Fishing for allergens hiding as prohaptens

Enzymes of the cytochrome P450 (CYP) superfamily are the most versatile and important class of drug-metabolizing enzymes, involved in the metabolism of xenobiotics to which human skin is exposed. Bergstr?m et al. used a recombinant human CYP cocktail in ratios that simulate those found in human skin for the purpose of identifying prohapten metabolites of otherwise inert molecules.     

Cytochrome P450, CYP26AI, is expressed at low levels in human epidermal keratinocytes and is not retinoic acid-inducible

Retinoids, and their synthetic analogues, are well-established regulators of the squamous differentiation programme both in vivo and in vitro. Despite this, very few studies have focused on the mechanism by which retinoid action is terminated, e.g. metabolism. Recently, a new cytochrome P450 family member (CYP26AI) was cloned. CYP26AI was reported to have substrate specificity for retinoids and to be retinoid-inducible. In this study, we have examined the expression and retinoic acid (RA) inducibility of CYP26AI in human epidermis and cultured keratinocytes. We found very low levels of CYP26AI mRNA expression in both epidermis and keratinocytes. Furthermore, we found no evidence for RA inducibility of CYP26 mRNA expression. This lack of RA inducibility was not due to inactivity of the retinoids, as we show that transglutaminase was still repressed by RA in the same cultures. Despite the low levels of CYP26AI expression in the keratinocytes, the keratinocytes were still capable of significant RA metabolism. In conclusion, our study reports, for the first time, that CYP26AI is unlikely to contribute to RA metabolism in keratinocytes. These studies also indicate that as yet unknown isoforms of cytochrome P450 may be involved in RA metabolism in keratinocytes.     

Evaluation of cytochrome P450 activity in vitro,using dermal and hepatic microsomes from four species and two keratinocyte cell lines in culture

The Cytochrome P450 (CYP450) enzymes are expressed in the skin, and despite a low activity, as compared to the hepatic counterpart, a role during transdermal delivery of a drug cannot be excluded. Additionally, the enzymes may play a role in local toxicity, and further knowledge of dermal CYP450 activity can contribute to elucidate this issue. To achieve this, a cocktail of six selective CYP450 probe substrates were incubated with dermal and hepatic microsomes isolated from mouse, rat, minipig and man. Different species were used to evaluate if a reliable substitute for human tissue was possible. Further, the hepatic microsomes were included in this study, to estimate if the hepatic CYP450 activity is predictive of dermal CYP450 activity. The CYP450 activity was determined in two keratinocyte cell lines as well, as this in vitro model is desirable due to the ease in handling, among other factors. Overall, the metabolism found in the dermal microsomes was very low, and major differences were observed between species. When comparing the activities in dermal and hepatic microsomes, the qualitative pattern was to some extent similar within species, but also a number of differences were observed. The CYP450 metabolic activity in the two keratinocyte cell lines was not comparable to metabolism in the human dermal microsomes.     

Regulation of cutaneous drug-metabolizing enzymes and cytoprotective gene expression by topical drugs in human skin in vivo

