POTENTIAL EFFECTS OF ALTERED SOLAR ULTRAVIOLET RADIATION ON HUMAN SKIN CANCER

There is highly significant evidence that non-melanoma skin cancers are primarily due to chronic repeated exposure to solar ultraviolet radiation, and that there is a significant, although somewhat different relationship between solar radiation and the development of cutaneous malignant melanoma. Recent experimental and epidemiologic studies show that the biologically most effective UVR wavelengths are in the segment of the solar UVR spectrum that would be significantly augmented by decreases in stratospheric ozone content. A recent report on measurements of column ozone changes in the stratosphere has shown that in the past 18 yrs, there has been an ozone decrease between 2 and 3%, greater in the winter months, and somewhat differing with latitude in the Northern Hemisphere. Calculations of the relationship of ozone decrease to increase in biologically effective UVR show great dependence on the biologic action spectrum assumed. Based on extensive epidemiologic studies of skin cancer incidence, it appears that the estimated increase in biologically effective UVR due to the measured ozone decreases in the past (almost) two decades are not likely to be the cause of the sharp increase in skin cancer incidence which have been observed. Most likely these increases in incidence are the result of increasing personal exposure, due to striking changes in personal behavior that have taken place for social reasons. However, there is every reason to believe that increases in biologically effective UVR due to stratospheric ozone decreases will have significant impact on human skin cancer incidence in the future.     

PROTECTIVE EFFECT OF SELENIUM AND ZINC ON UV-A DAMAGE IN HUMAN SKIN FIBROBLASTS

Ultraviolet A radiation participates in cytotoxicity and carcinogenesis of the skin by a mechanism involving the generation of reactive oxygen species. Endogenous antiradical defense systems utilize metalloenzymes including Se-dependent glutathione peroxidase and Cu and Zn superoxide dismutase. The aim of the present work was to determine the protective effect of two trace elements, Se and Zn, on cultured human diploid fibroblasts exposed to UV-A radiation (broad-spectrum source with a maximum intensity at 375 nm). Selenium in the culture medium (0.1 mg/L) in the form of sodium selenite increased the synthesis and activity of glutathione peroxidase by 60.5% in the absence of exposure to UV-A radiation and by 35% after irradiation with 5 J/cm² ( P = 0.043). The presence of this element significantly increased the survival of UV-A-irradiated fibroblasts ( P < 0.0001). This confirms the essential role of Se in the detoxifying activity of the enzyme. In addition, thiobarbituric acid-reacting substances (TBAR), which are lipid peroxidation markers, decreased in the presence of exogenous Se:—19% and -22% without irradiation and after irradiation with 5 J/cm² ( P = 0.056). When Zn was added at the dose of 6.5 mg/L as ZnCl₂, fibroblasts subjected to oxidizing stress induced by UV-A were protected from cytotoxicity ( P <0.0001). The TBAR production decreased significantly: -33% without irradiation and -34% after irradiation with 5 J/cm² ( P = 0.008). Superoxide dismutase activity, however, decreased after supplementing with Zn: - 26% without irradiation and - 20% after UV-A irradiation ( P = 0.017). The antioxidant properties of Zn are thus apparently independent of superoxide dismutase activity.     

ACTION SPECTRUM FOR ANTHRACENE-INDUCED PHOTOSENSITIZATION OF HUMAN SKIN

Abstract— The action spectrum for photosensitization by topically applied anthracene was determined in human volunteers. Spectral reactivity was demonstrated in the range between 320 and 380 nm, with peak activity at around 360 nm. Three distinct inflammatory responses viz. immediate transient erythema, delayed erythema, and wealing were evoked following exposure to effective wavelengths. The action spectra for these responses were similar but the threshold doses were different. Prior treatment with a mast cell degranulating agent (codeine) abolished anthracene-UVA induced wealing but did not influence the erythema response. These findings suggest that photosensitized damage to cutaneous mast cells may be partially responsible for some of the observed inflammatory responses, but other sites of photochemical injury are also involved.     

