Cyclosporin A induced augmentation of the beta-adrenergic adenylate cyclase response of pig epidermis

Summary It has been known that beta-adrenergic adenylate cyclase response is decreased in psoriatic-involved epidermis. Since the immunosuppressive agent, cyclosporin A, is reported to be effective on psoriasis clinically, the effect of cyclosporin A on beta-adrenergic adenylate cyclase response in pig skin was examined in vitro. Therapeutic serum levels of cyclosporin A (100–400 ng/ml) augmented the beta-adrenergic adenylate cyclase response of the epidermis. Highest levels of cyclosporin A (2–20 g/ml) did not have any effect on its response. Both low K _m and high K _m cyclic AMP phosphodiesterases were not affected by cyclosporin A. Therefore, it is suggested that the clinical efficacy of cyclosporin A on psoriasis can be explained partially by its direct effect on the keratinocyte itself.     

Effects of UVB irradiation on epidermal adenylate cyclase responses in vitro: its relation to sunburn cell formation

Summary UVB irradiation augmented the betaadrenergic adenylate cyclase response of pig skin epidermis in vitro. The effect was observed 2–4 h following the irradiation and lasted at least for 48 h. There was no significant difference in cyclic AMP phosphodiesterase activity between control and UVB-irradiated epidermis at lower irradiation dose (150 mJ/cm²), which is the dose of the most marked beta-adrenergic augmentation effect. The augmentation effect was specific to the beta-adrenergic system; adenosine and histamine adenylate cyclase responses were unchanged or decreased depending on the irradiation dose. Histologically, marked sumburn-cell formation was observed following the UVB irradiation.It has been suggested that oxygen intermediates generated by ultraviolet radiation participate in sunburncell formation. The addition of superoxide dismutase (SOD) in the incubation medium significantly inhibited sunburn-cell formation. On the other hand, the beta-adrenergic augmentation effect was not affected by the addition of SOD. Other scavengers of oxygen intermediates (catalase, catalase+SOD, xanthine, or mannitol) did not inhibit the UVB-induced beta-adrenergic augmentation effect. Further, superoxide-anion generating systems (hypoxanthine-xanthine oxidase system and acetaldehyde-xanthine oxidase system) revealed no stimulatory effect on the beta-adrenergic response of epidermis.These results indicate that (a) the UVB-induced beta-adrenergic augmentation effect is inherent to skin and does not depend on systemic factors such as inflammatory infiltrates following UVB irradiation; (b) in contrast to sunburn-cell formation, induction of the beta-adrenergic adenylate cyclase response is not directly associated with oxygen intermediates generated by UVB irradiation.     

Glucocorticoid-induced alteration of beta-adrenergic adenylate cyclase response of epidermis

Summary It has been reported that the beta-adrenergic adenylate cyclase system of the pig epidermis is regulated by glucocorticoids, resulting in the augmentation of epinephrine-induced cyclic-AMP accumulations. Using this phenomenon, we compared the glucocorticoidal potency of three typical glucocorticoids: hydrocortisone, prednisolone and dexamethasone. There was a considerable variation in the magnitude of the glucocorticoid-induced augmentation of the beta-adrenergic response when pig skin that had been obtained on different occasions was used. In each experimental series (using the same pig skin), however, the maximal augmentation effects obtained with these glucocorticoids were approximately the same. The potent glucocorticoid, dexamethasone, demonstrated its effect at lower concentrations than were required for prednisolone, while hydrocortisone required a much higher concentration before its effect was detectable. Thus, despite considerable variations in the magnitude of the glucocorticoid effects, the concentrations required for the glucocorticoid effect were closely associated with the established glucocorticoidal potency which has previously been described.     

