A decrease of mitochondrial ubiquitin ligase increases the secretion of matrix metalloproteinase-1 by dermal fibroblasts through the induction of ER stress.

BACKGROUND: We previously reported that the level of mitochondrial ubiquitin ligase (MITOL) protein in fibroblasts was decreased by UVA and that the knock-down (KD) of MITOL increased the secretion of matrix metalloprotease-1 (MMP-1) by fibroblasts. A recent study reported that MITOL suppresses endoplasmic reticulum (ER) stress by stabilizing the interaction between ER and mitochondria (MT) through the ubiquitination of mitofusin 2. These facts suggest that a decrease of MITOL would increase the secretion of MMP-1 through ER stress, but the detailed mechanism of that process in dermal fibroblasts remains unclear. Thus, this study was conducted to clarify the involvement of ER stress in the oversecretion of MMP-1 induced by the decreased MT quality caused by MITOL-KD. METHODS: MITOL-KD normal human dermal fibroblast (NHDFs) were prepared by treating them with MITOL-small interfering RNA, after which their MMP-1 protein levels were measured. ER stress in NHDFs was evaluated by measuring the mRNA levels of spliced X-box binding protein 1 (sXBP1) and the protein levels of inositol-requiring enzyme 1α (IRE1α). RESULTS: MITOL-KD NHDFs enhanced the secretion of MMP-1 via interleukin-6 (IL-6) elicited by the activation of nuclear factor-kappa B (NF-κB). The secretion of MMP-1 could be abrogated by a neutralizing IL-6 antibody and by JSH23, which is an inhibitor of NF-κB activation. Furthermore, MITOL-KD NHDFs as well as UVA-irradiated NHDFs showed increased ER stress levels. In addition, tunicamycin, which is an inducer of ER stress, also increased MMP-1 secretion. CONCLUSION: These results suggested that the decrease of MITOL caused the oversecretion of MMP-1 via NF-κB-IL-6 signaling through the activation of ER stress in fibroblasts.

Possible Involvement of Dermal Fibroblasts in Modulating Nrf2 Signaling in Epidermal Keratinocytes.

Epidermal keratinocytes protect themselves by cooperating with neighboring cells against internal and external stresses, which leads not only to the maintenance of cell homeostasis but also to the prevention of skin aging. Although it is known that nuclear factor (NF)-E2-related factor 2 (Nrf2) signaling plays a pivotal role in ameliorating oxidative stress and inflammatory responses under stress situations, it is unclear whether Nrf2 signaling in keratinocytes cooperates with neighboring cells such as dermal fibroblasts. Thus, this study was conducted to examine the influence of dermal fibroblasts on Nrf2 signaling in epidermal keratinocytes using a co-culture system. The results show that expression levels of Nrf2-regulated antioxidant factors, such as glutathione and heme oxygenase-1, in HaCaT keratinocytes (HaCaT KCs) are up-regulated in the presence of normal human dermal fibroblasts (NHDFs). In addition, the secretion of pro-inflammatory molecules, including interleukin-1α (IL-1α) and prostaglandin E2 (PGE2), is suppressed in co-cultures of NHDFs and UVB-irradiated HaCaT KCs. Interestingly, the localization of Nrf2 protein in HaCaT KCs was immediately translocated from the cytoplasm to the nucleus after the co-culture with NHDFs. These results suggest the possibility that Nrf2 signaling in keratinocytes is regulated in cooperation with dermal fibroblasts.

Decreased mitochondrial function in UVA-irradiated dermal fibroblasts causes the insufficient formation of type I collagen and fibrillin-1 fibers.

