Thymus vulgaris alleviates UVB irradiation induced skin damage via inhibition of MAPK/AP-1 and activation of Nrf2-ARE antioxidant system.

Solar ultraviolet (UV) radiation-induced reactive oxidative species is mainly responsible for the development of photoageing. Rosmarinic acid was one of the main bioactive components detected in Thymus vulgaris (TV) we extracted. In this study, UVB-induced skin damages have been shown to be ameliorated by treatment with TV in hairless mice (HR-1) skin, demonstrated by decreased matrix metalloproteinases (MMPs) and increased collagen production. However, the underlying molecular mechanism on which TV acted was unclear. We examined the photoprotective effects of TV against UVB and elucidated the molecular mechanism in normal human dermal fibroblasts. Thymus vulgaris remarkably prevented the UVB-induced reactive oxygen species and lactate dehydrogenase. Dose-dependent increase in glutathione, NAD(P)H: quinone oxidoreductase1 and heme oxygenase-1, by TV was confirmed by increased nuclear accumulation of Nrf2. Furthermore, 5-Methoxyindole-2-carboxylic acid was introduced as a specific inhibitor of dihydrolipoyl dehydrogenase (DLD). We demonstrated that Nrf2 expression was regulated by DLD, which was a tricarboxylic acid cycle-associated protein that decreased after UVB exposure. Besides, TV significantly diminished UVB induced phosphorylation of mitogen activated protein kinases pathway, containing extracellular signal-regulated kinase, Jun N-terminal kinase and p38, which consequently reduced phosphorylated c-fos and c-jun. Our results suggest that TV is a potential botanical agent for use against UV radiation-induced oxidative stress mediated skin damages.

EGF Functionalized Polymer-Coated Gold Nanoparticles Promote EGF Photostability and EGFR Internalization for Photothermal Therapy.

The application of functionalized nanocarriers on photothermal therapy for cancer ablation has wide interest. The success of this application depends on the therapeutic efficiency and biocompatibility of the system, but also on the stability and biorecognition of the conjugated protein. This study aims at investigating the hypothesis that EGF functionalized polymer-coated gold nanoparticles promote EGF photostability and EGFR internalization, making these conjugated particles suitable for photothermal therapy. The conjugated gold nanoparticles (100-200 nm) showed a plasmon absorption band located within the near-infrared range (650-900 nm), optimal for photothermal therapy applications. The effects of temperature, of polymer-coated gold nanoparticles and of UVB light (295nm) on the fluorescence properties of EGF have been investigated with steady-state and time-resolved fluorescence spectroscopy. The fluorescence properties of EGF, including the formation of Trp and Tyr photoproducts, is modulated by temperature and by the intensity of the excitation light. The presence of polymeric-coated gold nanoparticles reduced or even avoided the formation of Trp and Tyr photoproducts when EGF is exposed to UVB light, protecting this way the structure and function of EGF. Cytotoxicity studies of conjugated nanoparticles carried out in normal-like human keratinocytes showed small, concentration dependent decreases in cell viability (0-25%). Moreover, conjugated nanoparticles could activate and induce the internalization of overexpressed Epidermal Growth Factor Receptor in human lung carcinoma cells. In conclusion, the gold nanoparticles conjugated with Epidermal Growth Factor and coated with biopolymers developed in this work, show a potential application for near infrared photothermal therapy, which may efficiently destroy solid tumours, reducing the damage of the healthy tissue.

1950 MHz radiofrequency electromagnetic fields do not aggravate memory deficits in 5xFAD mice.

The increased use of mobile phones has generated public concern about the impact of radiofrequency electromagnetic fields (RF-EMF) on health. In the present study, we investigated whether RF-EMFs induce molecular changes in amyloid precursor protein (APP) processing and amyloid beta (Aß)-related memory impairment in the 5xFAD mouse, which is a widely used amyloid animal model. The 5xFAD mice at the age of 1.5 months were assigned to two groups (RF-EMF- and sham-exposed groups, eight mice per group). The RF-EMF group was placed in a reverberation chamber and exposed to 1950 MHz electromagnetic fields for 3 months (SAR 5 W/kg, 2 h/day, 5 days/week). The Y-maze, Morris water maze, and novel object recognition memory test were used to evaluate spatial and non-spatial memory following 3-month RF-EMF exposure. Furthermore, Aß deposition and APP and carboxyl-terminal fragment ß (CTFß) levels were evaluated in the hippocampus and cortex of 5xFAD mice, and plasma levels of Aß peptides were also investigated. In behavioral tests, mice that were exposed to RF-EMF for 3 months did not exhibit differences in spatial and non-spatial memory compared to the sham-exposed group, and no apparent change was evident in locomotor activity. Consistent with behavioral data, RF-EMF did not alter APP and CTFß levels or Aß deposition in the brains of the 5xFAD mice. These findings indicate that 3-month RF-EMF exposure did not affect Aß-related memory impairment or Aß accumulation in the 5xFAD Alzheimer's disease model. Bioelectromagnetics. 37:391-399, 2016. © 2016 The Authors Bioelectromagnetics published by Wiley Periodicals, Inc. on behalf of Bioelectromagnetics Society.

