Asunto(s)

Asma Ocupacional/inducido químicamente , Dermatitis Profesional/etiología , Detergentes/efectos adversos , Eugenol/efectos adversos , Adulto , Alérgenos/inmunología , Asma Ocupacional/diagnóstico , Asma Ocupacional/inmunología , Asma Ocupacional/fisiopatología , Pruebas de Provocación Bronquial , Dermatitis Profesional/diagnóstico , Dermatitis Profesional/inmunología , Dermatitis Profesional/fisiopatología , Eugenol/inmunología , Femenino , Humanos , Pruebas Cutáneas

RESUMEN

Cytokine release from dendritic cells in vitro is a useful marker to discriminate between sensitizing and irritant haptenic chemicals. Unfortunately, pro-haptens, which gain reactivity following metabolic/auto activation, yield negative results. To overcome this, we exposed human neutrophils and THP-1 cells to haptens/pro-haptens and measured IL-8 release. Haptenic compounds stimulated IL-8 release in neutrophils and THP-1 cells. In contrast, the pro-haptens eugenol, isoeugenol, and 2-aminophenol stimulated high levels of IL-8 release from neutrophils alone. Neutrophil cytokine release was reduced when glutathione was added. Cyp1A1/1B1/3A4 were not detectable in THP-1 cells or neutrophils; however, neutrophils expressed high levels of myeloperoxidase.

Asunto(s)

Aminofenoles/inmunología , Eugenol/análogos & derivados , Eugenol/inmunología , Interleucina-8/inmunología , Neutrófilos/inmunología , Línea Celular , Células Cultivadas , Haptenos/inmunología , Humanos

RESUMEN

Eugenol is a major volatile constituent of clove essential oil obtained through hydrodistillation of mainly Eugenia caryophyllata (=Syzygium aromaticum) buds and leaves. It is a remarkably versatile molecule incorporated as a functional ingredient in numerous products and has found application in the pharmaceutical, agricultural, fragrance, flavour, cosmetic and various other industries. Its vast range of pharmacological activities has been well-researched and includes antimicrobial, anti-inflammatory, analgesic, anti-oxidant and anticancer activities, amongst others. In addition, it is widely used in agricultural applications to protect foods from micro-organisms during storage, which might have an effect on human health, and as a pesticide and fumigant. As a functional ingredient, it is included in many dental preparations and it has also been shown to enhance skin permeation of various drugs. Eugenol is considered safe as a food additive but due to the wide range of different applications, extensive use and availability of clove oil, it is pertinent to discuss the general toxicity with special reference to contact dermatitis. This review summarises the pharmacological, agricultural and other applications of eugenol with specific emphasis on mechanism of action as well as toxicity data.

Asunto(s)

Eugenol/farmacología , Agroquímicos/química , Agroquímicos/farmacología , Analgésicos/química , Analgésicos/farmacología , Antiinfecciosos/química , Antiinfecciosos/farmacología , Antiinflamatorios/química , Antiinflamatorios/farmacología , Antimutagênicos/química , Antimutagênicos/farmacología , Antineoplásicos/química , Antineoplásicos/farmacología , Antioxidantes/química , Antioxidantes/farmacología , Eugenol/química , Eugenol/inmunología , Eugenol/toxicidad , Factores Inmunológicos/química , Factores Inmunológicos/farmacología

RESUMEN

The concept that thresholds exist for the induction of allergic contact dermatitis by chemicals with skin sensitizing properties has been used for a quantitative risk assessment approach. In this approach the potency of skin sensitizers as determined in the Local Lymph Node Assay is used to calculate the threshold for induction of sensitization. These are then used to estimate safe exposure levels for consumers. Whether these exposure levels will protect subjects that are already sensitized is unknown. The elicitation of allergic contact dermatitis supposedly occurs above a certain threshold as well and this threshold is most likely lower than that for the induction. It is unclear if induction thresholds can be extrapolated to elicitation thresholds. The aim of this study was to assess the potency of sensitizers with different sensitizing potencies in the elicitation phase in a mouse model for elicitation. Mice were sensitized by topical application on days 0 and 7 using equipotent concentrations of oxazolone, 2,4-dinitrochlorobenzene (DNCB) and eugenol to ensure that the sensitization strength would not influence the elicitation potency. Mice were challenged on day 21 by topical application on the ears in a dose-dependent manner and dose-response data were used to calculate the elicitation potency. Unexpectedly, sensitizers with different sensitizing potencies induced not the same dose-response curves in sensitized mice. The most potent sensitizer in the elicitation phase was oxazolone, followed by DNCB and eugenol. Similar to the induction phase, under equipotent sensitization conditions strong sensitizers such as oxazolone and DNCB elicit allergic reactions at lower concentrations than weak sensitizers such as eugenol. Our results indicate that elicitation thresholds cannot be readily deduced from sensitization thresholds.

