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R J Dearman

Publications and source records attributed to R J Dearman.

At least 73 records · Page 4Linked to original sources
PubMed1999

Immune responses to contact allergens: novel approaches to hazard evaluation.

Progress in our understanding of the immunobiological mechanisms that cause skin sensitization and allergic contact dermatitis has facilitated consideration of alternative approaches to hazard evaluation. One such is the murine local lymph node assay in which, in contrast to more traditional guinea pig tests, sensitizing activity is measured as a function of events associated with the induction, rather than the elicitation, phase of contact hypersensitivity. Activity in the local lymph node assay is dependent upon all of those immunological events that are initiated following first encounter with chemical allergen and which result in the stimulation of T lymphocyte proliferative responses in lymph nodes draining the site of exposure. In this respect the assay embraces in an holistic way the induction of skin sensitization. With the objective of developing in vitro approaches to hazard identification, consideration has been given to discrete immunological responses that characterize the induction of skin sensitization. Most attention has focused upon the changes induced by chemical allergens in the phenotype and function of epidermal Langerhans cells and in cytokine expression. In addition, attempts have been made to identify contact allergens as a function of their ability to provoke in vitro specific responses by unprimed T lymphocytes. These novel approaches to skin sensitization testing and their potential utility in the context of toxicological evaluations are reviewed in this article.

Allergens↗
PubMed1999

Analysis of interleukin 12 protein production and mRNA expression in mice exposed topically to chemical allergens.

Interleukin (IL) 12 is a heterodimeric cytokine which stimulates IFN-gamma production and promotes the development of type 1 T helper cells and T cytotoxic cells from their respective precursors. We have described previously that contact allergens such as 2,4-dinitrochlorobenzene (DNCB), and respiratory allergens such as trimellitic anhydride (TMA) induce discrete type 1 and type 2 cytokine secretion patterns, respectively, following repeated topical exposure of BALB/c strain mice. Under such conditions, we have now examined production by draining LNC of the inducible subunit of IL-12 (p40) and p40 and p35 subunit mRNA expression. Cultured LNC prepared from mice treated with DNCB secreted significantly more IL-12 p40 protein than did TMA- or vehicle-activated LNC, such differences becoming more pronounced as the duration of culture increased. Maximal levels of mRNA expression of the IL-12 p40 subunit were observed after 6-24 h of culture in all treatment groups and declined thereafter. Somewhat higher levels were induced following exposure to DNCB, these differences only reached statistical significance at 24 h of culture. Expression of this subunit by LNC from all treatment groups declined with time in culture. Levels of IL-12 p35 mRNA expression were comparable in cultured LNC prepared from allergen and vehicle treated mice and remained constant throughout the entire culture period. These data indicate that the divergent T cell cytokine responses seen in response to contact and respiratory allergens are associated with differential production of IL-12 p40 protein in the absence of substantial changes in the expression of mRNA for either subunit.

Acetone↗
PubMed1999

Cytokine fingerprinting: characterization of chemical allergens.

Chemical allergy is a common and important occupational health issue. Allergic sensitization induced by chemicals may take a variety of forms, including allergic contact dermatitis (skin sensitization) and allergic asthma and rhinitis (sensitization of the respiratory tract). There is a need to identify and characterize chemicals that have the potential to cause such sensitization reactions. Although a number of methods are available for the prospective analysis of skin sensitizing activity, there are currently no widely accepted tests for the identification of chemical respiratory allergens. We here describe a novel approach, cytokine fingerprinting, that has the potential to distinguish between chemical contact and respiratory allergens. The pattern of cytokine production by draining lymph node cells (LNCs) is evaluated following repeated topical exposure of mice to test chemicals. Experience to date reveals that contact allergens stimulate the selective development of type 1 immune responses associated with the secretion by draining LNCs of interferon gamma (IFN-gamma), but little interleukin-4 (IL-4) or interleukin-10 (IL-10). In contrast, chemical respiratory allergens are found to induce the appearance of preferential type 2 immune responses characterized by IL-4 and IL-10 production, but comparatively low levels of IFN-gamma. It is proposed that cytokine fingerprinting may permit the simultaneous identification and characterization of those chemicals that have the potential to cause allergic sensitization.

Allergens↗
PubMed1999

Inhibition by dexamethasone of Langerhans cell migration: influence of epidermal cytokine signals.