Photo-enhancement of macrophage phagocytic activity via Rac1-mediated signaling pathway: Implications for bacterial infection.

Phagocytosis and the subsequent destruction of invading pathogens by macrophages are indispensable steps in host immune responses to microbial infections. Low-power laser irradiation (LPLI) has been found to exert photobiological effects on immune responses, but the signaling mechanisms underlying this photobiomodulation of phagocytosis remains largely unknown. Here, we demonstrated for the first time that LPLI enhanced the phagocytic activity of macrophages by stimulating the activation of Rac1. The overexpression of constitutively activated Rac1 clearly enhanced LPLI-induced phagocytosis, whereas the overexpression of dominant negative Rac1 exerted the opposite effect. The phosphorylation of cofilin was involved in the effects of LPLI on phagocytosis, which was regulated by the membrane translocation and activation of Rac1. Furthermore, the photoactivation of Rac1 was dependent on the Src/PI3K/Vav1 pathway. The inhibition of the Src/PI3K pathway significantly suppressed LPLI-induced actin polymerization and phagocytosis enhancement. Additionally, LPLI-treated mice exhibited increased survival and a decreased organ bacterial load when challenged with Listeria monocytogenes, indicating that LPLI enhanced macrophage phagocytosis in vivo. These findings highlight the important roles of the Src/PI3K/Vav1/Rac1/cofilin pathway in regulating macrophage phagocytosis and provide a potential strategy for treating phagocytic deficiency via LPLI.

Overexpression of rice OsREX1-S, encoding a putative component of the core general transcription and DNA repair factor IIH, renders plant cells tolerant to cadmium- and UV-induced damage by enhancing DNA excision repair.

Screening of 40,000 Arabidopsis FOX (Full-length cDNA Over-eXpressor gene hunting system) lines expressing rice full-length cDNAs brings us to identify four cadmium (Cd)-tolerant lines, one of which carried OsREX1-S as a transgene. OsREX1-S shows the highest levels of identity to Chlamydomonas reinhardtii REX1-S (referred to as CrREX1-S, in which REX denotes Required for Excision) and to yeast and human TFB5s (RNA polymerase II transcription factor B5), both of which are components of the general transcription and DNA repair factor, TFIIH. Transient expression of OsREX1-S consistently localized the protein to the nucleus of onion cells. The newly generated transgenic Arabidopsis plants expressing OsREX1-S reproducibly displayed enhanced Cd tolerance, confirming that the Cd-tolerance of the initial identified line was conferred solely by OsREX1-S expression. Furthermore, transgenic Arabidopsis plants expressing OsREX1-S exhibited ultraviolet-B (UVB) tolerance by reducing the amounts of cyclobutane pyrimidine dimers produced by UVB radiation. Moreover, those transgenic OsREX1-S Arabidopsis plants became resistant to bleomycin (an inducer of DNA strand break) and mitomycin C (DNA intercalating activity), compared to wild type. Our results indicate that OsREX1-S renders host plants tolerant to Cd, UVB radiation, bleomycin and mitomycin C through the enhanced DNA excision repair.

Irradiance, but not fluence, plays a crucial role in UVB-induced immature pigment cell development: new insights for efficient UVB phototherapy.

Light exposure modulates development of living organisms. In the field of medicine, light has frequently been used for regenerative purposes. Excimer light (308 nm) has demonstrated superior efficacy in treating vitiligo, a condition requiring development of melanoblasts and a model for studying nerve cell regeneration, as compared to narrow-band ultraviolet B (NBUVB; 311 nm). Using mouse-derived melanoblast cells to examine the pro-differentiation effects of these two light sources, we demonstrated that at equivalent fluence, excimer light induces melanoblast differentiation, while NBUVB failed to so. Mechanistically, activation of aryl hydrocarbon receptor pathway and nuclear translocation of epidermal growth factor receptor are involved in pro-differentiation effects of excimer light. Reduction in irradiance by filter abrogated the effects of excimer light in melanoblasts, even when equivalent fluence was delivered by the same light source. As ultraviolet B (UVB) irradiation is closely associated pigment cell development, future therapy employing UVB for pigmentation purposes should incorporate irradiance as a crucial specification.

