In vivo PUVA and UVB sensitivity of various human epidermal Langerhans cell markers (ATPase, HLA-DR and T6)—Dose-response and time-sequence studies

To form a comprehensive view of the UV-sensitivity of human epidermal Langerhans cells (LC), the time-sequence and close response effects of single doses of UVB or 8-methoxypsoralen plus UVA (PUVA) radiation on three different LC surface markers were studied with histochemical and immunohistochemical staining. An increasing PUVA dose from 1 to 10 J/cm² caused an almost linear decrease in the surface enzyme (ATPase) positive LC count, whereas the cell surface antigens (HLA-DR and T6) were rather more resistant, up to a PUVA dose of 5 J/cm². A single dose of 5 J/cm² of PUVA induced an LC depletion that was similar during the 21 days of observation, irrespective of whether the cells were visualized with ATPase staining or with monoclonal antibodies against the cell surface antigens HLA-DR or T6. In each case, the nadir was reached 14 days after irradiation; the average residual LC count was then 57%. The cell counts 21 days after PUVA irradiation were still only approximately 74% of the nontreated skin counts. Langerhans cell depletion induced by an erythemagenic dose of UVB irradiation was swifter and more pronounced than that induced by 5 J/cm² of PUVA but, again, a similar time schedule was recorded with ATPase, HLA-DR and T6 staining.     

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.     

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.     

Assoziation zwischen UVA1 bzw. Bade-PUVA-Bestrahlung und Melanomentwicklung?

After 18 months of intensive UVA₁- and bath PUVA therapy for urticaria pigmentosa, a female patient developed a malignant melanoma on the right thigh. This can be taken as further indication that intense UVA₁ or PUVA therapy is capable of inducing malignant melanoma, even though this risk has not been completely proven. The total dose of UVA₁ was 910 J/cm², while the total dose of PUVA was 214,4 J/cm². It is unclear to what extent modified UVA application of long-wave UVA light (UVA₁) and modified PUVA therapy (bath PUVA) have a similar risk compared to known data.     

Erythemal and therapeutic response of psoriasis to PUVA using high-dose UVA

In PUVA treatment of psoriasis, clinical observation suggests that uninvolved skin is more susceptible to PUVA erythema than lesions of psoriasis. If this is the case, then the efficacy of PUVA treatment might be increased by using localized high-dose UVA restricted to lesional skin. We have therefore studied the erythemal and therapeutic response of psoriasis to PUVA using high-dose UVA and, for comparison, the erythemal response to UVB. In 14 patients, an area of psoriasis and adjacent uninvolved skin were exposed to a series of UVA doses (350 ± 30 nm, 1–16 J/cm²), using an irradiation monochromator. Six other patients were similarly phototested with a series of UVB doses (300 ± 5 nm, 20–112 mJ/cm²) to both uninvolved and lesional skin. Erythema was judged visually at 72 h for psoralen–UVA, and at 24 h for UVB, and measured using a scanning laser–Doppler velocimeter. In 10 patients, PUVA therapy using high-dose UVA was subsequently given to lesional skin (8–16 J/cm² twice weekly) in addition to conventional whole-body PUVA. For psoralen–UVA, the minimal phototoxic dose within psoriasis was increased by a factor of 4 compared with non-lesional skin (P < 0.01, Wilcoxon signed-rank test). For UVB, the minimal erythema dose within psoriasis was higher than that for non-lesional skin (medians > 112 and 28 respectively, P < 0.05). Laser–Doppler measurements confirmed that the reduced erythemal sensitivity was not due to masking of response by pre-existing increased blood flux within psoriasis. In six patients, the sites subsequently treated twice weekly with PUVA, using high-dose UVA, cleared faster (median number of treatments 3), but with a similar cumulative UVA dose, compared with adjacent lesional skin treated with conventional PUVA (median number of treatments 12). This study demonstrates that psoriasis may clear rapidly, without burning, using high-dose UVA. Availability of a suitable irradiation apparatus would allow rapid and effective PUVA treatment to be used for localized, resistant disease.     

