DNA damage after acute exposure of mice skin to physiological doses of UVB and UVA light

Solar ultraviolet (UV) radiation is an important risk factor in skin carcinogenesis. This has been attributed mainly to the UVB waveband because the high-energetic photons are capable of interacting with DNA and inducing DNA damage. Recently, UVA light has also gained increasing interest in relation to DNA alteration. Although UVA photons are less energetic than UVB, they comprise a major fraction of sunlight UV radiation and penetrate deep into the skin. The study was carried out to compare the acute effects of UVA and UVB light on SKH-1 mice in relation to DNA damage and associated parameters. Mice were exposed to UVA (10 and 20 J/cm(2)) or UVB (200 and 800 mJ/cm(2)) radiation. The number of DNA single-strand breaks (SSB) in lymphocytes, amount of phosphorylated histone H2AX (gamma-H2AX) and apoptosis or DNA fragmentation (TUNEL-positive cells) in skin sections and level of gamma-H2AX, activated caspase-3 and phosphorylated p53 in skin were evaluated after 4 and 24 h. SSB analyzed by alkaline comet assay were found to be 4 and 24 h following UVB and UVA treatment, respectively. TUNEL and gamma-H2AX-positive cell were observed only in UVB exposed animals at both time intervals. The level of activated caspase-3 and phospho-p53 was increased 24 h after UVA and UVB radiation and was more apparent in UVB treated mice. The results indicate that the mechanism of DNA damage caused by acute UVA exposure includes formation of SSB (oxidative damage), but not double-strand breaks.     

UV-induced DNA damage in human keratinocytes: quantitation and correlation with long-term survival

Ultraviolet (UV) radiation has a major role in the pathogenesis of skin cancer due to its capacity to induce immunosuppression and DNA damage in cells. In this study, we describe the use of a novel extra-long polymerase chain reaction (XL-PCR) assay for detection of UV-inducible DNA lesions in a human keratinocyte line (HaCaT cells). Ultraviolet B (UVB), in doses from 4 to 50 mJ/cm2 resulted in a linear increase in the number of DNA lesions in the genome [range 0.3 +/- 0.2 lesions-3.6 +/- 0.7 lesions (mean +/- SD)/10 kb]. At lower doses of UVB (<10 mJ/cm2), 89 +/- 13% lesions were repaired within 24 h of culture. At higher doses, more lesions remained unrepaired, but the repair efficacy expressed as a proportion of repaired lesions to the total amount of DNA lesions remained constant in the range 0-50 mJ/cm2. Moreover, we demonstrated a correlation between the dose of UV and cell survival. The D37 (dose that reduced clonogenic survival to 37%) of UVB equaled 19 mJ/cm2, corresponding to the introduction of 1.4 lesions/10 kb. In contrast to UVB, UVA1 irradiation neither induced measurable DNA damage nor induced cell death in the doses up to 15 J/cm2. In conclusion, the non-radioactive extra-long (XL)-based real-time (RT)-PCR assay system can be used to quantify the UV-induced DNA damage in intact cells. The DNA lesions detected by this assay are mainly induced by short-waved radiation in the UVB range, and unrepaired DNA lesions cause keratinocyte death or permanent cell-cycle block.     