Asunto(s)

Alérgenos/farmacología , Dermatitis Alérgica por Contacto/etiología , Dinitroclorobenceno/farmacología , Eugenol/farmacología , Oxazolona/farmacología , Piel/efectos de los fármacos , Administración Tópica , Alérgenos/inmunología , Animales , Proliferación Celular/efectos de los fármacos , Dermatitis Alérgica por Contacto/inmunología , Dinitroclorobenceno/inmunología , Eugenol/inmunología , Femenino , Ensayo del Nódulo Linfático Local , Ratones , Ratones Endogámicos BALB C , Oxazolona/inmunología , Proyectos Piloto , Piel/inmunología , Organismos Libres de Patógenos Específicos

RESUMEN

Skin exposure to sensitizing chemicals can induce allergic reactions. Certain chemicals, so called pro-sensitizers, need metabolic activation to become allergenic. Their metabolic activation occurs in skin cells such as keratinocytes or dendritic cells. These cell types are also incorporated into dermal in vitro test systems used to assess the sensitizing potential of chemicals for humans. In vitrosystems range from single cell cultures to organotypic multi-cellular reconstructed skin models. Until now, their metabolic competence to unmask sensitizing potential of pro-sensitizers was rarely investigated. This review aims to summarize current information on available skin in vitro models and the relevance of xenobiotic metabolizing enzymes for the activation of pro-sensitizers such as eugenol, 4-allylanisole, and ethylendiamine. Among others, these chemicals are discussed as performance standards to validate new coming in vitro systems for their potential to identify pro-sensitizers.

Asunto(s)

Sistema Enzimático del Citocromo P-450/metabolismo , Dermatitis Alérgica por Contacto/enzimología , Oxidorreductasas/metabolismo , Piel/enzimología , Piel/inmunología , Xenobióticos/metabolismo , Alérgenos/química , Alérgenos/inmunología , Alérgenos/metabolismo , Animales , Biotransformación , Sistema Enzimático del Citocromo P-450/inmunología , Dermatitis Alérgica por Contacto/diagnóstico , Dermatitis Alérgica por Contacto/inmunología , Eugenol/análogos & derivados , Eugenol/química , Eugenol/inmunología , Eugenol/metabolismo , Haptenos/química , Haptenos/inmunología , Haptenos/metabolismo , Humanos , Inmunización , Técnicas de Cultivo de Órganos/métodos , Oxidorreductasas/inmunología , Patología Molecular/tendencias , Xenobióticos/inmunología

RESUMEN

Many attempts have been made to develop in vitro sensitization tests that employ dendritic cells (DCs), DC-like cell lines or keratinocytes. The aim of the present investigation was to establish a co-culture of THP-1 cells and keratinocytes for evaluation of skin sensitization potential of chemicals. Co-cultures were constructed by THP-1 cells cultured in lower compartments and keratinocytes cultured in upper compartments of cell culture inserts. After 24 h exposure to sensitizers (2, 4-dinitrochlorobenzene, p-phenylenediamine, formaldehyde, nickel sulfate, isoeugenol and eugenol) and non-sensitizers (sodium lauryl sulfate, benzalkonium chloride and lactic acid), the expression of CD86 and CD54 on THP-1 cells were evaluated by flow cytometry, and cell viabilities were determined. The sensitizers induced the augmentation of CD86 and CD54 expression, but the non-sensitizers had no significant effect. Compared with mono-cultures of THP-1 cells, the augmentation of CD86 and CD54 could be detected even at a non-toxic concentration of sensitizers in THP-1 cell/keratinocyte co-cultures. Moreover, isoeugenol was distinguished as a sensitizer in co-cultures, but failed to be identified in mono-cultures. These results revealed that the co-cultures of THP-1 cells and keratinocytes were successfully established and suitable for identifying sensitizers using CD86 and CD54 expression as identification markers.

Asunto(s)

Alternativas a las Pruebas en Animales/métodos , Dermatitis Alérgica por Contacto/inmunología , Haptenos/inmunología , Queratinocitos/inmunología , Monocitos/inmunología , Antígeno B7-2/metabolismo , Compuestos de Benzalconio/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Técnicas de Cocultivo , Dinitroclorobenceno/inmunología , Dinitroclorobenceno/farmacología , Eugenol/análogos & derivados , Eugenol/inmunología , Eugenol/farmacología , Formaldehído/inmunología , Formaldehído/farmacología , Haptenos/farmacología , Humanos , Molécula 1 de Adhesión Intercelular/metabolismo , Queratinocitos/citología , Ácido Láctico/inmunología , Ácido Láctico/farmacología , Monocitos/citología , Monocitos/metabolismo , Níquel/inmunología , Níquel/farmacología , Fenilendiaminas/inmunología , Fenilendiaminas/farmacología , Sensibilidad y Especificidad , Pruebas Cutáneas/métodos , Dodecil Sulfato de Sodio/farmacología