Inhibitory effect of Astragalus membranaceus root on matrix metalloproteinase-1 collagenase expression and procollagen destruction in ultraviolet B-irradiated human dermal fibroblasts by suppressing nuclear factor kappa-B activity.

OBJECTIVES: The root of Astragalus membranaceus, regarded as a tonic in traditional Korean medicine, has been prescribed for long periods to treat chronic illness by boosting the immune system. Ultraviolet (UV) irradiation causes damage to skin connective tissue by degrading collagen, which is a major structural component of the extracellular matrix. Such damage is considered to be a cause of the wrinkling observed in premature ageing of the skin. This study has investigated the photo-protective effect of A. membranaceus on UVB radiation-induced activation of nuclear factor kappa-B (NF-κB) activity in human dermal fibroblasts. METHODS: HS68 fibroblast cells cultured with various concentrations of A. membranaceus were exposed to UVB (40 mJ/cm²). Activation of NF-κB P65 and expression of matrix metalloproteinase-1 (MMP-1) and type 1 procollagen were measured by Western blotting. Translocation of NF-κB P65 and MMP-1 regulation were also examined by immunocytochemistry. KEY FINDINGS: Western blotting and immunocytochemistry results showed that A. membranaceus inhibited UVB-induced translocation of NF-κB P65 and MMP-1 expression. The data suggested that A. membranaceus restored type 1 procollagen synthesis by inhibiting NF-κB P65 activity and MMP-1 expression in UVB-exposed human dermal fibroblasts. CONCLUSION: A. membranaceus is a candidate for use in skin protection from UVB-induced skin inflammation and photoageing.

Ion-beam irradiation, gene identification, and marker-assisted breeding in the development of low-cadmium rice.

Rice (Oryza sativa L.) grain is a major dietary source of cadmium (Cd), which is toxic to humans, but no practical technique exists to substantially reduce Cd contamination. Carbon ion-beam irradiation produced three rice mutants with <0.05 mg Cd⋅kg(-1) in the grain compared with a mean of 1.73 mg Cd⋅kg(-1) in the parent, Koshihikari. We identified the gene responsible for reduced Cd uptake and developed a strategy for marker-assisted selection of low-Cd cultivars. Sequence analysis revealed that these mutants have different mutations of the same gene (OsNRAMP5), which encodes a natural resistance-associated macrophage protein. Functional analysis revealed that the defective transporter protein encoded by the mutant osnramp5 greatly decreases Cd uptake by roots, resulting in decreased Cd in the straw and grain. In addition, we developed DNA markers to facilitate marker-assisted selection of cultivars carrying osnramp5. When grown in Cd-contaminated paddy fields, the mutants have nearly undetectable Cd in their grains and exhibit no agriculturally or economically adverse traits. Because mutants produced by ion-beam radiation are not transgenic plants, they are likely to be accepted by consumers and thus represent a practical choice for rice production worldwide.

Far-infrared therapy induces the nuclear translocation of PLZF which inhibits VEGF-induced proliferation in human umbilical vein endothelial cells.

Many studies suggest that far-infrared (FIR) therapy can reduce the frequency of some vascular-related diseases. The non-thermal effect of FIR was recently found to play a role in the long-term protective effect on vascular function, but its molecular mechanism is still unknown. In the present study, we evaluated the biological effect of FIR on vascular endothelial growth factor (VEGF)-induced proliferation in human umbilical vein endothelial cells (HUVECs). We found that FIR ranging 3∼10 µm significantly inhibited VEGF-induced proliferation in HUVECs. According to intensity and time course analyses, the inhibitory effect of FIR peaked at an effective intensity of 0.13 mW/cm(2) at 30 min. On the other hand, a thermal effect did not inhibit VEGF-induced proliferation in HUVECs. FIR exposure also inhibited the VEGF-induced phosphorylation of extracellular signal-regulated kinases in HUVECs. FIR exposure further induced the phosphorylation of endothelial nitric oxide (NO) synthase (eNOS) and NO generation in VEGF-treated HUVECs. Both VEGF-induced NO and reactive oxygen species generation was involved in the inhibitory effect of FIR. Nitrotyrosine formation significantly increased in HUVECs treated with VEGF and FIR together. Inhibition of phosphoinositide 3-kinase (PI3K) by wortmannin abolished the FIR-induced phosphorylation of eNOS and Akt in HUVECs. FIR exposure upregulated the expression of PI3K p85 at the transcriptional level. We further found that FIR exposure induced the nuclear translocation of promyelocytic leukemia zinc finger protein (PLZF) in HUVECs. This induction was independent of a thermal effect. The small interfering RNA transfection of PLZF blocked FIR-increased PI3K levels and the inhibitory effect of FIR. These data suggest that FIR induces the nuclear translocation of PLZF which inhibits VEGF-induced proliferation in HUVECs.