SPECTROSCOPIC AND MICROSCOPIC CHARACTERISTICS OF HUMAN SKIN AUTOFLUORESCENCE EMISSION

To improve the understanding of human skin autofluorescence emission, the spectroscopic and microscopic characteristics of skin autofluorescence were studied using a combined fluorescence and reflectance spectroanalyzer and a fiber optic microspectrophotometer. The autofluorescence spectra of in vivo human skin were measured over a wide excitation wavelength range (350–470 nm). The excitation–emission matrices of in vivo skin were obtained. An excitation–emission maximum pair (380 nm, 470 nm) was identified. It was revealed that the most probable energy of skin autofluorescence emission photons increases monotonically and near linearly with increasing excitation photon energy. It was demonstrated that the diffuse reflectance, R, can be used as a first order approximation of the fluorescence distortion factor f to correct the measured in vivo autofluorescence spectra for the effect of tissue reabsorption and scattering. The microscopic in vitro autofluorescence properties of excised skin tissue sections were examined using 442 nm He–Cd laser light excitation as an example. It was demonstrated that the fluorophore distribution inside the skin tissue is not uniform and the shapes of the autofluorescence spectra of different anatomical skin layers vary. The result of this study confirms that the major skin fluorophores are located in the dermis and provides an excellent foundation for Monte Carlo modeling of in vivo autofluorescence measurements.     

PHOTOREPAIR OF PYRIMIDINE DIMERS IN HUMAN SKIN IN VIVO

Abstract— The exposure of human skin in vivo to UV radiation emitted from a sunlamp induces the formation of pyrimidine dimers. The number of dimers, as detected by UV-endonuclease, decreases following exposure of the UV–irradiated skin to visible wavelengths of light. These results suggest that humans possess a mechanism by which pyrimidine dimers are photorepaired upon illumination of human skin in vivo with visible light.     

THE VASCULAR RESPONSE OF HUMAN SKIN TO ULTRAVIOLET RADIATION

The vascular response of human skin to 300 nm (UV-B) and 254 nm (UV-C) ultraviolet radiation was assessed using the reflectance measurement of erythema and the technique of laser Doppler velocimetry. For both wavelengths, the increase in measured Doppler blood flux varied with the increase in erythema in a quadratic manner predicted by a simple model based on the principles of fluid mechanics. This suggests that the mean red blood cell velocity increases significantly in areas of UV-B and UV-C erythema. No qualitative difference in response to these two wavelengths was demonstrated, suggesting that the same blood vessels are involved in the causation of both UV-B and UV-C erythema.     

PYRIMIDINE DIMER FORMATION IN HUMAN SKIN

Cyclobutyl pyrimidine dimers are major photoproducts formed upon irradiation of DNA with ultraviolet light. We have developed a method for detecting as few as one pyrimidine dimer per million bases in about 50 ng of non-radioactive DNA, and have used this method to quantitate dimer yields in human skin DNA exposed in situ to UV. We found that UVA radiation (320–400 nm) produces detectable levels of dimers in the DNA of human skin. We also measured UVB-induced dimer yields in skin of individuals of differing sun sensitivity and found higher yields in individuals with higher UVB minimal erythema doses and greater sun sensitivity. These approaches should provide important information on damage induced in human skin upon exposure to natural or artificial sources of ultraviolet radiation.     

COLLAGEN ALTERATIONS IN CHRONICALLY SUN-DAMAGED HUMAN SKIN

Abstract The major histological characteristic of sun-damaged skin is the accumulation of an elastotic material that appears to replace collagen. This elastotic material consists primarily of elastin and histological studies suggest a large loss of collagen in the dermis of chronically sun-damaged skin. In this study, we examine the content and distribution of collagen and procollagen in sun-damaged human skin. The total collagen content of sun-damaged skin was 20% less than nonsolar-exposed skin (524 μ g collagen per mg total protein in sun-damaged skin and 667 μg collagen per mg total protein in nonsolar-exposed skin). In addition, there was a 40% decrease in the content of intact amino propeptide moiety of type III procollagen in sun-damaged skin (0.68 U per 50 mg wet weight) as compared to nonsolar-exposed skin (1.12 U per 50 mg wet weight). The data suggest that this change in collagen content is due to increased degradation. The distribution of collagen in sun-damaged skin was examined by indirect immunofluo-rescence. Mild digestion of sun-damaged skin with elastase removed the elastin and revealed the presence of collagen in the elastotic material. Therefore, the elastin appears to mask the presence of collagen fibers in the dermis of sun-damaged skin.     