BACKGROUND: Decreases of collagen fibers and the disappearance of oxytalan fibers are typical symptoms of photoaged skin. Although a low quality of mitochondria (MT) in photoaged skin cells has been observed, it is unknown whether the decreased quality of MT is responsible for the insufficient formation of dermal fibers. OBJECTIVE: To identify the role of mitochondrial quality in skin photoaging focusing on the formation of dermal fibers. METHODS: Type I collagen and fibrillin-1 fibers in normal human dermal fibroblasts (NHDFs) were observed by immunostaining. Type I collagen and fibrillin-1 proteins in NHDFs were quantified by ELISA. Mitochondrial quality was evaluated by measuring levels of intracellular ATP and MITOL, which regulate mitochondrial quality. RESULTS: UVA-irradiated NHDFs formed insufficient type I collagen and fibrillin-1 fibers and had a decreased ratio of extracellular versus intracellular levels of those proteins. Although expression levels of motor proteins that transport those proteins intracellularly were not affected by UVA, intracellular ATP levels, which is the driving force of motor proteins, were decreased by UVA along with decreased MITOL protein. Knockdown of MITOL in NHDFs decreased the level of intracellular ATP and caused the insufficient formation of type I collagen and fibrillin-1 fibers due to interfering with the secretion of those proteins. CONCLUSION: These results indicate that a low quality of MT with ATP depletion in dermal fibroblasts caused by irradiation with UVA induces the insufficient formation of type I collagen and fibrillin-1 fibers due to the decreased extracellular secretion of those proteins.

Pyridoxine Has a Potential to Prevent the Appearance of Pigmented Spots: Effects on the Phagocytosis and Differentiation of Keratinocytes.

Pyridoxine (VB6) is a vitamin that is essential to maintain the homeostasis of the human body by contributing to various metabolic reactions. In the skin, although some studies have shown that VB6 is involved in regulating homeostasis through the attenuation of intracellular oxidative stress, there are few reports regarding the effects of VB6 on the prevention or improvement of skin aging. Thus, we conducted this study to determine the potential anti-skin pigmentation effect of VB6 focusing on the phagocytosis of melanosomes (MSs) by keratinocytes. The phagocytosis of MSs by keratinocytes is activated by oxidative stress and is an important factor of skin pigmentation and the eventual appearance of pigmented spots. First, we confirmed the antioxidant property of VB6 that enhanced the expression of several intracellular antioxidants via nuclear erythroid factor 2-related factor 2 (Nrf2). Although the incorporation of fluorescent beads (FBs), which are used as pseudo-MSs, into keratinocytes was increased under higher oxidation conditions caused by UVB and by the depletion of intracellular glutathione, treatment with VB6 suppressed the increased incorporation of FBs into those keratinocytes via Nrf2 activation. Furthermore, VB6 restored the decreased expression of differentiation marker proteins in keratinocytes caused by FB incorporation. Taken together, the results show that VB6 has the potential to prevent the appearance of pigmented spots by suppressing the activation of phagocytosis in keratinocytes caused by oxidative stress, and by restoring the differentiation of keratinocytes disrupted by FB incorporation.

Decreased levels of endocytic collagen receptor Endo180 in dermal fibroblasts lead to decreased production of type I collagen and increased expression of matrix metalloproteinase-1.

BACKGROUND: Endo180 is involved in collagen remodeling by incorporating extracellular degraded collagen. Ultraviolet irradiation of dermal fibroblasts reduces Endo180 expression, which affects collagen fiber remodeling. However, it is unclear whether the decrease in Endo180 is directly related to the decrease in type I collagen fibers during photoaging. We aimed to clarify the relationship between Endo180 reduction and the decrease in type I collagen fibers observed in photoaged dermis. METHODS: Endo180 was reduced in normal human dermal fibroblasts using RNAi. Endo180 knockdown cells were inoculated into collagen gels. The influence of Endo180 knockdown was evaluated by measuring mRNA expression of collagen fiber remodeling-related factors and collagen gel contraction. The collagen state and oxidative stress in the collagen gels were also measured. RESULTS: Endo180 knockdown cells, which were confirmed by gelatin uptake inhibition, showed upregulation of matrix metalloproteinase-1 and downregulation of type I collagen mRNA expression when cultured in collagen gels. The contractility of the collagen gel was reduced by Endo180 knockdown. The collagen state in the extracellular matrix of the collagen gels containing Endo180 knockdown fibroblasts showed increased amounts of 3/4 fragmented collagen and denatured collagen and decreased type I collagen synthesis. In addition, an increase in intracellular oxidative stress was observed. CONCLUSIONS: This study confirmed that the decrease in Endo180 caused a failure in collagen fiber formation and a decrease in collagen production, reproducing the photoaging dermal structural changes. This suggests that the decrease in Endo180 may be involved in wrinkle formation, which is a characteristic of photoaged skin.

