Interspecies assessment factors and skin sensitization risk assessment.

For many endpoints in toxicology, an interspecies safety factor remains a standard requirement. However, for skin sensitization, the hazard and potency predictions, notably from the local lymph node assay (LLNA) have been shown to correlate well with human data. Despite this, there are always exceptions, both over and under predictions. For this reason it has been suggested that an interspecies factor of 15 would accommodate potential "errors". An alternative approach is suggested in which an evidence-based strategy is taken: the large majority of the information indicates a human:LLNA ratio of 1, therefore a corrective factor would best be applied where our knowledge of the underlying chemistry of sensitization indicates that it is necessary.

Skin sensitization: Implications for integration of clinical data into hazard identification and risk assessment.

Skin sensitization associated with allergic contact dermatitis is a common health problem and is an important consideration for toxicologists in safety assessment. Historically, in vivo predictive tests have been used with good success to identify substances that have the potential to induce skin sensitization, and these tests formed the basis of safety evaluation. These original tests are now being replaced gradually either by in vitro assays or by further refinements of in vivo methods such as the local lymph node assay. Human data have also been available to inform classification decisions for some substances and have been used by risk managers to introduce measures for exposure reduction. However, humans encounter hazards in the context of exposure rather than in the form of intrinsic hazards per se, and so in this article, we have examined critically the extent to which human data have been used to refine classification decisions and safety evaluations. We have also evaluated information on the burden of human allergic skin disease and used this to address the question of whether, and to what extent, the identification and evaluation of skin sensitization hazards has led to an improvement of public and/or occupational health.

T helper cell 2 immune skewing in pregnancy/early life: chemical exposure and the development of atopic disease and allergy.

During the last 50 years there has been a significant increase in Western societies of atopic disease and associated allergy. The balance between functional subpopulations of T helper cells (Th) determines the quality of the immune response provoked by antigen. One such subpopulation - Th2 cells - is associated with the production of IgE antibody and atopic allergy, whereas, Th1 cells antagonize IgE responses and the development of allergic disease. In seeking to provide a mechanistic basis for this increased prevalence of allergic disease, one proposal has been the 'hygiene hypothesis', which argues that in Westernized societies reduced exposure during early childhood to pathogenic microorganisms favours the development of atopic allergy. Pregnancy is normally associated with Th2 skewing, which persists for some months in the neonate before Th1/Th2 realignment occurs. In this review, we consider the immunophysiology of Th2 immune skewing during pregnancy. In particular, we explore the possibility that altered and increased patterns of exposure to certain chemicals have served to accentuate this normal Th2 skewing and therefore further promote the persistence of a Th2 bias in neonates. Furthermore, we propose that the more marked Th2 skewing observed in first pregnancy may, at least in part, explain the higher prevalence of atopic disease and allergy in the first born.

The hapten-atopy hypothesis III: the potential role of airborne chemicals.

One explanation for the large increase in the prevalence of atopic disease in developed countries during the last 50 years is the 'hygiene hypothesis'. This proposes that a reduced exposure to pathogenic microorganisms at a key period(s) during development results in the maintenance or acquisition of an atopic phenotype. Alternatively, or additionally, we have postulated that increased exposure to chemicals generally, and to irritant/haptenic chemicals in particular, during critical windows of maternal pregnancy/early life have also contributed to changes in the prevalence of atopic disease. Having previously reviewed the potential roles of oral and cutaneous exposure to chemicals on the subsequent diagnosis of atopic disease, we here consider possible evidence of a role for exposure to airborne chemicals as a contributory factor in acquired susceptibility to atopic allergy. After controlling for known confounders, five specific maternal occupations during pregnancy have been implicated as being associated with subsequent atopic disease in the offspring. Each of these occupations is characterized by high and persistent exposure to airborne chemicals. High-level exposure to volatile organic compounds in the domestic environment, either during pregnancy or in early life, is also associated with development of childhood atopic disease. Similarly, sustained exposure to airborne chlorinated chemicals from swimming pools during childhood has been associated with the development of atopic allergy. A possible immunological basis for these associations is that exposure to certain airborne chemicals, even at low levels, can result in the delivery of 'danger' signals that, in turn, bias the immune response towards the selective induction or maintenance of preferential T helper 2-type immune responses consistent with the acquisition of allergic sensitization.

