Ebselen oxide attenuates mechlorethamine dermatotoxicity in the mouse ear vesicant model.

Mechlorethamine (HN2) is an alkylating agent and sulfur mustard mimetic. Topical exposure to HN2 is associated with tissue blistering. Previous work in our laboratory has shown that ebselen (EB-1) possesses anti-vesicant, anti-inflammatory, anti-bacterial, anti-fungal, and cytoprotective properties, both in vivo and in vitro. We recently reported that ebselen oxide (EB-2), an analog of EB-1 with a tetravalent selenium atom, also possesses anti-bacterial and anti-fungal activity and confers cytoprotection against HN2 in vitro. The purpose of the present study was to determine the vesicant countermeasure potential of EB-2 using the mouse ear vesicant model (MEVM). Compared to control ears, mouse ears exposed to a single dose of HN2 (0.500 µmol/ear) showed an increase in wet weights, ear thickness, hyperplasia, vesication, and inflammatory cell infiltration after 24 h. Fluorescence microscopy of terminal deoxynucleotidyl transferase (TdT) dUTP nick end labeling (TUNEL)-stained sections showed that the occurrence of apoptosis extended from the epidermis of the HN2-treated side, all the way to the contralateral epidermis. In contrast, HN2-exposed ears treated topically with EB-2 at a test dose of 0.250 mg/ear showed a significant decrease in wet weight (12% less vs. HN2 alone), morphometric thickness (13% less vs. HN2 alone), and vesication. In addition, TUNEL staining revealed that HN2 ears treated with EB-2 (0.250 mg/ear) showed a decrease in apoptosis as compared to the HN2 group. EB-2 also reduced the abundance of matrix metalloproteinase-9 (MMP-9) in ear tissues exposed to HN2. Taken together, our study demonstrates that EB-2 is an efficacious countermeasure to HN2.

Counter irritant activity of Carthamus oxycantha.

Many locally occurring species of Asteraceae are used as medicinal plants by various tribal and ethnic communities in Pakistan. Carthamus oxycantha is often occurs as weed in cultivated fields. Folk medicines indicated its use as an anti inflammatory and wound healing plant. It is used for wound healing by the local population in the form of powder paste. No scientific Report, about the behavior of this plant has so far been published. The counter irritant studies of locally occurring Carthamus oxycantha was carried out. The main objectives of the project were to evaluate its wound healing effects on animal skin and the identity and characterization of chromatographically isolated fractions. For this purpose, different solvents with a broad range of polarity were successively used to extract non-polar compounds (petroleum ether extract), constituents intermediate polarities (chloroform extract) and polar constituents (methanol extract) from the whole herb of Carthamus oxycantha. The counter irritant activity of the crude extracts and isolated fractions was evaluated on rabbit's skin. Five fractions Co-1 to Co-5 were isolated from the active chloroform extract by column and thin layer chromatography. Co-1, Co-3 and Co-5 appeared to be the most potent counter irritant than others. A possible structure-activity relationship of these active compounds was investigated by using spectroscopy (UV and FTIR analysis).

Cytokine regulation by MAPK activated kinase 2 in keratinocytes exposed to sulfur mustard.

Uncontrolled inflammation contributes to cutaneous damage following exposure to the warfare agent bis(2-chloroethyl) sulfide (sulfur mustard, SM). Activation of the p38 mitogen activated protein kinase (MAPK) precedes SM-induced cytokine secretion in normal human epidermal keratinocytes (NHEKs). This study examined the role of p38-regulated MAPK activated kinase 2 (MK2) during this process. Time course analysis studies using NHEK cells exposed to 200µM SM demonstrated rapid MK2 activation via phosphorylation that occurred within 15 min. p38 activation was necessary for MK2 phosphorylation as determined by studies using the p38 inhibitor SB203580. To compare the role of p38 and MK2 during SM-induced cytokine secretion, small interfering RNA (siRNA) targeting these proteins was utilized. TNF-α, IL-1ß, IL-6 and IL-8 secretion was evaluated 24h postexposure, while mRNA changes were quantified after 8h. TNF-α, IL-6 and IL-8 up regulation at the protein and mRNA level was observed following SM exposure. IL-1ß secretion was also elevated despite unchanged mRNA levels. p38 knockdown reduced SM-induced secretion of all the cytokines examined, whereas significant reduction in SM-induced cytokine secretion was only observed with TNF-α and IL-6 following MK2 knockdown. Our observations demonstrate potential activation of other p38 targets in addition to MK2 during SM-induced cytokine secretion.