Adenylate cyclase-cyclic AMP system in pure epidermis isolated by use of dispase

Epidermal adenylate cyclase systems following dispase treatment were investigated. Dispase is a bacterial neutral protease obtained from Bacillus polymyxa. Following the treatment with dispase, the epidermal sheet is easily peeled off the dermis. Dispase-treated pure epidermal sheets were shown to contain three major (beta-adrenergic-, adenosine-, and histamine-) receptor adenylate cyclase systems. Without phosphodiesterase inhibitors, the intracellular cyclic AMP (cAMP) level reached the maximal level at 3 min. This effect was markedly enhanced by the addition of cAMP phosphodiesterase inhibitor. Among these epidermal adenylate cyclase systems, the most marked cAMP accumulation was observed by histamine, followed by adenosine, and then by epinephrine. The separation of epidermis and dermis following dispase treatment revealed that epidermis contained most of the beta-adrenergic response (87%), whereas the dermis retained a significant proportion of adenosine (26%) and histamine (40%) responses when 0.3 mm thickness skin was studied. Specific antagonists of epinephrine, adenosine, and histamine inhibited the effects of these agents completely. The simultaneous addition of two stimulators into the incubation medium resulted in an additive effect. Beta-augmentations by hydrocortisone, colchicine, and retinoid all remained in the dispase-treated pure epidermal sheets, but beta-augmentations by these drugs were spoiled by trypsin treatment. These results indicate that dispase-treated pure epidermis contains three major (beta-adrenergic-, adenosine-, and histamine-) specific and independent receptor adenylate cyclase systems. Dispase is a very useful tool for investigating the metabolism and regulatory system of keratinocytes without any significant damage to epidermal membrane receptor systems.     

Inhibitory guanine nucleotide binding protein in pig epidermis: regulation of epidermal adenylate cyclase

Summary Islet-activating protein (IAP), one of the pertussis toxins, serving [-³²P]nicotinamide adenine dinucleotide (NAD) as a substrate for ADP ribosylation, radiolabelled a specific pig epidermal membrane protein. The IAP-specific substrate was detectable by sodium dodecyl sulphate-polyacrylamide gel electrophoresis as a single band corresponding to a molecular weight of 40 kDa. The ADP ribosylation catalysed by IAP was inhibited by the addition of Mg²⁺ to the reaction mixture. IAP is known to work on intact cell systems resulting in the ADP ribosylation using intracellular NAD as the ADP ribose donor. Following IAP pretreatment of intact pig epidermis, the epidermal receptor adenylate cyclase responses were markedly increased; all the stimulatory receptor adenylate cyclase reponses (beta-adrenergic, prostaglandin E, adenosine and histamine responses) were significantly increased. Cholera toxin-induced cyclic AMP accumulation was also significantly increased. Forskolin-induced cyclic AMP accumulation was slightly increased after IAP pretreatment, but this was not statistically significant. The IAP-dependent ADP ribosylation of the epidermal 40 kDa membrane protein, which was prepared from the IAP pretreated epidermis, was significantly decreased. It is known that the tumour promoter, phorbol 12-myristate,13-acetate (PMA), decreases stimulatory receptor adenylate cyclase responses of the epidermis. Following the PMA pretreatment, IAP-dependent ADP ribosylation of the epidermal membrane protein was unaffected. Furthermore, following the PMA pretreatment, the IAP-induced increase in the epidermal receptor adenylate cyclase responses still remained. Our results indicate that pig epidermis contains 40 kDa membrane substrate for IAP-dependent ADP ribosylation, which has an inhibitory tonus on the epidermal adenylate cyclase until its ADP ribosylation by IAP. The results are consistent with the view that the epidermis contains an IAP-sensitive inhibitory guanine nucleotide binding protein (Gi) of the adenylate cyclase system. Although PMA decreases epidermal stimulatory receptor adenylate cyclase responses, it is unlikely that PMA reveals its effect through the modulation of Gi.     

Reversible inhibition of keratinocyte thymidine incorporation by the calmodulin antagonist, W-7

Although calmodulin has been suggested as an important regulator of keratinocyte proliferation, its precise role remains unknown. We employed a calmodulin antagonist, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7), to examine the role of calmodulin on keratinocyte proliferation. N-(6 aminohexyl-1-naphthalenesulfonamide (W-5), a chlorine-deficient analogue of W-7 with little anti-calmodulin activity, was used as the control. W-7 markedly inhibited thymidine incorporation of pig epidermis at concentrations close to its anti-calmodulin activity; W-5 had no effect on the thymidine incorporation. The inhibitory effect of W-7 was reversible; the removal of W-7 from the incubation medium resulted in the reinitiation of the thymidine incorporation, suggesting that W-7 is not a cytotoxic agent. These results are consistent with the view that calmodulin is an essential regulator of keratinocyte proliferation. The epidermal beta-adrenergic response, which is decreased in various hyperproliferative epidermal abnormalities, was increased in W-7-treated hypoproliferative epidermis. The epidermal SOD activity, which is also decreased in the hyperproliferative epidermis, however, was not affected by the W-7 treatment.     