RESUMEN

BACKGROUND: Perfumes are complex mixtures composed of many fragrance ingredients, many of which are known to be only weak allergens when tested individually. It is therefore surprising that fragrance contact allergy is one of the most common forms of contact allergy. OBJECTIVES: To investigate whether mixing different fragrance allergens leads to increased sensitization potency, and to examine the difference in the challenge response to one chemical in mice sensitized either with the mixture of allergens or with only the relevant allergen. METHODS: CBA mice were sensitized with three different concentrations of three fragrance allergens alone or as a mixture. The sensitization and elicitation responses were measured by ear thickness plus infiltration of B and T cells and T cell proliferation in the draining lymph nodes. RESULTS: We found a dose-dependent sensitization response for each of the allergens. An increased response was seen when the allergens were mixed. A stronger challenge response to cinnamal was seen in mice sensitized with the allergen mixture than in mice sensitized with cinnamal alone. CONCLUSIONS: Our findings suggest that mixtures of allergens increase the primary response that potentiates the generation of memory T cells in response to the specific allergen. Thus, allergen mixtures enhance both induction and elicitation of contact allergy.

Asunto(s)

Alérgenos/toxicidad , Dermatitis Alérgica por Contacto/etiología , Perfumes/toxicidad , Acroleína/análogos & derivados , Acroleína/inmunología , Acroleína/toxicidad , Aldehídos/inmunología , Aldehídos/toxicidad , Animales , Recuento de Linfocito CD4 , Linfocitos T CD8-positivos , Proliferación Celular , Células Cultivadas , Ciclohexenos/inmunología , Ciclohexenos/toxicidad , Relación Dosis-Respuesta Inmunológica , Eugenol/análogos & derivados , Eugenol/inmunología , Eugenol/toxicidad , Femenino , Citometría de Flujo , Ratones , Ratones Endogámicos CBA , Perfumes/química

RESUMEN

The dysregulation of immune functions by some pesticides leads to various immune disorders, including immunodeficiency, tumorigenesis, allergies, and autoimmunity. This study's primary objective was to examine the relationship between immune disorders and the immunosuppression induced by immunosuppressive pesticides. We focused on the modulation of allergic potential by the organophosphorus pesticide parathion, organochlorine pesticide methoxychlor, phenoxyacetic acid herbicide 2,4-d-butyl, and benzoic acid fungicide eugenol, as detected by a local lymph node assay (LLNA), which was developed initially for hazard identification of skin sensitization. Parathion and methoxychlor are immunosuppressive chemicals, and 2,4-d-butyl and eugenol are contact allergens. After the immunosuppressive characteristics of parathion and methoxychlor were confirmed in a pilot study, 4-week-old mice were orally administered parathion (0, 0.4, 1.2mg/kg) or methoxychlor (0, 100, 300 mg/kg). Four weeks after the last administration, an LLNA was conducted using 2,4-d-butyl (0%, 2.5%, 5%, and 10%) and eugenol (0%, 5%, 10%, and 25%). In addition, detailed analysis of their auricular lymph nodes for number of surface antigen expression of T cells and local cytokine production were performed using 5% 2,4-d-butyl and 5% eugenol treatment groups. EC3 values (estimated concentration to yield a stimulation index of 3) of 2,4-d-butyl and eugenol decreased markedly in parathion- and methoxychlor-pretreated groups. Parathion- and methoxychlor-pretreated groups induced marked increase in number of surface antigen expression of T cells and levels of Th1 cytokines (IFN-γ, TNF-α, and IL-17) produced by ex vivo restimulated lymph node cells. According to our results, the allergic potentials of 2,4-d-butyl and eugenol are increased by prior exposure to parathion and methoxychlor.

Asunto(s)

Ácido 2,4-Diclorofenoxiacético/análogos & derivados , Alérgenos/inmunología , Eugenol/inmunología , Metoxicloro/toxicidad , Paratión/toxicidad , Plaguicidas/toxicidad , Ácido 2,4-Diclorofenoxiacético/inmunología , Animales , Citocinas/biosíntesis , Relación Dosis-Respuesta a Droga , Femenino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos CBA