Ebselen reduces the toxicity of mechlorethamine in A-431 cells via inhibition of apoptosis.

A series of test compounds were evaluated for an ability to reduce the toxicity of the nitrogen mustard mechlorethamine (HN2) in vitro. The test compounds included resveratrol, pterostilbene, vitamin C, ebselen, ebselen diselenide, and ebselen-sulfur. Among them, ebselen demonstrated the highest degree of protection against HN2 toxicity. To this end, pretreatment of the cells with ebselen offered protection against the toxicant whereas no protection was observed when cells were first incubated with HN2 and then treated with ebselen. Significant increases in caspase 3 and caspase 9 activities were observed in response to HN2, and ebselen was found to reduce these effects. Taken together, the data presented here indicate that ebselen is an effective countermeasure to nitrogen mustard in vitro, which is worthy of future investigation in vivo.

Identification and evaluation of counter-irritant potential of crude extract of Malva parviflora L. by WHO recommended methods.

Plenty of medicinal plants are available in Pakistan and are in human use as herbal medicines from ancient time. Present work is based on the evaluation of the use of Malva parviflora in skin irritation problems. For this purpose, powdered plant material (The aerial part and roots separately) was extracted by using successive solvent extraction method using petroleum ether, chloroform and methanol. Resulting three crude fractions were subjected to counter-irritant investigations on rabbit's ear. Petroleum ether fraction exhibited prominent counter-irritant potential. Five compounds named, as MP-1, MP-2, MP-3, MP-4 and MP-5 were isolated from petroleum ether extract by column and thin layer chromatography. These compounds were subjected to UV spectrophotometer for detection of absorption of light, then FTIR for specific functional group identification and counter-irritant potentials was evaluated on rabbit's ear skin. The MP-1 and MP-2 exhibited excellent counter-irritant activity in different dilutions than others. However, dilution 100 µg/ml showed maximum activity.

Menthol attenuates respiratory irritation responses to multiple cigarette smoke irritants.

Menthol, the cooling agent in peppermint, is added to almost all commercially available cigarettes. Menthol stimulates olfactory sensations, and interacts with transient receptor potential melastatin 8 (TRPM8) ion channels in cold-sensitive sensory neurons, and transient receptor potential ankyrin 1 (TRPA1), an irritant-sensing channel. It is highly controversial whether menthol in cigarette smoke exerts pharmacological actions affecting smoking behavior. Using plethysmography, we investigated the effects of menthol on the respiratory sensory irritation response in mice elicited by smoke irritants (acrolein, acetic acid, and cyclohexanone). Menthol, at a concentration (16 ppm) lower than in smoke of mentholated cigarettes, immediately abolished the irritation response to acrolein, an agonist of TRPA1, as did eucalyptol (460 ppm), another TRPM8 agonist. Menthol's effects were reversed by a TRPM8 antagonist, AMTB. Menthol's effects were not specific to acrolein, as menthol also attenuated irritation responses to acetic acid, and cyclohexanone, an agonist of the capsaicin receptor, TRPV1. Menthol was efficiently absorbed in the respiratory tract, reaching local concentrations sufficient for activation of sensory TRP channels. These experiments demonstrate that menthol and eucalyptol, through activation of TRPM8, act as potent counterirritants against a broad spectrum of smoke constituents. Through suppression of respiratory irritation, menthol may facilitate smoke inhalation and promote nicotine addiction and smoking-related morbidities.

Protective effect of ß-(1,3 → 1,6)-D-glucan against irritant-induced gastric lesions.

ß-(1,3)-D-Glucan with ß-(1,6) branches has been reported to have various pharmacological activities, such as anti-tumour and anti-infection activities, which result from its immunomodulating effects. Gastric lesions result from an imbalance between aggressive and defensive factors. In the present study, we examined the effect of ß-(1,3)-D-glucan with ß-(1,6) branches isolated from Aureobasidium pullulans on the gastric ulcerogenic response in mice. Oral administration of ß-glucan ameliorated gastric lesions induced by ethanol (EtOH) or HCl. This administration of ß-glucan also suppressed EtOH-induced inflammatory responses, such as infiltration of neutrophils and expression of pro-inflammatory cytokines, chemokines and cell adhesion molecules (CAM) at the gastric mucosa. Of the various defensive factors, the levels of heat shock protein (HSP) 70 and mucin but not PGE(2) were increased by the administration of ß-glucan. ß-Glucan-dependent induction of the expression of HSP70 and mucin proteins and suppression of the expression of pro-inflammatory cytokines, chemokines and CAM were also observed in cultured cells in vitro. The results of the present study suggest that ß-glucan protects the gastric mucosa from the formation of irritant-induced lesions by increasing the levels of defensive factors, such as HSP70 and mucin.

