Dorsal Skin Of Hairless Mice Research Articles

Photodegradation enhances the toxic effect of anthracene on skin

Anthracene, a polycyclic aromatic hydrocarbon (PAH), is a widespread environmental pollutant that poses potential risks to human health. Exposure to anthracene can result in various adverse health effects, including skin-related disorders. Photo exposure sufficiently removes the anthracene from the environment but also generates more degradation products which can be more toxic. The goal of this study was to assess the change in anthracene dermotoxicity caused by photodegradation and understand the mechanism of this change. In the present study, over 99.99% of anthracene was degraded within 24 h of sunlight exposure, while producing many intermediate products including 9,10-anthraquinone and phthalic acid. The anthracene products with different durations of photo exposure were applied to 2D and 3D human keratinocyte cultures. Although the non-degraded anthracene significantly delayed the cell migration, the cell viability and differentiation decreased dramatically in the presence of the photodegraded anthracene. Anthracene photodegradation products also altered the expression patterns of a number of inflammation-related genes in comparison to the control cells. Among these genes, il1a, il1b, il8, cxcl2, s100a9, and mmp1 were upregulated whereas the tlr4 and mmp3 were downregulated by the photodegraded anthracene. Topical deliveries of the photodegraded and non-degraded anthracene to the dorsal skin of hairless mice showed more toxic effects by the photodegraded anthracene. The 4-hour photodegradation products of anthracene thickened the epidermal layer, increased the dermal cellularity, and induced the upregulation of inflammatory markers, il1a, il1b, s100a9, and mmp1. In addition, it also prevented the production of a gap junction protein, Connexin-43. All the evidence suggested that photodegradation enhanced the toxicities of anthracene to the skin. The 4-hour photodegradation products of anthracene led to clinical signs similar to acute inflammatory skin diseases, such as atopic and contact dermatitis, eczema, and psoriasis. Therefore, the potential risk of skin irritation by anthracene should be also considered when an individual is exposed to PAHs, especially in environments with strong sunlight.

Effects of the Higenamine, a Potent Compound from Aconitum, on UVB-Induced Photoaging in Hairless Mice.

Aim Higenamine [1-[(4-hydroxyphenyl) methyl]-1, 2, 3, 4-tetrahydroisoquinoline-6, 7-diol], a potent cardiotonic compound from Aconitum, contributes to vascular relaxation and bronchodilation. However, the effects and mechanisms of action of higenamine on skin aging remain poorly understood. In this study, the effects of higenamine on UVB-induced photoaging were examined in the hairless mouse model. Methods The dorsal skin of hairless mice (CrlOri : SKH1) was exposed to chronic UVB irradiation (100–300 mJ/cm2 for 6 weeks), with subsequent administration of higenamine (1–20 mg/kg, p.o.) for 2 weeks. TGF-β, Smad3 DNA-binding phosphorylation, and COL1A1 levels were analyzed by immunohistochemistry, and histological analysis of the skin was performed via H&E and MT staining. Results Higenamine increased TGF-β, Smad3 DNA-binding phosphorylation, and COL1A1 expression in primary human fibroblast cells and mouse skin. Higenamine suppressed UVB-induced photoaging via skin recovery, improved epidermal thickness, and prevented Smad3, DNA-binding phosphorylation, and COL1A1 depletion via TGF-β signaling. Conclusion Higenamine enhances collagen production in the skin through TGF-β/Smad3 signaling and potentially suppresses UVB-induced skin aging.