Impact of protein carbonylation on the chemical characteristics of the hair surface.

OBJECTIVE: The purpose of this study was to clarify the impact of protein carbonylation on the chemical characteristics of the hair surface focusing on hydrophobicity. METHODS: First, we examined the validity of methods to evaluate hydrophobicity, one that utilizes the fluorescence of 1-anilinonaphtalene-8-sulfonic acid (1,8-ANS) compared with the contact angles against H2 O, of the hair surface chemically modified by alkaline hydrolysis or treated with stearyl ammonium chloride. We measured hairs bleached with H2 O2 or treated with acrolein for fluorescence originating from 1,8-ANS, for the contact angle and for changes of functional groups, aldehydes (the degree of carbonylation), NH2 , COOH and SH, using fluorescence labelling methods. RESULTS: The fluorescence intensity of 1,8-ANS of the hair surface modified chemically correlated well with the contact angles against H2 O. The results indicated that 1,8-ANS is suitable for evaluating the hydrophobicity of the hair surface. The hydrophobicity of hairs bleached with H2 O2 or carbonylated with acrolein was decreased. In addition, changes of functional groups in hairs carbonylated with acrolein increased as did those of hairs bleached with H2 O2 . CONCLUSION: The results suggest that the carbonylation of proteins at the hair surface with aldehydes decreases hydrophobicity and promotes further damage as does bleaching.

OBJECTIF: l'objectif de cette étude était de clarifier l'impact de la carbonylation des protéines sur les caractéristiques chimiques de la surface des cheveux en se concentrant sur l'hydrophobicité. MÉTHODES: nous avons d'abord examiné la validité de la méthode d'évaluation de l'hydrophobicité, une méthode qui utilise la fluorescence de l'acide 1-anilinonaphtalène-8-sulfonique (1,8-ANS) par rapport aux angles de contact avec l'H2O, de la surface des cheveux chimiquement modifiés par hydrolyse alcaline ou traités par chlorure d'ammonium stéarylique. Nous avons mesuré la fluorescence provenant du 1,8-ANS, l'angle de contact et les modifications des groupes fonctionnels, aldéhydes (le degré de carbonylation), NH2, COOH et SH des cheveux décolorés à l'H2O2 ou traités par acroléine, à l'aide de méthodes de marquage par fluorescence. RÉSULTATS: l'intensité de la fluorescence du 1,8-ANS de la surface des cheveux modifiés chimiquement était bien corrélée aux angles de contact avec l'H2O. Les résultats ont indiqué que le 1,8-ANS était adapté à l'évaluation de l'hydrophobicité de la surface des cheveux. L'hydrophobicité des cheveux décolorés à l'H2O2 ou carbonylés à l'acroléine a diminué. De plus, les modifications des groupes fonctionnels des cheveux carbonylés par l'acroléine ont augmenté, tout comme celles des cheveux décolorés à l'H2O2. CONCLUSION: les résultats suggèrent que la carbonylation des protéines à la surface des cheveux par des aldéhydes diminue l'hydrophobicité et favorise d'autres dommages, tout comme la décoloration.

Intracellular oxidative stress induced by calcium influx initiates the activation of phagocytosis in keratinocytes accumulating at S-phase of the cell cycle after UVB irradiation.