Dimethylthiourea protects against chlorine induced changes in airway function in a murine model of irritant induced asthma.

BACKGROUND: Exposure to chlorine (Cl2) causes airway injury, characterized by oxidative damage, an influx of inflammatory cells and airway hyperresponsiveness. We hypothesized that Cl2-induced airway injury may be attenuated by antioxidant treatment, even after the initial injury. METHODS: Balb/C mice were exposed to Cl2 gas (100 ppm) for 5 mins, an exposure that was established to alter airway function with minimal histological disruption of the epithelium. Twenty-four hours after exposure to Cl2, airway responsiveness to aerosolized methacholine (MCh) was measured. Bronchoalveolar lavage (BAL) was performed to determine inflammatory cell profiles, total protein, and glutathione levels. Dimethylthiourea (DMTU;100 mg/kg) was administered one hour before or one hour following Cl2 exposure. RESULTS: Mice exposed to Cl2 had airway hyperresponsiveness to MCh compared to control animals pre-treated and post-treated with DMTU. Total cell counts in BAL fluid were elevated by Cl2 exposure and were not affected by DMTU treatment. However, DMTU-treated mice had lower protein levels in the BAL than the Cl2-only treated animals. 4-Hydroxynonenal analysis showed that DMTU given pre- or post-Cl2 prevented lipid peroxidation in the lung. Following Cl2 exposure glutathione (GSH) was elevated immediately following exposure both in BAL cells and in fluid and this change was prevented by DMTU. GSSG was depleted in Cl2 exposed mice at later time points. However, the GSH/GSSG ratio remained high in chlorine exposed mice, an effect attenuated by DMTU. CONCLUSION: Our data show that the anti-oxidant DMTU is effective in attenuating Cl2 induced increase in airway responsiveness, inflammation and biomarkers of oxidative stress.

Capsaicin-induced joint inflammation is not blocked by local anesthesia.

The purpose of this study was to evaluate the effect of local anesthetic blockade of afferent innervation on the development of capsaicin-induced edema in the rat temporomandibular joint (TMJ) region and on reflex jaw muscle activity. Under halothane anesthesia, 64 male Sprague-Dawley rats were prepared for monitoring of edema development by lateral movement of a needle overlying the left TMJ region and for acute recording of electromyographic activity in ipsilateral digastric and masseter muscles. A double-barrel catheter was inserted into the TMJ region for delivery of saline or 0.5% bupivacaine from 1 needle, followed with the injection of 1% capsaicin, 0.1% capsaicin, or vehicle control from the other needle 5 minutes later. Application of capsaicin into the saline pretreated TMJ region led to dose-dependent edema development and reflex jaw muscle activity; however, only 1% capsaicin solution resulted in significant tissue expansion and muscle activity when compared with the vehicle control. Pretreatment of the rat TMJ region with bupivacaine failed to inhibit capsaicin-induced edema development, although successful blockade of nerve conduction was confirmed with the absence of reflex jaw muscle activity. Capsaicin-induced edema of the rat TMJ region developed independent of axonal conduction, suggesting neurogenic inflammation may arise regardless of functional nerve conduction.

Coprecipitation of nonoxynol-9 with polyvinylpyrrolidone to decrease vaginal irritation potential while maintaining spermicidal potency.