Why does allergic contact dermatitis exist?

The skin immune system's propensity to produce allergic contact dermatitis (ACD) to harmless chemicals, while otherwise being an efficient defence system, represents a dermatological paradox. We postulate that a major role in signalling in ACD is played by Toll-like receptor (TLR)2 and TLR4, and arises from their activation by extracellular danger-associated molecular patterns (DAMPs). Ligand activation of TLR4/2 results in the expression of interleukins (ILs) IL-1ß, IL-6, IL-12, IL-18 and IL-23, tumour necrosis factor-α and interferon-α. These cytokines promote acquisition of sensitization, and facilitate elicitation of contact allergy via multiple mechanisms, including the recruitment of CD4+ Th1 and Th17 cells. As Th1 cells secrete large amounts of DAMPs, a DAMP immune circuit (positive-feedback loop) is created. This is an important driver of skin sensitization and skin inflammation. Pathogenic extracellular bacteria, but not commensal bacteria, produce pathogen-associated molecular pattern molecules, which stimulate the expression of Th1- and Th17-promoting cytokines via TLR2 and TLR4. This also induces an immune circuit. The ability of the skin immune system to activate host defence mechanisms and to distinguish between pathogenic bacteria and commensals provides an explanation for why skin sensitization and ACD develop, as they are processes that rely on the same biological pathways. These pathways may also shed light on the pathogenesis of chronic pustular inflammatory dermatoses (e.g. acne vulgaris). The existence of safety signals from commensal bacteria, which prevent initiation of these pathways, may provide opportunities for novel therapeutic approaches to the treatment of inflammatory skin diseases.

Inter-relationships between different classes of chemical allergens.

Although allergic sensitization of the respiratory tract induced by chemicals is not as common as skin sensitization, it is nevertheless an important occupational health issue. Respiratory allergy to chemicals, characterized typically by rhinitis and asthma, is associated with considerable morbidity and with related socioeconomic costs. Several experimental approaches have been proposed for the prospective identification of chemical respiratory allergens, but none of these has yet been validated formally. In the absence of a widely accepted method for respiratory allergen identification, it is appropriate and relevant to explore their relationship with skin-sensitizing chemicals. A series of chemicals known to cause immune-mediated respiratory allergy in humans has been examined. The majority of the respiratory allergens tested were found to elicit positive responses in one or more standard tests used for the identification of skin-sensitizing potential (guinea pig maximization test, the Buehler test and/or the local lymph node assay). We suggest that this observation might form the basis of a potentially useful paradigm for initial characterization of the respiratory-sensitizing potential of chemicals. Specifically, chemicals that fail to elicit positive responses in accepted skin-sensitization test methods might also be regarded as lacking the inherent potential to cause allergic sensitization of the respiratory tract.

Allergic contact dermatitis: epidemiology, molecular mechanisms, in vitro methods and regulatory aspects. Current knowledge assembled at an international workshop at BfR, Germany.

Contact allergies are complex diseases, and one of the important challenges for public health and immunology. The German 'Federal Institute for Risk Assessment' hosted an 'International Workshop on Contact Dermatitis'. The scope of the workshop was to discuss new discoveries and developments in the field of contact dermatitis. This included the epidemiology and molecular biology of contact allergy, as well as the development of new in vitro methods. Furthermore, it considered regulatory aspects aiming to reduce exposure to contact sensitisers. An estimated 15-20% of the general population suffers from contact allergy. Workplace exposure, age, sex, use of consumer products and genetic predispositions were identified as the most important risk factors. Research highlights included: advances in understanding of immune responses to contact sensitisers, the importance of autoxidation or enzyme-mediated oxidation for the activation of chemicals, the mechanisms through which hapten-protein conjugates are formed and the development of novel in vitro strategies for the identification of skin-sensitising chemicals. Dendritic cell cultures and structure-activity relationships are being developed to identify potential contact allergens. However, the local lymph node assay (LLNA) presently remains the validated method of choice for hazard identification and characterisation. At the workshop the use of the LLNA for regulatory purposes and for quantitative risk assessment was also discussed.