BACKGROUND: Phagocytosis is an essential process that maintains cellular homeostasis. In the epidermis, the phagocytosis of melanosomes into keratinocytes is important to protect their DNA against damage from ultraviolet B (UVB) radiation. Furthermore, it is considered that UVB activates the phagocytosis by keratinocytes but the detailed mechanism involved is not fully understood. OBJECTIVE: To clarify the mechanism of UVB-enhanced phagocytosis in keratinocytes, we investigated the relationship between the phagocytic ability of keratinocytes and the cell cycle stage of keratinocytes. METHODS: The phagocytic ability of keratinocytes was evaluated using the incorporation of fluorescent beads after exposure to UVB or oxidative stress. S-phase was evaluated by BrdU incorporation and immunostaining of cyclin D1. Intracellular calcium levels of keratinocytes were measured using the probe Fluo-4AM. RESULTS: The phagocytosis of fluorescent beads into keratinocytes was enhanced by UVB and also by oxidative stress. We found that keratinocytes exposed to UVB or oxidative stress were at S-phase of the cell cycle. Furthermore, keratinocytes synchronized to S-phase showed a higher phagocytic ability according to the increased intracellular ROS level. The UVB-enhanced phagocytosis and entrance into S-phase of keratinocytes was abolished by ascorbic acid, a typical antioxidant. Keratinocytes synchronized to S-phase and exposed to UVB or oxidative stress had increased levels of intracellular calcium and their enhanced phagocytic abilities were diminished by the calcium ion chelator BAPTA-AM. CONCLUSION: Taken together, intracellular oxidative stress induced by intracellular calcium influx mediates the UVB-enhanced phagocytic ability of keratinocytes accumulating at S-phase of the cell cycle.

Xanthophyll Carotenoids Reduce the Dysfunction of Dermal Fibroblasts to Reconstruct the Dermal Matrix Damaged by Carbonylated Proteins.

Although extracellular carbonylated proteins (CPs) are found at higher levels in sun-exposed skin, their impact on the cellular functions of fibroblasts and their involvement in the progression of photoaging skin are not fully clarified. In our previous study, we reported that extracellular CPs increase levels of intracellular oxidative stress and result in the accumulation of newly synthesized CPs in normal human dermal fibroblasts (NHDF). Furthermore, fibroblasts exposed to CP-BSA, which is a model of extracellular CPs, had upregulated expression levels of mRNAs encoding matrix metalloproteinase-1 (MMP-1) and interleukin-8/CXCL8 (IL-8/CXCL8). These facts suggested the possibility that extracellular CPs induce a fragile structure in the dermis through the degradation of collagen and elastin. The purpose of this study was to characterize the efficacy of natural carotenoids, such as astaxanthin analogs, produced by Hematococus pluvialis (CHPs) to improve the impaired functions of fibroblasts exposed to CPs. CHPs suppressed the intracellular CP levels elevated by CP-BSA, restored mRNA expression levels of factors involved in the formation and assembly of collagen and elastin fibers and improved the formation of those fibers impaired by CP-BSA. We conclude that CHPs function as antiaging substances due to their restoration of the impaired formation of collagen and elastin fibers caused by extracellular soluble CPs.

Oil Thickening with Organoclay Enhances the Ultraviolet Absorption Ability of Sunscreen on a Skin-mimicking Substrate.

The performance of sunscreen products depends on their ultraviolet (UV) absorption ability through the film formed on the skin surface upon their application. Therefore, it is important that a uniform film is formed on the uneven skin surface for effective sunscreen performance. Because most UV filters are oil soluble, we hypothesized in this study that increasing the viscosity of the oil phase of a sunscreen product can improve the performance of the sunscreen. We first examined the association between the concentration of the oil thickener and the UV absorption ability of the sunscreen product using a skin-mimicking substrate (SMS). Among all thickeners examined (petrolatum, dextrin palmitate, silica silylate, and organoclay), organoclay and silica silylate significantly increased the UV absorbance of sunscreen on the SMS in a concentration-dependent manner. Thereafter, we examined film uniformity to elucidate the mechanism underlying the observed increase in UV absorption. The uniformity of film thickness on the SMS increased with increasing organoclay content, based on decreased standard deviations of film thickness. Our results showed that increasing the viscosity of the oil phase with organoclay resulted in the formation of a uniform film by preventing the sunscreen from flowing into the grooves when applied on the SMS, thereby increasing UV absorbance by more than two-fold that of sunscreen without organoclay. Thus, the use of thickeners, such as organoclay, increases the viscosity of the oil phase at a low shear rate after the high shear of application. This is an effective strategy for improving the overall quality and performance of sunscreen products.