The aim of this study was to test the hypothesis that polyvinylpyrrolidone (PVP) would increase the critical micelle concentration (CMC) of nonoxynol-9 (N-9), providing a reduction in its irritation potential, while maintaining essential spermicidal activity. Solid coprecipitates of N-9 with PVP were manufactured with the use of a modified lyophilization process. The irritation potential of N-9 was estimated by an in vitro assay, monitoring the extent of hemolysis of red blood cells. CMCs of N-9 were measured in the presence of various concentrations of PVP. A modified Sander-Cramer assay was implemented to measure the spermicidal activity of N-9 and the N-9/PVP coprecipitates. With the use of the lyophilization process and more suitable solvents, solid coprecipitates of N-9/PVP were manufactured with no residual organic solvents. The irritation potential of N-9 was reduced when in the presence of PVP-50% hemolysis values increased from 0.054 mM to more than 0.2mM. N-9 CMC values increased in the presence of PVP from 0.085 mM (0% PVP) to 0.110 mM (3.5% PVP) and 0.16 6mM (10% PVP). However, spermicidal activities ranged from 0.213 mM to 0.238 mM, N-9 remaining steady regardless of the amount of PVP. By use of N-9/PVP coprecipitates, the self-association properties and irritation potentials of N-9 were altered. This result suggests a process to produce a spermicidal product that reduces the detrimental implications to the vaginal epithelium while maintaining the essential spermicidal activity.

Oral irritation by mustard oil: self-desensitization and cross-desensitization with capsaicin.

We investigated the temporal pattern of oral irritation elicited by sequential application of mustard oil (allyl-isothiocyanate), and whether it exhibits self-desensitization and cross-desensitization with capsaicin. Mustard oil (0.125%, 40 micro l) was sequentially applied to one side of the tongue at 1 min intervals, and subjects rated the intensity of the irritant sensation elicited by each stimulus. Ratings successively declined across trials, indicating desensitization. In contrast, sequential application of capsaicin (10 ppm) elicited irritation that increased in intensity across trials (sensitization). To test for self-desensitization by mustard oil, a 10 min hiatus was imposed following the series of unilateral mustard oil stimuli, after which mustard oil was applied to both sides of the tongue. In a two-alternative forced-choice paradigm, subjects chose which side had stronger irritation and also independently rated the irritant intensity on each side. A significant majority of subjects chose the side not previously receiving mustard oil as more intense, and assigned significantly higher intensity ratings to that side, indicating self-desensitization. In two additional sessions, the same paradigm was used to show mustard oil cross-desensitization of irritation elicited by capsaicin, and capsaicin cross-desensitization of irritation from mustard oil. In a final session, sequential application of mustard oil at faster (20 s) intervals initially evoked a sensitizing pattern followed by desensitization. The temporal patterns of oral irritation exhibited by mustard oil, and its reciprocal cross-desensitization with capsaicin, are similar to those of menthol and nicotine.

Synthesis and structure--activity relationships of aroylpyrrole alkylamide bradykinin (B2) antagonists.

The synthesis and structure-activity relationships of a novel series of aroylpyrrole alkylamides as potent selective bradykinin B(2) receptor antagonists are described. Several members of this series display nanomolar affinity at the B(2) receptor and show activity in an animal model of antinociception.

The use of different concentrations of betaine as a reducing irritation agent in soaps monitored visually and non-invasively.

BACKGROUND/AIMS: Products containing detergents can damage the skin and give rise to irritant contact dermatitis. Therefore, attempts have been made to find less irritating detergents as well as substances decreasing undesired side-effects of detergents, and a novel approach is offered by betaine. The aim of the study has been to determine the irritating properties of some liquid soaps for personal hygiene and to map the effect of different concentrations of betaine using electrical impedance, trans-epidermal water loss and visual inspection. METHODS: Twenty-eight healthy subjects were patch tested with different commercial soaps with and without betaine and sodium lauryl sulphate on both volar forearms for 24 h. A site with distilled water and an unoccluded area were used as references. Responses of the skin reactions were evaluated by visual inspection and by measuring trans-epidermal water loss and electrical impedance before application and 24 h after removal of the chambers. RESULTS/CONCLUSIONS: Significant skin reactions were found for all soaps tested but the soaps containing betaine were the least irritating. However, the skin irritation did not decrease with increasing concentrations of betaine in the tested range. On the whole the differences between the products were not large. The non-invasive methods used were more sensitive than visual assessment for evaluation of invisible or barely visible skin responses.

The ability of betaine to reduce the irritating effects of detergents assessed visually, histologically and by bioengineering methods.

