Phloroglucinol attenuates ultraviolet B radiation-induced matrix metalloproteinase-1 production in human keratinocytes via inhibitory actions against mitogen-activated protein kinases and activator protein-1

Excessive amounts of reactive oxygen species (ROS) induced by ultraviolet (UV) radiation cause skin aging via basement membrane/extracellular matrix degradation resulting from the action of matrix metalloproteinases (MMPs). Recently, phloroglucinol (1,3,5-trihydroxybenzene) was demonstrated to attenuate the cell damage induced by oxidative stress by quenching ROS and stimulating antioxidant systems. In the current study, the effect of phloroglucinol on UVB-induced photoaging was investigated in human HaCaT keratinocytes. Phloroglucinol significantly inhibited the UVB-induced (1) upregulation of MMP-1 mRNA, protein and activity; (2) augmentation of intracellular Ca(2+) levels; (3) phosphorylation of mitogen-activated protein kinases (MAPKs); (4) expression of c-Fos and phospho c-Jun; and (5) enhancement of activator protein-1 (AP-1) binding to the MMP-1 promoter. In addition, the knockdown of MAPKs significantly inhibited UVB-induced MMP-1 expression. The results of this study suggest that phloroglucinol may be useful as a photoprotective compound for the skin.     

Bucillamine Inhibits UVB-Induced MAPK Activation and Apoptosis in Human HaCaT Keratinocytes and SKH-1 Hairless Mouse Skin

Ultraviolet B (UVB) radiation is known as a culprit in skin carcinogenesis. We have previously reported that bucillamine (N-[2-mercapto-2-methylpropionyl]-L-cysteine), a cysteine derivative with antioxidant and anti-inflammatory capacity, protects against UVB-induced p53 activation and inflammatory responses in mouse skin. Since MAPK signaling pathways regulate p53 expression and activation, here we determined bucillamine effect on UVB-mediated MAPK activation in vitro using human skin keratinocyte cell line HaCaT and in vivo using SKH-1 hairless mouse skin. A single low dose of UVB (30 mJ cm⁻²) resulted in increased JNK/MAPK phosphorylation and caspase-3 cleavage in HaCaT cells. However, JNK activation and casaspe-3 cleavage were inhibited by pretreatment of HaCaT cells with physiological doses of bucillamine (25 and 100 µm ). Consistent with these results, bucillamine pretreatment in mice (20 mg kg⁻¹) inhibited JNK/MAPK and ERK/MAPK activation in skin epidermal cells at 6–12 and 24 h, respectively, after UVB exposure. Moreover, bucillamine attenuated UVB-induced Ki-67-positive cells and cleaved caspase-3-positive cells in mouse skin. These findings demonstrate that bucillamine inhibits UVB-induced MAPK signaling, cell proliferation and apoptosis. Together with our previous report, we provide evidence that bucillamine has a photoprotective effect against UV exposure.     

Pomegranate fruit extract inhibits UVB-induced inflammation and proliferation by modulating NF-κB and MAPK signaling pathways in mouse skin

There is considerable interest in the identification of natural agents capable of affording protection to skin from the adverse effects of solar ultraviolet B (UVB) radiation. Pomegranate (Punica granatum L.) fruit possesses as strong antioxidant, anti-inflammatory and antiproliferative properties. Recently, we have shown that oral feeding of pomegranate fruit extract (PFE) to mice afforded substantial protection from the adverse effects of single UVB radiation via modulation in early biomarkers of photocarcinogenesis. This study was designed to investigate the photochemopreventive effects of PFE (0.2%, wt/vol) after multiple UVB irradiations (180 mJ cm(-2), on alternative day, for a total of seven treatments) to the skin of SKH-1 hairless mice. Oral feeding of PFE to SKH-1 mice inhibited UVB-induced epidermal hyperplasia, infiltration of leukocytes, protein oxidation and lipid peroxidation. Immunoblot analysis demonstrated that oral feeding of PFE to mice inhibited UVB-induced (1) nuclear translocation and phosphorylation of nuclear factor kappa B/p65, (2) phosphorylation and degradation of IκBα, (3) activation of IKKα/ΙΚΚβ and (4) phosphorylation of mitogen-activated protein kinase proteins and c-Jun. PFE consumption also inhibited UVB-induced protein expression of (1) COX-2 and iNOS, (2) PCNA and cyclin D1 and (3) matrix metalloproteinases-2,-3 and -9 in mouse skin. Taken together, these data show that PFE consumption afforded protection to mouse skin against the adverse effects of UVB radiation by modulating UVB-induced signaling pathways.     

