Studies on contact hypersensitivity in the guinea pig The cumulative contact enhancement test

A method to determine the quantitative induction and challenge of the allergenicity of externally applied toiletories and cosmetics, including their components, is described.
The experiment used oil-soluble cinnamic aldehyde and water-soluble formalin as allergens, and guinea pigs as the experimental animals. A high sensitization method resulted, carried out as follows. A 24-h closed patch is attached to the skin every other day over a period of 2 weeks (a total of 4 applications). Freund's complete adjuvant is administered intradermally just before the 3rd application of the patch. The challenge step is performed by directly applying the test material.
This method was compared with other allergenicity evaluation methods. As a result, this method was found to be in no way inferior in sensitization performance to the other methods. The method was used cm perfume mixtures and tested for its evaluation effectiveness. It proved to be satisfactory.     

Studies of new short-period method for delayed contact hypersensitivity assay in the guinea pig

The enhancement effect of cyclophosphamide on the delayed contact hypersensitivity reaction of chemical compounds was studied in Hartley albino guinea pigs. A series of assay procedures. combining the AP2 test (adjuvant and 24-h occlusive patch 2× test, as previously reported) with intraperitoneal cyclophosphamide administration, were examined. The newly developed method was as follows; cyclophosphamide 200 mg/kg intraperitoneal administration 3 days before the 1st sensitization of the AP2 test (cyclophosphamide. adjuvant and 24-h occlusive patch 2× test: CAP2 test). Comparing the CAP2 test with the AP2 test, the cumulative contact enhancement test (CCET) and the guinea pig maximization test (GPMT), the CAP2 test equally and/or better enabled the detection of allergenicities not only of strong allergens such as bromostyrol, citronellal, p -phenylendediamine and formaldehyde, but also of weak allergens such as benzyl salicylate and p -aminobenzoic acid ethyl ester. Acanthosis and spongiosis in the epidermis and mononuclear cell infiltration into the dennis at the skin reaction site were histopathologically observed. Cyclophosphamide effectively enhanced the delayed contact hypersensitivity reaction of weak allergens.     

Studies of new short-period method for delayed contact hypersensitivity assay in the guinea pig

A new method for delayed contact hypersensitivity assay of chemical compounds in guinea pigs, a short-period method (14 days) with a high detection sensitivity, has been developed. The new method was as follows; a combination of a Freund's complete adjuvant (FCA, undiluted) intradermal injection and a 24–h occusive patch on a guinea pig was performed 2x at an interval of 4 days and challenged by non-occlusive topical application II days after the first sensitization (with benzyl alcohol during test development). Acanthosis and spongiosis in the epidermis and mononuclear cell infiltration into the dermis were observed histopathologically at the skin reaction site. This newly developed method (adjuvant and 24–h occusive patch 2 test: AP2 test) could equally and/or better detect the allergenicities of 6 other chemical compounds (bromostyrol, citronellal, benzyl salicyfate. p -aminobenzoic acid ethyl ester, phenylenediamine and formaldehyde) as compared with the cumulative contact enhancement test (CCET) and the guinea pig maximization test (GPMT).     

Reduced skin threshold to irritation in the presence of allergic contact dermatitis in the guinea pig

The skin is more susceptible to irritation when an active eczematous process is present. This reduced threshold to irritation occurs in skin distant from the site of the eczematous skin. Data is presented to demonstrate the appearance of irritant dermatitis to lower concentrations of sodium lauryl sulfate during the presence of an allergic contact dermatitis in the guinea pig.     

Effects of short-wave ultraviolet light (UVB) on delayed hypersensitivity in the guinea pig

Guinea pigs, previously sensitized to dinitrochlorobenzene (DNCB), were exposed to varying doses of UVB radiation on the right flank for a period of 12 days. The response to an elicitation dose of DNCB was diminished in the irradiated skin immediately after UV treatment. This effect was dose-dependent. No reduction in the response could be demonstrated in unexposed skin. One week after UVB treatment the response to DNCB was increased, and after 2 weeks there was a normal eczematous reaction after application of DNCB.     

Modified short-term guinea pig sensitization tests for detecting contact allergens as an alternative to the conventional test

The conventional adjuvant and patch test (APT) method of guinea pig sensitization testing was modified in 2 ways, s-APT and s-APT(2), in order to shorten the test period. These short-term test methods consist of 72-h closed application of test material with intradermal injection of emulsified Freund's complete adjuvant (E-FCA) for 1st induction, 48-h closed application of test material with (s-APT) or without (s-APT(2)) intradermal injection of E-FCA on the 7th day for 2nd induction, and open application on the 14th day for challenge. They were compared with conventional APT by using 8 allergenic chemicals (formaldehyde, nickel sulfate, cobalt sulfate, ethyl-p-aminobenzoate (benzocaine), isoeugenol, 2-mercaptobenzothiazole, 2,4-dinitrochlorobenzene (DNCB) and 1-phenylazo-2-naphthol (Sudan I)). The short-term methods gave similar results to those of conventional APT in terms of mean response, sensitization rate and sensitization potency (challenge concentration that induces a mean response equal to 1.0). Thus, our short-term methods, which are capable of evaluating skin sensitization within 17 days, are sufficiently sensitive to detect potentially hazardous contact allergens.     