BACKGROUND/AIMS: A novel approach for reducing the undesired irritating properties of detergents on skin might be offered by betaine, which is a natural product derived from the sugar beet. The aim of the study was to explore the ability of betaine to reduce the irritating effects of two surfactants, sodium lauryl sulphate (SLS) and cocoamidopropylbetaine (CAPB). For evaluation of changes in skin reactions visual scoring, electrical impedance, transepidermal water loss and histology were used. METHODS: Twenty-one healthy subjects were patch tested for 24 h with SLS and CAPB alone and together with betaine, betaine alone, and the two controls distilled water and an unoccluded test site on both volar forearms. Responses were evaluated by measuring electrical impedance and transepidermal water loss before exposure and 24 h after the removal of the test substances, and also by visual inspection and histology. The electrical impedance device enables measurements at 31 frequencies and relevant information was extracted from the spectra using four indices. RESULTS: CAPB was found to be less irritating than SLS. The used detergents gave rise to distinctive impedance patterns also reflected by different types of histopathological skin responses. After the adding of betaine, the irritant reaction decreased for both detergents. CONCLUSIONS: Betaine is a promising ingredient to reduce the side effects of detergents and electrical impedance is a suitable tool both to quantify the degree of irritation as well as to differentiate between various types of reactions.

Differences among moisturizers in affecting skin susceptibility to hexyl nicotinate, measured as time to increase skin blood flow.

BACKGROUND/AIMS: A wide range of branded and generic moisturizers is frequently used for the prevention and treatment of dry skin. The influence of moisturizers on the skin permeability is pertinent to the understanding of their therapeutic efficacy. The aim of the present study was to compare the effect of two moisturizers on the skin permeability barrier, assessed as skin reactivity to a vasodilating substance. METHODS: The study was parallel, randomized and double blind on 53 healthy volunteers. One of the creams contained 5% urea, whereas the other contained no humectant but had a high lipid content. The participants were instructed to apply the cream twice daily for three weeks on the volar aspect of one of their forearms. The skin was then exposed to hexyl nicotinate, which induces vasodilation. The time-course and magnitude of the microvascular changes in the two skin areas were monitored with a non-invasive optical technique (laser Doppler flowmetry) with two measuring probes. RESULTS: The lag-time between application and initial response was significantly longer for the urea-treated site compared with the other cream. Furthermore, the time for maximum response was shorter for the lipid-rich cream than for its placebo. CONCLUSION: The study shows differences in action between moisturizers, which may influence the skin susceptibility to other irritants and allergens in the environment.

Cholecystokinin secretagogue-induced gastroprotection: role of nitric oxide and blood flow.

This study was done to examine the role of CCK in gastric mucosal defense and to assess the gastroprotective roles of nitric oxide and blood flow. In rats, the CCK secretagogues oleate and soybean trypsin inhibitor augmented gastric mucosal blood flow and prevented gastric injury from luminal irritants. Type A CCK receptor blockade negated CCK secretagogue-induced gastroprotection and exacerbated gastric injury from bile and ethanol but did not block adaptive cytoprotection. CCK secretagogue-induced gastroprotection and hyperemia were negated by nonselective nitric oxide synthase (NOS) inhibition (N(G)-nitro-L-arginine methyl ester) but not by selective inducible NOS inhibition (aminoguanidine). Gastric mucosal calcium-dependent NOS activity, but not calcium-independent NOS activity, was increased following CCK and CCK secretagogues. The release of endogenous CCK plays a role in the intrinsic gastric mucosal defense system against injury from luminal irritants. The protective mechanism appears to involve increased production of nitric oxide from primarily the constitutive isoforms of NOS and a resultant increase in blood flow.

Il-1-related cytokine responses of nonimmune skin cells subjected to CEES exposure with and without potential vesicant antagonists.