Low-dose UVB irradiation stimulates matrix metalloproteinase-1 expression via a BLT2-linked pathway in HaCaT cells

Skin exposure to low-dose ultraviolet B (UVB) light up-regulates the expression of matrix metalloproteinase-1 (MMP-1), thus contributing to premature skin aging (photo-aging). Although cyclooxygenase-2 (COX- 2) and its product, prostaglandin E(2) (PGE((2))), have been associated with UVB-induced signaling to MMP expression, very little are known about the roles of lipoxygenases and their products, especially leukotriene B((4)) (LTB((4))) and 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE), in MMP-1 expression in skin keratinocytes. In the present study, we demonstrate that BLT2, a cell surface receptor for LTB((4)) and 12(S)-HETE, plays a critical role in UVB-mediated MMP-1 upregulation in human HaCaT keratinocytes. Moreover, our results demonstrated that BLT2-mediated MMP-1 upregulation occurs through a signaling pathway dependent on reactive oxygen species (ROS) production and the subsequent stimulation of ERK. Blockage of BLT2 via siRNA knockdown or with the BLT2-antagonist LY255283 completely abolished the up-regulated expression of MMP-1 induced by low-dose UVB irradiation. Finally, when HaCaT cells were transiently transfected with a BLT2 expression plasmid, MMP-1 expression was significantly enhanced, along with ERK phosphorylation, suggesting that BLT2 overexpression alone is sufficient for MMP-1 up-regulation. Together, our results suggest that the BLT2-ROS- ERK-linked cascade is a novel signaling mechanism for MMP-1 upregulation in low-dose UVB- irradiated keratinocytes and thus potentially contributes to photo-aging.     

Super-highly hydroxylated fullerene derivative protects human keratinocytes from UV-induced cell injuries together with the decreases in intracellular ROS generation and DNA damages

Polyhydroxylated fullerenes (fullerenols: C(60)(OH)(n)) are known as the major water-soluble fullerene derivatives which possess particular significance as free radical scavengers or antioxidants in biological systems. Recently, the novel polyhydroxylated fullerene (C(60) (OH)(44)·8H(2)O: SHH-F) was successfully synthesized. In the present study, we investigated the radical-scavenging effects and cytoprotective effects of three types of fullerenols (C(60)(OH)(6-12): LH-F, C(60) (OH)(32-34)·7H(2)O: HH-F, and C(60) (OH)(44)·8H(2)O: SHH-F) on UV-irradiation-induced cell injuries. HH-F and SHH-F exerted hydroxyl-radical scavenging activities as shown by DMPO-spin trap/ESR method, more markedly than LH-F. UVA or UVB irradiation-induced injuries in human skin keratinocytes HaCaT were significantly suppressed by HH-F and SHH-F, but scarcely by LF-H. The cytoprotective effects of SHH-F had a tendency to be superior to that of HH-F. And the cytoprotective effects of SHH-F against UVB-induced injuries were more effective than those of UVA. Irradiation with UVB to HaCaT cells was shown to cause rapid increases in cell-injury-associated symptoms such as intracellular oxidative stress levels, the formation of cyclobutane pyrimidine dimers and chromatin condensation, all of which were repressed by SHH-F. Thus, UVB-induced diverse harmful effects could be prevented by SHH-F, which was suggested to exert the cytoprotective effects through intracellular reactive oxygen species-scavenging in the keratinocytes.     