Comparison of turbo‐PUVA and conventional American‐style PUVA in the treatment of psoriatic patients

Background: Ultraviolet (UV)A protective properties of dihydroxyacetone (DHA) have been used as a topical UV‐resisting barrier to optimize psoralens and UVA (turbo‐PUVA). Starting doses and increments were based on the DHA diffuse reflectance spectroscopy‐derived protection factor. Objective: To evaluate the efficacy of turbo‐PUVA in psoriatic patients using a simpler method for determining starting doses and increments, in comparison to the conventional American‐style PUVA photochemotherapy. Methods: Thirty psoriasis patients (15 on American‐style PUVA and 15 on turbo‐PUVA) were evaluated, each receiving PUVA twice weekly. Starting UVA dose was determined according to skin phototype for the American‐style PUVA group and according to the patient's skin phototype × DHA SPF 3 in turbo‐PUVA group. UVA increments used were 0.5–1.5 J/cm² per treatment in American‐style PUVA and 25% of the previous dose in turbo‐PUVA. Results: Turbo‐PUVA group showed a significantly lower mean cumulative dose, a significantly higher psoriasis area and severity index score reduction, lesser mean number of treatment sessions, and less duration of treatment till remission (188.44±106.2 J/cm², 92.164±1.975%, 11.2±3.52 session, and 1.4±0.44 months, respectively) than conventional American‐style PUVA group (255.13±18.304 J/cm², 74.725±10.976%, 30±0.00 sessions, and 3.75±0.00 months, respectively). Conclusion: Turbo‐PUVA is more effective and time convenient for the treatment of psoriasis with less cumulative dose than the conventional American‐style PUVA.     

Melanin granula distribution and phagocytosis in psoriasis vulgaris after PUVA therapy

Summary Melanin-containing basal cells of the epidermis, melanin-containing macrophages, mast cells, eosinophilic granulocytes and plasma cells were quantitatively investigated with the purpose of gaining an understanding of the quantitative changes in these cell systems under PUVA therapy. This patients have been exposed to solar radiation some weeks ore months before the begin of the PUVA-treatment. Different dying-processes were used to investigate biopsy samples of psoriatically healthy and psoriatically affected skin, from 28 patients before, and 39 patients after PUVA therapy, using a 2 d m with a field of view of 0.1 mm². Altogether more than 9,000 fields of view have been analysed. The average radiation amount was 12 irradiations with an average total energy of 21.5 J/cm². It was found that the count of granula-containing basal layer cells decreases in the psoriatic healthy region due to pigment incontinence and increase in the psoriatically affected region. The subepidermal melanincontaining phagocytes increase in both regions to a similar extent. In the case of the mast cells there was no trend to degranulation. The count of eosinophilic granulocytes and plasma cells was unchanged.     

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.     

Exploring the role of a nonablative laser (1320 nm cooltouch laser) in skin photorejuvenation

Background: Nonablative laser has been used for the treatment and prevention of skin aging for many years. Although the mechanism has not been elucidated, histological evaluation showed that the dermal fibroblasts and its collagen production should be the main target for this rejuvenation. In order to determine the effects of a 1320 nm nonablative laser on the human dermal fibroblasts, the two main secretion factors, basic fibroblast growth factor (bFGF) and transforming growth factor β1(TGF‐β1), in vitro were detected. Methods: The human dermal fibroblasts were cultured and irradiated with a 1320 nm laser at the dose of 15, 20, and 24 J/cm² respectively. The number of fibroblasts was counted and the levels of bFGF and TGF‐β1 were detected by enzyme‐linked immunosorbent assay at the time points of 0, 24, 48, and 72 h after irradiation. Results: The results showed that both the number of fibroblasts and the secretion of bFGF increased after the irradiation at the dose of 20 and 24 J/cm² (P<0.05) compared with that of the control cells. The bFGF secretion in the group exposed to 20 J/cm² was more significant than that of 24 J/cm², and the peak level was 24 h after irradiation. The level of TGF‐β1 secretion decreased after irradiation in a dose‐dependent manner (15 and 20 J/cm², both P<0.05; 24 J/cm², P<0.01), and reached a nadir at 24 h. Conclusion: Our results suggested that the 1320 nm nonablative laser accelerates the vitality of fibroblasts, promotes the secretion of bFGF, and inhibits TGF‐β1 secretion by fibroblasts.     