Effect of ultraviolet irradiation on mast cell-deficient W/Wv mice

The effect of UV irradiation on the skin was investigated in (WB-W/+) X (C57BL/6J-Wv/+)F1-W/Wv mice, which are genetically deficient in tissue mast cells. Their congenic littermates (+/+) and normal albino mice (ICR or BALB/c) were used as controls. Mice were irradiated with 500 mJ/cm2 of UVB and the increment of ear thickness was measured before and 6, 12, and 24 h after irradiation. Ear swelling in W/Wv mice at 12 and 24 h after irradiation was significantly smaller than that in +/+ and ICR mice. In contrast, the number of sunburn cells formed 24 h after UVB irradiation (200 or 500 mJ/cm2) was similar in W/Wv, +/+ and ICR mice. On the other hand, when mice were treated with 8-methoxy-psoralen (0.5%) plus UVA irradiation (4 J/cm2) (topical PUVA), ears of W/Wv and BALB/c mice, which were both white in color, were thickened similarly 72 h after treatment, but less swelling was observed in +/+ mice, which were black in skin color. The amount of prostaglandin D2 (PGD2) in ears, determined by radioimmunoassay specific for PGD2, was elevated 3-fold in +/+ and ICR mice at 3 h after irradiation with 500 mJ/cm2 of UVB in comparison with basal level without irradiation. However, such elevation was not observed in W/Wv mice. These results suggest that mast cells play an important role in UVB-induced inflammation, and PGs from mast cells are responsible at least in part for the development of this reaction. However, neither mast cells nor PGs contribute to the sunburn cell formation and ear swelling response by PUVA treatment.     

Direct effects on proliferation,antigen expression and melanin synthesis of cultured normal human melanocytes in response to UVB and UVA light

BACKGROUND/PURPOSE: Ultraviolet (UV) radiation; induces a variety of responses in the skin, including tanning and inflammation, and may also act as a carcinogen. As epidermal melanocytes are seen as the major targets of UV light, the present study was conducted to evaluate the direct effects of UVA and UVB irradiation on melanocytes in vitro. METHODS: Normal human epidermal melanocytes (NHM) were exposed on 3 consecutive days to UVA (0.072-7.2 J/cm2) and UVB (7.2-48 mJ/cm2), respectively, and changes of morphology, cell number, melanin synthesis and antigen expression (APAAP technique) were determined 5 days after the first exposure. RESULTS: UVA radiation caused only minimal effects on NHM by slightly inducing expression of the activation marker HMB-45 and decreasing expression of the proliferation marker Ki-67. No changes of morphology, cell number or melanin synthesis were detectable with any of the applied doses. On the other hand, UVB radiation significantly induced dendrite formation and decreased the number of NHM in a dose-dependent manner (74% of the controls at 7.2 mJ/cm2, 64% at 14.4 mJ/cm2 and 28% at 36 mJ/cm2). Significant induction of the activation marker HMB-45 was found in parallel to decreased expression of the differentiation marker K.1.2.58. UVB doses >or=9.6 mJ/cm2 also resulted in significant downregulation of the proliferation marker Ki-67, confirming the data of the cell counts, and melanin content was increased in NHM (20% over the controls, P<0.01) after applying 7.2 mJ/cm2 UVB. CONCLUSION: Our results may suggest that the effect of UVB radiation in skin is due to direct activation of melanocytes, whereas skin tanning caused by UVA is mediated rather in an indirect way.     

Higher susceptibility to apoptosis following ultraviolet B irradiation of xeroderma pigmentosum fibroblasts is accompanied by upregulation of p53 and downregulation of Bcl-2

Apoptosis plays an important part as a defence mechanism in eliminating damaged cells. Among the complex factors which regulate apoptosis, the p53 tumour suppressor protein which is induced by DNA damage has been suggested to play a crucial part. Cells from xeroderma pigmentosum (XP) patients, which are defective in nucleotide excision repair, express higher levels of p53 and are highly susceptible to cell death after ultraviolet (UV) irradiation. To examine the relationships between DNA damage, p53 and apoptosis, normal and XP group A fibroblasts were exposed to UVB, and expressions of molecules involved in apoptosis were examined. Apoptosis of XP and normal cells was clearly detected at 48 h after irradiation with UVB at doses of 5 and 40 mJ/cm2, respectively. Cells were positive by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labelling (TUNEL) staining under these exposure conditions. At 6 h after irradiation, p53 protein expression was induced in normal and XP cells at minimal doses of 10 and 2.5 mJ/cm2, respectively. Bcl-2 protein, an inhibitor of apoptosis, was downregulated prior to cell death following UVB exposure at doses that induced apoptosis in both cell types. These results suggest that DNA damage due to UVB induces apoptosis by upregulating proapoptotic molecules such as p53, and by downregulating anti-apoptotic molecules such as Bcl-2.     