The Hapten-Atopy hypothesis II: the 'cutaneous hapten paradox'.

One explanation for the striking increase in atopic disease in developed countries over the last 50 years has been the 'Hygiene Hypothesis'; a reduced exposure to pathogenic microorganisms. We have postulated previously that oral and cutaneous exposure to chemicals generally and to haptens in particular, may have also contributed to the increased prevalence of atopic disease; the 'Hapten-Atopy Hypothesis'. The purpose here is to extend further that hypothesis by consideration of the impact interplay between the innate and adaptive immune systems may have on the development of atopic allergy. It is clear that experimental cutaneous exposure to hapten can generate immune responses of different types with regard to T-helper (Th) cell phenotype. Allergic contact dermatitis is frequently associated with a selective Th1 (and Tc1)-driven inflammation, whereas atopic dermatitis is characterized by preferential Th2 cell responses. We postulate here that initial innate immune responses to chemical haptens result in the promotion of Th1 cell responses secondary to stimulation of Toll-like receptor. However, we argue also that under conditions where there is prolonged skin exposure to hapten there will be a shift of Th cell phenotype to selective Th2-type responses. The significance of such interactions is the possibility that repeated low-level skin exposure to certain types of hapten may result in the creation of an immunological environment in which the development of Th2 immune responses to third party antigens is favoured. The hypothesis is advanced that the nature and conditions of skin exposure to common haptens may impact on the quality of cutaneous immune responses such that in some circumstances the development atopic disease is favoured.

Defining occupational and consumer exposure limits for enzyme protein respiratory allergens under REACH.

A wide range of substances have been recognized as sensitizing, either to the skin and/or to the respiratory tract. Many of these are useful materials, so to ensure that they can be used safely it is necessary to characterize the hazards and establish appropriate exposure limits. Under new EU legislation (REACH), there is a requirement to define a derived no effect level (DNEL). Where a DNEL cannot be established, e.g. for sensitizing substances, then a derived minimal effect level (DMEL) is recommended. For the bacterial and fungal enzymes which are well recognized respiratory sensitizers and have widespread use industrially as well as in a range of consumer products, a DMEL can be established by thorough retrospective review of occupational and consumer experience. In particular, setting the validated employee medical surveillance data against exposure records generated over an extended period of time is vital in informing the occupational DMEL. This experience shows that a long established limit of 60 ng/m(3) for pure enzyme protein has been a successful starting point for the definition of occupational health limits for sensitization in the detergent industry. Application to this of adjustment factors has limited sensitization induction, avoided any meaningful risk of the elicitation of symptoms with known enzymes and provided an appropriate level of security for new enzymes whose potency has not been fully characterized. For example, in the detergent industry, this has led to general use of occupational exposure limits 3-10 times lower than the 60 ng/m(3) starting point. In contrast, consumer exposure limits vary because the types of exposure themselves cover a wide range. The highest levels shown to be safe in use, 15 ng/m(3), are associated with laundry trigger sprays, but very much lower levels (e.g. 0.01 ng/m(3)) are commonly associated with other types of safe exposure. Consumer limits typically will lie between these values and depend on the actual exposure associated with product use.

Does hapten exposure predispose to atopic disease? The hapten-atopy hypothesis.

Contact allergy data indicates that atopics have heightened oral tolerance to haptens (chemical allergens). We speculate here, that artificially increased oral exposure to chemicals compete with dietary proteins for the development of oral tolerance, predisposing to the acquisition of food protein allergy and representing one driver for the increasing prevalence of protein allergy and/or atopy. Hapten exposure via other surfaces such as the skin and airways might also be important in promoting atopic disease. Consistent with this hypothesis it is notable that over 40 years, with the huge increase in atopic disease, there has also been an increase in dietary hapten exposure through processed food, formula milk and oral antibiotic and drug use.