Inhibition of Aß(25-35)-induced cell apoptosis by low-power-laser-irradiation (LPLI) through promoting Akt-dependent YAP cytoplasmic translocation.

Deposition of amyloid-ß-peptide (Aß) in the brain is considered a pathological hallmark of Alzheimer's disease (AD). Our previous studies show that Yes-associated protein (YAP) is involved in the regulation of apoptosis induced by Aß(25-35) through YAP nuclear translocation and its pro-apoptotic function is mediated by its interaction with p73. In the present study, we first found that Low-power laser irradiation (LPLI) promoted YAP cytoplasmic translocation and inhibited Aß(25-35)-induced YAP nuclear translocation. Moreover, the cytoplasmic translocation was in an Akt-dependent manner. Activated Akt by LPLI phosphorylated YAP on ser127 (S127) and resulted in decreasing the interaction between YAP and p73, and in suppressing the proapoptotic gene bax expression following Aß(25-35) treatment. Inhibition of Akt expression by siRNA significantly abolished the effect of LPLI. More importantly, LPLI could inhibit Aß(25-35)-induced cell apoptosis through activation of Akt/YAP/p73 signaling pathway. Therefore, our findings first suggest that YAP may be a therapeutic target and these results directly point to a potential therapeutic strategy for the treatment of AD through Akt/YAP/p73 signaling pathway with LPLI.

GATA3 expression is decreased in psoriasis and during epidermal regeneration; induction by narrow-band UVB and IL-4.

Psoriasis is characterized by hyperproliferation of keratinocytes and by infiltration of activated Th1 and Th17 cells in the (epi)dermis. By expression microarray, we previously found the GATA3 transcription factor significantly downregulated in lesional psoriatic skin. Since GATA3 serves as a key switch in both epidermal and T helper cell differentiation, we investigated its function in psoriasis. Because psoriatic skin inflammation shares many characteristics of epidermal regeneration during wound healing, we also studied GATA3 expression under such conditions.Psoriatic lesional skin showed decreased GATA3 mRNA and protein expression compared to non-lesional skin. GATA3 expression was also markedly decreased in inflamed skin of mice with a psoriasiform dermatitis induced with imiquimod. Tape-stripping of non-lesional skin of patients with psoriasis, a standardized psoriasis-triggering and skin regeneration-inducing technique, reduced the expression of GATA3. In wounded skin of mice, low GATA3 mRNA and protein expression was detected. Taken together, GATA3 expression is downregulated under regenerative and inflammatory hyperproliferative skin conditions. GATA3 expression could be re-induced by successful narrow-band UVB treatment of both human psoriasis and imiquimod-induced psoriasiform dermatitis in mice. The prototypic Th2 cytokine IL-4 was the only cytokine capable of inducing GATA3 in skin explants from healthy donors. Based on these findings we argue that GATA3 serves as a key regulator in psoriatic inflammation, keratinocyte hyperproliferation and skin barrier dysfunction.

The Usher 1B protein, MYO7A, is required for normal localization and function of the visual retinoid cycle enzyme, RPE65.

Mutations in the MYO7A gene cause a deaf-blindness disorder, known as Usher syndrome 1B.  In the retina, the majority of MYO7A is in the retinal pigmented epithelium (RPE), where many of the reactions of the visual retinoid cycle take place.  We have observed that the retinas of Myo7a-mutant mice are resistant to acute light damage. In exploring the basis of this resistance, we found that Myo7a-mutant mice have lower levels of RPE65, the RPE isomerase that has a key role in the retinoid cycle.  We show for the first time that RPE65 normally undergoes a light-dependent translocation to become more concentrated in the central region of the RPE cells.  This translocation requires MYO7A, so that, in Myo7a-mutant mice, RPE65 is partly mislocalized in the light.  RPE65 is degraded more quickly in Myo7a-mutant mice, perhaps due to its mislocalization, providing a plausible explanation for its lower levels.  Following a 50-60% photobleach, Myo7a-mutant retinas exhibited increased all-trans-retinyl ester levels during the initial stages of dark recovery, consistent with a deficiency in RPE65 activity.  Lastly, MYO7A and RPE65 were co-immunoprecipitated from RPE cell lysate by antibodies against either of the proteins, and the two proteins were partly colocalized, suggesting a direct or indirect interaction.  Together, the results support a role for MYO7A in the translocation of RPE65, illustrating the involvement of a molecular motor in the spatiotemporal organization of the retinoid cycle in vision.