A Peptide YGDEY from Tilapia Gelatin Hydrolysates Inhibits UVB‐mediated Skin Photoaging by Regulating MMP‐1 and MMP‐9 Expression in HaCaT Cells

In this study, we investigated the protective effects of a peptide (YGDEY, Tyr‐Gly‐Asp‐Glu‐Tyr) isolated from tilapia skin gelatin hydrolysates (TGHs), against UVB‐induced photoaging in human keratinocytes (HaCaT) cells. Results showed that YGDEY significantly decreased levels of intracellular reactive oxygen species (ROS), increased antioxidant factors (Superoxide Dismutase, SOD and Glutathione, GSH) expression and maintained balance between GSH and GSSG in HaCaT cells. Comet assay shows that YGDEY can protect DNA from oxidative damage. Furthermore, it significantly inhibited MMP‐1 (collagenase) and MMP‐9 (gelatinase) expression and increased Type I procollagen production. In addition, the molecular docking study showed that YGDEY may form active sites with MMP‐1 and MMP‐9. Moreover, Western blot analysis was utilized to measure the protein levels of UVB‐induced mitogen‐activated protein kinase (MAPK) and nuclear factor‐kappa B (NF‐κB) signaling pathways. Therefore, these results suggested that YGDEY has a therapeutic effectiveness in prevention of UVB‐induced cellular damage, and it is a candidate worthy of being developed as a potential natural antioxidant and food additive.     

Protective Effect of Curcumin Against Acute Ultraviolet B Irradiation‐induced Photo‐damage

Ultraviolet B (UVB) irradiation is one of the most dangerous insults for skin and causes sunburn, erythema, photoaging and photocarcinogenesis. Curcumin (diferuloylmethane), a yellow spice derived from dried rhizomes of Curcuma longa, has been shown to possess significant anti‐inflammatory, antioxidant, anticarcinogenic, antimutagenic, anticoagulant and anti‐infective effects. However, the protective effects of curcumin against acute photo‐damage are poorly understood. In this study, we investigated the photoprotective effects of curcumin against UVB‐induced acute photo‐damage in hairless mice and immortalized human keratinocytes (HaCaT). Topical application of curcumin significantly inhibited acute UVB (540 mJ cm^?2, for 3 successive days)‐induced inflammatory cells, collagen accrementition derangement and lipid peroxidation, and effectively induced NF‐E2‐related factor 2 (Nrf2) nuclear accumulation in uncovered (Uncv) hairless mice skin. Treatment of HaCaT cells with curcumin significantly attenuated acute UVB (300 mJ cm^?2)‐induced lactate dehydrogenase release, intracellular reactive oxygen species production and DNA damage, activated the expression of the phase II detoxifying enzymes and promoted DNA repair activity. The photoprotective effect provided by curcumin was potential associated with modulation of Nrf2‐dependent antioxidant response. Our study suggested that curcumin is a potential agent for preventing and/or treating UV radiation‐induced acute inflammation and photoaging.     

Cordycepin inhibits UVB-induced matrix metalloproteinase expression by suppressing the NF-��B pathway in human dermal fibroblasts

Cordycepin (3''-deoxyadenosine) has been shown to exhibit many pharmacological activities, including anti-cancer, anti-inflammatory, and anti-infection activities. However, the anti-skin photoaging effects of cordycepin have not yet been reported. In the present study, we investigated the inhibitory effects of cordycepin on matrix metalloproteinase-1 (MMP-1) and -3 expressions of the human dermal fibroblast cells. Western blot analysis and real-time PCR revealed cordycepin inhibited UVB-induced MMP-1 and -3 expressions in a dose-dependent manner. UVB strongly activated NF-κB activity, which was determined by IκBα degradation, nuclear localization of p50 and p65 subunit, and NF-κB binding activity. However, UVB-induced NF-κB activation and MMP expression were completely blocked by cordycepin pretreatment. These findings suggest that cordycepin could prevent UVB-induced MMPs expressions through inhibition of NF-κB activation. In conclusion, cordycepin might be used as a potential agent for the prevention and treatment of skin photoaging.     

