UVA Exposure Research Articles

Seaberry (<i>Hippophae rhamnoides L.</i>) and Water Lily (<i>Nymphaeaceae</i>) Extracts Protect Human Skin against Blue Light, Environmental Pollutants and UV-A Irradiations in an <i>Ex Vivo</i> Model System

Background: The skin is the outer shell of the mammalian body and it is continuously exposed to a large spectrum of external stimuli including sun irradiation and atmospheric pollutants. These factors present deleterious effects on the cutaneous compartment by altering the skin barrier functions and accelerating the aging of the skin. Objectives: The goal of this study was to investigate the activity of Seaberry and Water Lily extracts, here called Dermina complex, against different external stresses. Methods: Human skin explants were exposed to different stimuli including delipidation by organic solvents, blue light, atmospheric pollutants and UV-A. The activity of the Seaberry and Water Lily extracts was assessed by immunohistochemistry and by biochemical assays. Results: We showed that Dermina complex prevents the delipidation-induced filaggrin decrease, suggesting that these plant extracts exhibited barrier function protecting properties. Also, we observed that Dermina complex showed an antioxidant and DNA protection activity by decreasing the activated form of Nrf2, the oxidized proteins and the formation of γ-H2AX induced upon stress conditions. The Dermina complex also decreased the pro-inflammatory cytokine IL-1 alpha released in the culture medium following atmospheric pollutants and UV-A exposure confirming its anti-inflammatory activity. Moreover, Dermina complex reduced the blue light-induced overexpression of opsin 3, indicating that its skin protection activities may be due, in part, to filter property against environmental stresses. Conclusions: Dermina complex shows a protective activity of the skin against different environmental stresses and these extracts may be able to slow down the aging process of the cutaneous compartment.

Exogenous Polyamine Treatment Prevents UV-A Induced Cellular Damage and Photoinhibition of Photosystem II in Lichen Physcia semipinnata (J. F. Gmel.) Moberg

In this study the protection ability of polyamines on Photosystem II (PSII), the proline content of the cells and necrosis formation induced by Ultraviolet-A irradiation were studied. Although 24 h Ultraviolet-A exposure decreased the photosynthetic quantum yield (Fv/Fm) ratio in Physcia semipinnata (J. F. Gmel.) Moberg, thalli that treated with 1 mM polyamine, especially spermidine and spermine, were slightly influenced by the UV-A exposure. It was also found that spd and spm treated samples had lower proline content than putrescine (put)-treated samples. Moreover, It was found that the samples that were treated with spd and spm had lower percentage of DNA damage than put-treated samples and non-treated groups. In this study, Ultraviolet-A induced DNA damage detected by acridine orange/ethidium bromide staining assay. The DNA damage in thalli of P. semipinnata was detected in situ in phycobiont layer after 24 h of Ultraviolet-A exposure, in contrast, polyamine-treated samples had a lower rate of necrotic cells.

Synthesis of antibacterial N‐halamine acryl acid copolymers and their application onto cotton

ABSTRACTA series of N‐halamine copolymers are synthesized by reacting hydantoinylacrylamide (HA) with acrylic acid (AA) in several mole ratios. The synthesized copolymers are characterized by Fourier transform infrared (FTIR) and1H NMR and coated onto cotton fabrics. The coated cotton fabrics are rendered antibacterial by chlorinating with dilute household bleach solution. The ultraviolet (UVA) light stabilities of the resulting copolymers on the coated cotton fabrics are investigated; the results show that UVA exposure has minimal effect on the structure of the copolymers but can cause partial loss of the chlorine loading on the coated cotton fabrics. The coated cotton fabrics exhibit excellent antibacterial efficacies achieving inactivations of about six logs ofStaphylococcusaureusandEscherichiacoliO157:H7 within 1 min of contact time. The cotton fabrics coated with the copolymers would best be employed in disposable fabric applications because of lack of washing fastness due to the weak bonding interaction between the AA unit and cellulose. © 2018 Wiley Periodicals, Inc. J. Appl. Polym. Sci.2019,136, 47426.