Background Individuality in the expression and regulation of hepatic drug‐metabolizing enzymes (DMEs) and cytoprotective (CP) genes is an important determinant of treatment response. There is increasing evidence that many DMEs and CP genes are also expressed in human skin. Responses to topical drugs used to treat common skin diseases, such as psoriasis, are unpredictable and may potentially be rationalized, at least in part, by interindividual differences in cutaneous DME and CP gene expression. Objectives We investigated whether three topical drugs [coal tar, all‐trans retinoic acid (atRA) and clobetasol 17‐propionate] used in routine clinical practice modulated the expression of a variety of DME and CP genes [cytochrome P450s, glutathione S‐transferases (GSTs) and drug transporters] in healthy human skin in vivo. Methods Healthy adult volunteers (n = 30) were invited to participate in the study. Each subject was randomly allocated to receive two of the three study chemicals and one control site application. Crude coal tar (n = 13), atRA (n = 14) or clobetasol 17‐propionate (n = 10) was applied under occlusion to photoprotected buttock skin for 96 h. A vehicle control (white soft paraffin) was also applied under the same conditions at an adjacent site in all subjects. Full‐thickness punch biopsies (4‐mm diameter) were then taken from treated and control sites. Total RNA was extracted and reverse transcribed into cDNA, which was used as a template in subsequent real‐time polymerase chain reaction analysis, where fluorescent output was directly proportional to input cDNA concentration. Triplicate measurements of skin mRNA expression were made from each sample, and the arithmetic mean values taken. After logarithmic transformation, the paired t‐test was used to compare values between treated and control skin. Results Cytochrome P450s CYP1A1, CYP1A2, CYP1B1, CYP2C18, quinone reductase (NQO‐1), GSTP1, γ‐glutamyl cysteine synthetase (γ‐GCS), glutathione peroxidase‐1 (GPx‐1), cyclooxygenase‐2 (COX‐2) and haem oxygenase‐1 (HO‐1) were induced by coal tar; CYP26, NADPH P450 reductase (CPR), GSTP1 and HO‐1 by atRA; and CYP3A5 by clobetasol 17‐propionate. In contrast, CYP1A1 and CYP1A2 expression was suppressed by atRA, and γ‐GCS and MRP1 by clobetasol 17‐propionate. Marked interindividual variation in gene regulation by topical drugs was seen for the majority of genes examined. Conclusions These data demonstrate that topical drugs can modulate DME gene expression in human skin in vivo and indicate that variation in the expression and regulation of these genes may be a determinant of individuality in response to topical therapies for common skin diseases.     

Neue Aspekte zur Bedeutung des Vitamin-D3-Stoffwechsels in der Haut

Epidermal keratinocytes are the site of both UVB-induced photochemical conversion of 7-dehydrocholesterol to vitamin D(3) (25 OHD(3)) and the enzymatically controlled hydroxylation via 25-hydroxyvitamin D(3) to the biologically active final product 1alpha,25-dihydroxy vitamin D(3) (1alpha,25(OH)(2)D(3), calcitriol). The epidermal synthesis of calcitriol is of fundamental relevance because calcitriol regulates important cellular functions in keratinocytes and dermal immunocompetent cells. Calcitriol and other vitamin D-analogues are effective in the treatment of psoriasis because of their anti-proliferative and pro-differentiation effects. One mechanism for UVB-light therapy in psoriasis could be the induction of calcitriol synthesis. A better understanding of the metabolism of vitamin D(3) in the skin opens new perspectives for potential therapeutic applications of vitamin D analogues in inflammatory skin diseases. Further studies investigating the role of vitamin D(3) metabolism in the prevention of malignant skin disorders are needed.     

The spectrum of cutaneous reactions associated with calcium antagonists: a review of the literature and the possible etiopathogenic mechanisms

Calcium antagonists (CAs) or calcium-channel blockers, are a common group of antihypertensive medications. These drugs have the property of blocking the calcium channels of the vascular and cardiac smooth-muscle fibers. They have been associated with cutaneous reactions ranging from exanthems to severe adverse events. The frequency of these reactions may be as high as 48 percent. The most common are ankle or pedal edema (up to 30 %), gingival hyperplasia (up to 21 %), and flushing (up to 10 %). Less common are facial or truncal telangiectasia, photosensitivity reactions, new-onset psoriasis (as well as exacerbation of it), purpuric exanthems, pemphigoid manifestations, subacute cutaneous lupus erythematosus, gynecomastia, erythromelalgia, and oral ulcers. Particular adverse manifestations relate to drug potency, degree of vasodilatation, patient age, coexistence of other diseases, co-administration of other cytochrome P450 CYP3A-metabolized medications, fibroblast stimulation, and blood cell effects. Calcium antagonists are associated with a wide range of skin reactions, and the dermatologist should include these in the differential diagnosis of cutaneous diseases.     