(-)-Epicatechin-3-gallate, a green tea polyphenol is a potent agent against UVB-induced damage in HaCaT keratinocytes

(-)-Epicatechin-3-gallate (ECG) is a polyphenolic compound similar to (-)-epigallocatechin-3-gallate (EGCG) which is abundant in green tea. Numerous workers have proposed that EGCG protects epidermal cells against UVB-induced damage. However, little has been known about whether ECG protects keratinocytes against UVB-induced damage. We decided to investigate the protective effects and underlying mechanisms of ECG on UVB-induced damage. Cell viability was determined by the MTT assay. Activation of ERK1/2, p38 and JNK was analyzed by Western blotting. Intracellular H2O2 production and DNA content was analyzed by flow cytometry. Lipid peroxidation was assayed by colorimetry. In our study, we found that ECG dose-dependently attenuated UVB-induced keratinocyte death. Moreover, ECG markedly inhibited UVB-induced cell membrane lipid peroxidation and H2O2 generation in keratinocytes, suggesting that ECG can act as a free radical scavenger when keratinocytes were photodamaged. In parallel, H2O2-induced the activation of ERK1/2, p38 and JNK in keratinocytes could be inhibited by ECG. UVB-induced pre-G1 arrest leading to apoptotic changes of keratinocytes were blocked by ECG. Taken together, we provide here evidence that ECG protects keratinocytes from UVB-induced photodamage and H2O2-induced oxidative stress, possibly through inhibition of the activation of ERK1/2, p38 and JNK and/or scavenging of free radicals.     

In vitro and in vivo transfer of bcl-2 gene into keratinocytes suppresses UVB-induced apoptosis.

Bcl-2 is a member of the large Bcl-2 family and protects cells from apoptosis. Ultraviolet B (UVB) irradiation induces apoptosis of keratinocytes that is known as "sunburn cells." Previously we reported that UVB irradiation induces apoptosis accompanied by sequential activation of caspase 8, 3 and 1 in keratinocytes, and that the process is inhibited by various caspase inhibitors. Using bcl-2-expressing adenovirus vector we investigated the effect of Bcl-2 on UVB-induced apoptosis. Adenovirus vector efficiently introduced bcl-2 gene in cultured normal mouse keratinocytes (NMK cells); almost all NMK cells (1 x 10(6)) were transfected at 1 x 10(8) plaque-forming unit (PFU)/mL. Bcl-2-transfected NMK cells were significantly resistant to UVB-induced apoptosis with the suppressive effect dependent on the Bcl-2 expression level. Following UVB irradiation caspase 8, 3 and 9 activities were stimulated in NMK cells, whereas in bcl-2-transfected cells only caspase 8, but not caspase 3 or 9, activity was stimulated. In order to investigate the effect of Bcl-2 in vivo topical application of Ad-bcl-2 on tape-stripped mouse skin was performed. Following the application Bcl-2 was efficiently overexpressed in almost all viable keratinocytes. The expression was transient with the maximal expression of Bcl-2 on the first day following the application of 1 x 10(9) PFU in 200 microL. The introduced Bcl-2 remained at least for 6 days. UVB irradiation (1250 J/m2) induced apoptosis within 12 h and the maximal effect was observed at 24 h in control mouse skin. Both bcl-2-transfected and topical caspase 3 inhibitor-treated mice skin were resistant to UVB-induced apoptosis. The suppressive effect of Bcl-2 was more potent than that of caspase 3 inhibitor application. Topical application of empty adenovirus vector alone had no effect on Bcl-2 expression or UVB-induced apoptosis. These results indicate that adenovirus vector is an efficient gene delivery system into keratinocytes and that Bcl-2 is a potent inhibitor of UVB-induced apoptosis both in vitro and in vivo.     

Photoprotective Effect of Black Tea Extracts Against UVB-induced Phototoxicity in Skin