The Impact of Sunlight on Skin Aging

In this review, we summarize and discuss the current literature on photoaging. We focus on the molecular effects of UV radiation, and the histological and clinical features of chronically sun-exposed skin. The expression of numerous proteins is altered in habitually sun-exposed skin, including sirtuins and hedgehog signaling proteins. Pharmacological manipulation of the levels of these proteins can potentially downregulate the photoaging process. Mitochondrial and nuclear DNA damage, reactive oxygen species (ROS) production, and altered gene expression patterns are critical components of both photoaging and chronological aging. Chronic UVA and UVB exposure result in the photoaged phenotype through distinct molecular mechanisms, with UVA being primarily ROS-mediated and UVB being DNA damage-mediated. Molecular effects result in an altered dermal and epidermal environment that manifests as the photoaged phenotype. Importantly, skin phototype and environmental influences impact individual responses to chronic UV radiation.

Cytoprotective effect of a functional antipollutant blend through reducing B[a] P-induced intracellular oxidative stress and UVA exposure.

Benzo[a]pyrene (B[a]P) is a ubiquitous environmental pollutant that reacts with skin and induces intracellular oxidative stress through reactive oxygen species (ROS) accumulation. The antipollution properties of natural extracts, especially including antioxidants, for inhibiting ROS in cells are gaining importance, in addition to the anticancer effects attributed to them. In this study, a commercial functional antipollutant blend of plant extracts consisting of ellagic acid standardized Punica granatum peel extract, Sambucus nigra fruit extract, Prunus cerasus seed extract, and hydrolyzed wheat protein with high antioxidant properties and UV damage-protective properties attributed to each one was investigated. The cytoprotective effect of this functional antipollutant blend was determined by ROS assay through reducing the level of intracellular ROS induced by B[a]P as an oxidative stress factor in human neonatal keratinocytes and fibroblast cells. In addition, the cytoprotective effect of the functional antipollutant blend after UVA exposure was also determined. It is shown that the oxidative damage induced by B[a]P and UVA, which are the most abundant factors of chemical and physical pollution, would be prevented by the functional antipollutant blend. Thus, it can be concluded that this antipollutant functional blend may offer a promising ingredient for the cosmetic industry's skincare products.

Photoreactivation of fungal spores in water following UV disinfection and their control using UV-based advanced oxidation processes

The occurrence of repair system in microorganisms after ultraviolet (UV)-induced damage to them evokes concern regarding the effectiveness of UV disinfection. Most studies focus on the repair of bacteria, but little research has been conducted on the repair of fungi in water. This study aimed to investigate the photoreactivation and dark repair properties of three dominant genera of fungal spores (Trichoderma harzianum, Aspergillus niger, and Penicillium polonicum) isolated from groundwater. UV-based advanced oxidation processes (AOPs) (including UV/peroxymonosulfate and UV/hydrogen peroxide) were used to control their photoreactivation. The results demonstrated that the three genera of fungal spores inactivated by UV (254 nm) exhibited different levels of photoreactivation under UVA (365 nm) exposure, and the photoreactivation percentage showed that T. harzianum (51.35%) ＞A. niger (29.07%) ＞P. polonicum (9.01%). The photoreactivation process of fungal spores was well described by the first-order model. T. harzianum had lower photoreactivation percentage but a more rapid initial photoreactivation process than E. coli. Higher UV dosages significantly decreased the photoreactivation percentage of fungal spores. However, dark repair was insignificant following UV disinfection for all the three genera of fungal spores. After treatment by UV-based AOPs, the fungal spores exhibited the same photoreactivation trend as those treated by UV alone. However, both the maximum survival ratios and photoreactivation rate constants were reduced to varying degrees. This study revealed the photoreactivation rule of dominant genera of fungi isolated from groundwater following UV treatment alone and UV-based AOPs, which is effective for controlling fungi in water.