Drug-induced pellagra

In industrialized countries pellagra may nowadays be induced by inappropriate diet schedules, alcohol abuse, endogenous disorders of metabolism, and also intake or abuse of drugs. In particular, INH, hydantoine, and other anticonvulsive drugs may induce pellagra by causing disorders in vitamin B metabolism. On this pathway disorders of tryptophan metabolism may also occur. We report here on a woman who developed pellagra after long-term abuse of analgetics and psychopharmaca obviously due to disorders in gastrointestinal resorption and liver metabolism. Phenobarbital, salicylic amide, phenacetin, possible also diazepam, induced the disease most likely. The clinical diagnosis was confirmed by low blood levels of niacinamide. Discontinuation of drug medication and systemic application of vitamin preparations were followed by full remission in 6 weeks.     

Do Women Have More Adverse Drug Reactions?

Up to 5% of all hospital admissions are the result of adverse drug reactions (ADRs). Identifying those factors which may predispose to ADRs is essential for risk management. Amongst the known risk factors for adverse reactions are increasing age, polypharmacy, liver and renal disease as well as being female. Female patients have a 1.5- to 1.7-fold greater risk of developing an ADR, including adverse skin reactions, compared with male patients. The reasons for this increased risk are not entirely clear but include gender-related differences in pharmacokinetic, immunological and hormonal factors as well as differences in the use of medications by women compared with men. Women generally have a lower lean body mass, a reduced hepatic clearance, have differences in activity of cytochrome P450 (CYP) enzymes (40% increase in CYP3A4, varied decrease in CYP2D6, CYP2C19 and CYP1A2), and metabolize drugs at different rates compared with men. Other important factors include conjugation, absorption, protein binding and renal elimination, which may all have some gender-based differences. However, how these differences result in an increased risk of ADRs is not clear. There are pharmacodynamic differences between men and women, seen particularly with cardiac and psychotropic medications. There is no doubt that chlorpromazine, fluspirilene and various antipsychotics appear more effective in women than men for the same dosage and plasma concentration. Similarly, women are at increased risk of QT prolongation with certain anti-arrhythmic drugs compared with men even at equivalent serum concentrations. The mechanisms are unknown. Increasingly the evidence is that idiosyncratic drug reactions, particularly cutaneous reactions, appear to have an immunological etiology. It is possible that gender difference in T cell activation and proliferation account for this as well as the increased prevalence of skin diseases such as systemic lupus erythematosus and photosensitivity. Whatever the mechanism(s), it is important to be aware that gender is a significant factor in ADRs.     

Research in practice: the second barrier of the human skin

A major function of human skin is to form an effective barrier between the environment and the inside of the organism. Especially important for this function is the activity of the physical barrier of the skin, which is mainly located in the stratum corneum. To improve this barrier function of the skin, skin protection agents are used. Recent studies have revealed that application of skin protection agents before exposition to xenobiotics does not generally reduce the percutaneous uptake of these compounds. These findings indicate that besides new study designs and improved test systems, there seems to be a need for new therapeutic approaches for more effective skin protection. In this light, new findings regarding a second barrier function of the human skin, the biochemical/toxicological barrier, could be of interest. A crucial part of this barrier function are members of the cytochrome P450 (CYP) family and efflux‐transport proteins of the multidrug resistance‐related protein family (MRPs), which are mainly expressed by basal layer keratinocytes. Recent studies have revealed that besides the physiological and protective function of these transport proteins and CYP enzymes in skin cells, the same proteins also play a role in the transport of contact allergens and activation of prohaptens to haptens causing contact dermatitis. Inhibition of this metabolism mediated activation of prohaptens and stimulation of the active elimination of contact allergens from skin cells could represent novel mechanisms improving the established tools for skin protection.     

Important drug interactions and reactions in dermatology

A constantly expanding pharmacological armamentarium increases the concern for serious drug interactions. This article discusses drug metabolism and how the cytochrome P-450 family facilitates drug biotransformation. Clinically significant drug interactions involving antifungal drugs, antibiotics, retinoids, and immunosuppressive agents, as well as topical anesthetics and various foods, are included.     