Dietary Collagen Hydrolysates Ameliorate Furrowed and Parched Skin Caused by Photoaging in Hairless Mice

sCollagen hydrolysates have been suggested as a favorable antiaging modality in skin photoaged by persistent exposure to ultraviolet radiation (UV). The current study evaluated the beneficial effect of collagen hydrolysates (fsCH) extracted from Pangasius hypophthalmus fish skin on wrinkle formation and moisture preservation in dorsal skin of hairless mice challenged with UV-B. Inter-comparative experiments were conducted for anti-photoaging among fsCH, retinoic acid (RA), N-acetyl-D-glucosamine (NAG), and glycine-proline-hydroxyproline (GPH). Treating human HaCaT keratinocytes with 100−200 μg/mL fsCH reciprocally ameliorated the expression of aquaporin 3 (AQP3) and CD44 deranged by UV-B. The UV-B-induced deep furrows and skin thickening were improved in parched dorsal skin of mice supplemented with 206–412 mg/kg fsCH as well as RA and GPH. The UV-B irradiation enhanced collagen fiber loss in the dorsal dermis, which was attenuated by fsCH through enhancing procollagen conversion to collagen. The matrix metalloproteinase expression by UV-B in dorsal skin was diminished by fsCH, similar to RA and GPH, via blockade of collagen degradation. Supplementing fsCH to UV-B-irradiated mice decreased transepidermal water loss in dorsal skin with reduced AQP3 level and restored keratinocyte expression of filaggrin. The expression of hyaluronic acid synthase 2 and hyaluronidase 1 by UV-B was remarkably ameliorated with increased production of hyaluronic acid by treating fsCH to photoaged mice. Taken together, fsCH attenuated photoaging typical of deep wrinkles, epidermal thickening, and skin water loss, like NAG, RA, or GPH, through inhibiting collagen destruction and epidermal barrier impairment.

Alleviation of Ultraviolet B-Induced Photoaging by 7-MEGATM 500 in Hairless Mouse Skin

The purpose of this study was to investigate the effect of 7-MEGA™ 500 on the improvement of skin aging in an UVB-induced photo-aging model of hairless mice. The dorsal skin of hairless mice was exposed to UVB three times a week for 12 weeks to induce skin wrinkle. After inducing the wrinkle, 7-MEGA™ 500 was orally administered once a day for 4 weeks. Skin thickness, skin barrier function, and wrinkle indicators were improved by treatment with 7-MEGA™ 500. Both gene and protein expression levels of MMP-3 and c-Jun in skin were significantly decreased by 7-MEGA™ 500. Therefore, the intake of 7-MEGA™ 500 is thought to have a positive effect on the improvement of skin aging, although further studies are needed.

Orally administered collagen peptide protects against UVB-induced skin aging through the absorption of dipeptide forms, Gly-Pro and Pro-Hyp.

Collagen hydrolysate is a well-known nutritional supplement for the improvement of healthy skin. Here, collagen peptide NS (CPNS) from fish scale was prepared, and its physicochemical properties were investigated. Gly-Pro was revealed as a representative low molecular weight peptide of CPNS, by performing prep-HPLC and LC-MS/MS. CPNS treatment attenuated matrix metalloproteinase-1 production and increased the synthesis of type 1 procollagen in HDF cells. After orally administering CPNS to rats, the plasma concentrations of Gly-Pro and Pro-Hyp increased dramatically. To examine the protective effects of CPNS against ultraviolet B (UVB)-induced photoaging in vivo, the dorsal skins of hairless mice were exposed to UVB and supplemented with CPNS for 12weeks. The CPNS consumption significantly attenuated UVB-induced wrinkle formation, transepidermal water loss, and epidermis thickness, and increased skin hydration. Collectively, these results suggest that bioactive peptides of CPNS, Gly-Pro and Pro-Hyp, exert beneficial effects on skin health.

5-aminolaevulinic acid-based photodynamic therapy inhibits ultraviolet B-induced skin photodamage.