Sub-lethal UV radiation during early life stages alters the behaviour, heart rate and oxidative stress parameters in zebrafish (Danio rerio)

Environmental UV radiation in sufficient doses, as a possible consequence of climate change, is potent enough to affect living organisms with different outcomes, depending on the exposure life stage. The aim of this project was to evaluate the potentially toxic effects of exposure to sub-lethal and environmentally relevant doses of UVA (9.4, 18. 7, 37.7 J/cm2) and UVB radiation (0.013, 0.025, 0.076 J/cm2) on the development and behaviour in early life stages (4.5–5.5 h post fertilization, hpf) of the zebrafish (Danio rerio). The used doses were all below the median lethal dose (LD50) and caused no significant difference in survival, deformities, or hatching between exposed and control groups. Compared to controls, there were transient UVA and UVB exposure effects on heart rate, with dose dependent reductions at 50 hpf, and at 60 hpf for UVA only. The UVB exposure caused an increasing trend in reactive oxygen species (ROS) formation at the two highest doses, even though only significant at 120 hpf for the second highest dose. Both UVA and UVB caused an increasing trend in lipid peroxidation (LPO) at the highest doses tested at 72 hpf. Furthermore, UVA exposure led to significant reductions in larval movement following exposure to the two highest doses of UVA, i.e., reduction in the time spent active and the total distance moved compared to control at 100 hpf, while no effect on the swimming speed was observed. The lowest dose of UVA had no effect on behaviour. In contrast, the highest dose of UVB led to a possible increase in the time spent active and a slower average swimming speed although these effects were not significant (p = 0.07). The obtained results show that UV doses below LD50 levels are able to cause changes in the behaviour and physiological parameters of zebrafish larvae, as well as oxidative stress in the form of ROS formation and LPO. Further testing is necessary to assess how this type of radiation and the effects observed could affect fish population dynamics.

Effects and Mechanism of Nicotinamide Against UVA‐ and/or UVB‐mediated DNA Damages in Normal Melanocytes

Melanoma incidences are increasing rapidly, and ultraviolet (UV) radiation from the sun is believed to be its major contributing factor. UV exposure causes DNA damage in skin which may initiate cutaneous skin cancers including melanoma. Melanoma arises from melanocytes, the melanin-producing skin cells, following genetic dysregulations resulting into hyperproliferative phenotype and neoplastic transformation. Both UVA and UVB exposures to the skin are believed to trigger melanocytic hyperplasia and melanomagenesis. Melanocytes by themselves are deficient in repair of oxidative DNA damage and UV-induced photoproducts. Nicotinamide, an active form of vitamin B3 and a critical component of the human body's defense system has been shown to prevent certain cancers including nonmelanoma skin cancers. However, the mechanism of nicotinamide's protective effects is not well understood. Here, we investigated potential protective effects and mechanism of nicotinamide against UVA- and/or UVB- induced damage in normal human epidermal melanocytes. Our data demonstrated an appreciable protective effect of nicotinamide against UVA- and/or UVB- induced DNA damage in melanocytes by decreasing both cyclobutane pyrimidine dimers and 8-hydroxy-2'-deoxyguanosine levels. We found that the photoprotective response of nicotinamide was associated with the activation of nucleotide excision repair genes and NRF2 signaling. Further studies are needed to validate our findings in in vivo models.

In Vitro Amoebicidal Activity of Titanium Dioxide/UV-A Combination Against Acanthamoeba.

To assess the amoebicidal effect of titanium dioxide (TiO2)/UV-A combination against Acanthamoeba sp trophozoites and cysts. The amoebicidal effect of the TiO2/UV-A combination was tested on trophozoites and cysts of clinical isolates of Acanthamoeba hatchetti and Acanthamoeba sp genotype T4, obtained from two severe cases of ulcerative keratitis. Samples of cultured Acanthamoeba were transferred to a 96-well plate. We tested the effect of sterile water (blank control), TiO2 alone, UV-A alone, TiO2 and additional UV-A exposure, chlorhexidine 0.02% alone, chlorhexidine 0.02% and TiO2, chlorhexidine and UV-A, chlorhexidine 0.02% and TiO2, and additional UV-A exposure. Cell viability assessment was done using the trypan blue dye exclusion method. The combination of TiO2 with UV-A demonstrated antitrophozoite and anticyst activity (P < 0.05). This in vitro study showed a synergistic effect of the association of chlorhexidine with TiO2 and UV-A on cysts (P < 0.001). Given the in vitro synergistic effectiveness of the association of chlorhexidine with TiO2 and UV-A against cysts, the treatment of Acanthamoeba keratitis could be improved by this new therapeutic approach.