LONG PERSISTENCE OF MONOFUNCTIONAL 8-METHOXYPSORALEN-DNA ADDUCTS IN HUMAN SKIN in vivo

Human skin can be persistently photosensitized by topical application of aqueous 8-methoxypsoralen plus immediate irradiation with a non-erythemogenic dose of wavelengths above 380 nm. Re-exposure of skin thus sensitized to broadband UV-A produces phototoxic erythema 72-120 h later. The persistence of the photosensitization was demonstrated by phototoxic erythema after re-exposure up to 15 days after the first sensitizing irradiation. According to the concept that the first exposure induces primarily psoralen monoadducts, we consider this an investigation of psoralen monoadduct persistence. In contrast to several earlier studies, this sensitive method indicates that psoralen monoadducts may remain in human skin in vivo for more than 2 weeks after formation.     

UVB-INDUCED PHOTOPEROXIDATION OF LIPIDS OF HUMAN LOW and HIGH DENSITY LIPOPROTEINS. A POSSIBLE ROLE OF TRYPTOPHAN RESIDUES

Ultraviolet radiation of the UVB region readily destroy tryptophan (Trp) residues of low (LDL) and high (HDL) density lipoproteins. The photooxidation of tryptophan residues is accompanied by the peroxidation of low and high density lipoproteins unsaturated fatty acids, as measured by the thiobarbituric acid assay. Moreover, low and high density lipoproteins are natural carriers of vitamin E and carotenoids. These two antioxidants are also rapidly bleached by UVB. The UVA radiation promotes neither tryptophan residue destruction nor lipid photoperoxidation. The redox cycling Cu2+ ions considerably increase lipid photoperoxidation. The synergistic action of photo and auto (Cu2(+)-induced) peroxidation induces marked post-irradiation modifications of apolipoproteins as illustrated by the degradation of most tryptophan residues after overnight incubation in the dark of pre-irradiated samples.     

药物抗氧化作用对DNA发光动力学行为的影响

本文应用CuSO₄-Phen-VC-H₂O₂-DNA化学发光体系观察药物对发光动力学行为的影响,研究了九种药物对DNA损伤的保护作用,并对其药物结构对抗氧化能力的影响进行了分析。发现药物在该系统中的抗氧化作用有三种类型,提示它们可能在体内不同的反应过程中起不同的作用。在该系统中具有抑制DNA损伤发光能力的药物,按其作用强弱依次为阿魏酸、丹参素、芦丁、茶多酚、SOD、丹参酮Ⅱ-A、丹皮酚、五味子乙素,具有延迟DNA损伤发光能力的药物,按其作用强弱依次为阿魏酸、芦丁、茶多酚、Trolox、丹参素、丹皮酚。分子结构中具有共轭大π键、羟基、羧基的药物具有较强抑制DNA损伤的能力,这可能与药物分子的氧化还原电位或自由基生成热等多种因素有关。     

ENHANCEMENT OF MUTAGENESIS AND HUMAN SKIN CANCER RATES RESULTING FROM INCREASED FLUENCES OF SOLAR ULTRAVIOLET RADIATION

Abstract— We present a new method for calculating the effects of reduction of atmospheric ozone upon induction of nonmelanoma skin cancer. These estimates are based upon several recent experimental improvements: a model for the atmospheric penetration of UV-B; measurements of the transmission of this radiation by human epidermis; a precise action spectrum for genetic effects (mutation) in Escherichia coli , which was corrected for finite slit width. The calculated radiation amplification factor or percent increase in exposure per one percent decrease in atmospheric ozone is constant at 1.7 for solar zenith angles = 70° and decreases only with larger values of this angle. Thus the estimated increase applies to all heavily populated areas. of the globe. The value is robust: it is almost the same when the albedo is reduced from 0.2 to 0.1 or when the epidermal transmission is assumed to be about fourfold greater.     