In previous studies, we showed that green tea and black tea extracts and their major polyphenolic constituents protect against UVB light-induced carcinogenesis in murine skin. All of these studies required chronic administration of tea extracts or specific constituents either topically or orally. However, it is not known whether acute or subchronic administration of black tea extracts or constituents can ameliorate UVB-induced early effects in skin. In the present study, cultured keratinocytes and mouse and human skin were employed to assess the effect of both oral and topical administration of standardized black tea extract (SBTE) and its two major polyphenolic subfractions namely BTF1 and BTF2 against UVB-induced photodamage. In SKH-1 hairless mice, topical application of SBTE (0.2 mg/cm2) prior to UVB exposure (180 mJ/cm2) resulted in 40% reduced incidence and 64% reduced severity of erythema and 50% reduction in skinfold thickness by day 6 when compared to nontreated UVB-exposed animals. The SBTE was also effective in protecting against UVB-induced erythema in human volunteers. Administration of SBTE 5 min after UVB irradiation was similarly effective in reducing UVB-induced inflammation in both murine and human skin. The major polyphenolic subfractions, BTF1 and BTF2, were also effective in protecting in mouse skin. The SBTE subfractions inhibited UVB-induced tyrosine phosphorylation of epidermal growth factor receptor (EGFR). The UVB irradiation of human epidermoid carcinoma cells resulted in 3.3-fold induction of tyrosine phosphorylation of EGFR. Pretreatment with BTF1 and BTF2 reduced tyrosine phosphorylation of EGFR by 53% and 31%, respectively. The UVB-mediated enhanced expression of the early response genes, c-fos and c-jun in human epidermal keratinocytes was reduced in a dose-dependent manner by SBTE. Topical application of SBTE was also effective in reducing accumulation of c-fos and p53 proteins by 82% and 78%, respectively, in UVB-exposed mouse skin. These data provide evidence that constituents of black tea can abrogate UVB-induced erythema and associated early events in murine and human skin.     

12(S)-Hydroxyheptadeca-5Z,8E,10E-trienoic acid suppresses UV-induced IL-6 synthesis in keratinocytes, exerting an anti-inflammatory activity

12(S)-Hydroxyheptadeca-5Z,8E,10E-trienoic acid (12- HHT) is an enzymatic product of prostaglandin H(2) (PGH(2)) derived from cyclooxygenase (COX)-mediated arachidonic acid metabolism. Despite the high level of 12-HHT present in tissues and bodily fluids, its precise function remains largely unknown. In this study, we found that 12-HHT treatment in HaCaT cells remarkably down-regulated the ultraviolet B (UVB) irradiation-induced synthesis of interleukin-6 (IL-6), a pro-inflammatory cytokine associated with cutaneous inflammation. In an approach to identify the down-stream signaling mechanism by which 12-HHT down-regulates UVB-induced IL-6 synthesis in keratinocytes, we observed that 12-HHT inhibits the UVB-stimulated activation of p38 mitogen-activated protein kinase (MAPK) and nuclear factor kappa B (NF-κB). In addition, we found that 12-HHT markedly up-regulates MAPK phosphatase-1 (MKP-1), a critical negative regulator of p38 MAPK. When MKP-1 was suppressed by siRNA knock-down, the 12-HHT-mediated inhibitory effects on the UVB-stimulated activation of p38 MAPK and NF-κB, as well as the production of IL-6, were attenuated in HaCaT cells. Taken together, our results suggest that 12-HHT exerts anti-inflammatory effect via up-regulation of MKP-1, which negatively regulates p38 MAPK and NF-κB, thus attenuating IL-6 production in UVB-irradiated HaCaT cells. Considering the critical role of IL-6 in cutaneous inflammation, our findings provide the basis for the application of 12-HHT as a potential anti-inflammatory therapeutic agent in UV-induced skin diseases.     

Collagen-EGCG Combination Synergistically Prevents UVB-Induced Skin Photoaging in Nude Mice

Ultraviolet (UV) radiation is a major cause of skin photoaging through generating excessive oxidative stress and inflammation. One of the strategies is to use photo-chemoprotectors, such as natural products with antioxidant and anti-inflammatory properties, to protect the skin from photo damage. The present study investigates the photoprotective potentials of topical administration of unhydrolyzed collagen, epigallocatechin gallate (EGCG), and their combination against ultraviolet B (UVB)-induced photoaging in nude mice. It is found that both the solo and combined pretreatments could recover UVB-induced depletion of antioxidative enzymes, including superoxide dismutase and glutathione peroxidase (GSH-Px), as well as an increase of lipid peroxide malondialdehyde and inflammatory tumor necrosis factor-α. Meanwhile, the UVB-stimulated skin collagen degradation is attenuated significantly with drug treatments, which is evidenced by expression analysis of matrix metalloproteinase-1 and hydroxyproline. Additionally, the mouse skin histology shows that the drug-pretreated groups possess decreased epidermis thickness and normal collagen fiber structure of the dermis layer. These results demonstrate that both EGCG and collagen can protect the skin against UVB-induced skin photoaging. Synergistically, the combination of them shows the maximum prevention to skin damage, showing its potential in the application of anti-photoaging formulation products.     