RESUMEN

BACKGROUND: Ethosomes and liposomes are ultra-small vesicles capable of encapsulating drugs and cosmetic ingredients for topical use, thereby potentially increasing bioavailability and clinical efficacy. So far, few reports have suggested that formulation of cosmetic ingredients in vesicular carrier systems may increase the allergenicity potential. OBJECTIVES: To investigate the effect of ethosome formulation of isoeugenol and methyldibromo glutaronitrile on the elicitation response under patch test conditions and by repeated open applications. PATIENTS/MATERIALS/METHODS: A total of 27 volunteer patients with a previous positive patch test reaction to either isoeugenol or methyldibromo glutaronitrile were included in the study. In all patients, a serial dilution patch test was performed with the allergen in question formulated in ethosomes and in an ethanol/water solution. In addition, a repeated open application test (ROAT) was performed in a subset of 16 patients, and lag time until a positive response was recorded. RESULTS: Both contact allergens encapsulated in ethosomes showed significantly enhanced patch test reactions as compared with the allergen preparation in ethanol/water without ethosomes. No significant difference in the median lag time was recorded between preparations in the ROAT. CONCLUSIONS: Encapsulating potential contact allergens in ethosomes may increase the challenge response as compared with the same concentrations in an ethanol/water base without ethosomes.

Asunto(s)

Alérgenos/inmunología , Eugenol/análogos & derivados , Liposomas , Nitrilos/química , Pruebas del Parche , Conservadores Farmacéuticos/química , Dermatitis Alérgica por Contacto/diagnóstico , Dermatitis Alérgica por Contacto/etiología , Sistemas de Liberación de Medicamentos/métodos , Eugenol/química , Eugenol/inmunología , Humanos , Nitrilos/inmunología , Conservadores Farmacéuticos/efectos adversos

RESUMEN

Vesicular systems, such as liposomes and ethosomes, are used in cosmetic and pharmaceutical products to encapsulate ingredients, to protect ingredients from degradation, to increase bioavailability, and to improve cosmetic performance. Some reports have suggested that formulation of cosmetic ingredients in vesicular carrier systems may increase their contact allergy elicitation potential in humans. However, no sensitization studies have been published. We formulated two model contact allergens (isoeugenol and dinitrochlorobenzene) in ethosomes and investigated the sensitization response using a modified local lymph node assay (LLNA). The results were compared with those for the same allergens in similar concentrations and vehicles without ethosomes. Both allergens encapsulated in 200-300 nm ethosomes showed increased sensitizing potency in the murine assay compared with the allergens in solution without ethosomes. Empty ethosomes were non-sensitizing according to LLNA. The clinical implications are so far uncertain, but increased allergenicity from ethosome-encapsulated topical product ingredients cannot be excluded.

Asunto(s)

Alérgenos/administración & dosificación , Proliferación Celular/efectos de los fármacos , Dermatitis Alérgica por Contacto/etiología , Dinitroclorobenceno/administración & dosificación , Eugenol/análogos & derivados , Inmunización , Ganglios Linfáticos/efectos de los fármacos , Alérgenos/inmunología , Animales , Química Farmacéutica , Dermatitis Alérgica por Contacto/inmunología , Dinitroclorobenceno/inmunología , Relación Dosis-Respuesta a Droga , Composición de Medicamentos , Eugenol/administración & dosificación , Eugenol/inmunología , Femenino , Liposomas , Ensayo del Nódulo Linfático Local , Ganglios Linfáticos/inmunología , Ganglios Linfáticos/patología , Ratones , Ratones Endogámicos CBA , Tamaño de la Partícula

RESUMEN

The present study was undertaken to investigate the anti-arthritic activity of hydroxychavicol (HC) a major phenolic compound isolated from the aqueous extract leaves of plant Piper betle (Piperaceae). The compound showed significant lowering of pro-inflammatory (Th1) cytokine levels in arthritic paw tissue homogenate supernatant viz. IL-2, IFN-gamma, and TNF-alpha with maximum inhibition at higher dose levels of 2 and 4 mg/kg p.o. and enhanced the production of anti-inflammatory (Th2) cytokines IL-4 and IL-5 estimated by cytometric bead array immunoassay. Cytometric bead array uses the sensitivity of amplified fluorescence detection by flowcytometer to measure soluble analytes in a particle based immune assay. This assay can accurately quantitate five cytokines in a 50-microl sample volume. The T-helper (Th1) deviated cells produce detectable level of tumor necrosis factor (TNF-alpha), interleukin-2 (IL-2), and interferon-gamma (IFN-gamma), while the Th2 deviated cells produce significant amount of interleukin-4 (IL-4) and interleukin-5 (IL-5). HC at graded doses also significantly decreased the expression of IL-1beta, PGE(2), LTB(4), and nitric oxide levels showing significant inhibition of these parameters. Elevated levels of CD4(+) T cell specific interferon-gamma (IFN-gamma) in splenocytes of arthritic animals was also inhibited in treated animals. The oral LD(0) in both mice and rats was more than 1000 mg/kg.