Low concentrations of formaldehyde induce DNA damage and delay DNA repair after UV irradiation in human skin cells

Long-term occupational exposure to formaldehyde (FA) increases the risk for nasopharyngeal squamous cell carcinoma. As the skin is also in contact with FA by environmental exposure, we tested the genotoxic properties of appropriate low concentrations (<100 microM) of FA on cultured keratinocytes and fibroblasts of human skin. The initial DNA damage was assessed by comet assay. The induction of DNA protein crosslinks was measured by the ability of FA to reduce DNA migration induced by methyl-methane-sulfonate. Upon 4 h of exposure to FA, significant (P < 0.05) crosslink formations were observed in fibroblasts (50 microM FA) and in keratinocytes (25 microM FA). Upon 8 h of exposure to FA (25 microM FA), significant crosslink formations were observed in both the cell types. FA is known to inhibit different DNA repair pathways. Therefore, we studied the effect of FA on UV-induced repair. Human keratinocytes and fibroblasts exposed to 10 microM FA prior to UV irradiation showed disturbed repair kinetics after UVC and UVB, but not after UVA irradiation. Single-strand breaks (SSBs) derived from nucleotide excision repair disappeared 6 h after solely UVC (3 mJ/cm2) or 3 h solely UVB (30 mJ/cm2) exposure in both the cell types. In the presence of FA, SSBs were still present at these time points containing a reference to a delay in DNA resynthesis/ligation. FA at a concentration not inducing micronuclei (12.5 microM) caused significant increase of UVC-induced (4 mJ/cm2) chromosomal damage. Proliferation of keratinocytes and fibroblasts was in parallel to observed DNA damages. In conclusion, our data suggest that environmental exposure to FA may contribute to UV-induced skin carcinogenesis.     

Effect of ultraviolet (UV) A, UVB or ionizing radiation on the cell cycle of human melanoma cells

BACKGROUND: One important component of the cellular response to irradiation is the activation of cell cycle checkpoints. It is known that both ultraviolet (UV) radiation and ionizing radiation (IR) can activate checkpoints at transitions from G(1) to S phase, from G(2) phase to mitosis and during DNA replication. OBJECTIVES: To evaluate the effects of irradiation with different wavelengths on cell cycle alterations. METHODS: p53-deficient IPC-298 melanoma cells were irradiated with 10 J cm(-2) UVA, 40 mJ cm(-2) UVB, or with 7.5 Gy IR. Cell cycle effects were then determined by DNA/5-bromodeoxyuridine dual-parameter flow cytometry. RESULTS: IPC-298 cells irradiated in G(1) with UVA were not arrested at the G(1)/S transition, but at the G(2)/M transition. Despite p53 deficiency, the cells showed a G(1) arrest after UVB exposure. Furthermore, IR did not affect G(1) or S phase, but induced G(2) phase arrest. Hence, the effects of UVA, but not of UVB, on the cell cycle in p53-deficient melanoma cells are comparable with those of IR. CONCLUSIONS: UVA and IR induce radical-mediated strand breaks and DNA lesions, and UVB essentially induces thymine dimers that lead to excision repair-related strand breaks. Different cell cycle effects may be a consequence of different types of DNA damage. The results showed that UVB-irradiated p53-deficient cells are arrested in G(1). Irradiation with the solar radiation component UVB can therefore result in a beneficial retardation of tumour promotion in human skin carrying p53-mutated cell clones.     