To evaluate the photoprotective effect of 5-aminolaevulinic acid-based photodynamic therapy (ALA-PDT) on ultraviolet B (UVB)-induced skin photodamage. In vivo experiments, the dorsal skin of hairless mice were treated with ALA-PDT or saline-PDT, and then exposed to 180 mJ/m2 UVB. Results showed that the number of sunburn cells and apoptotic cells in the epidermis of ALA-PDT-treated groups at 24 h after UVB irradiation were significantly decreased compared with those in the UVB groups. And the removal rate of CPDs was obviously higher in ALA-PDT-treated groups. At 48 h, the number of Ki67 positive nuclei in ALA-PDT-UVB group was significantly fewer than that in UVB group. Further in vitro experiments, human keratinocyte cell line (HaCaT) cells of two groups (one treated with ALA-PDT, the other untreated), were exposed to 60 mJ/m2 UVB irradiation. We found 0.5 mmol/L of ALA and 3 J/cm2 of red light did not affect the vitality of cells, and could reduce UVB induced apoptosis, accelerate the clearance of CPDs, inhibit proliferation and activate p53. Thus, our data demonstrate that ALA-PDT pretreatment can induce a protective DNA damage response that protects skin cells from UVB-induced photodamages.

Sanshool improves UVB-induced skin photodamage by targeting JAK2/STAT3-dependent autophagy

Ultraviolet radiation is markedly increased because of pollution and the depletion of the stratospheric ozone layer. Excessive exposure to sunlight can negatively affect the skin, resulting in sunburn, photo-aging, or skin cancer. In this study, we first determined the photoprotective effect of sanshool, a major component in Zanthoxylum bungeanum, on UVB-irradiated responses in human dermal fibroblasts (HDFs) and nude mouse. We found that sanshool treatment can protect cells against the effects of UVB irradiation by (i) increasing cell viability, (ii) inhibiting MMP expression, and (iii) inducing autophagy. We also used the recombinant CSF2 or anti-CSF2 antibody co-cultured with human dermal fibroblasts (HDFs) and found that CSF2 contributes to sanshool-induced autophagy. Sanshool hindered the UVB-induced activation of JAK2-STAT3 signaling in HDFs, thereby inhibiting the expression of MMPs and activation of autophagic flux. Exposure of the dorsal skin of hairless mice to UVB radiation and subsequent topical application of sanshool delayed the progression of skin inflammation, leading to autophagy and inhibiting the activation of JAK2-STAT3 signaling. These results provide a basis for the study of the photoprotective effect of sanshool and suggest that it can be potentially used as an agent against UVB-induced skin damage in humans.

Topical treatment with a cathepsin G inhibitor, β-keto-phosphonic acid, blocks ultraviolet irradiation-induced basement membrane damage in hairless mouse skin.

Ultraviolet light (UV) exposure contributes various effects to skin including damage of the basement membrane. Cathepsin G (CTSG) belongs to serine protease family, and its upregulation is involved in wrinkle formation by chronic UV irradiation. However, the effect of CTSG on the basement membrane damage in skin remains unclear. To investigate the effects of topical treatment with a CTSG inhibitor, β-keto-phosphonic acid (KPA), on basement membrane damage in chronically UV-irradiated hairless mouse skin. The dorsal skin of hairless mice was exposed to UV three times per week for 8weeks. KPA was applied immediately after each session of UV irradiation. The basement membrane components, CTSG expression, and neutrophil infiltration were analyzed by immunofluorescence staining. The basement membrane structures were visualized by transmission electron microscope. CTSG and MMP-13 protein levels were analyzed by Western blotting. Assessment of wrinkle formation was examined using a skin replica assay. β-keto-phosphonic acid prevented UV irradiation-induced decrease in type VII collagen, laminin 332, and perlecan at the basement membrane zone and prevented UV-induced breakage of lamina densa and UV-induced shortening of hemidesmosome. KPA prevented UV-induced CTSG and MMP-13 expressions in chronically UV-irradiated hairless mice. Increase in neutrophil infiltration by UV irradiation and UV-induced wrinkle formation was also prevented by KPA. Our present study showed the possible involvement of CTSG in UV-induced basement membrane damage in skin through topical treatment with a CTSG inhibitor, KPA. Thus, inhibition of CTSG may be a useful strategy for the prevention of UV-induced basement membrane damage and photoaging.