Keratinocyte stem cells are more resistant to UVA radiation than their direct progeny.

The epidermis undergoes constant renewal during its lifetime. This is possible due to a special population of keratinocyte stem cells (KSCs) located at the basal layer. These cells are surrounded by their direct progeny, keratinocyte progenitors or transient amplifying cells (TAs), which arise from cell division. Skin is exposed every day to sun radiation; in particular, UVA radiation penetrates through the epidermis and induces damage to KSCs and TAs. Although keratinocytes in the basal layer are the most likely skin carcinomas and/or photoaging cells of origin, surprisingly few studies have addressed the specific responses of these cells to UV radiation. In this study, we showed for the first time that keratinocyte stem cells were more resistant to UVA irradiation than their direct progeny, transient amplifying cells. Using both the MTT assay and clonogenic assay, we found that KSCs were more photo-resistant compared to TAs after exposure to different doses of UVA (from 0 to 50 J/cm2). Moreover, KSCs had a greater ability to reconstruct human epidermis (RHE) after UVA exposure compared with TAs. Finally, investigations of DNA repair using the comet assay showed that DNA single-strand breaks and thymine dimers were repaired quicker and more efficiently in KSCs compared with TAs. In a previous work, we showed that the same stem cell population was more resistant to ionizing radiation, another carcinogenic agent. Collectively, our results combined with other observations demonstrate that keratinocyte stem cells, which are responsible for epidermal renewal throughout life, are equipped with an efficient arsenal against several genotoxic agents. Our future work will try to identify the factors or signaling pathways that are responsible for this differential photo-sensitivity and DNA repair capacity between KSCs and TAs.

TLR4-mediated activation of the ERK pathway following UVA irradiation contributes to increased cytokine and MMP expression in senescent human dermal fibroblasts.

Exposure to ultraviolet (UV) radiation is a major contributing factor to premature aging (photoaging) and skin cancer. In vitro models of cellular senescence have proven to be very useful for the study of slow and progressive accumulation of damage resulting in the growth arrest of aging skin cells. In this study, we compared UVA-induced cellular responses in non-senescent (NS) vs. senescent (S) human dermal fibroblasts (HDFs). HDFs were irradiated with a single dose of UVA (7.5 J/cm2) and QuantSeq 3' mRNA sequencing was performed to assess differential gene expression. Both NS and S HDFs expressed similar numbers of differentially expressed genes, although distinct sets of genes were differentially expressed between the two groups. Higher expression of matrix metalloproteinases (MMPs) and Toll-like receptor (TLR) pathway genes, such as TLR4, MyD88, and CXCL-8, was detected in S HDFs as compared with NS HDFs, and UVA exposure led to a downregulation of collagen genes, such as COL8A2 and COL5A3. Consistent with gene expression profiling, enhanced IL-6 and IL-8 secretion was observed in S HDFs compared with NS HDFs, in response to UVA. Furthermore, we show that TLR4-mediated ERK pathway is responsible for the UVA-mediated mitochondrial dysfunction as well as increased secretion of MMP-1 and IL-8 in S HDFs. Taken together, our results demonstrate the UVA-induced common and distinct molecular patterns of cellular responses between NS and S HDFs and suggest TLR4/ERK pathways as candidate targets to reduce senescent phenotypes.

Effects of titanium dioxide nanoparticles on the inhibition of cellular activity in human Tenon's fibroblasts under UVA exposure.