Skin photosensitizing and Langerhans' cell depleting activity of topical (bath) PUVA therapy: comparison of trimethylpsoralen and 8-methoxypsoralen

The skin photosensitizing and Langerhans' cell (LC) depleting effects of a single bath PUVA exposure were studied in 22 healthy young volunteers. The photosensitizing effect of bathing for 15 min in a 0.2 mg/1 trioxsalen-water solution was about 30 times as great as a similar treatment in an equipotent methoxsalen solution. The skin erythema induced by methoxsalen bath PUVA peaked on day 2 and diminished thereafter, whereas the trioxsalen reaction showed a broad plateau on days 2-5 after the irradiation. A reduction in LC density to about 30-40% of the starting value was seen in both trioxsalen and methoxsalen bath PUVA treated skin sites on day 4 after irradiation, and low or diminishing LC counts prevailed until day 10-11. The amount of UVA needed to produce a similar degree of LC depletion was 15-30 times as great in the case of methoxsalen, compared with trioxsalen. A perceptible erythema reaction, however, was, not a prerequisite for a reduction in LC density.     

Chronic exposure of Sk-1 hairless mice to narrow-band ultraviolet A (320–355 nm)

Several recent investigations collectively suggest that the role of ultraviolet A (UVA) in chronic actinic skin damage may be greater than originally thought. In the present work, the output of a xenon-arc solar-simulator passed through a Bausch & Lomb monochromator in conjunction with a 2-mm Schott WG-320 filter produced narrow-band UVA centered at 338 nm, half-band width 24 nm, I_o=3.4±0.3 mW/cm². We chronically irradiated 10 Sk-1 albino hairless mice 5 times per week for 18 weeks, starting with 1.25 J/cm², for 33 irradiation days, sequentially followed by 1.50 J/cm² (34 days), 1.8 J/cm² (10 days), 2.0 J/cm² (22 days) to afford a total UVA dose of 154.3 J/cm² over 99 irradiation days. Erythema was noted clinically by day 6, which persisted throughout the irradiation. During the irradiation period, some scaling, consistent with mild epidermal hyperplasia was noted during irradiation days 37–56. This response later regressed despite continued chronic irradiation. Hematoxylin and eosin examination immediately after the final irradiation revealed a mild inflammatory response, with some dermal restructuring. At the end of the experiment, no significant signs of epidermal hyperplasia or (pre)malignant lesions were seen, although some stratum corneum thickening was noted. Marked dermal collagen damage and moderate elastosis was also evident. We believe that the observed differences in results reported in previous studies are in large part due to differences in light sources and irradiation protocols.     

Successful Treatment of Scleroderma with PUVA Therapy

PUVA therapy was carried out on four patients with scleroderma; three of them had cutaneous manifestations of progressive systemic sclerosis and one other exhibited generalized morphea. PUVA therapy was given with daily doses of 0.25 J/cm² or 0.4 J/cm² for 3–8 weeks, resulting in total doses between 3.5 J/cm² and 9.6 J/cm². All four patients responded well to this treatment; improvements of hand closure, skin sclerosis index, and flexion of fingers or knee joints were obtained. Thus, PUVA appeared to be beneficial for treating scleroderma.     

Suppressive effect of ultraviolet B radiation on contact sensitization in mice. II. Systemic immunosuppression is modulated by ultraviolet irradiation and hapten application