Successful treatment of lichen myxoedematosus with PUVA photochemotherapy

Lichen myxoedematosus is an unusual disorder of unknown etiology and pathogenesis. Although several treatments have reportedly been tried, therapeutic efficacies are variable and unsatisfactory. A patient with severe skin changes of this disease was successfully treated with PUVA photochemotherapy. PUVA therapy was carried out using 30 mg of 8-methoxypsoralen orally, and subsequent exposure to UVA starting at 4 J/cm2. The eruption disappeared almost completely after 35 treatments at a cumulative dose of 202 J/cm2. Histologically, mucin deposition was greatly diminished after the therapy. The photochemotherapy may exert its effect directly by inhibiting proliferation of fibroblasts and synthesis of mucopolysaccharides, and also indirectly by immunomodulating action.     

即时型PUVA对郎格罕细胞和接触过敏的影响

在实验动物中研究了即时型光化学疗法对表皮郎格单细胞数量和形态的影响及对接触过敏反应的抑制，并与常规使用的光化学疗法进行了对比，结果表明二种方法间无差异，３Ｊ/ｃｍ２的即时型光化学疗法无红斑反应并能引起郎格罕细胞数量和形态的变化，并且通过诱导抑制性淋巴细胞抑制接触过敏反应。还探讨了有关致癌的可能性。     

PUVA照射与青黛丸口服对银屑病患者血cAMP、cGMP、SOD及LPO含量的影响

目的：研究ＰＵＶＡ及青黛丸口服治疗寻常型银屑病对患者血液中ＳＯＤ活力、ＬＰＯ水平、ｃＡＭＰ及ｃＧＭＰ含量的影响。方法：寻常型银屑病患者７４例，随机分成ＰＵＶＡ照射组（３６例）及青黛丸服用组（３８例），按常规行治疗方案。采用微量指血超氧化物歧化酶（ＳＯＤ）快速测定法测定红细胞内ＳＯＤ活力，荧光分光光度法测定血清脂质过氧化物（ＬＰＯ）值、放射免疫分析法测定血浆ｃＡＭＰ及ｃＧＭＰ含量。结果：治疗前患者ＳＯＤ活力皆低于对照组（Ｐ＜０．０１），ＬＰＯ水平无明显差异，ｃＡＭＰ低于对照组，而ｃＧＭＰ高于对照组。青黛丸口服后，ＳＯＤ活力改善，ｃＡＭＰ有所回升，但ＬＰＯ及ｃＧＭＰ变化不明显；ＰＵＶＡ照射后，ＳＯＤ活力进一步低于疗前水平，ＬＰＯ值则高于疗前，ｃＡＭＰ的改善略优于青黛丸，ｃＧＭＰ明显改善。对两种疗法的疗效、改变上述指标的可能作用机理及其与临床应用的相互关系作了分析。     

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.     

A case of Schamberg's disease responding dramatically to PUVA treatment

Pigmented purpuric dermatoses are a group of chronic, recurrent disorders characterized by purpuric lesions mainly involving the lower extremities. Their etiology is unknown. Treatment options are limited and none of them have proven benefit. Phototherapy has been reported to be effective in a small number of patients in the literature. We present a case of Schamberg's disease showing a dramatic response to psoralen plus ultraviolet A therapy and discuss the current therapeutic options focusing mainly on phototherapy. We believe that phototherapy is a valuable alternative, especially for patients with long-standing and widespread pigmented purpuric dermatitis.     