Tranexamic Acid Improves the Disrupted Formation of Collagen and Fibrillin-1 Fibers Produced by Fibroblasts Repetitively Irradiated with UVA.

The dermis is mainly constructed by type I collagen fibers, which provide mechanical strength to the skin by building a frame-like structure, and by elastic fibers, which provide elasticity to respond to movements of the skin. The depletion of collagen fibers and the disappearance of oxytalan fibers, which are a type of elastic fiber, are characteristic changes in photoaged skin. Prostaglandin E2 (PGE2) is one of the chemical mediators involved in inflammation and is responsible for sunburn. Furthermore, it has been reported that PGE2 attenuates the production of collagen and the expression of elastic fiber-related factors in fibroblasts. Tranexamic acid (TXA), which is an anti-inflammatory medicine that inhibits plasmin, reduces the level of PGE2 secreted following UV exposure or after inflammatory stimulation. However, few reports have verified TXA as an anti-skin aging agent. In this study, we examined the potential of TXA as an anti-skin aging agent using repetitively UVA-irradiated fibroblasts as a model for fibroblasts located in chronically sun-exposed dermis. Repetitively UVA-irradiated fibroblasts had higher secretion levels of PGE2. In addition, fibroblasts repetitively irradiated with UVA or treated with PGE2 produced disrupted collagen and fibrillin-1 fibers. Treatment with TXA improved the formation of both types of fibers by repetitively UVA-irradiated fibroblasts by restoring the expression of fiber-related proteins at the mRNA and protein levels. Thus, these results demonstrate that TXA has potential as an anti-photoaging agent.

Safety and Efficacy of Oral Intake of Ceramide-Containing Acetic Acid Bacteria for Improving the Stratum Corneum Hydration: A Randomized, Double-Blind, Placebo-Controlled Study over 12 Weeks.

The barrier function of the skin protects it from external stresses to which it is constantly exposed, such as dryness, ultraviolet rays, and chemicals. Lipids, in particular a type of sphingolipid known as ceramides, play a central role in the barrier function of the skin by preventing dryness. The number of ceramides in the skin is known to decrease with age, which has led to the development of a large number of anti-aging cosmetic products that contain ceramides. Recently, it has become evident that oral intake of ceramides can also improve the quality of the skin. To elucidate the effects of oral ceramide intake on skin moisture content, we conducted a randomized, double-blinded, placebo-controlled parallel comparative study in which males and females between 20 and 60 years of age who were worried about dry skin ingested a food with acetic acid bacteria containing 0.8 mg of dihydroceramide or a placebo for 12 weeks. Concurrently, we investigated the safety of continuous ingestion of the ceramide-containing food over 12 weeks. Oral intake of ceramide over the 12 weeks significantly improved stratum corneum hydration, i.e. the moisture content of the skin, and did not result in harmful effects in any of the participants.

An Ocimum basilicum Extract Containing Rosmarinic Acid Restores the Disruption of Collagen Fibers Caused by Repetitive UVA Irradiation of Dermal Fibroblasts.

Photoaged skin is characterized by the appearance of pigmented spots such as solar lentigos, deep wrinkles and sags, and progresses due to chronic sun exposure. Among the wavelengths of sunlight, UVA is responsible for the appearance of wrinkles and sags that originate from structural alterations in the dermis of photoaged skin such as the depletion of collagen fibers. Thus, improving and restoring collagen fibers is an effective approach to reduce skin photoaging and maintain a youthful appearance. This study was conducted to evaluate the potential of an extract of Ocimum basilicum (OC), which contains rosmarinic acid (RA), as an anti-photoaging material focusing on the capacity to restore collagen fibers that are disrupted due to intracellular oxidative stress. In spite of their relatively low capacities for chemical scavenging of reactive oxygen species (ROS), both OC and RA showed efficient removal of biological oxidative stress by reducing levels of intracellular ROS and carbonylated proteins (CPs) in fibroblasts following exposure to single or repetitive UVA irradiations. Fibroblasts irradiated with repetitive UVA as a model for chronic sun-exposed cells showed significant increases in matrix metalloproteinase-1 and decreases in type I collagen synthesis and formed reduced numbers of collagen fibers. Since both OC and RA restored the adverse phenomena caused by repetitive UVA irradiation, we conclude that OC containing RA is an effective anti-photoaging material.