5-Aminolevulinic Acid-Based Photodynamic Therapy Pretreatment Mitigates Ultraviolet A-Induced Oxidative Photodamage.

Aim To determine whether 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT) is effective in combating ultraviolet A- (UVA-) induced oxidative photodamage of hairless mice skin in vivo and human epidermal keratinocytes in vitro. Methods In in vitro experiments, the human keratinocyte cell line (HaCaT cells) was divided into two groups: the experimental group was treated with ALA-PDT and the control group was left untreated. Then, the experimental group and the control group of cells were exposed to 10 J/m2 of UVA radiation. ROS, O2− species, and MMP were determined by fluorescence microscopy; p53, OGG1, and XPC were determined by Western blot analysis; apoptosis was determined by flow cytometry; and 8-oxo-dG was determined by immunofluorescence. Moreover, HaCaT cells were also treated with ALA-PDT. Then, SOD1 and SOD2 were examined by Western blot analysis. In in vivo experiments, the dorsal skin of hairless mice was treated with ALA-PDT or saline-PDT, and then, they were exposed to 20 J/m2 UVA light. The compound 8-oxo-dG was detected by immunofluorescence. Conclusion In human epidermal keratinocytes and hairless mice skin, UVA-induced oxidative damage can be prevented effectively with ALA-PDT pretreatment.

Anti-photoaging effects of chitosan oligosaccharide in ultraviolet-irradiated hairless mouse skin

Skin photoaging (SP) is a premature skin-aging damage after repeated exposure to ultraviolet (UV) radiation, mainly characterized by oxidative stress and inflammatory disequilibrium, which makes skin show the typical symptoms of photoaging such as coarse wrinkling, dryness, irregular pigmentation and laxity. Chitosan oligosaccharide (COS), a natural polysaccharide with good humectant property, is the depolymerized product of chitosan with various biological activities, among which the antioxidant and anti-inflammatory effects have been frequently reported in recent years. However, no existing invivo study indicates whether COS has direct protective effect on UV-induced SP. In the current research, we investigated the potential preventive effect of COS against UV-caused damage in hairless mouse dorsal skin. The data showed that COS, by topical application after each UV-radiation for 10weeks, effectively inhibited the undesirable changes on the skin induced by UV. To be specific, COS obviously alleviated the macroscopic and histopathological damages of mice skin, via mitigating the disrupted collagenous fibers, as well as improving the relative content of type I collagen and the amount of total collagen. Furthermore, COS effectively inhibited the levels of pro-inflammatory cytokines such as TNF-α, IL-1β and IL-6, and markedly improved the activities of antioxidant enzymes (SOD, GSH-Px, CAT), as well as the content of skin hydroxyproline and moisture. These findings demonstrated that this natural polysaccharide attenuated UV-induced SP, at least in part, by virtue of favorable regulation of antioxidant and anti-inflammatory status, which presumably worked in concert to maintain the morphology and level of dermal collagen.

Formulation of a microemulsion-based hydrogel containing celecoxib

The aims of this study are to prepare and characterize a microemulsion-based hydrogel containing celecoxib (CLX) for transdermal delivery. Pseudo-ternary phase diagrams containing 0.74% (w/w) CLX were constructed using a water titration method. The mixture exhibiting the largest area of the microemulsion region was selected. The effect of the ratio of oil to the mixture of surfactant and cosurfactant (O/Smix) on in vitro skin permeation was investigated. Attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR) was employed to investigate the effects of the CLX microemulsion on skin. Rheological measurements were performed, and in vitro skin permeation of the CLX microemulsion-based hydrogel was tested. Based on pseudo-ternary phase diagrams, the Capryol PGMC-Tween 80-Transcutol P mixture was selected because it had the largest microemulsion area. A CLX microemulsion with O/Smix ratio of 3/7 was chosen because it had the greatest in vitro skin permeation. ATR-FTIR demonstrated that CLX microemulsion led to a transition in the alkyl chain of SC lipids and permeated hairless mouse dorsal skin. The CLX microemulsion-based hydrogel was elastic and suitable for application to skin. The incorporation of a gelling agent controlled the drug release of CLX, implying improved therapeutic effects of CLX for arthritis.