CD11b+ cells are the major source of oxidative stress in UV radiation-irradiated skin: possible role in photoaging and photocarcinogenesis

Exposure of skin to solar UV radiation induces oxidative stress and suppression of cell-mediated immune responses. These effects are associated with the greater risk of several skin disorders including photoaging and photocarcinogenesis. We have shown that UV-induced infiltrating leukocytes contribute in developing oxidative stress in UV-irradiated skin. The peak period of UV-induced infiltrating leukocytes lies between 48 and 72 h after UV exposure of the skin. In this study we demonstrated that UV (90 mJ/cm2)-induced infiltrating CD11b+ cells in C3H/HeN mice skin were the major source of oxidative stress. Hydrogen peroxide (H2O2) was determined as a marker of oxidative stress. Flow cytometric analysis of viable cells revealed that the number of CD11b+H2O2+ cells were significantly higher (31.8%, P < 0.001) in UV-irradiated skin in comparison with non-UV-exposed skin (0.4%). Intraperitoneal administration of monoclonal antibodies to CD11b (rat IgG2b) to C3H/HeN mice inhibited UVB-induced infiltration of leukocytes, as evidenced by reduction in myeloperoxidase activity (64-80%, P < 0.0005), concomitant with significant reduction in H2O2 production both in epidermis and dermis (66-83%, P < 0.001-0.0005) when compared with the administration of rat IgG2b isotype of anti-CD11b. Furthermore, CD11b+ and CD11b- cell subsets were separated by immunomagnetic cell isolation technique from total epidermal and dermal single cell suspensions obtained 48 h after UV irradiation of the skin and analyzed for H2O2 production. Analytical data revealed that CD11b+ cell population from UV-irradiated skin resulted in significantly higher production of total H2O2 in both epidermis and dermis (87-89%, P < 0.0001) in comparison with CD11b- cell population (11-13% of total H2O2). These data revealed that infiltrating CD11b+ cells were the major source of oxidative stress in UV-irradiated skin and thus may contribute to photoaging and promotion of skin tumor growth within the UV-irradiated skin. Together, these data suggest that reduction in UV-induced skin infiltration of CD11b+ cells may be an alternative and effective strategy to reduce solar UV light-induced oxidative stress-mediated skin disorders including photoaging and photocarcinogenesis.     

Pomegranate fruit extract modulates UV-B-mediated phosphorylation of mitogen-activated protein kinases and activation of nuclear factor kappa B in normal human epidermal keratinocytes paragraph sign