2-Hydroxy-4-methoxybenzophenone Enhances the Suppression of Superoxide Anion Radicals Generated via UVA-induced Photosensitizing by t-Butyl Methoxydibenzoylmethane.

4-tert-Butyl-4'-methoxydibenzoylmethane (BMDM) is widely used throughout the world as a highly effective UVA absorber that can prevent the progression of photoaging in skin. However, due to its low photostability, BMDM is also known for the disadvantage of having a reduced capability to absorb UVA during prolonged exposure to sunlight. Although many studies have been carried out to overcome this disadvantage of BMDM, little attention has been paid to how the radicals generated from BMDM during UV exposure influence the skin. Therefore, the purpose of this study was twofold: One goal was to clarify the influence of radicals on human skin using cytotoxicity as a parameter. The second was to propose a solution that could reduce the radical formation while taking photostability into consideration. Using ESR spin trapping and superoxide dismutase (SOD) treatment, the radicals produced by the UV exposure of BMDM were shown to be superoxide anion radicals (•O2-). HaCaT keratinocytes exposed to UVA in the presence of BMDM showed a significant reduction in cell viability, indicating that the radicals produced from BMDM have a harmful influence on the skin. UVA exposure coincidently led to a reduction of UVA absorbance by BMDM. Interestingly, 2-hydroxy-4-methoxybenzophenone (Benzophenone-3; BP3) reduced both the radical formation and the cytotoxicity resulting from the UVA-exposure of BMDM, while also restoring its UVA absorbance. In conclusion, the results show that BMDM and BP3 is an effective combination to reduce the influence of UVA-exposed BMDM on the skin and to prevent the loss of UVA absorbance by BMDM during UV exposure.

An Extract of Young Olive Fruit Residues Attenuates Oxidative Stress in HaCaT Keratinocytes through the Ativation of Nrf2 Signaling.

Residues of olive fruit (ROF) after the extraction of oils are an increasing source of industrial waste, because olive oil is becoming more popular as a healthy food. It has been reported that olives have some polyphenols that have an antioxidation capability. On the other hand, excess oxidative stress disrupts epidermal barrier function. This study was conducted to determine whether ROF could be utilized as an antioxidant source to reduce industrial wastes and to identify possible active materials to maintain healthy skin. Olive fruits are categorized into two groups depending on the time of harvest, young fruit (YF) and mature fruit (MF). Thus, we examined the antioxidant potentials of extracts from YF and from MF to remove reactive oxygen species (ROS) from biological and chemical aspects. HaCaT keratinocytes cultured with extracts of YF or MF had reduced levels of intracellular ROS in spite of the relatively low chemical capability against ROS scavenging. The biological effects of the YF extract were superior to those of the MF extract. The YF extract showed effective reductions of intracellular ROS and carbonylated proteins that were elevated by the stress-related hormone cortisol. In addition, the YF extract reinforced the intracellular antioxidation capability through the activation of Nrf2 signaling. Taken together, the YF extract was an effective source to reinforce the intracellular antioxidation capability. We conclude from these results that utilizing ROF would lead to the reduction of industrial wastes and would supply active materials to maintain healthy skin.

Impairment of the autophagy system in repetitively UVA-irradiated fibroblasts.

