Effects of in vitro PUVA treatment on adenylate cyclase responses of epidermis

We investigated the effect of 8-methoxypsoralen (8-MOP) plus long wave ultraviolet irradiation (PUVA) on the pig skin epidermal cyclic AMP (cAMP) system. Following PUVA treatment in vitro, pig skin squares were incubated in RPMI 1640 medium, and the cAMP accumulation by various adenylate cyclase stimulators was determined. A significant increase of the beta-adrenergic adenylate cyclase response was observed as early as 6-h following PUVA treatment; the maximal effect was reached at 24-h and lasted at least for 72-h. This augmentation effect of PUVA treatment was potentiated by increasing the 8-MOP concentration (0.5–160 μg/ml) and UVA irradiation dose (0.05–1.4 J/cm²). At higher irradiation doses (2.1–2.8 J/cm²), the beta-adrenergic augmentation effect was less marked. There were no significant differences in the adenosine-and histamine-adenylate cyclase response up to 1.4 J/cm² irradiation; however, the latter was decreased at a higher irradiation dose (2.8 J/cm²). Although UVA irradiation alone had no effect on the beta-adrenergic response, 8-MOP treatment alone increased this receptor response at higher concentrations; this effect was much weaker than that induced by PUVA treatment. Cyclic AMP phosphodiesterase activities were not affected by PUVA treatment. These results indicate that epidermal adenylate cyclase response is affected by in vitro PUVA treatment.     

DNA repair elicited by UVB during PUVA therapy for psoriasis

Summary The skin of patients receiving psoralen and UVA (PUVA) therapy for psoriasis is exposed to trace amounts of UVB radiation emitted by PUVA irradiators in addition to UVA. DNA repair activity was measured using autoradiography in the uninvolved skin of PUVA-treated patients in order to determine whether 8-methoxypsoralen (8-MOP) plus UVA elicits repair, inhibits the skin repair response to UVB, or protects epidermal-cell DNA from UVB damage by promoting a tan. Epidermal-DNA repair activity was observed in 27 out of 37 patients following the first PUVA treatment. Phototesting with multiples of the initial UV dose elicited a linear increase in repair activity. Glass-filtered radiation failed to stimulate repair, indicating that the reaction was due to UVB, not to 8-MOP plus UVA. The same amount of repair activity was observed in the skin of patients irradiated either before or after 8-MOP ingestion, demonstrating that the drug did not interfere with the response of the skin to UVB. At clearing, however, the repair activity was never greater than that elicited at the initial treatment and was often undetectable despite a tenfold increase in UV exposure. It is proposed that DNA damage should be measured to determine whether epidermal cells are entirely protected from UVB radiation at the completion of therapy.     

In vitro photoinhibition by psoralen and ultraviolet A radiation of human hematopoietic progenitors

The in vitro sensitivity of human hematopoietic progenitors to PUVA, 8-MOP and UVA alone was investigated. 8-MOP alone at final concentrations of 150, 200, 600 and 1000 ng/ml did not modify colony growth of circulating and bone marrow erythroid (BFU-E), myeloid (CFU-GM) and immature (CFU-GEMM) hematopoietic progenitors obtained from normal controls. The exposure of the same progenitors to increasing doses of UVA, up to 12 J/cm², progressively decreased hematopoietic colony growth (with estimated 50% inhibition occurring at about 5 J/cm²). In vitro PUVA treatment (8-MOP 200 ng/ml followed by UVA 5 J/cm²) caused 90% growth inhibition of circulating and bone marrow hematopoietic progenitors. In addition, the treatment completely inhibited the formation of spontaneous erythroid colonies, obtained from 5 polycythemic patients, that are considered to be a marker of this neoplastic disease. PUVA cytotoxicity was assessed by the colorimetric MTT assay. The percentage of cell death after PUVA exposure was 29 ± 10% for both peripheral and bone marrow mononuclear cells. Our findings indicate that 8-MOP alone is not toxic to hematopoietic progenitors whereas UVA treatment determines in vitro a dose-dependent inhibition of the clonogenic capacity of normal hematopoietic cells. PUVA treatment enhances this effect, causing a quite complete inhibition of hematopoietic progenitors colony formation from normal donors and spontaneous BFU-E colony formation from polycythemic patients.     