Asunto(s)

Antirreumáticos/uso terapéutico , Artritis Experimental/tratamiento farmacológico , Citocinas/inmunología , Eugenol/análogos & derivados , Mediadores de Inflamación/inmunología , Células TH1/inmunología , Células Th2/inmunología , Animales , Antirreumáticos/química , Antirreumáticos/inmunología , Antirreumáticos/aislamiento & purificación , Artritis Experimental/inmunología , Eugenol/química , Eugenol/inmunología , Eugenol/aislamiento & purificación , Eugenol/uso terapéutico , Femenino , Inmunoensayo/instrumentación , Inmunoensayo/métodos , Dosis Máxima Tolerada , Ratones , Ratones Endogámicos BALB C , Estructura Molecular , Piper betle/química , Extractos Vegetales/química , Extractos Vegetales/inmunología , Extractos Vegetales/uso terapéutico , Bazo/citología , Bazo/inmunología

RESUMEN

Establishing of alternatives to animal tests is ethically desirable and gains in importance in context of new European Union regulations such as REACH. We have refined our new in vitro assay for prediction of the sensitizing potency of xenobiotics. Monocytes cocultured with primary human keratinocytes develop to a novel class of in vitro generated dendritic cells after treatment with transforming growth factor beta and Interleukin-4 in serum-free medium. These dendritic cell-related cells (DCrc) are the key players in the loose-fit coculture-based sensitization assay (LCSA). Assay duration and cytokine consumption could be cut down without impairing the assay's functionality. DCrc showed a dose-dependent upregulation of CD86 after treatment with the contact allergens 2,4,6-trinitrobenzenesulfonic acid, the prohapten isoeugenol, and alpha-hexyl cinnamic aldehyde. The metal allergens nickel and cobalt could be detected by measuring Interleukin-6 and macrophage inflammatory protein 1-beta (MIP-1beta, CCL-4) in coculture supernatants. The irritant zinc elicited no reaction. Lipopolysaccharide produced upregulation of CD86, IL-6 and MIP-1beta. Determination of tolerable concentrations of an allergen in consumer products requires a widely accepted sharp quantitative assay. Animal-based assays do not meet this requirement. The LCSA provides dose-response information, thereby allowing prediction of the relative ability of a substance to induce sensitization.

Asunto(s)

Alérgenos/inmunología , Células Dendríticas/inmunología , Dermatitis por Contacto/inmunología , Alérgenos/toxicidad , Antígeno B7-2/inmunología , Supervivencia Celular/efectos de los fármacos , Medios de Cultivo , Ensayo de Inmunoadsorción Enzimática , Eugenol/análogos & derivados , Eugenol/inmunología , Eugenol/toxicidad , Citometría de Flujo , Factor Estimulante de Colonias de Granulocitos y Macrófagos/farmacología , Humanos , Indicadores y Reactivos , Interleucina-4/farmacología , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Lipopolisacáridos/inmunología , Lipopolisacáridos/toxicidad , Metales/inmunología , Metales/toxicidad , Monocitos/efectos de los fármacos , Factor de Crecimiento Transformador beta/farmacología

RESUMEN

Axillary dermatitis is common and overrepresented in people with contact allergy to fragrances. Many people suspect their deodorants to be the incriminating products. In order to investigate the significance of isoeugenol in deodorants for the development of axillary dermatitis when used by people with and without contact allergy to isoeugenol, patch tests with deodorants and ethanol solutions with isoeugenol, as well as repeated open application tests (ROAT) with roll-on deodorants with and without isoeugenol at various concentrations, were performed in 35 dermatitis patients, 10 without and 25 with contact allergy to isoeugenol. A positive ROAT was observed only in patients hypersensitive to isoeugenol (P<0.001) and only in the axilla to which the deodorants containing isoeugenol had been applied (P<0.001). Deodorants containing isoeugenol in the concentration range of 0.0063-0.2% used 2 times daily on healthy skin can thus elicit axillary dermatitis within a few weeks in people with contact allergy to isoeugenol.

Asunto(s)

Desodorantes/efectos adversos , Dermatitis Alérgica por Contacto/diagnóstico , Eugenol/análogos & derivados , Hipersensibilidad Inmediata/diagnóstico , Pruebas del Parche , Adulto , Estudios de Casos y Controles , Dermatitis Alérgica por Contacto/epidemiología , Eugenol/efectos adversos , Eugenol/inmunología , Femenino , Humanos , Incidencia , Persona de Mediana Edad , Probabilidad , Factores de Riesgo , Muestreo , Sensibilidad y Especificidad