BACKGROUND: Autophagy is known as an intracellular cleanup system necessary to maintain homeostasis of the skin. Many studies have pointed out the relationship between aging and the inactivation of autophagy function, which suggests that the inactivation of autophagy occurs in aged skin. However, the aging process of the skin is complicated compared with other organs, because the skin is localized at the border between the inside of the body and the environment. Thus, skin aging is strongly affected by environmental factors, and it is well recognized that ultraviolet (UV) radiation is an important environmental factor that promotes skin aging. Therefore, characterizing the autophagic phenotypes induced by environmental factors is important to understand the process of skin aging. METHODS: In order to demonstrate the status of autophagy during environment-induced aging of the skin, we investigated the autophagy profiles of normal human dermal fibroblasts (NHDFs) treated with repetitive UVA irradiation as model fibroblasts in photoaged skin. RESULTS: Repetitively UVA-irradiated NHDFs showed increased numbers of autophagosomes, which coincided with the accumulation of p62 and increased levels of LAMP-1 and lysosomes. The behavior of repetitively UVA-irradiated NHDFs on autophagy was similar to that of NHDFs treated with hydroxychloroquine (HCQ), which is an inhibitor of lysosomal proteinase. CONCLUSION: In summary, these results demonstrate that repetitively UVA-irradiated fibroblasts have reduced autophagy function due to the dysfunction of lysosomes.

Interleukin-1 alpha derived from ultraviolet B-exposed keratinocytes is associated with a decrease of endocytic collagen receptor Endo180.

BACKGROUND: Endo180 contributes to the remodeling of the collagen fibers that comprise the dermal matrix due to the internalization of extracellular collagen fragments. In the sun-exposed elder skin, an accumulation of collagen fragments was observed in the dermal matrix which was associated with a reduction in Endo180 in the dermal fibroblasts. This suggests that the loss of Endo180 results in the accumulation of collagen fragments in the surrounding fibroblasts and causes interference with dermal matrix remodeling via collagen fibers. The purpose of the study was to identify a mechanism by which ultraviolet B (UVB) exposure induces a loss of Endo 180 with a specific focus on the crosstalk between keratinocytes and fibroblasts. METHODS: Endo180 from normal human dermal fibroblasts, which were cultured with a conditioned medium (CM) of UVB-exposed keratinocytes, was examined using mRNA expression, protein levels and collagen internalization by quantitative RT-PCR, ELISA, and flow cytometry, respectively. RESULTS: Although UVB irradiation to fibroblasts failed to reduce Endo180, the CM of UVB-exposed keratinocytes reduced Endo180 in the fibroblasts. Collagen internalization into the fibroblasts was decreased and was associated with a loss of Endo180. Among cytokines secreted from UVB-exposed keratinocytes, IL-1α solely reduced Endo180, and the reduction induced by the CM of UVB-exposed keratinocytes was abolished by the presence of IL-1RA. CONCLUSIONS: These results indicate that a substance secreted from UVB-exposed keratinocytes regulates Endo180 expression and that IL-1α may play an important role in the maintenance of Endo180.

A Potato Peel Extract Stimulates Type I Collagen Synthesis via Akt and ERK Signaling in Normal Human Dermal Fibroblasts.

The ability of dermal fibroblasts to synthesize collagen decreases with ages. The integrity of collagen fibers severely decreases in aged skin, causing its characteristic morphological changes such as wrinkles and sagging. To prevent and improve skin aging, the stimulation of collagen synthesis in dermal fibroblasts is important. Potato peels contain many biofunctional compounds, but not much is known about their effects on human skin physiology. To characterize the potential effects of a potato peel extract (PPE) against skin aging, we examined its effects on the synthesis of type I collagen by normal human dermal fibroblasts (NHDFs). Treatment with the PPE significantly increased the expression of type I collagen mRNA in NHDFs and their secretion of type I collagen. To elucidate the mechanism involved, we examined the signaling pathway controlled by transforming growth factor-ß (TGF-ß), which regulates the synthesis of type I collagen. Treatment of NHDFs with the PPE significantly increased the expression of TGF-ß receptor mRNA. TGF-ß signaling involves Smad-dependent and Smad-independent pathways, like phosphatidylinositol-3 kinase (PI3K)/Akt and mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK). The PPE did not activate Smad, but significantly activated Akt and ERK. These results demonstrate that the PPE activates PI3K/Akt and MAPK/ERK signals via TGF-ß receptors, which stimulate the synthesis of type I collagen in NHDFs. These results suggest that the PPE could be a novel and effective antiaging material.