To investigate the effect of titanium dioxide (TiO2) nanoparticles on the inhibition of the in vitro cellular activity of human Tenon's fibroblasts (HTFs) under UVA exposure. The effects of TiO2 nanoparticles on human Tenon's fibroblasts were evaluated after 1, 4, 6, and 24h of exposure to UVA at levels of 2.5, 5.0, and 10J/cm2. The methyl thiazolyl tetrazolium (MTT) assay was performed to measure the suppression of cellular metabolic activity. The lactate dehydrogenase (LDH) assay was performed to determine the extent of cell membrane damage. Flow cytometric analysis and inverted phase-contrast and electron microscopy were performed. The scratch wound assay was performed to visualize suppression of cellular migration. MTT assay values were similar between the UVA-exposed groups and the control group without UVA exposure. However, the combined exposure of TiO2 nanoparticles and UVA exposure induced significant dose-dependent inhibition of cellular viability and damage to HTFs, especially at concentrations of TiO2 equal to or greater than 100μg/mL and 2.5J/cm2 of UVA irradiation. Changes in cellular morphology increased in a dose-dependent pattern with a TiO2 concentration greater than 100μg/mL under UVA exposure. At a TiO2 concentration of 150μg/mL, damage to the cellular morphology of the HTFs was significantly increased, and nanoparticles were seen inside of the cytoplasm in the affected HTFs exposed to UVA. There was a significant reduction of cellular migration at TiO2 concentrations higher than 150μg/mL. TiO2 nanoparticles inhibited the cellular activity of HTFs under UVA irradiation and showed potential for use to prevent the wound scarring of Tenon's fibroblasts. Further studies will be necessary to determine the optimal concentration of TiO2 nanoparticles and UVA exposure dose for clinical applications.

Keratinocyte Motility Is Affected by UVA Radiation-A Comparison between Normal and Dysplastic Cells.

UVA radiation induces multiple and complex changes in the skin, affecting epidermal cell behavior. This study reports the effects of UVA exposure on normal (HaCaT) and dysplastic (DOK) keratinocytes. The adherence, spreading and proliferation were investigated by time-lapse measurement of cell layer impedance on different matrix proteins. Prior to UVA exposure, the time required for adherence and spreading did not differ significantly for HaCaT and DOK cells, while spreading areas were larger for HaCaT cells. Under UVA exposure, HaCaT and DOK cells behavior differed in terms of movement and proliferation. The cells’ ability to cover the denuded surface and individual cell trajectories were recorded by time-lapse videomicroscopy, during wound healing experiments. Dysplastic keratinocytes showed more sensitivity to UVA, exhibiting transient deficiencies in directionality of movement and a delay in re-coating the denuded area. The actin cytoskeleton displayed a cortical organization immediately after irradiation, in both cell lines, similar to mock-irradiated cells. Post-irradiation, DOK cells displayed a better organization of stress fibers, persistent filopodia, and new, stronger focal contacts. In conclusion, after UVA exposure HaCaT and DOK cells showed a different behavior in terms of adherence, spreading, motility, proliferation, and actin cytoskeleton dynamics, with the dyplastic keratinocytes being more sensitive.

The role of phototherapy in the surgical treatment of vitiligo: a systematic review.

Vitiligo is frequently treated with the combination of phototherapy and melanocyte transplantation. However, the additional benefit of phototherapy is unclear. Moreover, the optimal type and regimen of phototherapy are unknown. The objective of this systematic review was to identify whether phototherapy improves the outcome of melanocyte transplantation in vitiligo. We searched and screened for eligible studies in the databases of MEDLINE, EMBASE and CENTRAL. We included all clinical studies investigating melanocyte transplantation combined with phototherapy. After screening and selection of abstracts and full-texts, we found 39 eligible clinical studies with 1624 patients. The eligible studies investigated several phototherapy modalities, such as NBUVB (n=9), PUVA (n=19), UVA (n=1), MEL (n=4) and active sunlight exposure (n=9). Four studies directly compared phototherapy versus no phototherapy and two studies confirmed the benefit of phototherapy for melanocyte transplantation. We found no significant differences in repigmentation in studies directly comparing phototherapy modalities. The overall quality of the studies was moderate to poor and high heterogeneity between studies was found. We found limited evidence that phototherapy improves the outcome of melanocyte transplantation in vitiligo. There is insufficient evidence to recommend a specific type or regimen of phototherapy. More studies should be performed investigating the additional benefit of different phototherapies and the preferred moment of phototherapy.