Prevention of UV-Induced Melanin Production by Accumulation of Redox Nanoparticles in the Epidermal Layer via Iontophoresis.

UV rays induce melanin production in the skin, which, from a cosmetic point of view, is problematic. Reactive oxygen species (ROS) generated in the skin upon UV irradiation are thought to be responsible for melanin production. Thus, effective antioxidants are recognized as useful tools for prevention of UV-induced melanin production. Redox nanoparticles (RNPs) containing nitroxide radicals as free radical scavengers were previously developed, and shown to be effective ROS scavengers in the body. RNPs are therefore expected to be useful for effective protection against UV-induced melanin production. However, as the sizes of RNPs are typically larger than the intercellular spaces of the skin, transdermal penetration is difficult. We recently demonstrated effective transdermal delivery and accumulation of nanoparticles in the epidermal layer via faint electric treatment, i.e., iontophoresis, suggesting that iontophoresis of RNPs may be a useful strategy for prevention of UV-induced melanin production in the skin. Herein, we performed iontophoresis of RNPs on the dorsal skin of hairless mice that produce melanin in response to light exposure. RNPs accumulated in the epidermal layer upon application of iontophoresis. Further, the combination of RNPs with iontophoresis decreased UV-induced melanin spots and melanin content in the skin. Taken together, we successfully demonstrated that iontophoresis-mediated accumulation of RNPs in the epidermis prevented melanin production.

Formulation and in vitro/in vivo evaluation of chitosan-based film forming gel containing ketoprofen

The film forming gel, adhered to skin surfaces upon application and formed a film, has an advantage onto skin to provide protection and continuous drug release to the application site. This study aimed to prepare a chitosan-based film forming gel containing ketoprofen (CbFG) and to evaluate the CbFG and film from CbFG (CbFG-film). CbFG were prepared with chitosan, lactic acid and various skin permeation enhancers. The physicochemical characteristics were evaluated by texture analysis, viscometry, SEM, DSC, XRD and FT-IR. To identify the mechanism of skin permeation, in vitro skin permeation study was conducted with a Franz diffusion cell and excised SD-rat and hairless mouse dorsal skin. In vivo efficacy assessment in mono-iodoacetate (MIA)-induced rheumatoid arthritis animal model was also conducted. CbFG was successfully prepared and, after applying CbFG to the excised rat dorsal skin, the CbFG-film was also formed well. The physicochemical characteristics of CbFG and CbFG-film could be explained by the grafting of oleic acid onto chitosan in the absence of catalysts. In addition, CbFG containing oleic acid had a higher skin permeation rate in comparison with any other candidate enhancers. The in vivo efficacy study also confirmed significant anti-inflammatory and analgesic effects. Consequently, we report the successful preparation of chitosan-based film forming gel containing ketoprofen with excellent mechanical properties, skin permeation and anti-inflammatory and analgesic effects.

Effects of the Ultra-High-Frequency Electrical Field Radiofrequency Device on Mouse Skin: A Histologic and Molecular Study.