Protective effect of dl-alpha-tocopherol on the cytotoxicity of ultraviolet B against human skin fibroblasts in vitro

The effect of dl-alpha-tocopherol on ultraviolet light, 280-320 nm (UVB)-induced damage of human skin fibroblasts was studied by measuring the colony-forming ability, unscheduled DNA synthesis (UDS) and malondialdehyde (MDA) production. Regarding the cell toxicity, the values of the mean lethal dose (D0) of UV in fibroblast strains from 5 normal subjects were examined. D0 increased dose-dependently when the cells were cultured in the presence of dl-alpha-tocopherol at the concentration of 10-1000 micrograms/ml. UDS induced by 500 J/m2 UVB irradiation was not altered by treatment of 100 micrograms/ml dl-alpha-tocopherol. MDA did not increase after 500 J/m2 UVB irradiation in the fibroblasts cultured with 100 micrograms/ml dl-alpha-tocopherol, while MDA in the fibroblasts cultured without dl-alpha-tocopherol increased after irradiation. These results suggest that dl-alpha-tocopherol protects human skin fibroblasts against the cytotoxic effect of UVB, and its mechanism seems to be related to inhibition of UV-induced lipid peroxidation or to the antioxidation effect of dl-alpha-tocopherol.     

A mineral sunscreen affords genomic protection against ultraviolet (UV) B and UVA radiation: in vitro and in situ assays

Ultraviolet (UV) radiation has been shown to be responsible for different biological effects on human skin, including the initiation of photocarcinogenesis. Both UVB and UVA have been described as mutagenic, but the processes by which they alter the DNA are different. Although cells can repair DNA damage, some deleterious mutations nevertheless appear and can promote cancer. The risk of photocarcinogenesis is acknowledged and the frequency of photogenodermatosis is increasing. In order to evaluate the protection efficacy of a high sun protection factor (SPF) mineral sunscreen against UVB- and UVA-induced genomic alterations, we have followed two approaches. First, we have tested the sunscreen for its ability to decrease the unscheduled DNA synthesis response in vitro in human fibroblasts, as an indirect measure of UVB-induced lesions (0.005 and 0.01 J/cm2), and second, we have verified its ability to reduce the in situ end-labelling intensity in human skin as a direct measure of UVA-induced single-strand breaks (10 J/cm2). Microscopic analysis clearly demonstrated the protective effect of the sunscreen against UVB and UVA. A dose-dependent effect of mineral sunscreens was observed. There was also a relationship between the SPF and genomic protection. By limiting the accumulation of UV-induced lesions on DNA, this mineral sunscreen could limit the mutation frequency.     

中波紫外线辐射损伤角质形成细胞的p53信号传导通路研究

目的 探讨中波紫外线(UVB)辐射对角质形成细胞p53信号传导通路的影响.方法 以20、60和120mJ/cm²UVB辐射培养的取自健康儿童包皮的正常人表皮角质形成细胞,应用RT-PCR和免疫印迹方法,分别从mRNA水平和蛋白质水平,检测分析UVB辐射后2h、24h和48h,p53及其下游分子MDM2、p21、Bax和GADD45的表达.结果 不同剂量UVB辐射角质形成细胞后均可见p53表达水平持续性显著升高(P<0.05).低剂量UVB辐射角质形成细胞后,MDM2、p21、GADD45升高不明显(P>0.05),Bax不表达;UVB辐射剂量升至60mJ/cm²后,MDM2于辐射后2h短暂性显著升高,p21和Bax持续性显著升高.结论 UVB辐射激活了角质形成细胞的p53信号传导通路,p53下游分子的表达具有剂量依赖性和时相性.     

Ultraviolet A irradiation stimulates collagenase production in cultured human fibroblasts.