The influence of dexamethasone (DEX), a synthetic glucocorticoid, on the induction in mice of Langerhans cell (LC) migration has been investigated. Systemic treatment of mice with DEX was found to inhibit significantly the ability of a topically applied contact allergen (oxazolone) to induce the migration of LC from the epidermis and their subsequent accumulation as dendritic cells (DC) in draining lymph nodes. The stimulation of LC migration during skin sensitization is dependent upon signals provided by the epidermal cytokines tumour necrosis factor alpha (TNF-alpha) and interleukin 1beta (IL-1beta). It was found that treatment with DEX was unable to inhibit either LC migration or DC accumulation induced by the intradermal injection of TNF-alpha. In contrast, LC migration provoked by similar exposure of mice to IL-1beta (the action of which is dependent upon the de novo synthesis of TNF-alpha) was inhibited by DEX as was the arrival of DC in draining lymph nodes induced by this cytokine. Taken together, the data reported here indicate that DEX is able to inhibit very markedly the stimulation of LC migration during skin sensitization and it is proposed that such inhibition may represent an important aspect of the immunosuppressive properties of glucocorticoids and of their proven utility in the treatment of cutaneous inflammatory disorders. The results also indicate that DEX does not inhibit LC migration secondary to direct effects on cell motility. The proposal is that impaired LC migration results from the regulation by DEX of the de novo synthesis and/or release of TNF-alpha, an inducible epidermal cytokine that provides one important signal for LC to traffic from the skin.

Animals↗
PubMed1999

Threshold for classification as a skin sensitizer in the local lymph node assay: a statistical evaluation.

For more than 15 years, the murine local lymph node assay (LLNA) has undergone development, evaluation and validation as an alternative approach to the predictive identification of skin sensitizing chemicals. The criteria by which sensitizing chemicals are distinguished from those without significant skin sensitising hazard were developed empirically and were based on experience rather than a mathematical formula or statistical method. The current practice is to classify, as skin sensitizers, those chemicals which at one or more test concentrations stimulate a threefold or greater increase in the proliferative activity in draining lymph node cells. Despite the apparent confirmation of the utility of this approach from the extensive data available, there has not previously been any attempt to substantiate the accuracy of this criterion. In this present investigations, data from 134 chemicals tested in the LLNA and in the guinea pig and/or for which there exists clear evidence relating to human skin sensitization potential, have been subjected to a rigorous statistical evaluation using Receiver Operating Characteristic (ROC) curves. Whether the analysis is based on a comparison with guinea pig or human data, the results indicate that the empirically derived threefold threshold is an acceptable practical value for hazard identification.

Animal Testing Alternatives↗
PubMed1999

Tumour necrosis factor-alpha induces Langerhans cell migration in humans.

The role of tumour necrosis factor (TNF)-alpha in the mobilization and migration of human epidermal Langerhans cells (LC) has been investigated. Intradermal injection of normal human volunteers with homologous recombinant TNF-alpha was found to cause a dose-dependent reduction in the frequency of LC within epidermal sheets 2 h later. Equivalent results were obtained when epidermal LC were identified on the basis of either CD1a or HLA-DR expression. At the dose of TNF-alpha used routinely (500 U), treatment resulted in an average reduction in LC density of approximately 24%. Treatment with TNF-alpha was associated with a perivascular polymorphonuclear infiltration at 2 h, but the epidermis appeared normal with neither fibrinoid necrosis nor vasculitis, and LC morphology was not affected significantly. These results demonstrate that TNF-alpha provides an important signal for LC migration in humans and is likely therefore to play a crucial part in the induction of cutaneous immune responses.

Adult↗
PubMed1999

Comparison of cytokine secretion profiles provoked in mice by glutaraldehyde and formaldehyde.