Radiofrequency technology is one of the most recently developed methods for noninvasive skin tightening and facial contouring, and works by generating thermal energy in the deep dermis. Although clinical improvements have been reported using radiofrequency devices, there are few histologic and molecular studies about the mechanisms of dermal collagen remodeling. The authors investigated the histologic effects of an ultra-high-frequency electrical field (40.68 MHz) radiofrequency device (Polargen) on collagen remodeling in hairless mouse skin and evaluated its relative molecular mechanism. The radiofrequency was applied to the dorsal skin of hairless mice three times per week for 2 weeks. At 21 days after initial treatment, treated skin and nontreated control skin samples were excised for semiquantitative analysis of histologic features, including collagen. The authors also checked the mRNA expression levels of collagen type 1, transforming growth factor (TGF)-β, matrix metalloproteinase-1, vascular endothelial growth factor, tumor necrosis factor-α, and interleukin-1. Histologic examination revealed epidermal hyperplasia, increased collagen staining, and fat atrophy in treated skin area compared with the nontreated skin area. In addition, mRNA expression of collagen type І, TGF-β, and vascular endothelial growth factor in radiofrequency-treated areas was significantly increased compared with that in untreated control areas (p < 0.05, p < 0.05, and p < 0.01, respectively). These results suggest that the device may facilitate replacement of subcutaneous fat tissue with new collagen in association with the increased mRNA levels in TGF-β and vascular endothelial growth factor. Therefore, this device may effectively reduce adipose tissue and achieve facial contouring in addition to skin tightening.

Naringenin targets ERK2 and suppresses UVB-induced photoaging.

A number of natural phytochemicals have anti‐photoaging properties that appear to be mediated through the inhibition of matrix metalloproteinase‐1 (MMP‐1) expression, but their direct target molecule(s) and mechanism(s) remain unclear. We investigated the effect of naringenin, a major flavonoid found in citrus, on UVB‐induced MMP‐1 expression and identified its direct target. The HaCaT human skin keratinocyte cell line and 3‐dimensional (3‐D) human skin equivalent cultures were treated or not treated with naringenin for 1 hr before exposure to UVB. The mechanism and target(s) of naringenin were analysed by kinase assay and multiplex molecular assays. Dorsal skins of hairless mice were exposed to UVB 3 times per week, with a dose of irradiation that was increased weekly by 1 minimal erythema dose (MED; 45 mJ/cm2) to 4 MED over 15 weeks. Wrinkle formation, water loss and water content were then assessed. Naringenin suppressed UVB‐induced MMP‐1 expression and AP‐1 activity, and strongly suppressed UVB‐induced phosphorylation of Fos‐related antigen (FRA)‐1 at Ser265. Importantly, UVB irradiation‐induced FRA1 protein stability was reduced by treatment with naringenin, as well as with a mitogen‐activated protein kinase (MEK) inhibitor. Naringenin significantly suppressed UVB‐induced extracellular signal‐regulated kinase 2 (ERK2) activity and subsequently attenuated UVB‐induced phosphorylation of p90RSK by competitively binding with ATP. Constitutively active MEK (CA‐MEK) increased FRA1 phosphorylation and expression and also induced MMP‐1 expression, whereas dominant‐negative ERK2 (DN‐ERK2) had opposite effects. U0126, a MEK inhibitor, also decreased FRA1 phosphorylation and expression as well as MMP‐1 expression. The photoaging data obtained from mice clearly demonstrated that naringenin significantly inhibited UVB‐induced wrinkle formation, trans‐epidermal water loss and MMP‐13 expression. Naringenin exerts potent anti‐photoaging effects by suppressing ERK2 activity and decreasing FRA1 stability, followed by down‐regulation of AP‐1 transactivation and MMP‐1 expression.

CSP0510 Lotion as a Novel Moisturizer Containing Citric Acid and Trisodium Phosphate Relieves Objective and Subjective Symptoms of Atopic Dermatitis