QUANTITATIVE ASSESSMENT OF CUMULATIVE DAMAGE FROM REPETITIVE EXPOSURES TO SUBERYTHEMOGENIC DOSES OF UVA IN HUMAN SKIN

Abstract— Daily exposures to relatively small suberythemogenic fluences of UVA (50–200 kj/m²) for 8 days resulted in cumulative morphological skin alterations indicative of early tissue injury. Histologically, irradiated skin revealed epidermal hyperplasia, inflammation and deposition of lysozyme along the dermal elastic fiber network. Sunburn cells were also present within the epidermis. These changes were quantified by image analysis and were found to be related to the cumulative UVA fluence. A long UVA waveband (UVAI, 340–400 nm) was as effective as a broad UVA band (320–400 nm), suggesting that these changes are induced by longer UVA wavelengths.     

EFFECTS OF UV IRRADIATION ON A LIVING SKIN EQUIVALENT

Abstract— The Living Skin Equivalent (LSE™) is an organotypic coculture composed of human dermal fibroblasts interspersed in a collagen-containing matrix and overlaid with human keratinocytes forming a stratified epidermis. The LSE has a dry, air-exposed epidermal surface suitable for the application of oils, creams and emulsions. These features suggested its feasibility as an in vitro skin model for studying the protective effects of sunscreens. Using the thiazolyl blue (MTT) conversion assay as a measure of mitochondrial function, the extent of cytotoxicity induced by various doses of UV-R (280–400 nm) or UV-A (320–400 nm) was evaluated in the LSE. The doses of UV radiation that caused 50% reductions in MTT conversion (UV-R₅₀ or UV-A₅₀) in different lots of LSE were 0.053 ± 0.021 J/cm² (n = 29) and 11.6 ± 4.9 J/cm² (n = 17) for UV-R and UV-A, respectively. The protective effects of an 8% homosylate standard and of five UV-A sunscreens, topically applied to the LSE, were determined and compared with their reported protection factors in human skin. Morphological changes and the release of proinflammatory mediators (interleukin-1-α, tumor necrosis factor-α and prostaglandin E2) implicated in UV-induced erythema were also demonstrated in the LSE exposed to UV-A or UV-B. The data suggest that the LSE can be used for studying the effects of U V radiation on skin and may have utility for assessing the efficacy of certain sunscreens against UV-B and UV-A.     

SYSTEMIC INFLUENCE OF PRE-IRRADIATION OF A LIMITED SKIN AREA ON UV-TUMORIGENESIS

Abstract— It has been established that chronic UV irradiation of mice produces a systemic effect. The animals become incapable of rejecting an implanted UV-induced tumor. A possible consequence of the induction of this systemic effect could be an enhancement of the de novo formation of tumors by chronic UV irradiation. We have therefore carried out an investigation to determine whether such an effect is demonstrable in an animal model. Hairless mice (Skh hr 1) were pre-irradiated for a few months with UV radiation while certain skin areas of the animals were shielded from the radiation. Subsequently, the initially shielded skin areas were chronically exposed to UV radiation, which resulted in the development of tumors in these skin areas. It was found that the formation of tumors in the initially shielded skin areas was enhanced by the pre-irradiation of the other skin areas. Thus, a systemic effect appeared to have influenced the development of tumors in the initially shielded skin areas.     