Comparison of therapeutic efficacy of topical PUVA, oral etretinate, and combined PUVA and etretinate for the treatment of psoriasis and development of PUVA lentigines and antinuclear antibodies

Sixty-two and 38 psoriatic patients were treated with topical PUVA and combined etretinate and topical PUVA (Re-PUVA), respectively. In both groups, 50% of the patients showed initial recovery after 6 weeks and over 90% after 14 weeks. Re-PUVA was more effective than PUVA alone in obtaining complete clearance (p<0.05). To clear psoriasis in 50% of the patients, PUVA and Re-PUVA required 63 and 26 weeks, respectively. Furthermore, the integrated clearance rates after 70 weeks were 50% in PUVA and 63% in Re-PUVA. Each therapy showed a similar remission period; psoriasis recurred in 50% of the patients after 4 months. In addition, 17 patients were treated with oral etretinate, and Re-PUVA was found to be more effective than etretinate monotherapy. Another aim was to determine whether etretinate would inhibit the development of PUVA side effects. Adding etretinate failed to inhibit the production of PUVA lentigines but clearly suppressed antinuclear antibody (ANA) expression. Six of 56 patients treated with PUVA alone developed ANA during the treatment. In marked contrast (p=0.05), ANA was detected in none of 34 patients treated with Re-PUVA.     

The Effects of Non-Interval PUVA Therapy on the Plaque Stage of Mycosis Fungoides

The effectiveness of non-interval topical PUVA treatment was studied in four patients with mycosis fungoides at the plaque stage. Five regions of each patient were exposed to UVA immediately, 30 minutes, 60 minutes, 90 minutes, and 120 minutes, after topical application of 8-methoxypsoralen, respectively. The effects of these treatments were evaluated by clinical appearance and histological findings after the 20th treatment. All five regions were more improved clinically and histologically than the control region, which was not given PUVA therapy. There were no clear differences clinically among these five regions. Biopsy specimens from each region revealed the disappearance of epidermotropism and a marked decrease in atypical mononuclear cell infiltrations in the dermis. From these data, we concluded that there were no clear differences between these five treatments clinically or histologically and that non-interval PUVA therapy is useful for the early stages of mycosis fungoides. To our knowledge, this is the first report of non-interval PUVA therapy for mycosis fungoides.     

PUVA treatment in chromium hypersensitivity: Effect on skin reactivity and lymphocyte functions

Summary Two male patients with longstanding contact sensitivity to chromium were treated with PUVA. One patient, suffering from concomitant photosensitivity, reacted very favorably; his skin lesions cleared and light tolerance increased. This was paralleled by a decrease in the photopatch test reactivity and by the extinction of the patch-test reactivity on PUVA-exposed (pigmented) skin. Patch and photopatch tests on PUVA-shielded skin showed no decrease in skin test reactivity. PUVA-treatment caused a decrease in the number of rosette-forming T cells and an increase in lymphocyte stimulation in both patients. In one patient, abnormally high PHA-induced suppressor cell activities were recorded prior to treatment; after PUVA therapy the values were back to normal. In both patients, the PPD-induced suppressor cell activity of PWM response was clearly increased by PUVA-therapy. Other suppressor cell functions were not much affected. It is concluded that while PUVA-therapy may produce some systemic immunological effects, its abating effect on contact sensitivity and photosensitivity is mainly mediated through local mechanisms in the skin.     

Adenylate cyclase system in fetal rat keratinizing epidermal cells (FRSK cells) and SV40-transformed human keratinocytes