Atopic dermatitis and allergic reactions to individual fragrance chemicals.

BACKGROUND: Allergic contact dermatitis prevalence is reported as equal in atopic and nonatopic dermatitis. Atopic dermatitis is under-represented in those with allergic contact dermatitis to agents having cutaneous and dietary exposure. We compared rates of atopic dermatitis between patients with allergic contact dermatitis arising out of individual fragrance chemicals with known oral/cutaneous exposure against exclusively cutaneous exposure. METHODS: Between 1982 and 2007, 37 065 dermatitis patients were tested with Fragrance mix I. Those who were positive were tested for individual fragrance allergy. Chemicals were categorized according to whether their exposure pattern was solely cutaneous, oral or mixed. Current and past atopic dermatitis rates were compared between the whole population and groups allergic to individual fragrances. Age and gender were controlled. RESULTS: Cinnamic alcohol and cinnamal allergy groups had reduced rates of both 'current' [24/266 (9.0%) P = 0.0008, 38/364 (10.4%) P = 0.0005] and 'past' atopic dermatitis [44/266 (16.5%) P = 0.009, 70/346 (19.2%) P = 0.037]. Atopic dermatitis rates in groups allergic to Evernia prunastri and hydroxycitronellal (cutaneous exposure only) were not reduced [120/597 (20.1%) and 41/153 (26.8%)]. Groups allergic to cinnamic alcohol (P < 0.0001, P < 0.0001) and cinnamal (P < 0.0001, P < 0.004) had reductions in 'current' and 'past' atopic dermatitis, compared with Evernia prunastri. CONCLUSIONS: Patients allergic to individual fragrances with dietary exposure have reduced rates of atopic dermatitis. This suggests that patients with atopic dermatitis have heightened oral tolerance to dietary haptens, in contrast to the known close association of atopic dermatitis with food-protein allergy. Haptens may interfere with food protein tolerance by binding to soluble protein to alter its configuration and immunogenic profile.

Skin sensitization: strategies for the assessment and management of risk.

Allergic contact dermatitis (ACD) is to a considerable extent a preventable disease. Limitation can be achieved by correct identification of skin sensitizers, characterization of their potency, understanding human skin exposure and application of good risk assessment/management strategies. Various methods exist which are accurate for the predictive identification of chemicals that possess skin-sensitizing properties. These are enshrined in regulations that aim to provide a harmonized approach to hazard identification. One of the methods, the local lymph node assay, also delivers information on the relative potency of sensitizers. Efforts are continuing in the European Union and at the Organization for Economic Cooperation and Development to use elements of this information for regulatory categorization of skin sensitizers. However, greater use can be made of this potency information in the application of quantitative risk assessments. Such assessments depend also on the availability of accurate data on human skin exposure, one aspect where legislation has little role to play. Management of risks by restriction of skin exposure is, in contrast, a key point where legislation can play an important role, helping to establish a level playing field for industry and setting good standards based on the legislator's ability to access all data. Ultimately, the combination of accurate hazard identification, potency measurement, risk assessment and management, underpinned by enabling legislation, will lead to reduction of ACD. For individuals who do still develop contact allergy, avoidance of ACD should continue to be a goal, based on raising awareness of skin protection, allergen labelling and other skincare strategies.

Dendritic cells from control but not atopic donors respond to contact and respiratory sensitizer treatment in vitro with differential cytokine production and altered stimulatory capacity.

