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Cyclosporine, an immunosuppressant drug often administered to organ transplant recipients, is also effective against some forms of psoriasis (a chronic skin disease involving plaques and scales). To learn more concerning the optimal dose of cyclosporine for treating this disorder, a 16-week controlled study of 85 patients was carried out. The patients had at least one fourth of their body surface covered by plaque-type psoriasis, or severe psoriasis that was disabling, and an unsatisfactory response to standard treatments for the disease (ultraviolet light or psoralen). Patients were randomly assigned to receive daily doses of 7.5 milligrams (15 patients), 5 milligrams (20 patients), or 3 milligrams (25 patients) of oral cyclosporine per kilogram of body weight. Twenty-five patients received an inactive drug (the control group). After the first eight weeks, patients who did not improve (including controls) were adjusted to the next higher dose (those at 7.5 mg/kg went up to 10 mg/kg). Three specific lesions on each patient were rated at regular intervals during the study for scaling, redness, plaque thickness, and overall severity. In addition, overall severity and extent of involvement of the body were rated. Kidney function (which is affected by cyclosporine in many people), blood pressure, white blood cell count, and other physiological variables were also evaluated. All patients who received cyclosporine improved more than patients who received the vehicle; for the three dose levels (from lowest to highest), the decrease in the psoriasis global scores was 39, 58, and 71 percent, respectively. The best effect was in the highest-dose group (80 percent clear or almost clear of lesions). Adjustments to higher doses were required for the largest proportion of patients in the lowest-dose group (65 percent). Four patients left the study because of side effects, and four required dose reduction. Effects on renal and neurologic function were noted. The results indicate that the best initial cyclosporine dose for treating plaque-type psoriasis is 5 milligrams per kilogram per day. Additional long-term studies are needed to evaluate the drug's safety and effectiveness. (Consumer Summary produced by Reliance Medical Information, Inc.)

Oral lichen planus is a condition affecting the mouth and involving multiple areas of lacelike lesions or plaques that may be erosive. Oral lesions are more resistant to therapy and less likely to undergo remission than cutaneous forms of the disorder. Treatment of oral lichen planus is aimed primarily at relieving discomfort. A total of 16 patients with oral lichen planus and accompanying discomfort were studied; eight patients received cyclosporine, while eight received only the vehicle. Cyclosporine or the placebo was administered as a swish-and-spit medication three times a day for eight weeks. All eight patients who received the active drug experienced marked improvement in erosion, inflammation, pain, and the number of lesions. One patient who received the vehicle had moderate improvement and the remainder either were unaffected or experienced flare-ups of their ulcerations. Patients receiving the vehicle were later treated with cyclosporine and had clinical responses equivalent to those of the patients who received cyclosporine. Treatment of oral lichen planus is difficult and discontinuation of therapy frequently results in a relapse. Corticosteroids have been applied topically or injected, and retinoids have also been used to treat oral lichen planus; however, long-term use of these agents is not recommended because of their potential toxicity. The cause of oral lichen planus is unknown; however, treatment response from the agents used successfully indicates a possible role of the immune system. (Consumer Summary produced by Reliance Medical Information, Inc.)

Changes in the cyclic nucleotide modulation of the epidermis could be involved in the altered state psoriatic lesional tissue. Since several conflicting values for cyclic AMP levels in the diseased state have been reported and cyclic GMP levels have been determined in only a small number of psoriatic patients, we have attempted to define the changes in epidermal cyclic nucleotide levels that can occur between psoriatic and normal epidermis. In this study, 34 normal volunteers and 28 persons with classical psoriasis vulgaris were biopsied using a Castroviejo keratome. The cyclic AMP and cyclic GMP levels of the snap frozen sample, which consists almost entirely of epidermal tissue, were determined by acetylated RIA of partially purified cyclic nucleotide fractions obtained by ion-exchange chromatography. The results indicate that cyclic GMP levels are increased 132% (NDA date base) in the lesional tissue when compared to the uninvolved areas. This is in agreement with our previously reported values obtained from a small number of biopsies. However, in contrast to our previous work, no decrease in lesional cyclic AMP in comparison with uninvolved epidermis was detected, although the content of cyclic AMP in all psoriatic tissue (both involved and uninvolved) was now observed to be decreased when compared to the normal values that have been obtained in our past and this present study. We conclude that cyclic AMP and cyclic GMP assays of biopsied epidermal tissue obtained form a large and varied human population can only suggest the involvement of cyclic nucleotides in the pathophysiology of the disease. Other systems and techniques are presently being used and developed to define the role of cyclic nucleotides in epidermal function and to formulate hypothesis on which to base further investigations of the misregulation of cyclic AMP and cyclic GMP in psoriatic tissue. [ABSTRACT FROM AUTHOR]