Irradiation of mice with ultraviolet B (UVB) can suppress contact hypersensitivity “systemically”, even if hapten is applied to the non-irradiated skin site. We previously reported the factors influencing UVB-induced “local” immunosuppression. To obtain the most effective systemic immunosuppression, we further investigated the effect of the following factors on contact hypersensitivity to dinitrofluorobenzene (DNFB): UVB dose, dividing exposure, timing of sensitization after irradiation, area of exposure, hapten concentration, age, and genetic basis. The suppression was enhanced by increasing UVB dose. When 1 J/cm² of UVB was exposed, 4 daily divided exposures (0.25 J/cm²x4) was more suppressive than a single (1 J/cm²x1) or double divided (0.5 J/cm²x2) exposure. Five or 10 day intervals between irradiation and sensitization induced stronger suppression than 1 or 3 day intervals. When the total energy (Joule, J) was kept constant, the exposure of low dose-UVB to a large area (0.5 J/ cm² × 16.45 cm²) suppressed contact hypersensitivity more strongly than did high dose-UVB to a small area (2 J/cm²x4.11 cm²). When 25 ml of DNFB solution was applied, high concentration induced lower suppression. The stronger suppression was most prominant in the young (7 week) than in the old (22 week) mice. No difference was found in the systemic immunosuppression between C3H/HeN and Balb/c mice. These results suggest that not only UVB dose but also various factors should be taken into consideration to effectively induce systemic immunosuppression.     

Personal solar UV-A doses received by patients undergoing oral psoralen photochemotherapy for psoriasis

Patients undergoing oral psoralen photochemotherapy (PUVA) for psoriasis receive a known amount of UV-A in the treatment cubicle and an unknown amount of UV-A from sunlight. In order to measure the solar UV-A dose, fifty-six patients received a UV-sensitive film badge to be worn from the time they took their psoralen tablet throughout the course of the day, except during treatment. The UV-A doses measured in this way varied from less than 0.2 J cm^-2 to 7 J cm^-2, with a median of around 1 J cm^-2. These results show that the solar UV-A received by patients who have been photosensitized by 8-MOP may not be insignificant in relation to the treatment dose.     

Cell kinetic studies of the effect of 8-methoxypsoralen with long wave ultraviolet irradiation on human melanoma cell line (SEKI, II)

The growth inhibition and cell cycle phase distribution of a human malignant melanoma cell line (SEKI, II) treated with 8-methoxypsoralen (8MOP) and UVA irradiation (PUVA) were investigated in vitro. A growth inhibition effect was observed in 8 methoxypsoralen (8MOP) 0.1 μg/ml + UVA 1 J/cm² group, and a strong cytocidal effect was noted in the 8MOP 1 μg/ml + UVA 3 J/cm² groups. Continued accumulation was observed at 24 to 72 hours in the following phases: G₂M phase in the 8MOP 0.1 μg/ml + UVA 1 J/cm² group; middle S phase in the 8MOP 0.1 μg/ml + UVA 3 J/cm² group; and early S phase in the 8MOP 1 μg/ml + UVA 1 J/cm² group. It appeared that 8MOP with UVA irradiation acts mainly on the S phase to block the cell cycle progression between the S and G₂M phases. The variation of cell cycle phase distribution under the effect of 8MOP with UVA irradiation was similar to that observed with the alkylating agent, ACNU, suggesting a possible similarity of the mechanism of action between them. The reaction in the dark was vital for the onset of action of 8MOP with UVA irradiation; as is the case for DTIC with UVA irradiation (32).     

Antioxidant defence mechanism of the skin against UV irradiation: Study of the role of catalase using acatalasaemia fibroblasts

To clarify the role of catalase, an antioxidant enzyme, in response to UV irradiation, we compared the effects of irradiation on cytotoxicity, activities of antioxidant enzymes, total glutathione concentrations, lipid peroxidation and the rate of collagen synthesis in skin fibroblasts from a patient with acatalasaemia and in those from a normal individual. The cells were irradiated with UVA (6 and 12 J/cm² or UVB (0.5 and 1 J/cm²). Cell survival curves after UV irradiation were similar in cells from both subjects. Although superoxide dismutase activity in acatalasaemia cells was higher than in the control cells before irradiation, after irradiation the activity decreased in acatalasaemia cells (76% with 12 J/cm² UVA, 47% with 1 J/cm² UVB), but remained unchanged in control cells. Total glutathione concentrations also decreased in acatalasaemia cells (60% with 12 J/cm²) in response to UVA irradiation, but remained unchanged in control cells. Lipid peroxidation did not increase significantly in either cell type. The rate of collagen synthesis decreased to a similar extent in response to UV exposure in the two cell types (60–80% with 8.2 J/cm² UVA, 40–50% with 10 mJ/cm² UVB). We conclude from the results of cytotoxicity and lipid peroxidation that although acatalasaemia cells were killed by hydrogen peroxide at low concentrations with a single UV exposure, catalase functions only to a small degree as an antioxidant enzyme. There remains the possibility, however, that a deficiency of catalase may chronically damage the skin resulting in a reduced defence function of Superoxide dismutase and glutathione with repeated exposures to UV, which is becoming more common in our daily life.     