Detection of DNA-psoralen photoadducts in mammalian skin

An immunofluorescence (IF) method for the detection of 8-methoxypsoralen (8-MOP) photoadducts to DNA has been developed to assess nuclear damage in keratinocytes and melanocytes after psoralen plus UVA (PUVA) treatment, both under in vitro and in vivo conditions. Cryostat sections of the albino and pigmented guinea pig and human skin were used for in vitro studies to establish minimal and maximal drug concentration and UVA dosimetry for the detection of DNA-8-MOP photoadducts. Limits of detection were as low as 10 ng/cm2 8-MOP and 1 J/cm2 UVA for skin sections and sodium bromide-split epidermal sheets. Guinea pigs treated with topical PUVA revealed positive IF stain in epidermal cell nuclei at a threshold dose of 100 micrograms/cm2 8-MOP and 13 J/cm2 UVA. Pretreatments of cryostat cuts with ethanol and alkali before IF test enhanced the sensitivity of detection in vivo about 10-fold and enabled us to follow the repair of DNA damage after treating normal guinea pig skin with a dose of 50 micrograms/cm2 8-MOP plus 6 J/cm2 UVA. The most interesting findings were as follows: A sensitive method to detect PUVA-induced nuclear damage in epidermal and dermal cells was developed. PUVA treatment induced nuclear DNA damage to melanocytes as well as to adjacent keratinocytes, and melanocytes appeared to be 10 times less vulnerable to photo-damage than keratinocytes. There was a greater propensity for the proliferative cells to be damaged by PUVA. PUVA induced nuclear damage up to 700 micron depth in the dermis. The usefulness of the IF test in detecting DNA damage in microgram and ng amounts in vivo and in following the repair of damaged DNA induced by PUVA.     

Effects of 8-Methoxypsoralen (8-MOP) and UVA on human lymphocytes

Summary Peripheral lymphocytes of 37 psoriatic patients are tested before and under PUVA treatment using as parameter the non specific stimulation effect of HgCl₂ (10 µg/ml) in culture, measuring the³H-thymidine incorporation after the last 16 h of a 5-days culture. Oral 8-MOP in therapeutic doses is decreasing the lymphocyte stimulation as well as 8-MOP together with UVA irradiation during the first week of treatment. After 1 week, the stimulation is, on the contrary, significantly enhanced after irradiation. Lymphocytes isolated by centrifugation over Lymphoprep are submitted to PUVA conditions in petri dishes (Hank's solution 8-MOP 1 µg/ml, irradiation with 350 nm, 93-372 mJ/cm²). The total cell number, the E-rosette formation (as marker for T-Lymphocytes) and the EAC-rosette formation (as marker for B-Lymphocytes) are determined. PUVA conditions have an energy dependent decreasing effect on the cell number, while the T- and B-cell proportions remain constant. UVA irradiation alone has such an effect only with high energies. 8-MOP without UVA has no significant influence on the cell number.Offprint requests to: Dr. G. Lischka (address see above)     

Kinetics and dose-response of photosensitivity in cream psoralen plus ultraviolet A photochemotherapy: comparative in vivo studies after topical application of three standard preparations

BACKGROUND: Topical photochemotherapy with bath psoralen plus ultraviolet (UV) A irradiation (PUVA) has been developed to reduce possible side-effects of oral PUVA therapy. Although the efficacy of bath PUVA therapy appears to be similar to oral PUVA therapy, provision of bathing facilities has obvious economic, logistic and sanitary implications. Cream PUVA therapy has recently been developed as a variation of topical PUVA. OBJECTIVES: To understand the photobiological effects and to increase the safety and effectiveness of this novel topical PUVA therapy, we assessed the kinetics and dose-response of phototoxicity of 8-methoxypsoralen (8-MOP) cream in order to develop a treatment schedule for this treatment option. METHODS: Ninety-eight patients (63 men and 35 women) undergoing cream PUVA therapy were studied. The phototoxic properties of topically applied 8-MOP in three different water-in-oil creams as vehicles were assessed. In a dose-response study, four concentrations of 8-MOP cream (0.0006-0.005%) were used for determination of the minimal phototoxic dose (MPD). The kinetics of photosensitization were tested by determination of MPDs after different application times of 8-MOP cream (10, 20, 30 and 60 min). The persistence of phototoxicity was assessed by UVA exposure at defined time intervals after application of 8-MOP cream (0, 30, 60 and 120 min). RESULTS: The concentration required to produce sufficient but not undue photosensitization of the skin was 0.001% 8-MOP. The duration of application leading to the lowest MPD was 30 min. Greatest photosensitization was achieved when UVA irradiation was performed between 0 and 30 min after 8-MOP removal. These findings showed no significant difference between the three vehicles used. CONCLUSIONS: Based on our data we recommend application of 0.001% 8-MOP in a water-in-oil cream for 30 min. Irradiation with UVA should be performed within 30 min after removal of 8-MOP cream, as there is a rapid decrease in photosensitivity thereafter.     