Distribution of oil solubilized β-carotene in stabilized locust bean gum powders for the delivery of orange colorant to food products

Galactomannan gum (locust bean gum; LBG) was used to distribute and stabilize vibrantly orange colored oil-solubilized β-carotene (βC) into dry food powders. Powders were prepared by homogenization of oil solubilized βC into hydrated LBG, followed by addition of stabilizers (lecithin, soy protein hydrolysate, and α-tocopherol), freeze drying, and course grinding. This produced dry and light powders that display mild adherance to the skin and show no color or phase separation. ATR-IR spectra indicate the mannan powders create a highly dispersed lipophilic phase. Color stability testing showed LBG-only powders had short-term thermal stability (1 h at 80 °C) but little effect against UV exposure and storage. Incorporation of lecithin and soy protein hydrolysate both enhanced the orange color stability against UV-a exposure and inclusion of α-tocopherol resulted in full protection through the thermal, UV-a, UV-c, and storage. Formulation of similar powders inclusive of food flavorings may be of interest to industries targeting synthetic colorant replacement.

Light-enhanced antibiotic activity of Brazilian medical plants (Croton campestris A, Ocimum gratissimum L and Cordia verbenaceae DC)

Background: Although broad-band ultraviolet (UV)-A has been described as a therapeutic option for various skin diseases, there are few studies investigating the efficacy of UV-A irradiation in treating diseases related to infectious agents. Objective: Evaluate the light-enhanced antibacterial activity of Brazilian medical plants, Croton campestris A (Euphorbiaceae), Ocimum gratissimum L. (Lamiaceae), and Cordia verbenaceae DC (Boraginaceae). Methods: Hexane extracts of Croton campestris A., Ocimum gratissimum L., and Cordia verbenaceae DC were assayed using a UV-A exposure method against strains of Staphylococcus aureus and Escherichia coli. Assays were performed in triplicate with and without exposure to UV-A radiation to test for light-activated or lightenhanced antibacterial activity. Results: All extracts showed activity against the S. aureus strain. Extracts of O. gratissimum and C. verbenaceae were the most active after exposure to UV-A light, with an increase in antibacterial activity of 140 and 100%, respectively. No extract showed light-activated antibacterial activity against E. coli. Conclusion: C. campestris, O. gravissimum and C. verbenaceae showed light-enhanced antibiotic activity. This suggests that phytochemical investigations may be warranted.

Oocyte atresia in WT (HdrR) and P53 (-/-) medaka (Oryzias latipes) exposed to UVA

The negative effects of ambient ultraviolet (UVA) on the water environment have been recently highlighted; UVA can create deleterious effects by stimulating stress on pelagic organisms. Little is known about UVA effects on oocyte characteristics of female fish. In the present study we explored the effects of exposure to ecologically relevant levels of simulated UVA radiation on ovaries of two major strains WT (HdrR) and P53 (-/-) of medaka (Oryzias latipes) mature female. Fish were assigned to control and three UVA-exposed groups as (15 min, 30 min, and 60 min/day) for three days and sample selection was 24 h and 14 days after exposure. Histological alterations and oocyte atresia percentage were analyzed in the UVA-exposed fish compared to control. Alteration comprised hyperthrophied follicular cells with increased thickness, breakdown of egg chorion (zona radiata), damage of cortical alveoli, and distorted nucleus and cytoplasm. The atresia percentages significantly increased with higher UVA exposure dose and time for both the wild type and the p53 deficient fish. The wild type displayed significantly higher oocyte atresia percentage than the p53 mutant. These results suggested that UVA exposure provoked histological alterations in both p53 and WT medaka oocytes leading to follicular atresia, which reduce female reproductive ability and larval production. UVA oocyte response showed p53 dependent and independent histological alteration, however, the p53 mutant was less sensitive to UVA than the wild type in medaka fish.

Combined effect of Benzophenone-2 and ultraviolet radiation promote photogenotoxicity and photocytotoxicity in human keratinocytes

