Urban particulate matter in air pollution penetrates into the barrier-disrupted skin and produces ROS-dependent cutaneous inflammatory response in vivo.

BACKGROUND: Particulate matter (PM) is an integral part of air pollution, which is a mixture of particles suspended in the air. Recently, it has been reported that PM is associated with increased risks of skin diseases, especially atopic dermatitis in children. However, it is unclear if PM directly goes into the skin and what mechanisms are involved in response to PM. OBJECTIVE: To see whether PM could penetrate into the barrier-disrupted skin, produce reactive oxygen species (ROS), and elicit an inflammatory response. METHODS: We collected PMs during a winter in Seoul and used cultured keratinocytes for in vitro study and tape-stripped BALB/c mice for in vivo study. RESULTS: Keratinocyte cytotoxicity increased in a dose-dependent manner by PM treatment. IL-8 and MMP-1 mRNA expression and protein levels were significantly increased compared to control by qPCR and ELISA, respectively. Cellular ROS production was increased by PM treatment, and antioxidant N-acetyl cysteine pretreatment prevented induction of inflammatory cytokines IL-8 and MMP-1. In PM-treated keratinocytes, electron-dense subcellular particles were observed by transmission electron microscopy. PM was observed inside hair follicles in both intact and barrier-disrupted skin in vivo. Additionally, intercellular penetration of PM was seen in the barrier-disrupted skin. Repeated PM application induced epidermal thickening and dermal inflammation with neutrophil infiltration. Finally, N-acetyl cysteine could ameliorate skin inflammation induced by PM application. CONCLUSION: PM penetrates into the barrier-disrupted skin, causing inflammation, demonstrating detrimental effects in the skin.

Activation of Peroxisome Proliferator-Activated Receptor Alpha Improves Aged and UV-Irradiated Skin by Catalase Induction.

Peroxisome proliferator-activated receptor alpha (PPARα) is a nuclear hormone receptor involved in the transcriptional regulation of lipid metabolism, fatty acid oxidation, and glucose homeostasis. Its activation stimulates antioxidant enzymes such as catalase, whose expression is decreased in aged human skin. Here we investigated the expression of PPARα in aged and ultraviolet (UV)-irradiated skin, and whether PPARα activation can modulate expressions of matrix metalloproteinase (MMP)-1 and procollagen through catalase regulation. We found that PPARα mRNA level was significantly decreased in intrinsically aged and photoaged human skin as well as in UV-irradiated skin. A PPARα activator, Wy14643, inhibited UV-induced increase of MMP-1 and decrease of procollagen expression and caused marked increase in catalase expression. Furthermore, production of reactive oxygen species (ROS) was suppressed by Wy14643 in UV-irradiated and aged dermal fibroblasts, suggesting that the PPARα activation-induced upregulation of catalase leads to scavenging of ROS produced due to UV irradiation or aging. PPARα knockdown decreased catalase expression and abolished the beneficial effects of Wy14643. Topical application of Wy14643 on hairless mice restored catalase activity and prevented MMP-13 and inflammatory responses in skin. Our findings indicate that PPARα activation triggers catalase expression and ROS scavenging, thereby protecting skin from UV-induced damage and intrinsic aging.

Toll-like receptor 2 mediates a cutaneous reaction induced by repetitive ultraviolet B irradiation in C57/BL6 mice in vivo.

Toll-like receptors (TLRs) mediate not only innate immunity against infection and but also sterile inflammation triggered by endogenous molecules. We conducted a comparative study of the different inflammatory responses induced by repetitive ultraviolet (UV) B irradiation in wild-type (WT) and TLR2 knockout (KO) mice, to provide in vivo evidence of the role of TLRs in mediating UVB-induced responses. UVB-induced inflammatory responses were less severe in TLR2 KO mice than in WT mice after 6 weeks of repeated UVB irradiation. UVB-treated TLR2 KO mice displayed less prominent erythema and scaling, and histopathology showed significantly thinner skin and less inflammatory cell infiltration than that in WT mice. UVB-induced expression of heat-shock protein 70 (an endogenous ligand of TLR2) was lower in TLR2 KO mice. Quantitative RT-PCR revealed significantly lower gene expression levels of UVB-induced interleukin (IL)-1ß, IL-6 and matrix metalloproteinase (MMP)-13 in TLR2 KO mice. TLR2 KO mice also showed significantly lower protein level expression of UVB-induced IL-1ß in ELISA and MMP-13 in Western blots. Our study demonstrated that TLR2 was associated with inflammatory responses to repetitive UVB irradiation in C57/BL6 mice. Moreover, it suggests that the role of TLR2 in the cutaneous response of UV irradiation and in developing new agents for modulating the effects of UV irradiation should be considered.