Effect of psoralen and ultraviolet A on platelet functioning: an in vitro and in vivo study.

We investigated possible alterations induced by psoralen and ultraviolet A radiation (PUVA) on platelet function both in vitro and in vivo. In vitro, using conventional aggregometry and adenosine diphosphate (ADP), collagen, ristocetin and arachidonic acid as aggregating agents, platelet aggregation was determined on platelet-rich plasma (PRP) from normal subjects at basal conditions and following the addition of increasing concentrations of 8-methoxypsoralen (8-MOP) with and without exposure to ultraviolet A (UVA) light (5 J/cm2) and compared with UVA light exposure alone. At basal conditions and following exposure to UVA light alone, no changes in the normal platelet aggregation patterns were observed. Exposure to UVA light of PRP containing 8-MOP also demonstrated no abnormality in the platelet aggregation patterns at 8-MOP concentrations of 200 ng/ml. However, abnormal platelet aggregation as a response to ADP and collagen was observed at higher concentrations of 8-MOP, which was augmented upon exposure to UVA light. In vivo, platelet aggregometry was performed on PRP from 4 patients submitted to PUVA treatment at basal conditions, 2.5 h after oral ingestion of 8-MOP (0.6-0.8 mg/kg) and after 4 PUVA sessions. No patient showed modification of the platelet aggregation profile after either 8-MOP ingestion or PUVA treatment. Our study shows that 8-MOP at high concentrations in vitro impairs platelet aggregation by ADP and collagen augmented by UVA light exposure, but PUVA therapy causes no detectable abnormality in platelet function in vivo.     

Treatment of persistent severe atopic dermatitis in 113 Japanese patients with oral psoralen photo-chemotherapy

Oral administration of psoralen and whole body exposure to UVA (oral PUVA) has been used for the treatment of 113 patients with severe atopic dermatitis (AD). 8-Methoxypsoralen (8-MOP) was given at a dose of 0.5-0.6 mg/kg two hours prior to UVA (3-8 J/cm2) irradiation. Patients were treated three times a week while hospitalized. Other medications which had been given before PUVA therapy were permitted. At four and eight weeks after PUVA therapy, the severity score of AD had decreased by 51% and 80%, and the cumulative doses of UVA were 51.2 J/cm2 and 115.3 J/cm2, respectively. The amounts and strength of topical cortico-steroids were decreased during PUVA therapy. No adverse effects that required discontinuation of the PUVA therapy were observed. After discharge, maintenance therapy with UVB phototherapy and/or conventional treatment of AD kept the patients in remission in the outpatient clinic. The QOL of patients was greatly improved. Photochemotherapy with oral 8-MOP can be indicated in patients with severe, widespread AD, especially if standard therapy fails. This is the first report of oral PUVA therapy in a large series of Japanese patients with AD.     

Neoplastic transformation of C3H mouse embryo 10T1/2 cells by 8-methoxypsoralen plus UVA radiation

The effect of 8-methoxypsoralen plus UVA radiation (PUVA) on cell killing and induction of transformation was studied in the C3H mouse embryo 10T1/2 cell line. Dose-response data for both survival and transformation were obtained as a function of 8-methoxypsoralen (8-MOP) concentration and UVA dose. PUVA treatment caused cell death and induced transformation in a dose-dependent manner. Treatment of cells with 8-MOP alone (10 micrograms/ml) or UVA alone (90 J/m2) had no effect on either cell killing or transformation. The product of 8-MOP concentration and UVA dose calculated at 10% survival and 10(-3) transformation frequency levels were quite similar regardless of 8-MOP concentration or UVA dose. This suggests that there exists a simple reciprocal relationship between 8-MOP concentration and UVA dose. Both type II and type III foci induced by PUVA treatment were tumorigenic in vivo. These data provide further evidence for the carcinogenicity of PUVA treatment. In addition, the system described here could serve as a valuable model for studying the relationships between transformation and the specific cellular and molecular lesions induced by PUVA treatment.     