Experimental delayed contact sensitization to diazolidinyl urea (Germall II) in guinea pigs

Diazolidinyl urea (Germall II) is a new preservative recommended for use in certain consumer products. Although 2 reports document the human sensitization rates of this preservative, no publications quantify its sensitization potential in controlled animal experiments. Diazolidinyl urea induced mild sensitization (grade 2) under maximization test conditions. Further, there was evidence of cross-reactions with both imidazolidinyl urea (Germall 115) and formaldehyde in diazolidinyl-urea-sensitized animals. Rechallenge of diazolidinyl-urea-sensitized animals with diazolidinyl urea 4 weeks following the primary challenge only elicited a weak response (0.5) from 1 animal out of 8.     

Glucocorticoid effects on contact hypersensitivity and on the cutaneous response to ultraviolet light in the mouse

A single exposure to 254 nm ultraviolet irradiation (UV) can systemically suppress experimental sensitization to the simple allergen 2,4-dinitro, 1-chlorobenzene (DNCB) in the mouse. We show here that topical application at the site of irradiation of the 21-oic acid methyl ester derivative of the synthetic glucocorticoid triamcinolone acetonide (TAme) prevents UV suppression of sensitization. That is, mice painted with TAme at the site of UV exposure developed normal contact hypersensitivity (CH); mice exposed to UV only, like mice treated with the parent compound triamcinolone acetonide (TA), failed to be sensitized by DNCB applied to a distal site. TAme is inactivated rapidly by plasma esterases, so its effect is thought to be confined to the skin. Apparently, TAme blocked the cutaneous signal(s) for systemic suppression of CH. Histologically, irradiated skin exhibited mild inflammation and hyperproliferation, but these effects were greatly exaggerated and prolonged in the UV + TAme-treated skin, independent of sensitization at the distal site. The infiltrate consisted mostly of neutrophils and lacked the round cells characteristic of cell-mediated immunity. Apparently, normal immune suppression by UV prevented this vigorous reaction to irradiated skin. Applied together with DNCB. TAme blocked sensitization. It also prevented response to challenge by DNCB in previously sensitized animals. However, unlike the parent compound triamcinolone acetonide (TA), Budesonide or Beclomethasone diproprionate, each of which can penetrate the epidermis in active form, TAme had no effect on sensitization when applied at a distal site. Likewise, TAme did not affect plasma B (17-desoxycortisol) levels, whereas the other three compounds reduced plasma B tenfold, as expected of compounds causing adrenal-pituitary suppression. The results as a whole show that glucocorticoids can specifically inhibit cutaneous steps in induction of cell-mediated immunity or its suppression, and can, at the site of challenge, prevent its expression in CH.     

Influence of the vehicle on elicitation of contact allergic reactions to acrylic compounds in the guinea pig

Many factors can influence the elicitation of hypersensitivity reactions in guinea pigs and humans. The effect which the vehicle might have on the test response in guinea pigs sensitized with various acrylic compounds, using the "guinea pig maximization test", has been investigated. A marked decrease in the number of positive animals was seen when acetone was used as test vehicle, compared to petrolatum. The same result was seen with alcohol as vehicle, when neopentyl glycol diacrylate (NPGDA) was used as an acrylic monomer model. The patch test locations on the guinea pig flank, also affected the test response. Half of the animals did not react when challenged near the abdomen, compared to a test site near the back. By means of HPLC-analysis, the possible adsorption of the acrylic monomer to the aluminium chamber or filter paper disc, was analysed. Our findings did not indicate that adsorption occurs. A decrease in the amount of acrylic monomer in the chamber with increasing time, was noted. There was a marked difference in the monomer residue between solutions with (darkness) and without (daylight) inhibitor. The monomer decrease was also more affected by an aluminium surface than a glass or filter paper surface. Aluminium oxide probably enhances the polymerization process. The discrepancy between the test results in this study, when petrolatum and acetone were used as test vehicles, is due to a polymerization process of the acrylic compounds. Thus, the petrolatum vehicle probably prevents polymerization of the acrylic monomer.     