Supplementating with dietary astaxanthin combined with collagen hydrolysate improves facial elasticity and decreases matrix metalloproteinase-1 and -12 expression: a comparative study with placebo.

Photoaging accounts for most age-related changes in skin appearance. It has been suggested that both astaxanthin, a potent antioxidant, and collagen hydrolysate can be used as antiaging modalities in photoaged skin. However, there is no clinical study using astaxanthin combined with collagen hydrolysate. We investigated the effects of using a combination of dietary astaxanthin and collagen hydrolysate supplementation on moderately photoaged skin in humans. A total of 44 healthy subjects were recruited and treated with astaxanthin (2 mg/day) combined with collagen hydrolysate (3 g/day) or placebos, which were identical in appearance and taste to the active supplementation for 12 weeks. The elasticity and hydration properties of facial skin were evaluated using noninvasive objective devices. In addition, we also evaluated the expression of procollagen type I, fibrillin-1, matrix metalloproteinase-1 (MMP-1) and -12, and ultraviolet (UV)-induced DNA damage in artificially UV-irradiated buttock skin before and after treatment. The supplement group showed significant improvements in skin elasticity and transepidermal water loss in photoaged facial skin after 12 weeks compared with the placebo group. In the supplement group, expression of procollagen type I mRNA increased and expression of MMP-1 and -12 mRNA decreased compared with those in the placebo group. In contrast, there was no significant difference in UV-induced DNA damage between groups. These results demonstrate that dietary astaxanthin combined with collagen hydrolysate can improve elasticity and barrier integrity in photoaged human facial skin, and such treatment is well tolerated.

Long-term topical oestrogen treatment of sun-exposed facial skin in post-menopausal women does not improve facial wrinkles or skin elasticity, but induces matrix metalloproteinase-1 expression.

It is controversial whether treatment with oestrogen stimulates collagen production or accumulation in sun-exposed skin. The aim of this study was to determine the effect of long-term treatment with topical oestrogen on photoaged facial skin, with regard to wrinkle severity, and expression of procollagen and matrix metalloproteinase-1 enzyme. Two groups of 40 post-menopausal women applied either 1 g of 1% oestrone or vehicle cream once daily to the face for 24 weeks. Visiometer R1-R5 values (skin wrinkles) and Cutometer values (skin elasticity) were not significantly improved in the oestrone group after 24 weeks of treatment. Type I procollagen immunostaining did not increase in the oestrone group compared with the control group. However, levels of matrix metalloproteinase-1 mRNA increased robustly (10.3 times) in oestrone-treated skin compared with vehicle-treated skin. Thus, treatment with topical oestrogen may be deleterious in ultraviolet-induced skin ageing, at least in part, through induction of matrix metalloproteinase-1 (MMP-1) expression in human skin.

Blood type B antigen modulates cell migration through regulating cdc42 expression and activity in HaCaT cells.

ABO blood group is determined by carbohydrate antigens, called ABH antigens. It has been known that the change of carbohydrate antigen expression, including ABH antigens, has correlation with the tumor metastasis and survival; however, the exact mechanism remains to be elucidated. ABH antigens are expressed not only in blood cells but also in several tissues. In epidermis, ABH antigen is expressed in the uppermost spinous and granular layer. We investigated the role of ABH antigens on the cell migration of HaCaT keratinocytes, which express B antigen. Knock-down of B antigen expression by small interference RNA of FUT1 inhibited HaCaT cell migration. At that time, we found that lamellipodia and actin fiber were also reduced by knock-down of B antigen expression. The transcription of cdc42, a kind of Rho GTPase which plays a key role in actin polymerization, was reduced by down-regulated B antigen expression. Furthermore, the reduced B antigen expression also inhibited the interaction of cdc42 and N-WASP. Collectively, our data provide a clue how ABH antigens regulate the cell migration mechanism.