Cecropia obtusa extract and chlorogenic acid exhibit anti aging effect in human fibroblasts and keratinocytes cells exposed to UV radiation.

Cecropia obtusa is popularly used in the Amazonian region and exhibits antioxidant activity. Cosmetic formulations containing C. obtusa extract are commercially available for purchase; however, the chemical composition and the effects of the topical application of the extract are not described in the literature. Therefore, this study aimed to identify the main components of C. obtusa for the first time and to assess the anti aging effect in human fibroblasts and keratinocytes exposed to UVR. The main components in C. obtusa extract were identified by LC-DAD-MS/MS as chlorogenic acid (CGA), luteolin-C-hexoside, luteolin-C-hexose-O-deoxy-hexose, and apigenin-C-hexose-O-deoxy-hexose. C. obtusa extract and CGA decreased the metalloproteinase-1 and protein carbonyl levels and increased the collagen and hyaluronic acid contents. Overall, the extract exhibited better activity than CGA, and we demonstrated the ability of the extract to protect against the UV-induced increase in the pro inflammatory cytokines IL-1ß and IL-6, which are potential pathways of the antioxidant and anti aging effect. The chemical characterization added important data to broaden the knowledge related to C. obtusa, and the results suggest that the extract is a promising candidate to be incorporated in topical photochemoprotective formulations.

[Function of Sacran as an Artificial Skin Barrier and the Development of Skincare Products].

Sacran, a large molecular-weight polysaccharide isolated from algae, is composed of 11 types of saccharides, including sulfate and carboxylic acid groups. Because of its unique structure, sacran can form a gel-like sheet in the presence of polyols such as 1,3-butanediol. In addition, those sacran gel-like sheets prevent the evaporation of water and the penetration of chemicals. The results of our previous study suggested that sacran can work as an artificial barrier against external stimuli such as air pollutants which increase the stress on humans. Topically applied sacran was localized at the surface of reconstructed human epidermal equivalents. Those results suggested that sacran inhibits excessive water evaporation from the skin and protects against environmental stimuli by forming an artificial barrier at the skin surface. Then, in a clinical study, we examined the activity of sacran in improving skin problems caused by an impaired epidermal barrier. First, we conducted a use test on a serum formulated with sacran on human volunteers who had impaired skin barrier function. The results showed that sacran provided excellent benefits to improve the maturation of corneocytes. These results suggest that sacran could play an important role in providing optimal skin conditions for keratinocytes to progress through their differentiation.

Protective Effects of Sacran, a Natural Polysaccharide, Against Adverse Effects on the Skin Induced by Tobacco Smoke.

Recent increases in air pollution have raised concerns about its adverse effects on human health. Sacran is a natural polysaccharide isolated from a cyanobacterium. We previously reported that sacran improves skin conditions because of its effects as an artificial barrier against external stimuli, which suggested that sacran might protect the skin against air pollutants. The goal of this study was to characterize the potential of sacran to protect human skin against damage from air pollutants and to compare sacran with hyaluronic acid (HA). Sacran that was topically applied on the skin stayed on the surface or in the stratum corneum. Sacran-treated filters had a shielding effect against benzo[a]pyrene (BaP) and aldehyde compounds contained in tobacco smoke. Sacran suppressed the upregulation of cytochrome P4501A1 messenger ribonucleic acid (mRNA), which is a xenobiotic-metabolizing enzyme induced by BaP, and other responses against tobacco smoke in HaCaT keratinocytes. Furthermore, topical application of a serum containing 0.04% sacran on the skin reduced levels of carbonylated proteins in corneocytes of tobacco smokers. Sacran showed superior effects in every characteristic measured, compared with HA. We conclude that sacran ameliorates the oxidative stress initiated by tobacco smoke by shielding the skin surface and protects human skin.