An intraindividual comparative study of psoralen-UVA erythema induced by bath 8-methoxypsoralen and 4, 5', 8-trimethylpsoralen

BACKGROUND: Limited work has been conducted on the characteristics of topical trimethylpsoralen (TMP) psoralen-UVA (PUVA) erythema. OBJECTIVE: We sought to determine the time-course and dose-response characteristics of erythema induced by topical TMP, and to compare these parameters with those for topical 8-methoxypsoralen (MOP) within patients. METHODS: After photosensitization of one forearm with topical TMP, test sites were exposed to a UVA dose series. The procedure was repeated on the other forearm using 8-MOP solution. Erythema was assessed visually and with a reflectance instrument every 24 hours for 7 days. RESULTS: TMP PUVA erythema followed a similar time course to 8-MOP PUVA erythema. The majority of patients were at maximal erythema at or beyond 96 hours. TMP PUVA had a significantly steeper dose-response curve at 48, 72, and 96 hours compared with 8-MOP PUVA. CONCLUSION: On the basis of these data, the optimal time to read the TMP minimal phototoxic dose is 96 hours. In view of the steeper dose-response curve for TMP PUVA, a lower UVA incremental regimen should be considered compared with that for 8-MOP PUVA.     

In vitro studies on the antifungal effect of PUVA

The effect of PUVA therapy upon 12 dermatophyte strains, 3 of Candida albicans, and 4 of mould fungi was evaluated by the agar dilution method on Sabouraud agar. 8-MOP, 30 microgram/ml medium plus UVA, 4 Joule/cm2 proved to be fungicidal dose to all the strains studied, with the exception of three of the mould fungi. For dematophytes and Candida albicans the MIC was between 1 and 3 microgram, 8-MOP plus UVA, 4 J; and 3 microgram when the UVA dose was reduced to 1--2 J. For mould fungi it was higher. The MFC varied within the range 2 to 30 microgram, when the UVA dose was fixed at 4 J. It reached 16 J when the dose of 8-MOP was kept at 4 microgram. The wide range in the MFC of the dermatophytes could be due to resistant colonies. 8-MOP alone had a distinctly inhibitory effect upon dermatophytes, but not upon Candida albicans or mould fungi.     

Cutaneous carcinoma and PUVA lentigines in Thai patients treated with oral PUVA

A total of 113 Thai patients who were treated with oral PUVA from 1979 to 1992 were examined for long-term cutaneous side effects of PUVA. Two psoriatic patients developed PUVA keratosis on non-sun-exposed areas. Both were skin type IV and had had phototherapy with UVB and sunlight previously. The total doses of UVA were 909 J/cm² and 242 J/cm² respectively. One psoriatic patient developed Bowen's disease. He had had a cumulative dose of UVA 2207 J/cm². He also had a past history of arsenic intake and phototherapy with UVB and sunlight. PUVA lentigines were seen in 58 patients (51.4%). It was associated with older age at starting PUVA, higher cumulative UVA dose and greater number of PUVA treatment. This study suggests that previous exposure to other risk factors is important for inducing skin cancer in populations with skin phototype III, IV and V treated with oral PUVA.     

Alteration of lymphocyte functions by 8-methoxypsoralen and longwave ultraviolet radiation. I. Suppressive effects of PUVA on T-lymphocyte migration in vitro

We investigated the influence of 8-methoxypsoralen (8-MOP) plus long-wave ultraviolet radiation (PUVA) on lymphocyte migration in vitro. Nylon wool-purified, mouse splenic T lymphocytes showed locomotive responses to casein, normal mouse serum (NMS), and zymosan-activated mouse serum (ZAS). Migratory responses to casein and NMS, and to ZAS were remarkably suppressed in lymphocytes exposed to 0.5 J/cm2 UVA plus 0.1 micrograms/ml 8-MOP and to 0.8 J/cm2 UVA plus 8-MOP, respectively. The PUVA treatment used in the present study had no effect on random movement and lymphocyte viability. T lymphocytes cultured in the absence of mitogenic agent for 24 h demonstrated a greater increase in their migration activity than noncultured cells, while lymphocytes cultured after 1.0 J/cm2 PUVA pretreatment remained low. These findings suggest that the therapeutic effect of PUVA on inflammatory skin disorders may be due in part to the suppression of lymphocyte migration.     

PUVA and PUVB in vitiligo – are they equally effective?