Lipid ingredients in moisturizers can modulate skin responses to UV in barrier-disrupted human skin in vivo.

BACKGROUND: Chemicals with a molecular weight <500 and adequate lipid solubility can penetrate the intact human skin. As many lipid ingredients in moisturizers have molecular weights <500, the lipid ingredients may penetrate into the skin and affect skin responses to UV; however, little is known about this phenomenon. OBJECTIVE: To evaluate the effects of major lipid ingredients in moisturizers on skin responses to UV in tape-stripped human skin in vivo. METHODS: We evaluated the effects of three major lipid ingredients in moisturizers (cholesterol, linoleic acid, and a synthetic ceramide, N-oleoyl-phytosphingosine) on skin responses to UV in the tape-stripped skin of healthy volunteers. After 2 days of lipid-application, the areas were irradiated with UV, and skin samples were obtained 24h after irradiation. Histologic features and the expression of the markers of collagen metabolism and inflammatory mediators were evaluated. RESULTS: Compared to vehicle, topical cholesterol significantly decreased the degree of dermal inflammatory infiltrates and exocytosis, and also decreased the expression of MMP-1, IL-6, and IL-1ß mRNA. In contrast, topical linoleic acid increased the induction of apoptotic cells, and the expression of MMP-1 and IL-6 mRNA. N-oleoyl-phytosphingosine increased the expression of MMP-1 and IL-6 mRNA, while decreasing the expression of COX-2 mRNA. CONCLUSIONS: Topical cholesterol can protect the barrier-disrupted skin against UV-induced damage, while linoleic acid or N-oleoyl-phytosphingosine alone has the potential to aggravate the damage.

Differential effects of low-dose and high-dose beta-carotene supplementation on the signs of photoaging and type I procollagen gene expression in human skin in vivo.

BACKGROUND: Although the photoprotective effects of beta-carotene are thought to originate from its antioxidant properties, some studies documented pro-oxidant effects of beta-carotene. OBJECTIVE: Our purpose was to determine the effects of 2 different doses of dietary beta-carotene on wrinkles and elasticity, procollagen gene expression and ultraviolet (UV)-induced DNA damage in human skin. METHODS: Thirty healthy female subjects over the age of 50 years were randomized and received 2 different doses (30 and 90 mg/day) of beta-carotene for 90 days. The baseline status was used as control. At baseline and completion of the study, facial wrinkles and elasticity were measured objectively. Buttock skin was taken to determine the type I procollagen, matrix metalloproteinase-1 and fibrillin-1 mRNA levels, and UV-induced thymine dimer and 8-hydroxy-2'-deoxyguanosine formation. RESULTS: beta-Carotene improved facial wrinkles and elasticity significantly only in the low-dose group. The minimal erythema dose decreased significantly only in the high-dose group. Type I procollagen mRNA levels were significantly increased to 4.4 +/- 1.6 times the baseline level only in the low-dose group, and procollagen immunostaining increased accordingly. UV-induced thymine dimer staining was reduced in the low-dose group but tended to increase in the high-dose group. 8-hydroxy-2'-deoxyguanosine staining was significantly reduced in the low-dose group. CONCLUSIONS: 30 mg/day of beta-carotene supplementation is demonstrated to prevent and repair photoaging.

Red ginseng root extract mixed with Torilus fructus and Corni fructus improves facial wrinkles and increases type I procollagen synthesis in human skin: a randomized, double-blind, placebo-controlled study.