BackgroundMoisturizers with anti-inflammatory or anti-itch activity should be developed for the safe and effective management of atopic dermatitis (AD).ObjectiveThis study evaluated the efficacy of a newly developed moisturizer, CSP0510 lotion (Twolines Inc., Korea), containing citric acid (CA) and trisodium phosphate (TSP) as active ingredients, in mild to moderate AD.Methods and ResultsCSP0510 lotion applied twice daily for 4 weeks to eczematous lesions improved objective and subjective (itch) symptoms of AD. The physician's global assessment (PGA) score for objective symptoms decreased from 2.5±0.6 before application to 1.3±0.5 after application in the CSP0510-treated group (n=42, p<0.001). Also, the PGA score decreased from 2.3±0.6 to 1.9±0.5 by vehicle-treated (without CA and TSP) control group (p=0.001), but there was no statistical difference between CSP0510-treated and vehicle-treated groups (p=0.089). The visual analogue scale score for itch decreased from 4.8±1.3 to 2.0±0.9 in the CSP0510-treated group (p<0.001), and from 4.6±1.1 to 3.5±0.9 in the control group (p=0.075), showing a statistical significance between two groups (p=0.002). Our results in humans were further supported by in vitro and animal experiments. In HaCaT cells treated with compound 48/80 (7.5 µg/ml), CA:TSP (1:1, vol:vol) synergistically suppressed the compound 48/80-induced upregulation of thymic stromal lymphopoietin, nerve grow factor, and calcitonin gene-related peptide. Application of CSP0510 to the dorsal skin of hairless mice for 3 weeks suppressed the oxazolone-induced allergic skin inflammation.ConclusionIn conclusion, CSP0510 lotion has anti-itch and anti-inflammatory activity in the skin, which improves both objective and subjective symptoms of AD.

Photoprotective Potential of Penta-O-Galloyl-β-DGlucose by Targeting NF-κB and MAPK Signaling in UVB Radiation-Induced Human Dermal Fibroblasts and Mouse Skin.

Exposure of the skin to ultraviolet radiation can cause skin damage with various pathological changes including inflammation. In the present study, we identified the skin-protective activity of 1,2,3,4,6-penta-O-galloyl-β-D-glucose (pentagalloyl glucose, PGG) in ultraviolet B (UVB) radiation-induced human dermal fibroblasts and mouse skin. PGG exhibited antioxidant activity with regard to intracellular reactive oxygen species (ROS) generation as well as ROS and reactive nitrogen species (RNS) scavenging. Furthermore, PGG exhibited anti-inflammatory activity, inhibiting the activation of nuclear factor-kappaB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling, resulting in inhibition of the expression of pro-inflammatory mediators. Topical application of PGG followed by chronic exposure to UVB radiation in the dorsal skin of hairless mice resulted in a significant decrease in the progression of inflammatory skin damages, leading to inhibited activation of NF-κB signaling and expression of pro-inflammatory mediators. The present study demonstrated that PGG protected from skin damage induced by UVB radiation, and thus, may be a potential candidate for the prevention of environmental stimuli-induced inflammatory skin damage.

Noninvasive Optical Imaging of UV-Induced Squamous Cell Carcinoma in Murine Skin: Studies of Early Tumor Development and Vitamin D Enhancement of Protoporphyrin IX Production.

Better noninvasive techniques are needed to monitor protoporphyrin IX (PpIX) levels before and during photodynamic therapy (PDT) of squamous cell carcinoma (SCC) of the skin. Our aim was to evaluate (1) multispectral fluorescent imaging of ultraviolet light (UV)-induced cancer and precancer in a mouse model of SCC and (2) multispectral imaging and probe-based fluorescence detection as a tool to study vitamin D (VD) effects on aminolevulinic acid (ALA)-induced PpIX synthesis. Dorsal skin of hairless mice was imaged weekly during a 24-week UV carcinogenesis protocol. Hot spots of PpIX fluorescence were detectable by multispectral imaging beginning at 14 weeks of UV exposure. Many hot spots disappeared after cessation of UV at week 20, but others persisted or became visible after week 20, and corresponded to tumors that eventually became visible by eye. In SCC-bearing mice pretreated with topical VD before ALA application, our optical techniques confirmed that VD preconditioning induces a tumor-selective increase in PpIX levels. Fluorescence-based optical imaging of PpIX is a promising tool for detecting early SCC lesions of the skin. Pretreatment with VD can increase the ability to detect early tumors, providing a potential new way to improve efficacy of ALA-PDT.