BACKGROUND/AIMS: The combination of psoralens with different types of ultraviolet (UVL) sources in the treatment of vitiligo has led to different reports of success. The purpose of this trial is to compare in a random right-left comparison study the efficacy and side effects of oral 8-MOP plus UVA (PUVA) and oral 8-MOP plus UVB (broadband, 290-320 nm P-UVB) in the treatment of vitiligo. METHODS: The study included 24 cases of extensive vitiligo involving more than 30% of the body surface area in a bilateral symmetrical distribution. Each patient received 0.7 mg/kg 8-MOP orally 2 h before the light session. The right side of the body was exposed to UVA (320-400 nm), while the left half was exposed to UVB (290-320 nm). The patients received 3 sessions/week for a total of 30 sessions. RESULTS: Both PUVA and PUVB produced moderate (50-60%) improvement, with similar incidences of phototoxic reaction and skin thickening. However, the study revealed a significant difference in the number of sessions needed to improve produce erythema and perifollicular pigmentation as well as a moderate response, the response on the UVA side always being earlier. Furthermore, the amount of joules needed to achieve the same response was 10 times greater on the UVA side than on the UVB side. CONCLUSION: The use of psoralen plus broadband UVB is as effective as PUVA in the treatment of vitiligo. However, the long-term side effects of psoralen plus UVB are unknown.     

UVB/PUVA-induced suppression of human natural killer activity is reduced by superoxide dismutase and/or interleukin 2 in vitro

We have evaluated the effects of ultraviolet irradiation or PUVA treatment [8-methoxypsoralen (8-MOP) plus long-wave ultraviolet (UVA) irradiation] on natural killer (NK) activity of human peripheral blood mononuclear cells (PBMC). In vitro exposure of PBMC to UVB (280-320 nm, 1-30 mJ/cm2) or PUVA [8-MOP, 0.1 microgram/ml; UVA (320-400 nm), 0.5-5 J/cm2] resulted in a dose-dependent suppression of NK activity, whereas UVA (0.5-5 J/cm2) or 8-MOP (0.1 microgram/ml) treatment alone did not have the inhibitory effects. This suppressive effect of UVB/PUVA irradiation was successfully reduced in the presence of superoxide dismutase (SOD) (100 or 1000 U/ml) during the irradiation. The addition of interleukin 2 (IL-2) (100 U/ml) markedly enhanced the NK activity of both irradiated and nonirradiated PBMC. Combination treatment with both SOD and IL-2 to UVB/PUVA-irradiated PBMC resulted in a more remarkable improvement of NK suppression than with either SOD or IL-2 treatment alone.     

Plasma levels of 8-methoxypsoralen following PUVA-bath photochemotherapy

Administration of 8-methoxypsoralen (8-MOP) in a dilute bath water solution is an effective therapeutic alternative to oral PUVA therapy, avoiding systemic side effects, offering better bioavailability of the psoralen and requiring much smaller amounts of UVA for induction of therapeutic effects. To obtain exact data about the percutaneous absorption of 8-MOP during a psoralen bath, the plasma levels of the drug were determined in 26 patients with different skin diseases by a reverse high-performance liquid chromatographic method. Fifteen patients receiving oral PUVA therapy (0.8 mg 8-MOP/kg body weight) served as a positive control group. Bath solutions were prepared by diluting 15 ml of 0.5% stock solution of 8-MOP in 150 l of bath water (0.5 mg/l, 37°C). Blood samples were drawn from patients 5, 30, 60, 120 and 180 min after the bath. In the oral PUVA group, blood samples were obtained 1½ h after administration of the drug. In 23 of 26 patients, 8-MOP levels were undetectable in every blood sample. After 30 min, two patients showed detectable levels of 8-MOP (5 ng/ml, 7 ng/ml), while 60 min after the PUVA bath 8-MOP was detectable in only one volunteer (5 ng/ml). In patients receiving oral 8-MOP therapy, serum levels varied between 45 and 360 ng/ml 1½ h after drug administration. Our data confirm extremely low 8-MOP levels resulting from 8-MOP bath water treatments and provide confirmation of the absence of systemic side effects in patients who are undergoing PUVA-bath therapy.     