High fluence low-power laser irradiation induces apoptosis via inactivation of Akt/GSK3ß signaling pathway.

High fluence low-power laser irradiation (HF-LPLI) is a newly discovered stimulus through generating reactive oxygen species (ROS) to trigger cell apoptosis. Activation of glycogen synthase kinase 3ß (GSK3ß) is proved to be involved in intrinsic apoptotic pathways under various stimuli. However, whether the proapoptotic factor GSK3ß participates in HF-LPLI-induced apoptosis has not been elucidated. Therefore, in the present study, we investigated the involvement of GSK3ß in apoptosis under HF-LPLI treatment (120 J/cm2, 633 nm). We found that GSK3ß activation could promote HF-LPLI-induced apoptosis, which could be prevented by lithium chloride (a selective inhibitor of GSK3ß) exposure or by GSK3ß-KD (a dominant-negative GSK3ß) overexpression. We also found that the activation of GSK3ß by HF-LPLI was due to the inactivation of protein kinase B (Akt), a widely reported and important upstream negative regulator of GSK3ß, indicating the existence and inactivation of Akt/GSK3ß signaling pathway. Moreover, the inactivation of Akt/GSK3ß pathway depended on the fluence of HF-LPLI treatment. Furthermore, vitamin c, a ROS scavenger, completely prevented the inactivation of Akt/GSK3ß pathway, indicating ROS generation was crucial for the inactivation. In addition, GSK3ß promoted Bax activation by down-regulating Mcl-1 upon HF-LPLI treatment. Taken together, we have identified a new and important proapoptotic signaling pathway that is consisted of Akt/GSK3ß inactivation for HF-LPLI stimulation. Our research will extend the knowledge into the biological mechanisms induced by LPLI.

Subcellular translocation of the eGFP-tagged TRPL channel in Drosophila photoreceptors requires activation of the phototransduction cascade.

Signal-mediated translocation of transient receptor potential (TRP) channels is a novel mechanism to fine tune a variety of signaling pathways including neuronal path finding and Drosophila photoreception. In Drosophila phototransduction the cation channels TRP and TRP-like (TRPL) are the targets of a prototypical G protein-coupled signaling pathway. We have recently found that the TRPL channel translocates between the rhabdomere and the cell body in a light-dependent manner. This translocation modifies the ion channel composition of the signaling membrane and induces long-term adaptation. However, the molecular mechanism underlying TRPL translocation remains unclear. Here we report that eGFP-tagged TRPL expressed in the photoreceptor cells formed functional ion channels with properties of the native channels, whereas TRPL-eGFP translocation could be directly visualized in intact eyes. TRPL-eGFP failed to translocate to the cell body in flies carrying severe mutations in essential phototransduction proteins, including rhodopsin, Galphaq, phospholipase Cbeta and the TRP ion channel, or in proteins required for TRP function. Our data, furthermore, show that the activation of a small fraction of rhodopsin and of residual amounts of the Gq protein is sufficient to trigger TRPL-eGFP internalization. In addition, we found that endocytosis of TRPL-eGFP occurs independently of dynamin, whereas a mutation of the unconventional myosin III, NINAC, hinders complete translocation of TRPL-eGFP to the cell body. Altogether, this study revealed that activation of the phototransduction cascade is mandatory for TRPL internalization, suggesting a critical role for the light induced conductance increase and the ensuing Ca2+ -influx in the translocation process. The critical role of Ca2+ influx was directly demonstrated when the light-induced TRPL-eGFP translocation was blocked by removing extracellular Ca2+.

[Laser radiation to correct disorders of blood albumin transport in severe mechanical trauma]. / Korrektsiia lazernym izlucheniem narushenii transportnoi sposobnosti al'bumina krovi pri tiazheloi mekhanicheskoi travme.

The paper provides evidence for that it is expedient to perform multi-stage intravascular low-intensity laser blood radiation in patients with severe mechanical trauma and massive blood loss in the early posttraumatic period. The use of laser radiation at a wavelength of 632.0 nm by inserting a disposable light guide (its end power was 1.5-2.0 mW, and the duration of a session--30 min) as part of complex therapy in this group of patients romotes the increase of plasma albumin transport ability and the general stimulation of natural detoxifixation mechanisms.