Red ginseng contains many bioactive constituents, including various ginsenosides that are believed to have antioxidant, immunostimulatory, and anti-aging activities. Yet, no controlled human study has explored its effects on photoaged skin. This study determined whether long-term intake of a red ginseng extract-containing Torilus fructus and Corni fructus mixture reduces facial wrinkles and increases collagen synthesis in human skin. Healthy female volunteers over 40 years of age were randomized in a double-blind fashion to receive either red ginseng extract-containing herbal mixture at 3 g/day or placebo for 24 weeks. Facial wrinkles, elasticity, epidermal water content, erythema, and pigmentation were measured objectively. Facial skin samples were taken before and after treatment, and real-time polymerase chain reaction and immunohistochemical analyses were undertaken for expression of type I procollagen, matrix metalloproteinase (MMP)-9, and fibrillin-1, which are wrinkle-related biochemical markers. A total of 82 subjects completed the study. Facial wrinkles were significantly improved, type I procollagen gene and protein expression was increased, MMP-9 gene induction was prevented, and fibrillin-1 fiber length was elongated only in the treatment group. No changes were seen in the facial elasticity, epidermal water content, facial erythema and pigmentation, and epidermal thickness in either group. Thus a red ginseng extract-containing Torilus fructus and Corni fructus mixture improves facial wrinkles, a clinical sign of photoaging, and this improvement is associated with biochemical and histological evidence of increased collagen synthesis in the dermis. These results substantiate the alleged beneficial effects of red ginseng on photoaging and support its use as an effective "beauty food."

Dietary Aloe Vera Supplementation Improves Facial Wrinkles and Elasticity and It Increases the Type I Procollagen Gene Expression in Human Skin in vivo.

BACKGROUND: No studies have yet been undertaken to determine the effect of aloe gel on the clinical signs and biochemical changes of aging skin. OBJECTIVE: We wanted to determine whether dietary aloe vera gel has anti-aging properties on the skin. METHODS: Thirty healthy female subjects over the age of 45 were recruited and they received 2 different doses (low-dose: 1,200 mg/d, high-dose: 3,600 mg/d) of aloe vera gel supplementation for 90 days. Their baseline status was used as a control. At baseline and at completion of the study, facial wrinkles were measured using a skin replica, and facial elasticity was measured by an in vivo suction skin elasticity meter. Skin samples were taken before and after aloe intake to compare the type I procollagen and matrix metalloproteinase 1 (MMP-1) mRNA levels by performing real-time RT-PCR. RESULTS: After aloe gel intake, the facial wrinkles improved significantly (p<0.05) in both groups, and facial elasticity improved in the lower-dose group. In the photoprotected skin, the type I procollagen mRNA levels were increased in both groups, albeit without significance; the MMP-1 mRNA levels were significantly decreased in the higher-dose group. Type I procollagen immunostaining was substantially increased throughout the dermis in both groups. CONCLUSION: Aloe gel significantly improves wrinkles and elasticity in photoaged human skin, with an increase in collagen production in the photoprotected skin and a decrease in the collagen-degrading MMP-1 gene expression. However, no dose-response relationship was found between the low-dose and high-dose groups.

Phosphatidylserine prevents UV-induced decrease of type I procollagen and increase of MMP-1 in dermal fibroblasts and human skin in vivo.

In an effort to find topical agents that prevent or retard cutaneous aging, seven functional lipids were screened for their procollagen-upregulating and matrix metalloproteinase (MMP)-1-downregulating activities in human dermal fibroblasts by Western blotting. The preventive effect on ultraviolet (UV)-induced decrease of procollagen was demonstrated in phosphatidylserine (PS), lysophosphatidylserine (LPS), lysophosphatidic acid (LPA), N-acetyl phytosphingosine (NAPS), and tetraacetyl phytosphingosine (TAPS). Furthermore, PS, LPS, and LPA upregulated procollagen expression in unirradiated basal conditions. The inhibitory effect on UV-induced MMP-1 expression was seen in NAPS, TAPS, LPA, PS, lysophosphatidylglycerol, and LPS. PS was chosen as the most suitable candidate anti-aging chemical for the subsequent in vivo studies. We investigated the effects of PS on acute UV response and chronologic skin aging by topically applying it to young skin before UV irradiation and to aged human skin, respectively. Real-time PCR and Western blot revealed that in the young skin, PS treatment prevented UV-induced reduction in procollagen expression and inhibited UV-induced MMP-1 expression. PS also blocked UV-induced IL-6 and COX-2 gene expression in cultured fibroblasts dose-dependently. In the aged skin, PS caused increased procollagen transcription and procollagen immunostaining in the upper dermis, and a significant decrease in MMP-1 expression at both mRNA and protein levels. These results indicate that topical PS has anti-skin-aging properties and point to the potential use of PS as a therapeutic agent in the prevention and treatment of cutaneous aging.