Combination therapy of oral methoxypsoralen: photochemotherapy (PUVA) and an aromatic retinoid (etretinate, tigason) in the treatment of psoriasis

RePUVA is the combination of an oral aromatic retinoid, etretinate, with oral photochemotherapy, PUVA. This combination therapy has the advantage of faster clearance with fewer side effects. In Middle Road Hospital, we treated 29 patients with extensive psoriasis (more than 30% involvement) with a total 32 RePUVA treatments. There was complete clearance in 69% of patients with an average of 19 irradiations and an average total ultraviolet A (UVA) dose of 183 J/cm². This compared well with standard PUVA which needed 25 radiations and a total UVA dose of 346 J/cm². The main side effects were cheilitis, exfoliation of skin and pruritus. Biochemically, 41% of the patients showed elevations of serum triglyceride.     

Efficacy and safety of bexarotene combined with psoralen-ultraviolet A (PUVA) compared with PUVA treatment alone in stage IB-IIA mycosis fungoides: final results from the EORTC Cutaneous Lymphoma Task Force phase III randomized clinical trial (NCT00056056)

Background Psoralen plus ultraviolet A (PUVA) is the standard treatment for early stages of mycosis fungoides. There have been no adequate randomized controlled trials with sufficient power comparing this modality with other therapies. Objective To assess disease response and to compare the response rates of patients treated with PUVA alone or PUVA and bexarotene. Methods EORTC 21011 (NCT 00056056) was a randomized phase III study comparing combined bexarotene (Targretin^®) and PUVA vs. PUVA alone in patients with stage IB and IIA mycosis fungoides (MF). The primary endpoint was the overall response rate [complete clinical response (CCR) plus partial response (PR)]. Results The study was prematurely closed due to low accrual after 93 of 145 required patients (65%) were randomized. Of the 93 randomized patients, 87 started treatment, 41 received PUVA and 46 received PUVA + bexarotene. Total UVA doses received were 107 J cm^?2 (range 1·4–489·9) in the PUVA arm vs. 101·7 J cm^?2 (0·2–529·9) in the combination arm. The safety profile was acceptable with few grade 3–4 toxicities observed in either arm. More drop‐outs due to toxicity were observed in the combination arm compared with the PUVA‐alone arm. The best overall response (CCR + PR) rate was 71% for PUVA alone and 77% for the combination arm (P = 0·57). The median duration of response was 9·7 months for PUVA vs. 5·8 months for the combination arm (P = 0·33). CCR was seen in 25 patients of whom 10 received PUVA alone (CCR 22%) and 15 received combination therapy (CCR 31%) (P = 0·45). CCR was sustained in 25% of patients regardless of therapy. There was a trend towards fewer PUVA sessions needed to achieve CCR in the combination arm (median 22) compared with the PUVA arm (median 27·5) (P = 0·11). Similarly, a trend towards lower UVA dose required to achieve CCR in the combination arm (median 55·8 J cm^?2) compared with the PUVA arm alone (median 117·5 J cm^?2) (P = 0·5) was observed. Conclusions No significant difference in response rate or response duration was observed in this study. However, there was a trend towards fewer PUVA sessions and lower UVA dose required to achieve CCR in the combination arm (PUVA + bexarotene) but this did not achieve statistical significance due to insufficient power.     

A trial of accelerated PUVA in psoriasis

Ten psoriatic patients were treated with an accelerated PUVA regimen. Nine patients cleared in an average of 13·7 treatments (range 4-24) after an average total UVA dose of 83 J/cm² (range 18-186). One patient had to withdraw due to pruritus and burning and six others had burning problems. The high incidence of burning may limit the routine use of this irradiation regimen.