The effect of PUVA on langerhans cells in rat oral epithelium photosensitized with systemic methoxsalen or topical trioxsalen

BACKGROUND/PURPOSE: Ultraviolet-A radiation (UVA) of the oral mucosa after photosensitization with either systemic methoxsalen (8-MOP) or topical trioxsalen (TMP), i.e. mouth-PUVA, has been reported to be successful in the treatment of oral lichenoid lesions. In the case of PUVA treatment of skin disorders, local immune suppressive effects have been demonstrated, and the antigen presenting epithelial Langerhans cells (LCs) have been shown to be especially sensitive to ultraviolet treatments. Our aim was to compare the photobiological effects of PUVA in oral mucous membrane (OMM) using topical TMP or systemic 8-MOP photosensitization. METHODS: Rat OMM photosensitized with topical TMP or systemic 8-MOP was treated with PUVA using UVA doses of 1-8 J/cm2. The LCs were demonstrated in epithelial sheets of the treated OMM with ATPase staining. RESULTS: Both treatments caused a sim ilar, dose-dependent depletion of ATPase-positive LCs, with a maximal depletion of 80% or 73% with 8 J/cm2 at 2 days after irradiation as photosensitized with TMP or 8-MOP, respectively. This contrasts with earlier published findings in human skin, where topical TMP is an order of magnitude greater a sensitizer than 8-MOP, and PUVA-induced depletion of LCs occurs maximally 5 days after irradiation. CONCLUSION: The depletion of LCs of rat OMM after PUVA treatment is greater using topical TMP compared to systemic 8-MOP, but the difference is significantly smaller than reported earlier in human skin.     

Acute effect of 8-methoxypsoralen and ultraviolet light on sister chromatid exchange

Summary The acute effect of 8-methoxypsoralen (8-MOP) and 8-MOP+long wave ultraviolet light (UVA) on sister chromatid exchange (SCE) has been examined in an in vitro experiment. The SCE count was significantly increased by 8-MOP without light, but the effect was substantially greater (50%) by 8-MOP+UVA. In addition, mitoses with banded staining of the chromosomes were seen after 8-MOP and UVA. These changes were dose dependent, and they might be responsible for the reduced cell turnover in psoriasis plaque after PUVA.     

Protecting the eye from ultraviolet A radiation during photochemotherapy

Except for the skin, the eye is the only organ that is continuously exposed to solar radiation, including longwave ultraviolet irradiation (UVA). Since 8-methoxypsoralen (8-MOP) remains not only in the skin but also in the lens of the eye after 8-MOP + UVA (PUVA) treatment, wearing protective goggles just during the UVA irradiation is insufficient. It is wise to shield the eyes for several hours after 8-MOP ingestion, to avoid or reduce possible long-term side effects such as cataract formation. Adequate eye protection from UVA after PUVA can be provided by sunglasses that filter out the appropriate UVA spectrum from the sunlight. Nearly all the commercially available sunglasses are colored, which reduces the intensity of the visible light reaching the eye. In such cases, the diameter of the pupil can remain dilated, which it would not without sunglasses. However, the UVA intensity remains the same. As a result, more UVA can reach the lens. In this study, new uncolored glasses are evaluated and compared with commercially available sunglasses (all colored) to determine their UVA transmission.     

Penetration kinetics of 8-methoxypsoralen after 8-methoxypsoralen bath procedure with and without UVA irradiation.

Administration of 8-methoxypsoralen (8-MOP) in a dilute bath water solution is an effective therapeutic alternative for systemic application of 8-MOP, avoiding systemic side effects such as nausea and cataractogenesis. The aim of our study was to determine the epicutaneous penetration of 8-MOP in a dilute bath water solution with and without additional UVA irradiation in human skin under in vitro conditions. To simulate the PUVA bath procedure, 8 skin samples were exposed to radioactively labeled 8-MOP in a water solution. After 20 min, the test solution was removed and the skin surface was dried. Immediately after the bath procedure, 4 of the skin samples were irradiated with 0.5 J/cm2 UVA. During a test period of 15 h, the 8-MOP penetration was observed. In both test groups (with and without UVA irradiation) 8-MOP permeated through all skin layers between 30 min and 1 h after application. Compared to the unirradiated skin samples, the UVA-irradiated skin samples showed a significantly slower increase and a lower maximum of 8-MOP permeation. Following our results, UVA irradiation of 8-MOP-exposed skin samples led to a significantly decreased permeation rate. This might be due to UVA-induced links between 8-MOP molecules and human DNA. In addition, we investigated the levels of radioactivity emitted by tritium-labeled 8-MOP in stratum corneum, epidermis and dermis up to 30 min after 8-MOP bath in two further test groups with and without additional UVA irradiation. The statistical analysis revealed no significant differences between these two test groups. Thus, the levels of radioactivity remained constant in the epidermis and dermis during the test period of 30 min. Since the levels of radioactivity were constant up to 30 min after UVA irradiation, a previously supposed marked loss of 8-MOP concentration might not be responsible for the rapid extinction of observed in vivo photosensitivity within 1 h after PUVA bath observed in vivo in human skin.