RESUMEN

Dendritic cells (DC) have been shown to capture and process antigens and play an initiating role in contact sensitization. Cells with dendritic morphology can be generated in vitro either from CD34(+) cord blood cells or from CD14(+) peripheral monocytes. The aim of this study was to determine the state of maturation/activation of both populations after exposure to several concentrations of four well-established model allergens (nickel sulfate, eugenol, alpha-hexylcinnamaldehyde and 2,4,6-trinitrobenzene sulfonic acid) or the irritant sodium dodecyl sulfate. We analyzed the surface expression of CD86, CD83 and HLA-DR and the production of IL-1beta. DC from the two sources were generated separately in two laboratories, but challenged using identical test protocols. Using both DC populations it was possible to detect the allergens under investigation, though minor differences regarding effective concentrations were noted. The non-responsiveness of CD34-DC to CIN was probably due to non-optimal concentrations. Ni(2+), known as a moderate allergen in vivo, showed the most prominent effect in both cell systems. CD86 expression was the most reliable phenotypic marker for the in vitro identification of allergens. Due to substantial individual variations it was difficult to draw any definite conclusions as to the relevance of IL-1beta production as an activation endpoint. We conclude that both test systems are able to respond to allergens, but CD34-DC must be exposed to higher concentrations to demonstrate significant phenotypic changes. On the other hand, Mo-DC from only some of the donors reacted to allergens, in contrast to CD34-DC, which responded to allergens irrespective of the donor, thus necessitating the use of Mo-DC cultures from several blood donors.

Asunto(s)

Alérgenos/administración & dosificación , Células Dendríticas/inmunología , Interleucina-1/biosíntesis , Antígenos CD/metabolismo , Antígenos CD34/metabolismo , Antígeno B7-2 , División Celular , Células Dendríticas/citología , Eugenol/inmunología , Sangre Fetal/citología , Sangre Fetal/inmunología , Antígenos HLA-DR/metabolismo , Humanos , Inmunoglobulinas/metabolismo , Técnicas In Vitro , Recién Nacido , Glicoproteínas de Membrana/metabolismo , Modelos Inmunológicos , Monocitos/citología , Monocitos/inmunología , Níquel/inmunología , Fenotipo , Ácido Trinitrobencenosulfónico/inmunología , Antígeno CD83

RESUMEN

Fragrances are worldwide a major cause of allergic contact dermatitis (ACD), a delayed-type hypersensitivity reaction mediated by T lymphocytes. We investigated T cell responses to fragrances using peripheral blood mononuclear cells (PBMC) and T cells from skin lesions of fragrance-allergic patients. The components of a fragrance mixture (eugenol, isoeugenol, geraniol, oak moss, alpha-amyl cinnamic aldehyde, cinnamic aldehyde, cinnamic alcohol, and hydroxycitronellal) that is commonly used in the patch test were studied in vitro in the lymphocyte transformation test (LTT). PBMC from fragrance-allergic patients (n = 32) showed significant stimulations to all eight fragrances. The calculated stimulation indices (SI) varied between 2.1 and 21.8. The influence of metabolic enzymes on T cell stimulation was studied for two fragrances. Interestingly, stimulation of eugenol and isoeugenol was increased in the presence of antigen-modified human liver microsomes (CYP450) or recombinant CYP1A1 in five of seven cases. Furthermore, we established 18 T cell clones (TCC) from a skin lesion reacting specifically to eugenol. FACS analysis revealed that the majority (n = 15, 83%) of TCC were CD3(+), CD4(+), and HLA-DR(+). Seventeen percent (n = 3) of the clones were CD8(+). TCC (n = 4) released significant amounts of IL-2 and IFN-gamma but no IL-4 and IL-5. In addition, CD4(+) TCC (n = 3) showed antigen-induced cytotoxic activities against autologous B cells. In summary, we demonstrated for the first time that fragrance-specific CD4(+) and CD8(+) T lymphocytes are present in fragrance-allergic individuals. In addition, our results suggest that CYPs can be involved in the formation of the nominative antigen.

Asunto(s)

Alérgenos/inmunología , Perfumes , Linfocitos T/inmunología , Monoterpenos Acíclicos , Presentación de Antígeno , Complejo CD3/análisis , Antígenos CD4/análisis , Antígenos CD8/análisis , Cinamatos/inmunología , Citocromo P-450 CYP1A1/metabolismo , Dermatitis por Contacto/inmunología , Eugenol/análogos & derivados , Eugenol/inmunología , Eugenol/metabolismo , Citometría de Flujo , Antígenos HLA-DR/análisis , Humanos , Leucocitos Mononucleares/inmunología , Activación de Linfocitos , Microsomas Hepáticos/enzimología , Microsomas Hepáticos/inmunología , Odorantes , Proteínas Recombinantes/metabolismo , Piel/inmunología , Pruebas Cutáneas , Terpenos/inmunología