Topical treatment with CYP26 inhibitor talarozole (R115866) dose dependently alters the expression of retinoid-regulated genes in normal human epidermis

Background  An alternative approach to retinoid therapy is to inhibit the cytochrome P450 (CYP)-mediated catabolism of endogenous all- trans retinoic acid in the skin by applying retinoic acid metabolism blocking agents such as talarozole (R115866).
Objectives  To study the effects of topical talarozole on retinoid biomarkers in normal skin in a randomized phase I trial.
Methods  Gels containing talarozole (0·35% or 0·07%) and vehicle were applied once daily for 9 days on either buttock of 16 healthy volunteers. Epidermal shave biopsies (for mRNA analysis) and punch biopsies (for histology and immunofluorescence analysis) were collected from the treatment areas. Genes encoding the following were studied by quantitative real-time polymerase chain reaction: cellular retinoic acid binding protein 2 (CRABP2), cytokeratins (KRT2 and KRT4), CYP26A1, CYP26B1, CYP26C1 and CYP2S1, two enzymes in the retinol metabolism (retinal dehydrogenase-2 and retinol acyltransferase) and two proinflammatory cytokines [interleukin (IL)-1α and tumour necrosis factor-α].
Results  Talarozole treatment increased the mRNA expression of CRABP2, KRT4, CYP26A1 and CYP26B1 dose dependently, and decreased the expression of KRT2 and IL-1α compared with vehicle-treated skin. No mRNA change in retinol-metabolizing enzymes was obtained. There was no induction of epidermal thickness or overt skin inflammation in talarozole-treated skin. Immunofluorescence analysis confirmed an upregulation of KRT4 protein, but no upregulation of CYP26A1 and CYP26B1 expression was detected.
Conclusions  Talarozole influences the biomarker pattern consistently with increased retinoic acid stimulation. The low irritancy of talarozole at the two examined dosages is a possible advantage over topical retinoids.     

Mitochondrial dysfunction: a neglected component of skin diseases

Aberrant mitochondrial structure and function influence tissue homeostasis and thereby contribute to multiple human disorders and ageing. Ten per cent of patients with primary mitochondrial disorders present skin manifestations that can be categorized into hair abnormalities, rashes, pigmentation abnormalities and acrocyanosis. Less attention has been paid to the fact that several disorders of the skin are linked to alterations of mitochondrial energy metabolism. This review article summarizes the contribution of mitochondrial pathology to both common and rare skin diseases. We explore the intriguing observation that a wide array of skin disorders presents with primary or secondary mitochondrial pathology and that a variety of molecular defects can cause dysfunctional mitochondria. Among them are mutations in mitochondrial‐ and nuclear DNA‐encoded subunits and assembly factors of oxidative phosphorylation (OXPHOS) complexes; mutations in intermediate filament proteins involved in linking, moving and shaping of mitochondria; and disorders of mitochondrial DNA metabolism, fatty acid metabolism and heme synthesis. Thus, we assume that mitochondrial involvement is the rule rather than the exception in skin diseases. We conclude the article by discussing how improving mitochondrial function can be beneficial for aged skin and can be used as an adjunct therapy for certain skin disorders. Consideration of mitochondrial energy metabolism in the skin creates a new perspective for both dermatologists and experts in metabolic disease.     

Basic pharmacologic approach to immunosuppressive therapy for autoimmune blistering diseases

The pathophysiology of the most important bullous skin diseases, pemphigus and bullous pemphigoid, has been carefully defined by the use of experimental animal models of the diseases. This has shown that to successfully treat pemphigus, one must reduce autoantibody production, while in contrast, bullous pemphigoid can be treated by blocking any of several steps in an inflammatory cascade. This knowledge has led to rational therapeutic approaches to both disorders. This approach can be applied to other autoimmune bullous skin diseases, with an expected increase in efficacy of therapy and decreased morbidities from immunosuppressive drugs.     

Chronic Pruritus in the Absence of Specific Skin Disease

Chronic pruritus is a major and distressing symptom of many cutaneous and systemic diseases and can significantly impair the patient’s quality of life. Pruritus perception is the final result of a complex network involving dedicated nerve pathways and brain areas, and an increasing number of peripheral and central mediators are thought to be involved. Itch is associated with most cutaneous disorders and, in these circumstances, its management overlaps with that of the skin disease. Itch can also occur without associated skin diseases or primary skin lesions, but only with nonspecific lesions secondary to rubbing or scratching. Chronic itch with no or minimal skin changes can be secondary to important diseases, such as neurologic disorders, chronic renal failure, cholestasis, systemic infections, malignancies, and endocrine disorders, and may also result from exposure to some drugs. The search for the cause of pruritus usually requires a meticulous step-by-step assessment involving careful history taking as well as clinical examination and laboratory investigations. Few evidence-based treatments for pruritus are available. Topical therapy, oral histamine H1 receptor antagonists, and phototherapy with UV radiation can target pruritus elicitation in the skin, whereas antiepileptic drugs, opioid receptor antagonists, and antidepressants can block signal processing in the CNS.     