Benzophenone-2 (BP2), a common ingredient of sunscreens formulation is widely used as UV filter. We have assessed the photogenotoxic and photocytotoxic potential of BP2. Photostability test showed that BP2 is unstable under UV exposure. Cell proliferation assay revealed that viability of HaCaT cells significantly reduced under UVA, UVB and sunlight exposure. DCF fluorescence intensity proved intracellular ROS generation capacity of BP2 under sunlight, UVA and UVB irradiation. Photodynamic degradation of guanine base of DNA is promoted by BP2 under UV treatment. Genotoxicity assessed by comet assay, showed that photosensitized BP2 enhanced DNA damage, which is measured in term of % tail DNA and olive tail moment. Genotoxic potential of BP2 was further validated with photomicronuclei assay. Photogenotoxicity of BP2 was lastly confirmed by formation of CPDs (Cyclo butane pyrimidine dimmers). DNA damage induced by BP2 was irreversible and extended incubation periods (6–12 h) not favored the recovery from damaged DNA. JC 1 staining showed significant reduction in mitochondrial membrane potential. Membrane integrity compromisation of HaCaT cells was established by AO (Acridine orange), EtBr (Ethidium bromide) staining and confirmed with sub G1 population of cell cycle. Thus, results suggest that BP2 should be avoided in topical application for safe sunscreen practices.

Global Profiling of the Oxidative Stress Induced Effects on RNA Modifications by Liquid Chromatography‐Tandem Mass Spectrometry

The goal of this work was to profile the oxidative damage exerted by ultraviolet‐A radiation (UVA, 370 nm) or H2O2 on RNA modifications by liquid chromatography‐tandem mass spectrometry (LC‐MS/MS). Oxidative damage to RNA impacts biological systems due to its association with metabolic, mitochondrial, neurological diseases and cancer.1,2 In the presence of photosensitizer (e.g., riboflavin), UVA radiation induces the production of reactive oxygen species (ROS, O2•, HOO•, HO•) in vivo.3 Likewise, H2O2 generates ROS under physiological conditions, and formation of radicals is accelerated via Fenton reactions in the presence of catalysts (e.g., Fe2+, Cu2+).4Oxidation pathways of canonical nucleosides are well documented.5 However, the effect of ROS on modified ribonucleosides is not well known, and information about RNA damage is largely extrapolated from studies on DNA. More than 160 modified nucleosides have been reported in various types of RNA of different organisms.6 E. coli with 46 modifications in its tRNA serve as a good model system to investigate the stress‐induced effects. Apart from the known oxidative products arising from canonical nucleosides, we have documented a list of modified nucleosides that are adversely affected by UVA or H2O2. To understand the oxidative environment generated by UVA exposure, we exposed E. coli total tRNA with or without a photosensitizer for defined period of time. In a similar manner, the effect of H2O2 on RNA modifications was investigated by incubating E. coli total tRNA and H2O2, with or without the presence of transition metal ion catalysts. The oxidative damage observed on various modifications was measured by subjecting it to LC‐MS/MS following complete hydrolysis of tRNA. We will present data that describes the susceptibility of various functional groups of these modifications and underlying patterns. Further, these studies also document a range of RNA photoproducts and chemical alterations arising due to the oxidative conditions.Support or Funding InformationFinancial support of this work was provided by the NSF (CHE1507357) and DTRA (HDTRA1‐15‐1‐0033), both to Patrick A. Limbach, and the University of Cincinnati.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Effects of Repeated UVA Irradiation on Human Skin Fibroblasts Embedded in 3D Tense Collagen Matrix.

Skin photoaging is caused by cumulative UVA exposure that leads to dermal matrix alterations associated with impaired fibroblast functions. In this study, we evaluated the effects of repeated UVA irradiation on mechanically stressed fibroblasts which were embedded in 3D tense collagen matrix. By comparison to 2D monolayer culture, we investigated the expressions of alpha-smooth muscle actin (α-SMA) cytoskeleton and α2 subunit of integrin receptors, as well as the collagen metabolism, focusing to MMP-1 and collagen type-I expressions. We found that UVA exposure reduces collagen levels in both culture conditions. However, concerning integrin α2 and α-SMA expression, UVA irradiation had no effect on 2D culture, whereas in tense 3D culture, it had an inhibitory effect. In UVA-irradiated 3D culture, fibroblasts acquired elongated shape and lost their dynamic interaction with collagen fibers through a decrease in integrin α2 and α-SMA. Fibroblast responses to UVA irradiation were different in 2D versus 3D environment, highlighting the importance of collagen environment in the regulation of mechanical activities. The behavior of fibroblast upon mechanical stimulation closely mimics stressed extracellular environment. The model of UVA-irradiated fibroblasts cultured in tense 3D collagen gel illustrated the in vivo situation of both mechanically stressed and photoaged human skin.