BACKGROUND: Chemical haptens induce both contact and allergic respiratory disease with dendritic cells (DCs) controlling and directing immune responses in vivo. Contact and respiratory haptens may promote differential cytokine production yet distinguishing these effects in vitro remains difficult due to human donor variability. Objective We sought to determine the effect of atopic status on the ability of DC to respond to contact and respiratory sensitizer treatment in vitro as DC from atopic donors are believed to promote Th2-type responses. METHODS: Enriched DC from control or atopic donors were treated for 4 h with levels of the contact sensitizer 2,4-dinitrochlorobenzene (DNCB) or the respiratory sensitizer trimellitic anhydride (TMA) that did not reduce cell viability. A sensitive intracellular detection technique was used to measure cytokine production, while T cell responses were assessed in a mixed leucocyte reaction. RESULTS: DC from control, non-atopic, donors produced cytokines differentially in response to sensitizer treatment; DNCB treatment significantly increased the production of Th1 cytokines IL-12 and IFN-gamma while TMA induced the production of IL-13. Control donor DC treated with TMA stimulated less in a mixed leucocyte reaction than untreated cells with any response reduced further by blocking IL-13 in culture. However, DC from atopic donors showed no significant alteration in either cytokine production or T cell stimulatory capacity after sensitizer treatment. CONCLUSION: Haptens modulate DC by changing the production of cytokines that may play a role in T cell stimulation and subsequent polarization of the immune response. DC from atopic donors were unresponsive to chemical sensitizer treatment, and may be deficient in inducing divergent T cell responses.

Enzymes, detergents and skin: facts and fantasies.

In their raw state, enzymes of bacterial/fungal origin cause allergic reactions in the lung. Proteolytic enzymes also cause irritation to skin, eyes and the respiratory tract. For 40 years, encapsulated enzymes have been used worldwide in detergent products, especially laundry formulations, and have increasing importance due to biodegradability and functionality at low temperatures, offering environmental benefits. Uniquely to the U.K., for years it has been suggested that the inclusion of enzymes in such products leads to adverse skin reactions, including erythema, pruritus and exacerbation of eczema. In this review, we look at the facts, asking whether there is evidence that the hazards identified for enzymes translate into any risk for consumer health. By considering the actual exposures in consumer use and exaggerated product usage, it is concluded that the irritating and allergenic hazards of enzyme raw materials do not translate into a risk of skin reactions, either irritant or allergic. Investigations of numerous individuals with skin complaints attributed to laundry products demonstrate convincingly that enzymes were not responsible. Indeed, enzyme-containing laundry products have an extensive history of safe use. Thus, the supposed adverse effects of enzymes on skin seem to be a consequence of a mythology. The important practical lesson is that when primary or secondary care practitioners are presented with a skin complaint, it should not be dismissed as a result of using an enzyme-containing laundry product as the diagnosis will certainly lie elsewhere. Education for healthcare professionals could usefully be enhanced to take this on board.

Positive rates to propyl gallate on patch testing: a change in trend.

Propyl gallate (E310) has, until recently, been used as a major antioxidant in fatty food and, in the cosmetic industry, in the manufacture of lipsticks. Propyl gallate has a high sensitizing potential; however, the frequency of allergic contact dermatitis from antioxidants of the gallate type was previously thought to be surprisingly low. Previous exposure and orally induced tolerance, as suggested by Khan and colleagues, may have explained the low rates of allergic contact dermatitis to propyl gallate in the past. The objectives were to assess the prevalence of allergic contact dermatitis to propyl gallate in our centre from 1988 to 2005. From 1988 to 2005, 9529 patients were patch tested to the face series, 6973 were females and 2556 were males. Patch tests were read at 2 D and 4 D. Positive reactions were scored as per International Contact Dermatitis Research Group recommendations as negative, +, ++, and +++ reactions. Propyl gallate was used at a 1% petrolatum (pet.). A total of 55 patients had positive reactions to propyl gallate 1% pet. (0.57%), 46 were female (0.65%) and 9 were male (0.33%). Using chi-square, there was a significant difference (P < 0.05) in the positivity rates between the 1988-96 period (0.45%) and the 1997-2005 period (0.77%). A review of our face series performed in the last 18 years has shown a statistically significant increase in propyl gallate-positive rates on patch testing over the last decade. An increase in its use in the cosmetic industry may well be the explanation for this. Nevertheless, a concomitant reduction of propyl gallate as an antioxidant in food, with oral tolerance being less likely to develop, may also be a contributing factor in the increasing trend of allergic contact dermatitis caused by propyl gallate.