RESUMEN

International test guidelines, such as the Organisation for Economic Cooperation and Development (OECD) guideline #406, recommend 2 guinea pig methods for testing of the contact allergenic potential of chemicals: the Guinea Pig Maximization Test (GPMT) and the Buehler test. Previous comparisons between the methods suggested that the Buehler test was less sensitive than the GPMT although modified Buehler test protocols were used. Parallel GPMT and Buehler tests were conducted according to OECD guideline #406 using a multiple-dose design and test results were analysed using a standard logistic dose-response model. To compare the sensitivity of the 2 test procedures the test conditions were kept identical and the following chemicals with a range of sensitization potentials were tested: chloraniline, chlorhexidine, eugenol, formaldehyde, mercaptobenzothiazole and neomycin sulphate. Formaldehyde and neomycin sulphate were strong sensitizers in both tests. Mercaptobenzothiazole, eugenol and chloraniline were all strong sensitizers in the GPMT, eugenol and mercaptobenzothiazole were negative in the Buehler test and equivocal results were obtained with chloraniline. Chlorhexidine was negative in the GPMT and equivocal responses were obtained with the Buehler test. Higher induction concentrations were needed to show allergenicity in the Buehler test and for some allergens the Buehler test protocol was not sensitive enough to demonstrate allergenic potential.

Asunto(s)

Dermatitis Alérgica por Contacto/diagnóstico , Pruebas Cutáneas/métodos , Alérgenos , Animales , Benzotiazoles , Clorhexidina/inmunología , Eugenol/inmunología , Femenino , Formaldehído/inmunología , Cobayas , Hidroxilaminas/inmunología , Modelos Teóricos , Neomicina/inmunología , Sensibilidad y Especificidad , Tiazoles/inmunología

RESUMEN

BACKGROUND: Epidermal Langerhans cells (LCs) play a critical role in the induction of contact hypersensitivity. The LCs leave the skin, move to the regional lymph nodes and present the allergens embedded in the HLA-DR molecule to naive T-lymphocytes. To allow LC emigration from the epidermis, E-cadherin must be downregulated. In this study, we have examined the early events that occur in the human epidermis after exposure to three strong contact sensitizers and two commonly used fragrances by examining alterations of E-cadherin and HLA-DR expression. METHODS: To determine whether E-cadherin and HLA-DR levels were modulated by allergens, flow cytometry was utilized to evaluate E-cadherin and HLA-DR expression on human epidermal LCs exposed to the different chemicals for 4 h at 37 degrees C. RESULTS: In vitro stimulation with the contact sensitizers isoeugenol, cinnamaldehyde, 2,4, 6-trinitrobenzenesulfonic acid, Bandrowski'sbase, or p-phenylene diamine resulted in a dose-dependent decrease of HLA-DR expression on the surface of LCs without affecting the number of positive cells. These contact allergens induced a downregulation of E-cadherin expression as well as a significant decrease of the percentage of E-cadherin-positive cells. Incubation with an irritant, sodium lauryl sulfate, did not significantly change HLA-DR and E-cadherin expression. CONCLUSIONS: Based on the alteration of E-cadherin and HLA-DR expression of human LCs under short-term exposure conditions, there was a clear difference between contact sensitizers and a well-characterized irritant. For the first time, the ability of fragrance allergens in dipropylene glycol, a widely used vehicle in fragrance and cosmetic industries, was demonstrated to induce human LC phenotypic alterations. In combination with a series of in vitro tests, this rapid and simple method should help to detect the sensitizing potential of a substance to be applied onto the human skin as an alternative to animal testing.

Asunto(s)

Alérgenos , Cadherinas/metabolismo , Antígenos HLA-DR/metabolismo , Células de Langerhans/inmunología , Perfumes/efectos adversos , Acroleína/efectos adversos , Acroleína/análogos & derivados , Acroleína/inmunología , Dermatitis por Contacto/etiología , Dermatitis por Contacto/inmunología , Eugenol/efectos adversos , Eugenol/análogos & derivados , Eugenol/inmunología , Humanos , Técnicas In Vitro , Fenilendiaminas/efectos adversos , Fenilendiaminas/inmunología , Ácido Trinitrobencenosulfónico/efectos adversos , Ácido Trinitrobencenosulfónico/inmunología

RESUMEN

We examined sensitization and crossreaction by the guinea pig maximization test (GPMT) with phenolic compounds. Skin specimens were collected from earlobes of BALB/c mice on 24 h after elicitation phase of contact hypersensitivity reaction (CHR) with phenolic compounds. The expression of cytokines of skin specimens was examined by the reverse transcriptase/polymerase chain reaction (RT-PCR) method. Consequently, phenolic compounds which showed positive reaction in GPMT expressed IL-2 and IFN-gamma on 24 h after elicitation, and some phenolic compounds showed marked crossreaction. Therefore, it was found that on several phenolic compounds, dimerization of these compounds from monomer to dimer decrease sensitization.