Of the many major achievements in cell biology during the last 25 years, none is more important than the understanding of regulation of cell cycles. In 1953 two fundamental observations concerning DNA were made. Watson and Crick suggested that the three-dimensional structure of DNA exists as a double helix with specific base pairings, and Howard and Peic observed that DNA is replicated during a specific phase in the mitotic cycle. Thus developed the theory of cell cycles. Next, investigators explored which events occur during each phase of the cycle and what controls the readout of the genes for proliferation or differentiation. In 1961, Jacob and Monod proposed that for prokaryotic cells the operon is the mechanism which controls the readout of the genes; and by the end of the 1960s, several investigators had defined the role of cyclic AMP and its mechanism of action at the gene level. The control mechanisms for eukaryotic cells are less well defined. Basically there are two types of regulatory molecules: those that arrive at the cell surface and send messages inside the cell; and those that enter the cell, bind to receptors, and then enter the nucleus to interact with the genes. During the past five to ten years, the cell surface and its receptors have received considerable attention as the recognition and control areas for cell proliferation and differentiation, and currently the role of the cyclic nucleotides and prostaglandins is being investigated. Various model systems are now available for detailed studies of these control mechanisms. [ABSTRACT FROM AUTHOR]

How the constantly renewing epidermis maintains its homeostatic state remains a mystery. A "negative feedback" mechanism of control with chalone, a tissue-specific, species-nonspecific inhibitor of mitosis, has been suggested by Bullough and Laurence. Progress in isolating, purifying, and characterizing epidermal chalone has been slow and difficult because of the lack of a fast, accurate, and meaningful assay system. A G2 assay (depending upon mitotic counts) was used to obtain the data which served as the basis for postulating the existence of a chalone. Since a control point within the G1 phase of the cell cycle is considered to he the one with physiologic implications, in vivo and in vitro assays with pure primary epidermal cell cultures have been used to investigate a G1 block. The incorporation of tritiated thymidine into DNA is often used as a measure of the rate of proliferation. However, inhibition of thymidine transport and phosphorylation have been encountered in several systems and negate this method as an accurate measure of cell proliferation. [ABSTRACT FROM AUTHOR]

The formation of all-trans retinoic acid is an oxidative process whereby retinol is converted to retinaldehyde and then to retinoic acid. Because retinol causes qualitative molecular changes similar to those produced by retinoic acid, we compared potency of retinol, retinaldehyde, and retinyl palmitate to retinoic acid and assessed the effects of occlusion. Retinoids were prepared in an experimental vehicle of 95% ethanol:propylene glycol (7:3) with anti-oxidant. Induction of retinoic acid 4-hydroxylase activity was the end point for comparison. Retinoic acid concentrations from 0.001% to 0.05% under occlusion produced a linear dose-response induction of 4-hy- droxylase activity. The concentrations of the other retinoids under occlusion required to achieve significant induction of enzyme activity were 0.6% retinyl palmitate, 0.025% retinol, and 0.01% retinaldehyde. The linear dose-response was lost with retinoid concentrations in excess of 0.25% retinol or 0.5% retinaldehyde. Statistical analyses showed no difference in 4-hydroxylase activity between unocciuded and occluded retinol treated sites. By contrast, however, unoccluded sites treated with retinoic acid or retinyl palmitate had less induction of 4-hydroxylase activity than occluded sites. Retinol, retinaldehyde, and retinyl palmitate did not produce erythema but did increase epidermal thickness. Although retinol is a weaker retinoid than retinoic acid, the increased penetration of unoccluded retinol in comparison to unoccluded retinoic acid with this prototypic vehicle confers on retinol a more effective delivery of a retinoidal effect than unocciuded retinoic acid. Retinol at 0.25% may be a useful retinoid for application without occlusion because it does not irritate but does induce cellular and molecular changes similar to those observed with application of 0.025% retinoic acid. [ABSTRACT FROM AUTHOR]