Connexin 43 Antisense Therapy Modulates Connexin Expression and Wound Repair in Nitrogen Mustard Injured Mouse Skin

Connexin 43 (Cx43) antisense treatment accelerates diabetic skin wound repair. Cx43 expression and its phosphorylation (pCx43) alters gap junction communication during skin wound repair. Vesicant induced skin injury results in prolonged wound healing similar to diabetic wounds. We performed a time course study using Cx43 antisense oligodeoxynucleotides (asODN) on nitrogen mustard (NM) injured SKH-1 hairless mouse dorsal skin to evaluate wound progression and Cx expression. Immunofluorescent (IF) analysis of NM injured skin demonstrated increased keratin 17 (K17, skin wound marker), increased expression of Cx26 and pCx43, and minimal expression of Cx43 by day 1. All markers were up-regulated with time as the wounds progressed. Dual-labeling studies indicated strong expression of Cx26 near the leading edge of wounds. In contrast, Cx43 and pCx43 were highly expressed in the hyperplastic epithelium. Cx43asODN treatment improved NM wounded skin as indicated by minimal K17 expression at day 1. Western blotting showed reduced levels of Cx43 and pCX43 over time. Dual labeling demonstrated that Cx43 asODN treatment successfully reduced Cx43 and pCx43 expression and it was mostly restricted to the basal keratinocytes. Cx26 expression appeared more prevalent in both the wound margin and hyperplastic epithelium by day 3. We hypothesize that the increased epidermal Cx26 positive area is associated with migratory keratinocytes to improve epithelialization. In conclusion, Cx43asODN altered the Cx expression profile, modulated gap junction communication and accelerated vesicant wound repair. Supported by ES005022, EY09056, and NIAMS U54AR055073.

Modulating drug resistance by targeting BCRP/ABCG2 using retrovirus-mediated RNA interference.

BackgroundThe BCRP/ABCG2 transporter, which mediates drug resistance in many types of cells, depends on energy provided by ATP hydrolysis. Here, a retrovirus encoding a shRNA targeting the ATP-binding domain of this protein was used to screen for highly efficient agents that could reverse drug resistance and improve cell sensitivity to drugs, thus laying the foundation for further studies and applications.Methodology/Principal FindingsTo target the ATP-binding domain of BCRP/ABCG2, pLenti6/BCRPsi shRNA recombinant retroviruses, with 20 bp target sequences starting from the 270th, 745th and 939th bps of the 6th exon, were constructed and packaged. The pLenti6/BCRPsi retroviruses (V-BCRPi) that conferred significant knockdown effects were screened using a drug-sensitivity experiment and flow cytometry. The human choriocarcinoma cell line JAR, which highly expresses endogenous BCRP/ABCG2, was injected under the dorsal skin of a hairless mouse to initiate a JAR cytoma. After injecting V-BCRPi-infected JAR tumor cells into the dorsal skin of hairless mice, BCRP/ABCG2 expression in the tumor tissue was determined using immunohistochemistry, fluorescent quantitative RT-PCR and Western blot analyses. After intraperitoneal injection of BCRP/ABCG2-tolerant 5-FU, the tumor volume, weight change, and apoptosis rate of the tumor tissue were determined using in situ hybridization. V-BCRPi increased the sensitivity of the tumor histiocytes to 5-FU and improved the cell apoptosis-promoting effects of 5-FU in the tumor.Conclusions/SignificanceThe goal of the in vivo and in vitro studies was to screen for an RNA interference recombinant retrovirus capable of stably targeting the ATP-binding domain of BCRP/ABCG2 (V-BCRPi) to inhibit its function. A new method to improve the chemo-sensitivity of breast cancer and other tumor cells was discovered, and this method could be used for gene therapy and functional studies of malignant tumors.