Langerhans cells in contact dermatitis of the guinea pig

In the guinea pig, the epidermal Langerhans cells studied by adenosine triphosphatase and electron microscopic techniques in dinitrochlorobenzene-induced contact dermatitis showed early cellular vacuolar and granular changes and intraepidermal contact with mononuclear cells. At later periods of up to 48 hours, the Langerhans cells migrated to the surface of a thickened epidermis and were lost in the parakeratotic horny layer that was shed. Thus, the Langerhans cell probably has a macrophage-type role in the epidermal reaction of contact dermatitis, and as the sponglosis and the inflammatory reaction develop, these cells are shed with the degenerating keratinocytes.     

Induction of formaldehyde contact sensitivity: dose response relationship in the guinea pig maximization test

The sensitizing potential of aqueous formaldehyde was evaluated with the guinea pig maximization test (GPMT) in two laboratories (Copenhagen and Stockholm) using different guinea pig strains. Six intradermal (0.01%-3%), and 6 topical (0.5%-20%) concentrations were used for induction, and formaldehyde 1% and 0.1% was used for challenge. The incidence of contact sensitivity depended on the intradermal, but not on the topical induction dose. Statistical analyses showed a non-monotonous (non-linear) dose response relationship. The estimated maximal sensitization rate in Copenhagen was 80% after intradermal induction with 0.65% formaldehyde; in Stockholm it was 84% after induction with 0.34%. The data from the two laboratories could be described by parallel displaced dose response curves suggesting that the guinea pig strain used in Stockholm was significantly more susceptible to formaldehyde than the strain used in Copenhagen. The EC50 (formaldehyde concentration at which 50% of the guinea pigs were sensitized) at the 72 h scoring and a 1% challenge concentration, was 0.061% in Copenhagen and 0.024% in Stockholm.     

D & C Yellow Nos. 10 and 11: delayed contact hypersensitivity in the guinea pig

D & C Yellow No. 11 was found to be a skin sensitizer in guinea pigs at an elicitation concentration of 10.0% in ethanol but not at 1.0 and 3.0%. Sensitization was induced with a 50% suspension in ethanol. D & C Yellow No. 10, the disodium salt of the mono and disulfonic acids of D & C Yellow No. 11, was not a skin sensitizer nor was it capable of eliciting a response in the D & C Yellow No. 11-sensitized guinea pigs even at a challenge concentration of 10%. Two commercial products, a soap containing 0.015% D & C Yellow No. 11 and a shampoo containing 0.002% D & C Yellow No. 10 did not elicit a reaction in the D & C Yellow No. 11-sensitized guinea pigs.     

Studies of the quenching phenomenon in delayed contact hypersensitivity reactions

Studies in guinea pig and man have shown that eugenol can quench non-specifically contact urticarial responses, whereas limonene seems largely ineffective. In a comprehensive series of studies, there was little evidence of quenching of delayed contact hypersensitivity reactions to cinnamic aldehyde or citral, including in 'pre-quenched' material supplied by a perfume/flavour company, and in a similar mixture prepared in this laboratory, in the guinea pig model. In addition, there was no evidence of the quenching by eugenol of allergic reactions to cinnamic aldehyde in a panel of human subjects with a proven history of cinnamic-aldehyde-induced allergic contact dermatitis. Overall, the results lend little credibility to earlier literature reports of quenching phenomena in delayed contact hypersensitivity responses.     

迟发型超敏反应在药疹发病机制中的作用

迟发型超敏反应即Ⅳ型变态反应,由抗原特异性T细胞介导,根据炎症因子和效应细胞的不同,迟发型超敏反应可分为Ⅳa～Ⅳd四型.多数致敏药在人体内经代谢与蛋白结合后形成抗原,通过抗原提呈作用引起抗原特异性的皮肤固有T细胞及皮肤归巢T细胞活化,引发皮肤免疫反应.不同类型药疹的发生可涉及不同亚型的迟发型超敏反应以及不同的效应细胞与炎症因子.     

Experimental nickel sensitization in the guinea pig: comparison of different protocols

3 different sensitization protocols were compared for inducing delayed-type nickel contact hypersensitivity in guinea pigs. Open epicutaneous sensitization (OE) induced nickel allergy in 11/22 (50%) guinea pigs. When intradermal injections of Freund's complete adjuvant into the nickel-painted skin was added to the same protocol. 4/13 (31 %) became sensitized. The guinea pig maximization protocol induced nickel allergy in only 7/31 (23%) of the animals. Compared with the 2 other methods, animals sensitized with open epicutaneous applications reacted more rapidly (maximum at 6 h) and strongly (2+ reaction in 12/22 of animals) in previous patch lest sites upon systemic (i p.) nickel challenge. Open epicutaneous sensitization of guinea pigs should he a useful model for studying cellular and immunological mechanisms in nickel contact sensitivity.