Asunto(s)

Reacciones Cruzadas , Citocinas/metabolismo , Dermatitis por Contacto/inmunología , Eugenol/análogos & derivados , Eugenol/inmunología , Piel/inmunología , Animales , Citocinas/análisis , Dimerización , Femenino , Cobayas , Ratones , Ratones Endogámicos BALB C , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Pruebas Cutáneas/métodos

RESUMEN

Methyleugenol is an allylbenzene food flavoring which has been shown to form DNA and protein adducts, and to cause hepatotoxicity and carcinogenicity in rodents. In order to investigate the nature of the protein adducts, specific antisera were raised by immunizing rabbits with conjugates prepared by coupling 1'-acetoxymethyleugenol, or its acidic congener 3,4-dimethoxycinnamic acid, to rabbit serum albumin (RSA). These polyclonal antisera were shown by enzyme linked immunosorbent assay (ELISA) to contain antibodies which recognized the 3,4-dimethoxyphenyl ring portion of methyleugenol. Analysis of livers from rats given methyleugenol i.p. for 5 days, at doses between 10 and 300 mg/kg/day, revealed dose-dependent formation of novel protein adducts which were recognized by the antisera. The adducts were detected by ELISA and by immunoblotting and were concentrated in the microsomal fraction, and were shown in inhibition studies to be derived from methyleugenol. A 44 kDa adduct was the only protein adduct detected in livers of rats given low loses of methyleugenol (10 or 30 mg/kg/day) and was the major adduct detected in rats given high doses of the compound (100 and 300 mg/kg/day). This adduct was solubilized when microsomal fractions were extracted using 0.1 M sodium carbonate, implying that it is a peripheral membrane protein. A pattern of protein adducts which mirrored the in vivo situation was generated when rat hepatocytes were incubated with 1'-hydroxymethyleugenol in vitro, but could not be reproduced in experiments undertaken using liver microsomes or postmitochondrial supernatants. These findings imply that generation of protein adducts in livers of rats given methyleugenol in vivo proceeds via the 1'-hydroxy metabolite and requires crucial cofactors, and/or structural features, which are present in intact hepatocytes but not in broken cell preparations and which remain to be defined.

Asunto(s)

Eugenol/análogos & derivados , Aromatizantes/química , Aditivos Alimentarios/química , Microsomas Hepáticos/química , Albúmina Sérica/química , Animales , Relación Dosis-Respuesta a Droga , Electroforesis en Gel de Poliacrilamida , Ensayo de Inmunoadsorción Enzimática , Eugenol/química , Eugenol/inmunología , Eugenol/toxicidad , Aromatizantes/toxicidad , Aditivos Alimentarios/toxicidad , Haptenos/química , Haptenos/inmunología , Sueros Inmunes/inmunología , Immunoblotting , Hígado/química , Hígado/citología , Hígado/efectos de los fármacos , Hígado/inmunología , Masculino , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/inmunología , Proteínas/análisis , Conejos , Ratas , Ratas Endogámicas F344 , Juego de Reactivos para Diagnóstico , Albúmina Sérica/inmunología , Solubilidad , Espectrofotometría Ultravioleta

RESUMEN

Although many simple chemicals can give rise to the phenomenon of allergic contact dermatitis, it is rare that the mechanism of reaction between the chemical hapten and skin protein is known. A further complication is that metabolic processes may produce substantial changes to a chemical penetrating skin. Thus the skin contactant may be regarded as a prohapten which will give rise to the true hapten in vivo. In this study, the possible reaction mechanisms for a number of related simple aromatic chemicals have been investigated. The approach taken was to evaluate potential reaction mechanisms by assessing the degree to which chemicals could cross-react in sensitization tests. By careful choice of chemicals, it was then possible to confirm (or reject) options. Using this approach, a number of reaction schemes were investigated for eugenol, isoeugenol, dihydroeugenol, anethole and several related chemicals. The patterns of sensitization obtained and the cross-reactions observed indicated clearly that electrophile/nucleophile interactions were unlikely to provide a complete explanation of the sensitization processes. Eugenol and isoeugenol are not mutually cross-reactive, yet both cross-reacted with dihydroeugenol. Examination of the possible reaction mechanisms allows the speculation that eugenol reacts in part via a phenolic radical mechanism, whilst isoeugenol reacts largely via formation of an orthoquinone. Both reaction mechanisms are proposed for dihydroeugenol.

Asunto(s)

Eugenol/análogos & derivados , Piel/inmunología , Derivados de Alilbenceno , Animales , Anisoles/química , Anisoles/inmunología , Reacciones Cruzadas , Eugenol/química , Eugenol/inmunología , Cobayas