Excessive exposure of solar ultraviolet (UV) radiation, particularly its UV-B component, to humans causes many adverse effects that include erythema, hyperplasia, hyperpigmentation, immunosuppression, photoaging and skin cancer. In recent years, there is increasing use of botanical agents in skin care products. Pomegranate derived from the tree Punica granatum contains anthocyanins (such as delphinidin, cyanidin and pelargonidin) and hydrolyzable tannins (such as punicalin, pedunculagin, punicalagin, gallagic and ellagic acid esters of glucose) and possesses strong antioxidant and anti-inflammatory properties. Recently, we have shown that pomegranate fruit extract (PFE) possesses antitumor promoting effects in a mouse model of chemical carcinogenesis. To begin to establish the effect of PFE for humans in this study, we determined its effect on UV-B-induced adverse effects in normal human epidermal keratinocytes (NHEK). We first assessed the effect of PFE on UV-B-mediated phosphorylation of mitogen-activated protein kinases (MAPK) pathway in NHEK. Immunoblot analysis demonstrated that the treatment of NHEK with PFE (10-40 microg/mL) for 24 h before UV-B (40 mJ/cm(2)) exposure dose dependently inhibited UV-B-mediated phosphorylation of ERKl/2, JNK1/2 and p38 protein. We also observed that PFE (20 microg/mL) inhibited UV-B-mediated phosphorylation of MAPK in a time-dependent manner. Furthermore, in dose- and time-dependent studies, we evaluated the effect of PFE on UV-B-mediated activation of nuclear factor kappa B (NF-kappaB) pathway. Using Western blot analysis, we found that PFE treatment of NHEK resulted in a dose- and time-dependent inhibition of UV-B-mediated degradation and phosphorylation of IkappaBalpha and activation of IKKalpha. Using immunoblot analysis, enzyme-linked immunosorbent assay and electrophoretic mobility shift assay, we found that PFE treatment to NHEK resulted in a dose- and time-dependent inhibition of UV-B-mediated nuclear translocation and phosphorylation of NF-kappaB/p65 at Ser(536). Taken together, our data shows that PFE protects against the adverse effects of UV-B radiation by inhibiting UV-B-induced modulations of NF-kappaB and MAPK pathways and provides a molecular basis for the photochemopreventive effects of PFE.     

Plantamajoside Inhibits UVB and Advanced Glycation End Products‐Induced MMP‐1 Expression by Suppressing the MAPK and NF‐κB Pathways in HaCaT Cells

Photoaging and glycation stress are major causes of skin deterioration. Oxidative stress caused by ultraviolet B (UVB) irradiation can upregulate matrix metalloprotease 1 (MMP‐1), a major enzyme responsible for collagen damage in the skin. Advanced glycation end products (AGEs) accumulate via gradual formation from skin proteins, especially from long‐lived proteins such as dermal elastin and collagen. Plantamajoside (PM), isolated from Plantago asiatica, has various biological effects including anti‐inflammatory and antioxidant effects. In this study, we assessed the protective effects of PM on a human keratinocyte cell line (HaCaT) and primary human dermal fibroblasts (HDF) against stress caused by glyceraldehyde‐induced AGEs (glycer‐AGEs) with UVB irradiation. We found that PM attenuated UVB‐ and‐glycer‐AGEs‐induced MMP‐1 expression in HaCaT and HDF cells and proinflammatory cytokines expression by inhibiting the phosphorylation of mitogen‐activated protein kinases (MAPKs) activated by reactive oxygen species. Specific inhibitors of NF‐κB and MAPKs attenuated the induced expression of MMP‐1. PM also inhibited the phosphorylation of IκBα, and reduced nuclear translocation of NF‐κB in these cells. Furthermore, PM attenuated the upregulation of receptor for AGEs (RAGE) by glycer‐AGEs with UVB irradiation. Therefore, our findings strongly suggest that PM is a promising inhibitor of skin photoaging.     

Photo-Protective and Anti-Inflammatory Effects of Antidesma thwaitesianum Müll. Arg. Fruit Extract against UVB-Induced Keratinocyte Cell Damage

The main cause of most skin cancers is damage from UVB from sunlight, which penetrate the skin surface and induce inflammation. For this reason, this study aims to identify natural products with photo-protection properties and their mode of action by using the UVB-irradiated HaCaT keratinocyte model. Antidesma thwaitesianum fruit extracts at 25, 50, and 100 µg/mL recovered cell viability following UVB exposure in a dose-dependent manner. Cell survival was associated with the reduction in intracellular ROS and NO. In addition, we showed that the pre-treatment with the fruit extract lowered the phosphorylation level of two MAPK-signaling pathways: p38 MAPKs and JNKs. The resulting lower MAPK activation decreased their downstream pro-inflammatory cascade through COX-2 expression and subsequently reduced the PGE₂ proinflammatory mediator level. The photoprotective effects of the fruit extract were correlated with the presence of polyphenolic compounds, including cyanidin, ferulic acid, caffeic acid, vanillic acid, and protocatechuic acid, which have been previously described as antioxidant and anti-inflammation. Together, we demonstrated that the pre-treatment with the fruit extract had photo-protection by inhibiting oxidative stress and subsequently lowered stress-induced MAPK responses. Therefore, this fresh fruit is worthy of investigation to be utilized as a skincare ingredient for preventing UVB-induced skin damage.     