Does irritation potency contribute to the skin sensitization potency of contact allergens?

Chemicals that possess the capacity to cause skin sensitization have long been recognized to be reactive (electrophilic) or at least the precursor of an electrophile. The chemical species (hapten) covalently bound to skin protein then forms the antigen to which the immune system responds, with sufficient exposure ultimately leading to skin sensitization. However, for this process to occur, many have also considered that in addition to haptenation of skin protein, secondary stimuli (danger signals) are also necessary. Such signals might reasonably be expected to derive from keratinocytes and/or Langerhans cells perturbed by the chemical sensitizer. Whether this disturbance comes from the haptenation process itself or from other properties of the chemical is unknown. We hypothesized that chemicals that were stronger sensitizers might appear so, in part, as a consequence not only of greater (pro)electrophilic reactivity, but also if they were more able to produce inflammatory (danger) signals. To assess this, the sensitizing potency of 55 chemicals in the local lymph node assay was compared with their ability to produce pro-inflammatory signal release, measured as a function of their relative skin irritancy in guinea pigs. A limited trend was demonstrated, consistent with the hypothesis, but indicating that either skin irritation is a poor measure of danger signals, or that such signals are perhaps no more than a necessary requirement for the acquisition of skin sensitization rather than a key determinant of the relative potency of a skin sensitizing chemical. In addition, it is possible that irritancy alone does not represent a complete surrogate marker for the ability of a chemical to produce danger signals relevant to the induction of skin sensitization.

A general population from Thailand: incidence of common allergens with emphasis on para-phenylenediamine.

BACKGROUND: Most studies on the prevalence of allergy to the permanent hair dye chemical para-phenylenediamine (PPD) are reported from populations of eczema patients attending patch-test clinics, and are assumed to be much higher than in the normal population. No data exist on incidence of senitization to PPD resulting from the use of commercial hair dye preparations over a defined time period. METHOD: A total of 2545 healthy adult volunteers (Bangkok) were screened for PPD allergy through standard patch testing. Volunteers not allergic to PPD were then recruited into two groups: one group applying a commercial hair dye brand as instructed on a monthly basis for 6 months (n=548) and the other group (controls) (n=516) was instructed not to dye their hair for 6 months. Sensitization to PPD resulting from the use of hair dye over this period was then detected by repeat patch testing. RESULTS: The prevalence of PPD allergy in a normal adult population was 2.7% (m=2.4%, f=3.2%). Projected to the adult Thai population, at least 1,000,000 Thai individuals could be allergic to PPD. The incidence of sensitization through the monthly application of standard commercial hair dye preparations over a 6-month period was 1.3%, substantially higher than in controls (0.4%), although numbers were small and not statistically significant. INTERPRETATION: There is a higher prevalence of hair dye allergy among the normal population than previously thought. The incidence of new cases of PPD allergy would indicate that current regulations and practice of hair dye exposure lead to PPD sensitization and allergy, which is a public health problem.

An evaluation of selected global (Q)SARs/expert systems for the prediction of skin sensitisation potential.

Skin sensitisation potential is an endpoint that needs to be assessed within the framework of existing and forthcoming legislation. At present, skin sensitisation hazard is normally identified using in vivo test methods, the favoured approach being the local lymph node assay (LLNA). This method can also provide a measure of relative skin sensitising potency which is essential for assessing and managing human health risks. One potential alternative approach to skin sensitisation hazard identification is the use of (Quantitative) structure activity relationships ((Q)SARs) coupled with appropriate documentation and performance characteristics. This represents a major challenge. Current thinking is that (Q)SARs might best be employed as part of a battery of approaches that collectively provide information on skin sensitisation hazard. A number of (Q)SARs and expert systems have been developed and are described in the literature. Here we focus on three models (TOPKAT, Derek for Windows and TOPS-MODE), and evaluate their performance against a recently published dataset of 211 chemicals. The current strengths and limitations of one of these models is highlighted, together with modifications that could be made to improve its performance. Of the models/expert systems evaluated, none performed sufficiently well to act as a standalone tool for hazard identification.