The adenylate cyclase system of FRSK cells, a cultured cell line of fetal rat epidermal keratinocytes, and SV40-transformed human keratinocytes was investigated. Stimulators of the human epidermal adenylate cyclase, epinephrine, adenosine, and prostaglandin E2 increased cyclic AMP levels of these cells. There were marked differences in the stimulatory effects; while epinephrine revealed a much stronger effect than the other stimulators in FRSK cells, epinephrine and prostaglandin E2 revealed similarly marked effects in SV40-transformed cells. Histamine had little or only slight effect on the cyclic AMP levels of these cells. Cholera toxin and forskolin, which work on the stimulatory guanine nucleotide binding protein (Gs) and the catalytic component of adenylate cyclase, respectively, also increased cyclic AMP levels. Northern blot hybridization analysis revealed that both FRSK cells and SV40-transformed human keratinocytes express mRNAs for the beta 2-adrenergic receptor, as well as the stimulatory and inhibitory guanine nucleotide binding proteins (Gs and Gi, respectively). The presence of Gs as well as Gi were confirmed by cholera toxin-, and pertussis toxin (IAP)-induced ADP-ribosylation of membranous proteins of these cells. Our results indicate that both FRSK cells and SV40-transformed human keratinocytes express the fundamental components of the adenylate cyclase system. These cell lines might be useful tools for the analysis of the adenylate cyclase system in epidermal keratinocytes.     

NB-UVB联合8-MOP PUVA治疗小腿斑块状银屑病疗效评价

目的: 评价NB-UVB联合8-甲氧补骨脂素（8-MOP） PUVA治疗斑块状银屑病的疗效。方法: 分别对16例银屑病患者双侧小腿进行PASI评分,一侧给予NB-UVB照射,另一侧给予NB-UVB联合8-MOP PUVA,每周3次,共治疗20次。结果: NB-UVB治疗侧治疗前后PASI评分分别为8.21±2.97和2.31±1.01,差异有统计学意义(P＜0.05);NBUVB联合8-MOP PUVA治疗侧分别为8.33±2.54和1.20±0.93,差异有统计学意义(P＜0.05)。治疗后NBUVB联合8-MOP 治疗侧较NB-UVB治疗侧PASI更低,差异有统计学意义(P＜0.05)。结论：NB-UVB联合8-MOP PUVA可明显促进小腿顽固部位皮损的消退。     

Dimethyl sulfoxide-induced augmentation of adenosine-adenylate cyclase response of pig skin epidermis

Summary Adenosine-adenylate cyclase response in pig skin epidermis showed a specific increase after longterm (24 h) incubation in the presence of 0.5%–1% dimethyl sulfoxide (DMSO). There was no significant difference between control and DMSO-treated epidermis with regard to cyclic AMP (cAMP) phosphodiesterase activity. DMSO had no effect on the basal cAMP levels of epidermis; beta-adrenergic and histamine-adenylate cyclase responses were not affected. The direct addition of DMSO at the time of incubation with various adenylate cyclase stimulators (adenosine, epinephrine, and histamine) had no effect on agonist-induced cAMP accumulation effects. It was concluded that DMSO affected epidermal keratino-cytes during long-term incubation, resulting in a specific increase in the adenosine-adenylate cyclase response. Although the biological significance of this DMSO effect remains to be determined, it should be kept in mind when using DMSO as a solvent for various chemicals in the experiments dealing with epidermal keratinocytes in vitro.     

Cyclic AMP-dependent protein kinase in calf-snout epidermis

Summary The biochemical characteristics of cyclic AMP-dependent protein kinase in calf-snout epidermis were investigated. The activity of cyclic AMP-dependent protein kinase was higher in the lower layer than the upper layer of epidermis. The supernatant of homogenates of the lower layer of calf-snout epidermis was fractionated by DEAE-cellulose chromatography and contained two major peaks of protein kinase activity stimulated by cyclic AMP. This chromtographic pattern is similar to that referred to as Type I and Type II of cyclic AMP-dependent protein kinase in bovine muscle. Both peaks of cyclic AMP-dependent protein kinase in calf-snout epidermis could phosphorylate keratin polypeptides in vitro. The phosphorylation reaction was activated by cyclic AMP and inhibited by a heat-stable inhibitor of cyclic AMP-dependent protein kinase. When Type II enzyme of cyclic AMP-dependent protein kinase was incubated with [-³²P]ATP in the absence of substrates, such as histone or keratin polypeptides, the 54,000 dalton protein was phosphorylated and this autophosphorylation was inhibited by the addition of 10 M cyclic AMP. These results suggest that cyclic AMP-dependent protein kinase in calf-snout epidermis has properties similar to those in bovine muscle and plays an important role in the phosphorylation of keratin polypeptides in calf-snout epidermis.