This study was designed to investigate the biochemical mechanisms responsible for the connective tissue changes seen in actinically damaged skin, which is characterized histologically by diminution and ultrastructural alterations of collagen fibrils and deposition of elastotic material in the papillary dermis. We hypothesized that ultraviolet light could stimulate synthesis of interstitial collagenase in the skin, resulting in collagen degradation. Monolayer cultures of human fibroblasts or keratinocytes were irradiated with ultraviolet A (UVA) or ultraviolet B (UVB) radiation and interstitial collagenase or its inhibitor, TIMP (tissue inhibitor of metalloproteinases) assessed in the conditioned medium with Western immunoblots 24 h after irradiation. Northern blot analysis of the irradiated fibroblasts with a cDNA probe representing collagenase was also performed. Cell viability was greater than 90% with all doses of UV radiation studied. A dose-related increase in immunoreactive collagenase was detected in the medium of fibroblasts irradiated with 0-10 J/cm2 of UVA radiation as well as a parallel increase in the collagenase mRNA in the irradiated cells. UVA radiation stimulated collagenase synthesis in both neonatal and adult fibroblasts. TIMP production in UVA-irradiated fibroblasts increased to a lesser degree than did collagenase and its increase did not parallel the increase in collagenase. UVB (0-100 mJ/cm2) did not stimulate collagenase production by fibroblasts. In contrast to the stimulation of collagenase production by fibroblasts, a slight decrease in immunoreactive collagenase was seen in UVA-irradiated keratinocytes. These data suggest that direct stimulation of collagenase synthesis by human skin fibroblasts by UVA radiation may contribute to the connective tissue damage induced by ultraviolet radiation leading to photoaging.     

Low-dose ultraviolet B radiation synergizes with TNF-alpha to induce apoptosis of keratinocytes

High-dose ultraviolet B (UVB) irradiation is known to induce apoptosis of keratinocytes, but low-dose UVB dose not. In this paper we present evidence that low-dose UVB can induce TNF-alpha-dependent apoptosis of keratinocytes. In our study, 5 mJ/cm(2) doses of UVB were not sufficient by themselves to induce apoptosis of cultured human keratinocytes, but 20 mJ/cm(2) doses of UVB were. The combination of 5 mJ/cm(2) doses of UVB and exogenous TNF-alpha (15 ng/ml) induced significant apoptosis of keratinocytes, although exogenous TNF-alpha without UVB did not. This phenomenon was accompanied by enhanced clustering of tumor necrosis factor receptor 1 (TNFR1). TNF-alpha's promotion of the induction of apoptosis by low-dose UVB was seen until 30 min after irradiation but not at 1 h. We confirmed this finding using a skin organ culture system. UVB (20 mJ/cm(2)), which did not induce transformation of epidermal keratinocytes into sunburn cells, induced apoptosis when TNF-alpha was added to the culture medium. These results suggest that one of the possible mechanisms of inducing keratinocyte apoptosis by low-dose UVB and TNF-alpha is that low-dose UVB augments ligand-binding-induced TNFR1 clustering, resulting in increased apoptotic cell death.     

乌鲁木齐地区127例志愿者紫外线最小红斑量测定

目的 测定乌鲁木齐地区志愿者长波紫外线（UVA）、中波紫外线（UVB）的最小红斑量（MED）。方法 以SUV-1000型日光紫外线模拟器为光源,测定127例志愿者UVA-MED、UVB-MED。结果 Ⅲ型皮肤48例,UVA-MED中位数为38.10 J/cm2,UVB-MED中位数为31.80 mJ/cm2;Ⅳ型皮肤79例,UVA-MED中位数为59.16 J/cm2,UVB-MED中位数为48.00 mJ/cm2。男性UVA-MED中位数为59.16 mJ/cm2,女性为41.10 J/cm2;男性UVB-MED中位数为39.60 mJ/cm2,女性为35.55 mJ/cm2。男、女性Ⅲ型与Ⅳ型皮肤UVA-MED、UVB-MED中位数差异有统计学意义,Ⅲ型皮肤均显著低于Ⅳ型皮肤。男性UVA-MED显著高于女性（P < 0.05）,UVB-MED男女之间差异无统计学意义（P > 0.05）。Ⅲ型、Ⅳ型皮肤维族与汉族比较,UVA-MED和UVB-MED差异均无统计学意义（P值均 > 0.05）,男性、女性在不同年龄组间以及不同户外停留时间组之间差异也无统计学意义（P值均 > 0.05）。通过百分位数法确定UVA-MED的正常值 > 33.38 J/cm2,UVB-MED > 27.90 mJ/cm2。结论 皮肤光反应类型是决定MED的重要因素。     