Xenobiotic metabolism capacities of human skin in comparison with a 3D epidermis model and keratinocyte-based cell culture as in vitro alternatives for chemical testing: activating enzymes (Phase I)

Abstract: Skin is important for the absorption and metabolism of exposed chemicals such as cosmetics or pharmaceuticals. The Seventh Amendment to the EU Cosmetics Directive prohibits the use of animals for cosmetic testing for certain endpoints, such as genotoxicity; therefore, there is an urgent need to understand the xenobiotic metabolizing capacities of human skin and to compare these activities with reconstructed 3D skin models developed to replace animal testing. We have measured Phase I enzyme activities of cytochrome P450 (CYP) and cyclooxygenase (COX) in ex vivo human skin, the 3D skin model EpiDerm? (EPI‐200), immortalized keratinocyte‐based cell lines and primary normal human epidermal keratinocytes. Our data demonstrate that basal CYP enzyme activities are very low in whole human skin and EPI‐200 as well as keratinocytes. In addition, activities in monolayer cells differed from organotypic tissues after induction. COX activity was similar in skin, EPI‐200 and NHEK cells, but was significantly lower in immortalized keratinocytes. Hence, the 3D model EPI‐200 might represent a more suitable model for dermatotoxicological studies. Altogether, these data help to better understand skin metabolism and expand the knowledge of in vitro alternatives used for dermatotoxicity testing.     

Inhibition of retinoic acid catabolism by minocycline: evidence for a novel mode of action?

Retinoic acid (RA) represents an essential and highly potent endogenous retinoid with pronounced anti‐inflammatory properties and potent anti‐acne activity, and has recently been suggested to share a common anti‐inflammatory mode of action with tetracycline antibiotics. We hypothesized that tetracyclines may directly interfere with RA homeostasis via inhibition of its local cytochrome P450 (CYP450)‐mediated degradation, an essential component of tightly regulated skin RA homeostasis. To test this hypothesis, we performed controlled in vitro RA metabolism assays using rat skin microsomes and measured RA levels in a RA‐synthesizing human keratinocyte cell line, both in the presence and in the absence of minocycline, a tetracycline popular in acne treatment. Interestingly, minocycline potently blocked RA degradation in rat skin microsomes, and strikingly enhanced RA levels in RA‐synthesizing cell cultures, in a dose‐dependent manner. These findings indicate a potential role for CYP‐450‐mediated RA metabolism in minocycline's pleiotropic mode of action and anti‐acne efficacy and could account for the overlap between minocycline and RA‐induced effects at the level of their molecular mode of action, but also clinically at the level of the rare side effect of pseudotumor cerebri, which is observed for both, RA and minocycline treatment.     

Vitamin D and the skin

Vitamin D was originally discovered as a factor that regulates calcium and bone metabolism. Recent advances in investigation have shown that vitamin D also functions as a regulator of cellular growth and differentiation in various tissues. The skin is not an exception from such effects of vitamin D; it is regarded as a site of its activation and action. Evidence has accumulated showing that the active form of vitamin D and its analogs suppress growth and stimulate the terminal differentiation of keratinocytes. In psoriatic lesions, epidermal keratinocytes exhibit hyper-proliferation and impaired differentiation triggered by inflammation. Therefore, it is quite reasonable that vitamin D is effective on psoriasis. Indeed, within the past decade, analogs of vitamin D3 have been used as topical therapy for psoriasis. In this review, we summarize the fundamental features of vitamin D and the development of vitamin D therapy for psoriasis. Clinical application to other skin diseases and the future of vitamin D therapy in dermatology are also discussed.