Infrared plus visible light and heat from natural sunlight participate in the expression of MMPs and type I procollagen as well as infiltration of inflammatory cell in human skin in vivo.

BACKGROUND: Compared with the detailed characterization of the ultraviolet (UV) response in human skin, the effects of infrared (IR) and other regions of the sunlight are scarce. OBJECTIVES: To determine the participation of IR/visible light and heat components of the sunlight on matrix metalloproteinases (MMPs) and type I procollagen expression, and inflammatory cell infiltration in human skin in vivo. METHODS: The buttocks of 16 healthy volunteers (aged 24-43 years, 10 male and 6 female) were irradiated with a 1.1-3 minimal erythema dose (MED) of natural sunlight. To determine the differential effects of UV, IR/visible rays and solar heat alone, the exposed sites were covered with either a UV filter or black cloth, respectively, during irradiation. Skin samples were taken 24h later. RESULTS: IR/visible light spectrum of sunlight significantly increased MMP-1 and MMP-9 expression and decreased type I procollagen expression. Solar heat also contributed to the increased MMP-1 expression. Only the UV region recruited neutrophils into the dermis, while UV, IR/visible light and heat contributed to macrophage infiltration. CONCLUSIONS: IR/visible light and heat of natural sunlight, in addition to UV, play a role in modulating the expressions of MMPs and procollagen, and inflammatory cell infiltration in human skin.

Photoprotective and anti-skin-aging effects of eicosapentaenoic acid in human skin in vivo.

Skin aging can be attributed to photoaging (extrinsic) and chronological (intrinsic) aging. Photoaging and intrinsic aging are induced by damage to human skin attributable to repeated exposure to ultraviolet (UV) irradiation and to the passage of time, respectively. In our previous report, eicosapentaenoic acid (EPA) was found to inhibit UV-induced matrix metalloproteinase-1 (MMP-1) expression in human dermal fibroblasts. Therefore, we investigated the effects of EPA on UV-induced skin damage and intrinsic aging by applying EPA topically to young and aged human skin, respectively. By immunohistochemical analysis and Western blotting, we found that topical application of EPA reduced UV-induced epidermal thickening and inhibited collagen decrease induced by UV light. It was also found that EPA attenuated UV-induced MMP-1 and MMP-9 expression by inhibiting UV-induced c-Jun phosphorylation, which is closely related to UV-induced activator protein-1 activation, and by inhibiting JNK and p38 activation. EPA also inhibited UV-induced cyclooxygenase-2 (COX-2) expression without altering COX-1 expression. Moreover, it was found that EPA increased collagen and elastic fibers (tropoelastin and fibrillin-1) expression by increasing transformin growth factor-beta expression in aged human skin. Together, these results demonstrate that topical EPA has potential as an anti-skin-aging agent.

Ultraviolet radiation attenuates thrombospondin 1 expression via PI3K-Akt activation in human keratinocytes.

Thrombospondin 1 (TSP1) is an extracellular glycoprotein and a recognized inhibitor of angiogenesis. Recent studies have demonstrated that UV radiation induces an angiogenic switch, by which it alters the balance between pro- and anti-angiogenic factors in the skin. Here we describe the effects of acute UV exposure on TSP1 expression in human skin epidermis, primary keratinocytes and the epidermal cell line HaCaT. We found that protein and mRNA expressions of TSP1 are significantly reduced in human skin in vivo and in keratinocytes in vitro by a single UV exposure. In human skin and keratinocytes, UV exposure induced the phosphorylation of Akt, a downstream target of the PI3K pathways. Specific inhibitors of PI3K, wortmannin and LY294002, completely blocked Akt activation and UV-induced TSP1 downregulation in keratinocytes. We showed that a specific Akt phosphorylation inhibitor and small interfering RNA-mediated Akt depletion were also blocked by UV-induced TSP1 downregulation in keratinocytes. In conclusion, our findings demonstrate that acute UV exposure downregulates TSP1 expression via PI3K-Akt activation in human keratinocytes. These novel findings may help us understand the regulatory mechanisms of UV-induced skin angiogenesis.