DIFFERENTIAL EFFECTS OF PHOTOACTIVE FURANYL COMPOUNDS ON VIRUS FUNCTIONS

Abstract— Five photoactive furanyl compounds were investigated for their activities against viruses. The two furanocoumarins used were 8-methoxypsoralen (8-MOP) and angelicin; two furanochromones, visnagin and khellin, and the furanoquinoline, dictamnine, were also used. The DNA-containing herpes virus murine cytomegalovirus (MCMV) and the RNA-containing togavirus, Sindbis virus, were the target viruses. All five compounds inactivated both viruses in the presence of UVA, although Sindbis virus was much less sensitive. The relative order of antiviral potency was 8-MOP > dictamnine > visnagin > angelicin > khellin. Dictamnine however was slightly more effective than 8-MOP against Sindbis virus. None of the treatments affected the structural integrity of MCMV, nor did they interfere with the normal transit of the virus into host cells or the localisation of the viral genome in the cell nucleus. Some early viral gene functions were expressed but the viruses did not replicate.     

PROOXIDANT and ANTIOXIDANT EFFECTS OF ASCORBATE ON PHOTOSENSITIZED PEROXIDATION OF LIPIDS IN ERYTHROCYTE MEMBRANES

Abstract— Continuous blue light irradiation of resealed erythrocyte ghosts at 37°C in the presence of uroporphyrin or protoporphyrin results in ¹O₂-mediated (azide inhibitable) lipid peroxidation and membrane lysis. Lipid peroxidation was assessed by thiobarbituric acid reactivity and by quantitation of total hydroperoxides, while lysis was measured in terms of trappedglucose–6-P release. Low concentrations of ascorbate, AH^- (e.g. 0.5 m M ). present at the start of irradiation, significantly enhanced the rates of lysis and peroxidation, whereas relatively high concentrations of AH^- (e.g. 15 m M ) inhibited both processes. By way of contrast. AH^- produced only a dose-dependent inhibition of the photoinactivation of lysozyme, added as an extramembranous target. No significant AH^-induced lipid peroxidation was observed in dark or light controls, plus porphyrin or minus porphyrin, respectively. Stimulation of peroxidation and lysis by low levels of AH^- was enhanced by added Fe(III), abolished by EDTA. but unaffected by catalase or superoxide dismutase. A plausible explanation for these results is as follows. At low concentrations of AH^- prooxidant activity is favored. Redox metal-mediated breakdown of photoperoxides occurs, which tends to amplify lipid peroxidation. Neither O₂^- nor H₂O₂ appears to be involved. At significantly high concentrations, AH^- acts predominantly as an antioxidant by intercepting ¹O₂ and/or sensitizer triplet, or by scavenging free radical intermediates of lipid peroxidation.     

ACTIVATION OF NF-KB IN HUMAN SKIN FIBROBLASTS BY THE OXIDATIVE STRESS GENERATED BY UVA RADIATION

We have examined the role of the nucleus and the membrane in the activation of nuclear factor (NF)-KB by oxidant stress generated via the UVA (320–380nm) component of solar radiation. Nuclear extracts from human skin fibroblasts that had been irradiated with UVA at doses that caused little DNA damage contained activated NF-KB that bound to its recognition sequence in DNA. The UVA radiation-dependent activation of NF-KB in enucleated cells confirmed that the nucleus was not involved. On the other hand, UVA radiation-dependent activation of NF-KB appeared to be correlated with membrane damage, and activation could be prevented by a-tocopherol and butylated hydroxytol-uene, agents that inhibited UVA radiation-dependent peroxidation of cell membrane lipids. The activation of NF-KB by the DNA damaging agents UVC (200–290nm) and UVB (290–320nm) radiation also only occurred at doses where significant membrane damage was induced, and, overall, activation was not correlated with the relative levels of DNA damage induced by UVC/UVB and UVA radiations. We conclude that the oxidative modification of membrane components may be an important factor to consider in the UV radiation-dependent activation of NF-KB over all wavelength ranges examined.     

APPLICATION OF LIPIDS IN PHARMACEUTICALS

Recent development of the application of lipids in pharmaceutical utilization is reviewed. We have paid particular emphasis to the phospholipid-based dispersion systems, particularly liposome and lipid microsphere, which have high potency for the utilization in drug delivery systems. Physical chemical properties related to physiological characteristics are widely reviewed. The pharmaceutical effects of free fatty acids are also prersented.