Dosimetry of solar ultraviolet radiation. Daily and monthly changes in Paris

The intensity of ultraviolet A and B radiations was measured in Paris (48 degrees North) by means of silicon photoelectric cells (Osram Centra dosimeter) from December, 1984 till February, 1986. The results, which must be regarded as approximate, are expressed as physical units (mW/cm2) and biological units (minimal erythema dose/hour). For sunny days two curves are presented separately for UVB and UVA: daily variations in radiation (hourly measurements) and daily variations at 11 hours (solar time) during one year. Maximum irradiation was observed at noon in early July: UVB 0.15 mW/cm2, UVA 5.4 mW/cm2. Between December and July the amount of UVB radiation was multiplied by 14 and that of UVA radiation by 9. For subjects with clear photo-type and when the sun was at its zenith, an MED per hour was obtained from May 1 onwards. Within a day, 30 p. 100 (summer) and 50 p. 100 (winter) of erythema-producing UV intensity were delivered between 11 and 13 hours (solar time). This kind of study has numerous clinical applications: advice regarding exposure to sun rays, dosing of heliotherapy, epidemiological data concerning photodermatitis (circumstances of exposure, UV threshold dose) and photocarcinogenesis (determination of annual MED doses in relation to areas of uncovered skin and occupational exposure to sun rays). Other studies on the French territory will provide a map of UV irradiation.     

Early cell kinetic effects of a single dose of monochromatic ultraviolet B irradiation on hairless mouse epidermis

Hairless mice were exposed to a single erythemic (25 mJ/cm2) or suberythemic dose (12.5 mJ/cm2) of ultraviolet B (UVB) irradiation at 297 nm. The cell kinetic changes were observed at several times during the first 7 d after the irradiation. The mitotic count, the mitotic rate (stathmokinetic method), and the number of suprabasal and basal cells were scored in histologic sections. The incorporation of [3H]thymidine was measured after pulse labeling, and the DNA distribution pattern was studied by flow cytometry. Initially, both UVB-doses induced a block or delay in the cell proliferation. The rate of entrance of cells into mitosis and the uptake of [3H]thymidine were reduced, and cells accumulated in the S phase of the cell cycle. Hence, during the first period after irradiation, UVB seemed to interfere with the DNA synthesis by inducing a prolonged S phase duration. The DNA synthesis rate was reduced to the same degree after both UVB-doses. From 24 h after irradiation rapid regenerative proliferation took place, most pronounced after the highest UVB-dose. Waves of proliferation seemed to arise from partially synchronized cohorts of cells proceeding through the cell cycle at a higher speed than normal. Thus, the present study indicates that UVB irradiation is comparable with the cell kinetic effects following both chemical skin carcinogens and non-carcinogenic skin irritants. UVB induces an inhibitory effect on the DNA synthesis activity, in addition to regenerative cell proliferation subsequent to cell toxicity.     

