epidermal structure and a characteristic expression of claudin-1. Infrared spectroscopy showed well-ordered lipids with both orthorhombic and hexagonal packing and high-performance thin layer chromatography confirmed all lipid classes (including ceramide subclasses) in correct proportions. Taken together, SB represents a reliable sampling technique that can be utilized for multipurpose epidermal analyses in various studies.

A new unsaturated derivative of hyaluronic acid - Synthesis, analysis and applications.

Hyaluronic acid (HA) containing CC double bond in positions 4 and 5 of N-acetyl-glucosamine ring (ΔHA) is an unique material, which could be used for biomedical applications and cosmetics. The main advantage of the CC double bond is its ability to react with a wide range of oxidation agents. Location of the CC double bond directly on the glucopyranose ring allows to change the chemical capabilities and simultaneously to mimic the intrinsic physical properties of HA without introduction of linkers or other substances. The synthesis, structural analysis and basic chemical and biological characteristics of this novel biopolymer are described in details. In vitro cytotoxicity assays showed selective activity of ΔHA against cancer cell lines in comparison with standard human fibroblasts so this material has great potential in the field of anticancer drugs.

Arbutin Content and Tyrosinase Activity of Bergenia Extracts.

The total arbutin content in the leaves of all the studied Bergenia plants (B. crassifolia, B. ciliata and B. x ornata) was determined. The highest values of the arbutin content have been established for B. crassifolia (58.9 ± 0.7 mg.g-¹ DW) and B. x ornata (51.0 ± 1.21 mg.g-¹ DW), and the lowest for B. ciliata (5.9 ± 0.6 mg.g-¹ DW). Arbutin concentration in the Bergenia leaves was the lowest in spring, in the autumn, on the contrary it increased. All the tested aqueous extracts caused a dose-dependent increase in diphenolase activity of fungal tyrosinase in a similar way as arbutin. On the other hand, all the ethanol extracts inhibited the diphenolase activity of tyrosinase.

Synthesis and biological evaluation of guanidino analogues of roscovitine.

A series of 2,9-substituted 6-guanidinopurines, structurally related to the cyclin-dependent kinase (CDK) inhibitors olomoucine and roscovitine, has been synthesized and characterized. A new copper-catalyzed method for the synthesis of 2-substituted 6-guanidino-9-isopropylpurines under mild reaction conditions has been developed. All prepared compounds were screened for their CDK1 and CDK2 inhibitory activities, cytotoxicity and antiproliferative effects in the breast cancer-derived cell line MCF7. The most active derivative 16g possessed an identical side chain in the C2 position to roscovitine; this compound displayed approximately five fold higher inhibitory activity towards CDK2/cyclin E and more than ten fold increase in cytotoxicity in MCF7 cells. Interestingly and in contrast to previously described findings, (S)-6-guanidinopurine derivatives were generally more active than their (R)-counterparts. Kinase selectivity profiling of (R)- and (S)-enantiomers 16e and 16g, respectively, revealed that introduction of a guanidino group at the C6 position of the purine moiety decreased selectivity towards protein kinases compared to roscovitine. Nevertheless, increased inhibitory activity and decreased selectivity offer a good starting point for further development of new protein kinase inhibitors.

Antiproliferative and antiangiogenic effects of flavone eupatorin, an active constituent of chloroform extract of Orthosiphon stamineus leaves.

Flavone eupatorin is one of the constituents of Orthosiphon stamineus, a medicinal herb used in folk medicine in South East Asia for treatment of various disorders. In our study, we investigated the antiproliferative properties of a chloroform extract of the leaves of O. stamineus and of pure eupatorin. The compound was able to reduce the number of viable cancer cells to the same extent as the extract, with IC(50) values in micromolar range. Moreover, both the eupatorin standard and the extract caused cells to arrest in the G2/M phase of the cell cycle. This clearly demonstrates that eupatorin contributes significantly to the overall extract activity. Induction of mitotic catastrophe, accompanied by key molecular events defining apoptosis, is the mechanism of eupatorin-induced cell death. Importantly, eupatorin (at the doses cytotoxic to cancer cells) did not kill normal cells; it only limited migration of HUVEC endothelial cells and their ability to create tubes. The ability of eupatorin to nonspecifically inhibit many protein kinases was proven and is the probable cause of its cellular effects. In summary, eupatorin emerges as a promising agent in anticancer research.

Oncogenes do not Fully Override Cell-intrinsic Traits: Pronounced Impact of the Cellular Programme.

Overexpression of p53 tumor suppressor protein in malignant cells induces cell cycle arrest, or alternatively, apoptosis thereby indicating that additional factors may contribute to the p53-mediated outcome. Comparison of the experimental protocols revealed that the construct encoding wild-type (wt) p53 was expressed in cells of different origin. Therefore, we decided to determine whether the intrinsic cellular program of primary cells of the same genetic background could have any effect on the oncogenic potential of mutated c-Ha-RAS and TP53. Primary rat cells (RECs) isolated from rat embryos of different age: at 13.5 gd (y) and 15.5 gd (o), were used for transfection. Immortalized rat cell clones overexpressing temperature-sensitive (ts) p53(135val) mutant and transformed cell clones after co-transfection with oncogenic c-Ha-Ras, were generated. The ts p53(135Val) mutant, switching between wt and mutant conformation, offers the possibility to study the role of p53 in cell cycle control in a model of malignant transformation in cells with the same genetic background. Surprisingly, the kinetics of cell proliferation at non-permissive temperature and that of cell cycle arrest at 32°C strongly differed between cell clones established from yRECs and oRECs. Furthermore, the kinetics of the re-enter of G1-arrested cells in the active cell cycle strongly differed between distinct cell clones. Finally, the susceptibility of immortalized and transformed cells to the pharmacological inhibitors of cyclin-dependent kinases (CDKs) considerably differed. Our results clearly show that overexpression of genes such as mutated TP53 and oncogenic c-Ha-RAS is not able to fully override the intrinsic cellular programme.