Vitamin A, all-trans-retinol, is metabolized to retinoic acid in vivo by a tightly controlled two-step conversion. Retinoic acid then binds to nuclear receptors and modulates cellular proliferation and differentiation. Because only a small fraction of retinol applied topically can be metabolized to retinoic acid, alternative pathways of retinol metabolism in skin were investigated. Retinol (0.4%) was applied to adult human skin under occlusion for 6 h to 4 d. The conversion of retinal into various metabolites such as 14-hydroxy-4,14-retro-retinol, anhydroretinol, 4-oxoretinol, retinyl esters, and retinyl glucuronides was investigated. The level of 14-hydroxy-retro-retinol was increased from undetectable at time 0 to 326 ng/g wet weight of tissue at 6 h (6% of the retinol level) and maintained approximately the same concentration at 24 h to 409 ng/g wet weight (1.9% of the retinol level); it decreased to 48 ng/g wet weight of tissue (12% of its maximum level) by 4 d. Anhydroretinol was undetectable at time 0, increased only slightly at 6 h, and remained at the same level. We did not detect 4-oxoretinol. Because 14-hydroxy-retro-retinol was found in the retinol-treated areas, its effects on epidermis were compared with those of retinol. Topical application of trans-retinol (0.3%) significantly increased both epidermal thickness and cellular retinoic acid binding protein II mRNA, whereas 14-hydroxy-4,14-retro-retinol (0.3%) did not increase either of these well-characterized cutaneous retinoid responses. Retinol, when applied topically in pharmacologic doses to human epidermis, remained as free retinol, was metabolized primarily to retinol ester, and was metabolized to a lesser extent to retro-retinoids and didehydroretinol. [ABSTRACT FROM AUTHOR]

Metabolic inactivation of all-trans retinoic acid to 4-hydroxy retinoic acid occurs via a cytochrome P-450 enzyme. We investigated the effects of liarozole on the retinoic acid 4-hydroxylase activity of human epidermis and its ability to modify in vivo human skin responses to retinoic acid and all-trans retinol. Retinoic acid 4-hydroxylase activity induced in vivo by 4 d treatment with retinoic acid (0.1%) was inhibited in vitro by liarozole in a concentration-dependent manner. Comparable micromolar concentrations of liarozole were extracted from stratum corneum-free epidermis treated with 3% liarozole. Retinoic acid levels in liarozole-treated skin increased to 19 ± 5 ng/g wet wt (mean ± SEM, p < 0.002,n = 17) at 18 h and to 6 ± 2 ng/g wet wt (p = 0.38, n = 17) at 48 h as compared to vehicle (not detectable). At 48 h, retinoic acid 4-hydroxylase activity was induced 9-fold over vehicle (p < 0.03, n = 8). At 96 h, no significant erythema or increased epidermal thickness was found when either retinoic acid (0.001%), all-trans retinol (0.025%), or liarozole (3%) was applied individually, but when 0.001% retinoic acid and 3% liarozole were applied together, both erythema and increased epidermal thickness occurred. In contrast, 0.025% all-trans retinol and 3% liarozole together caused increased epidermal thickness but no erythema. These data demonstrate that, at doses used here, liarozole, although an effective inhibitor of retinoic acid 4-hydiroxylase, cannot function alone like a retinoid in vivo, probably because of retinoic acid 4-hydroxylase induction. In the presence of a low dose retinoic acid or all-trans retinol, however, liarozole can amplify human skin responses to each retinoid in a manner characteristic of the retinoid at a higher dose (erythema and hyperplasia with retinoic acid; no erythema but hyperplasia with all-trans retinol). [ABSTRACT FROM AUTHOR]