UV-induced isomerization of oral retinoids in vitro and in vivo in hairless mice

Ultraviolet (UV) irradiation causes isomerization and destruction of many vitamin A analogues (retinoids). Using high-performance liquid chromatography (HPLC), we investigated in vitro and in vivo the effects of UV irradiation on 2 all-trans aromatic retinoids (etretinate and acitretin) and on 13-cis retinoic acid (isotretinoin). When etretinate and acitretin dissolved in ethanol were irradiated with UVB (280-320 nm; 10-336 mJ/cm2) or UVA (320-400 nm; 1-5 J/cm2), extensive and reproducible cis-isomerizations occurred at the 13-position (cis/trans ratio approximately 1.6 in all experiments) but there was no progressive photodegradation of the molecules. Irradiation of isotretinoin produced only moderate trans-isomerization but the sum of HPLC peak heights fell with increasing UV doses, being 72% of the original value after 336 mJ/cm2 of UVB. Hairless mice were given etretinate (50 mg/kg bw), acitretin (200 mg/kg) or isotretinoin (50 mg/kg) on days 1, 4 and 7 and were irradiated daily for 8 d with 13 mJ/cm2 UVB plus 1 J/cm2 UVA. Samples of serum, dorsal skin and liver were collected and retinoids analyzed by HPLC. In the etretinate and acitretin-treated, irradiated animals the serum concentrations of the 13-cis isomers were 2-6 times higher than in nonirradiated controls. Irradiated epidermis also contained significantly higher concentrations of 13-cis etretinate and 13-cis acitretin than did control epidermis. The serum and epidermal concentrations of all-trans etretinate and acitretin were unchanged or even increased after irradiation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isoflavones in aglycone solution enhance ultraviolet B‐induced DNA damage repair efficiency

The isoflavones daidzein and genistein are natural compounds which have anti‐inflammatory and photoprotective activities, and may be effective in the repair of ultraviolet (UV)‐induced photodamage. In this study, an alcoholic solution of aglycone isoflavones with a genistein:daidzein ratio of 1:4 [Rottapharm (RPH)‐aglycone] was examined for its effects on the repair of DNA damage induced by a single dose of UVB irradiation (20 mJ/cm²). For this purpose, human skin cells were first UVB‐irradiated and then treated with RPH‐aglycone. Comet assay analysis was used to estimate the UVB‐induced DNA damage at different time points after treatment by measuring the tail moment parameter. We found that treatment with 10 μmol/L RPH‐aglycone solution resulted in a significantly reduced tail moment at 1 h after treatment, and 34–35% enhancement of damage repair at 4 h after treatment. These results suggest that isoflavone aglycones are protective against UVB‐induced DNA damage.     

A time-correlation study of ultraviolet B-induced erythema measured by reflectance spectroscopy and laser Doppler flowmetry.

Ultraviolet B (UVB)-irradiated skin pigmentation was quantified using 2 objective and noninvasive techniques. Ten healthy volunteers were exposed to increasing UVB doses in the interval from 6 mJ/cm2 to 120 mJ/cm2 and the resultant vascular and pigmentary changes were evaluated using laser Doppler flowmetry (LDF) and reflectance spectroscopy (RS). Measurements were made 0.25 h, 1 h, 4 h, 8 h, 24 h, 72 h and 192 h post-irradiation. Visual scores were determined 24 h post-irradiation. Both RS and LDF revealed an early and rapid erythematous response to UVB irradiation, peaking between 8 h and 24 h, with no single UVB dose showing any significant increase until 4 h post-irradiation. LDF showed a peak increase in blood flow at 8 h post-irradiation for sites with low UVB exposure (less than or equal to 36 mJ/cm2). Doses greater than or equal to 42 mJ/cm2 showed maximal increase at 24 h. After this increase in blood flow, a slow normalization began that was not complete at 192 h post-irradiation at sites exposed to greater than or equal to 60 mJ/cm2. The present RS analysis is able to distinguish between oxygenized (OH) and deoxygenized hemoglobin (DOH). The only significant increase in DOH was found for the averaged 6 mJ/cm2 and 12 mJ/cm2 UVB dose sites 24 h after irradiation. OH peaked 24 h post-irradiation and resolution was still incomplete after 192 h. RS revealed no dose, as did the LDF, below which the vascular response peaked earlier. LDF measures dermal blood flow and RS measures hemoglobin content in the skin.(ABSTRACT TRUNCATED AT 250 WORDS)