Sulfur mustard provokes an acute inflammatory response in skin. To determine if keratinocytes regulate this response and whether three potential vesicant antagonists can counteract adverse changes, specimens of EpiDerm (MatTek Corp., Ashland, MA), a human skin model of differentiating keratinocytes, were exposed 2 h to humidified air with or without 2-chloroethyl ethyl sulfide (CEES, 1.72-1.73 mg/L/min) with or without 10 mM niacinamide, a poly (ADP-ribose) polymerase (PARP) inhibitor, 25 microM CGS9343B (calmodulin antagonist), or 8.4 mM leupeptin (cysteine protease inhibitor). After a 22-h incubation, levels of interleukin-1 alpha (IL-1alpha), its receptor antagonist (IL-1Ra), soluble type II receptor (sIL-1RII) and prostaglandin-E(2) (PGE(2)) were determined. Methylthiazole tetrazolium (MTT) viability tests and histological observations were also conducted. PGE(2) levels were abundant but unaffected by CEES regardless of antagonist presence. Total amounts (media plus lysate) of IL-1alpha, IL-1Ra, and sIL-1RII were reduced with CEES irrespective of antagonist. CEES promoted the release of IL-1Ra. Exposure of EpiDerm to CEES in the presence of the vesicant antagonists did not improve viability or counteract histological damage. We conclude CEES depresses total IL-1alpha and related cytokines, does not affect PGE(2) release, and adverse changes associated with CEES-exposed EpiDerm are not ameliorated by these particular antagonists. Dramatically increased (5- to 10-fold) release of IL-1Ra may provide a useful marker for cytotoxicity. The high level of IL-1Ra and increased release with injury suggest a primary function in down-regulating IL-1 inflammatory responses in skin.

A modified HET-CAM assay approach to the assessment of anti-irritant properties of plant extracts.

Hen's egg--chorioallantoic membranes were used to screen for and assess anti-irritant properties among aqueous extracts of plants (HET-CAM tests), in connection with searches for plant-derived substances with topical anti-irritant action. The main question to be answered was whether CAM-assay screening of plant extracts could provide a useful route to identifying promising anti-irritant extracts for follow-up clinical testing. To be useful, the method would have to flag materials with strong anti-irritant properties, and would have to avoid registering false negatives. The tests conducted provided positive indications. We measured the delays in onset of three manifestations of membrane irritation-vascular hemorrhaging, membrane lysis and membrane coagulation-observed with test substances relative to positive controls. Aqueous 15% lactic acid, a commonly used irritant in direct tests on human skin, was employed as the test irritant in this study. The ratio [irritation onset times after test substance pre-treatment]:[onset times without test substance pretreatment] was used to measure the anti-irritant power of test substances. A scoring notation was devised for this which treats the delay parameters as independent effects. Most tested plant extracts showed no significant irritant or anti-irritant effects. Among the apparently anti-irritant plant extracts (approx. 10% of all those tested), most showed their greatest effect against hemorrhaging. Lesser but still readily measurable effects against membrane lysis and coagulation were also observed in nearly all the apparently anti-irritant extracts. Two of the tested extracts proved to be membrane irritants. Some key CAM assay results were compared with results obtained in direct tests on human skin using the same test irritant (15% lactic acid). In these comparative tests on skin, an essentially similar pattern of efficacy was obtained, with the plant extract deemed best in the CAM screenings, outperforming the benchmark anti-irritant hydrocortisone. From these initial results it appears that physiological CAM assays may prove useful in screening natural materials for anti-irritant properties, as alternatives to mechanism-dependent biochemical assays, or expensive direct screening tests on human subjects. Further work remains to extend the CAM screening approach to irritants other than lactic acid, and to assess its quantitative powers of prediction of topical anti-irritancy.

Inhibition of irritation and contact hypersensitivity by phenoxyacetic acid methyl ester in mice.

New anti-irritant treatments are required to prevent irritation and sensitization reactions to consumer medicines and dermatological drugs. We report here that phenoxyacetic acid methyl ester (PAME) is an effective agent to prevent and treat irritant and allergic contact dermatitis. Balb/c mice skin-treated with 1% PAME do not lose weight relative to vehicle-treated mice, nor is it irritating to mouse skin. Topical PAME prevents skin irritation to a wide variety of irritants including: arachidonic acid, capsaicin, sodium lauryl sulfate (SLS), disodium laureth sulfosuccinate and tetradecanoylphorbol-13-acetate. Histological studies showed that 1% PAME greatly diminished dermal neutrophilic infiltration and dermal capillary vessel dilation, and prevented epidermal hyperproliferation and hyperkeratosis that accompanies detergent (SLS)-induced skin irritation. Topical PAME inhibited ear swelling following ear challenge during the elicitation phase of contact hypersensitivity in mice sensitized with 1-chloro-2, 4-dinitrochlorobenzene (DNCB), oxazolone and the hair coloring dye rho-phenylenediamine (PPD). Finally, topical administration of 1% PAME prior to PPD or DNCB sensitization prevented the induction phase of contact hypersensitivity. These results indicate that PAME represents a potential new category of potent topical anti-inflammatory agents.