Application of all-trans retinoic acid to human skin for 4 d under occlusion produces a marked increase in retinoic acid 4-hydroxylase activity. In this study, the possible induction of other hydroxylases in response to 9-cis and 13-cis retinoic acid applications to adult human skin in vivo was determined. Application of 0.1% all-trans, 0.1% 9-cis, and 0.1% 13-cis retinoic acid to human skin for 2 d resulted in induction of only all-trans retinoic acid 4-hydroxylase activity. The 4-hydroxylase activity in microsomes from the treated tissue ranged from 383 ± 46 to 531 ± 59 pg of 4-hydroxy all-trans retinoic acid formed/min/mg protein (n = 6). These same preparations were unable to use 9-cis or 13-cis retinoic acid as substrate for the hydroxylation reaction. Extraction of the retinoic acid isomers from epidermis 48 h after application of 0.1% solution of each isomer yielded significant amounts of all-trans retinoic acid (36-72%) regardless of the isomer applied. The all-trans isomer produced by isomerization of both 9-cis and 13-cis retinoic acids is the likely inducer of the 4-hydroxylase. All-trans retinol and all-trans retinal were unable to compete with all-trans retinoic acid as substrate for 4-hydroxylase enzyme. The 4-hydroxylase induced in response to pharmacological doses of retinoic acids is specific for the all-trans isomer. The inability of 9-cis or 13-cis retinoic acid to induce their own hydroxylation and inactivation or act as substrate for the 4-hydroxylase in skin may have considerable implications in light of the clinical use of retinoids in the treatment of various diseases including cancers. [ABSTRACT FROM AUTHOR]

We investigated the clinical, histologic, and molecular responses of normal human skin to all-trans-retinol (ROL) application, compared to those induced by topical all-trans-retinoic acid (RA), and measured ROL-derived metabolites. Up to 1.6% ROL, 0.025% RA in vehicle (70% ethanol/30% propylene glycol), or vehicle alone were applied in a double-blind fashion to normal buttock skin and occluded for 4 d. ROL produced from none to only trace erythema, which was clinically and statistically insignificant, whereas RA induced a significant 3.7-fold increase in erythema score compared to vehicle (n = 10, p < 0.01). However, ROL induced significant epidermal thickening (1.5-fold at 1.6% ROL, p < 0.01), similar to RA (1.6-fold at 0.025% RA, p < 0.01), relative to the vehicle. ROL, compared with vehicle, also increased mRNA levels of cellular retinoic acid binding protein (CRABP-II) and cellular retinol binding protein (CRBP) genes as determined by Northern analysis (5- 6-fold and 6-7-fold, respectively) and riboprobe in situ hybridization. CRABP-II and CRBP protein levels were also higher following ROL than vehicle treatment, as measured by ligand binding (3.2-fold, p < 0.001; n = 7) and Western analysis (3.6-fold, p < 0.003; n = 6), respectively. Epidermal retinyl ester (RE) content, measured after removal of stratum corneum, rose 240-fold (p < 0.005, n = 5) by 24 h of ROL occlusion. RA content, however, was undetectable or detectable only at trace amounts in all samples obtained at 0, 6, 24, and 96 h after ROL occlusion. Detectability of RA was not correlated with ROL treatment (compared to untreated normal skin, p = 0.86) or baseline skin ROL levels (average r = -0.1, p> 0.3). [ABSTRACT FROM AUTHOR]

The polyunsaturated fatty acids linoleic acid (18:2, n-6) and arachidonic acid (20:4, n-6) are essential for normal skin function and structure, both as eicosanoid precursors and as components of lipids forming cell membranes. Adult human keratinocytes grow optimally in serum-free medium (MCDB 153) that contains no fatty acids. These keratinocytes expand rapidly and produce normal epidermis upon in vivo grafting. Analysis of lipid extracts of epidermis and of cultured keratinocytes was done to determine the fatty acid composition of cells grown in essential fatty acid (EFA) - deficient medium. Gas chromatography and high-performance liquid chromatography analyses were done of the fatty acids in the entire cell and in a thin-layer chromatography separated fraction containing those lipids that form cellular membranes. Comparison of snap-frozen epidermis and epidermal basal cell suspensions to passage 1 to 4 cultures shows that the cells are in an extreme essential fatty acid-deficient state by the first passage. The amount of the saturated fatty adds 16:0, 18:0, and 14:0 is unchanged by culture. The polyunsaturated fatty acids are found to be significantly decreased, the cells balancing their lack with a significant increase in the relative abundance of the monounsaturated fatty acids, 18:1 and 16:1. Greater than 85-90% of the fatty acids was found in lipids associated with membranes and no unusual fatty acids were detected. Because the serum-free medium is fatty acid free and the cells cannot synthesize essential fatty acids, the rapid division of the cells results in the predominance of an extreme EFA-deficient cell type. The essential fatty acid - deficient keratinocyte is an excellent adult, normal epidermal cell model that can be used to study EFA deficiency and the effect of the eicosanoid and fatty acids on cell function and structure. [ABSTRACT FROM AUTHOR]