Amauroderma rugosum Extract Suppresses Inflammatory Responses in Tumor Necrosis Factor Alpha/Interferon Gamma-Induced HaCaT Keratinocytes

Keratinocytes form the physical barrier of the skin and play an important role in the inflammatory process. Amauroderma rugosum is an edible mushroom; however, its pharmacological properties have seldom been studied. Although the anti-inflammatory effect of the organic solvent extract of Amauroderma rugosum has been previously reported, it is not known whether the aqueous extract has a similar effect. In addition, the effect of Amauorderma rugosum extract on skin has never been explored. Therefore, the objectives of the present study were to evaluate the anti-inflammatory effects of the aqueous extract of Amauroderma rugosum on HaCaT keratinocytes, to explore its mechanisms of action, and to study the possible active ingredients involved. The results showed that the aqueous extract of Amauroderm rugosum at a concentration of 1.5 mg/mL was non-toxic to HaCaT cells and inhibited the release of cytokine interleukin-1β, and chemokines interleukin-8 and monocyte chemoattractant protein-1 in tumor necrosis factor (TNF)-α- and interferon (IFN)-γ-stimulated HaCaT cells. Amauroderma rugosum extract reduced the intracellular levels of reactive oxygen species. In addition, Amauroderma rugosum extract reduced the total protein expression of nuclear factor-kappa B (NF-κB) and B-cells inhibitor alpha in HaCaT keratinocytes and inhibited the phosphorylation of mitogen-activated protein kinase kinase (MEK) 1/2, extracellular signal-regulated kinase (ERK) 1/2, protein kinase B (Akt), and mammalian target of rapamycin (mTOR) in TNF-α- and INF-γ-stimulated HaCaT keratinocytes. Chemical analysis revealed that the aqueous extract of Amauroderma rugosum contains polysaccharides, triterpenes, and phenolic compounds. Anti-inflammatory compounds, such as gallic acid, guanosine, and uridine, were also present. The anti-inflammatory effect of Amauroderma rugosum could be mimicked by a combination of gallic acid, guanosine, and uridine. In conclusion, our study suggests that the aqueous extract of Amauroderma rugosum exerts anti-inflammatory effects on keratinocytes through its antioxidant and inhibitory effects on MEK/ERK-, Akt/mTOR-, and NF-κB-dependent signaling pathways.     

Photoprotective properties of Prunella vulgaris and rosmarinic acid on human keratinocytes

UVA radiation provokes the generation of reactive oxygen species (ROS), which induce oxidative stress in the exposed cells leading to extensive cellular damage and cell death either by apoptosis or necrosis. One approach to protecting human skin against the harmful effects of UV radiation is by using herbal compounds as photoprotectants. This study evaluated the protective effects of Prunella vulgaris L. (Labiatae) and its main phenolic acid component, rosmarinic acid (RA), against UVA-induced changes in a human keratinocyte cell line (HaCaT). Human keratinocytes exposed to UVA (10-30 J/cm(2)) were treated with an extract of P. vulgaris (1-75 mg/l) or RA (0.9-18 mg/l) for 4h. P. vulgaris and RA exhibited ability to reduce the UVA-caused decrease in a cell viability monitored by neutral red retention and by LDH release into medium. The P. vulgaris extract and RA significantly suppressed UVA-induced ROS production, which manifests as a decrease in intracellular lipid peroxidation, elevation of ATP and reduced glutathione. Post-treatment with P. vulgaris extract and RA also significantly reduced DNA damage. In addition, UVA-induced activation of caspase-3 was inhibited by treatment with P. vulgaris and RA. The P. vulgaris extract and RA demonstrated a concentration-dependent photoprotection (maximum at 25-50 mg/l and 9 mg/l, respectively). These results suggest that P. vulgaris and RA, used in skin care cosmetics, may offer protection against UVA-induced oxidative stress and may be beneficial as a supplement in photoprotective dermatological preparations.