Cyclosporine A is efficacious in the treatment of psoriasis when taken orally or injected intralesionally but not topically. Lack of penetration to necessary locations or rapid metabolism during passage through the epidermis may account for the ineffectiveness. Cytochromes P-450III in the liver are known to be involved in cyclosporine metabolism and inactivation. This study was undertaken to determine if an epidermal cytochrome P-450III exists that can inactivate topical cyclosporine A. Rats were treated with the macrolide antibiotic erythromycin to induce the cytochrome P-450III family of enzymes. Microsomal fractions were prepared from liver and epidermis of rats and from lesional areas of psoriasis patients. NADPH cytochrome C reductase activity was determined as a positive control for microsomal enzymatic activity. Formation of metabolite 1, the predominant metabolite of cyclosporine A, by liver microsomes was increased 193% after 10 d erythromycin treatment. The cytochrome P-450 dependent activity in microsomes from the epidermis of control and erythromycin-treated rats and in microsomes from psoriatic tissue was at the detection limits of the assay system. Cytochrome P-450III gene family mRNA were detectable by polymerase chain reaction in liver but not in psoriatic or normal epidermis. The lack of detectable P-450III mRNA and the absence or minimal conversion of cyclosporine A to inactive metabolites by epidermal microsomes suggest that the ineffectiveness of topical cyclosporine A in psoriasis may not be due to inactivation of cyclosporine A by cytochrome P-450 in the skin. [ABSTRACT FROM AUTHOR]

We have simultaneously measured epidermal DNA synthesis, epidermal thickness and cyclic nucleotide levels in essential fatty acid (EFA) deficient hairless mice skin. In vitro epidermal DNA synthsis and epidermal thickness were significantly increased in EFA deficient epidermis. Cyclic AMP levels were significantly reduced when expressed using three different data bases. Cyclic GMP levels were increased using two data bases but these increases were not statistically significant. Cyclic AMP cyclic GMP ratios were significantly decreased in the EFA deficient skin. The results provide evidence that cyclic nucleotide levels are altered in an animal model of benign epidermal proliferation. [ABSTRACT FROM AUTHOR]

A retinoic acid (RA) inducible skin-specific gene transcript (RIS-1) was isolated by differential hybridization screening of a RA-treated human skin cDNA library. The library was constructed from pooled RNA derived from normal adult human skin treated with all trans-RA for 4 h (n=6) and 12 h (n=6) in vivo. RIS-1 cDNA corresponded to a 0.6 kb transcript that was barely detectable in normal adult human skin but was significantly induced by 8 h in RA-treated compared to vehicle-treated skin (range 1.1-3.6 fold). Prolonged RA treatment for up to 24 h further increased relative RIS-1 mRNA levels by 1.3-5.5 fold. HPLC analysis of the RA content of 0.1% RA-treated skin in vivo revealed significant levels at 6 h (18.8-120.6 ng RA/g wet weight tissue; approximately 240 nM), immediately preceding the time point at which the increased RIS-1 mRNA level was first seen. This concentration of RA also induced the mRNA levels for cellular RA binding protein II (1.6-19 fold), a marker of RA activity in human skin. RIS-1 mRNA was detected by Northern and dot blotting only in normal skin but not in any other normal human tissues examined, indicating a tissue-specific pattern of gene expression. RIS-1 transcripts were detected at very low levels in untreated cultured human epidermal keratinocytes, while no expression was seen in dermal fibroblasts and melanocytes, the other major cell types in skin. Southern analysis of human and mouse DNA indicated the existence of evolutionarily conserved sequences for RIS-1 between these two species. The polypeptide sequence derived from the partial RIS-1 cDNA was found to be identical to the calcium binding domain found in 'psoriasin', a gene whose expression appears to be increased in the skin of psoriasis patients. [ABSTRACT FROM AUTHOR]