BACKGROUND: Glutaraldehyde and formaldehyde are commonly used sterilizing agents that are known skin sensitizers. There is some controversy, however, regarding their capacity to cause respiratory allergy. The authors have demonstrated previously that topical exposure of mice to chemical contact allergens such as 2,4-dinitrochlorobenzene (DNCB) or respiratory allergens such as trimellitic anhydride (TMA) induces characteristic cytokine secretion profiles consistent with the selective activation of T helper1 (TH1)- and TH2-type cells, respectively. OBJECTIVE: To investigate the quality of immune response provoked following topical exposure of mice to these materials. METHODS: BALB/c strain mice were exposed topically to 50% formaldehyde or to various concentrations of glutaraldehyde in acetone. Control animals were treated concurrently with the reference contact allergen DNCB (1% in acetone:olive oil [AOO]) or with the reference respiratory sensitizer TMA (10% in AOO). Thirteen days after the initiation of exposure, draining lymph node cells (LNCs) were cultured for 12-120h and cytokine content of supernatants analysed by cytokine-specific enzyme-linked immunosorbent assay. RESULTS: DNCB-alpha and formaldehyde-activated LNCs produced high levels of the TH1-type cytokine interferon gamma, but little of the TH2-type products interleukins 4 and 10. TMA- and glutaraldehyde-stimulated LNCs displayed the converse TH2-type pattern of cytokine expression. CONCLUSIONS: These data are consistent with glutaraldehyde, but not formaldehyde, having significant potential to cause allergic sensitization of the respiratory tract.

Administration, Topical↗
PubMed1999

Reciprocal effects of interleukin-4 and interferon-gamma on immunoglobulin E-mediated mast cell degranulation: a role for nitric oxide but not peroxynitrite or cyclic guanosine monophosphate.

We report that cultured rat peritoneal cells spontaneously synthesize nitric oxide and this is associated with active suppression of mast cell secretory function. Addition of interleukin-4 (IL-4) or the nitric oxide synthase inhibitor N-monomethyl-l-arginine to peritoneal cells inhibited nitric oxide synthesis and enhanced anti-IgE-mediated mast cell degranulation, measured as serotonin release. Interferon-gamma (IFN-gamma) completely overcame the enhancement of serotonin release and suppression of nitrite production induced by IL-4. Over several experiments, with or without IL-4 and/or IFN-gamma, serotonin release correlated inversely with nitrite production. On a cell-for-cell basis, non-mast cells produced approximately 30 times more nitrite than mast cells in peritoneal cell populations, with or without IFN-gamma stimulation. The nitric oxide donor S-nitrosoglutathione inhibited anti-IgE-induced serotonin release from purified mast cells, whereas 8-bromo-cyclic GMP, the guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, superoxide dismutase and the peroxynitrite scavenger uric acid, were without effect. We conclude that IL-4 and IFN-gamma reciprocally regulate mast cell secretory responsiveness via control of nitric oxide synthesis by accessory cells; the nitric oxide effect on mast cells is direct but does not involve cyclic GMP or peroxynitrite.

Analysis of Variance↗
PubMed1999

Eotaxin and eotaxin receptor (CCR3) expression in Sephadex particle-induced rat lung inflammation.

The beta chemokine eotaxin is a potent eosinophil activator and chemoattractant. We examined immunohistochemically eotaxin protein expression in a range of normal rat tissues and in rat lung during Sephadex particle-induced pulmonary inflammation. The time course of eotaxin expression in lung at various time points after Sephadex administration was related to the appearance of eosinophils in the bronchoalveolar lavage fluid and tissue distribution of eotaxin receptor (CCR3) positive cells. Results showed that eotaxin protein was constitutively expressed by both lung airway epithelial cells and gut epithelial cells in normal tissues in the absence of inflammation. During Sephadex induced pulmonary inflammation, eotaxin expression increased in alveolar macrophages prior to the major increase in eosinophil numbers which reached a peak at 72 h. The pattern of eotaxin pulmonary expression and the location of CCR3 receptor positive cells suggest a chemoattractant gradient resulting in migration firstly into the tissue and subsequently through the airway epithelium into the airways. Treatment of rats with the glucocorticoid dexamethasone or the immunosuppressant cyclosporin A reduced eosinophil entry into lung tissue and airways but had no apparent effect on eotaxin expression in vivo, indicating that both these drugs inhibit eosinophil recruitment either by an eotaxin-independent mechanism, or by targetting factors that synergise with eotaxin, or an event post eotaxin expression.

Animals↗
PubMed1999

Identification of metal allergens in the local lymph node assay.

BACKGROUND: The murine local lymph node assay (LLNA) has recently been endorsed as a validated alternative to guinea pig methods for the identification of skin sensitization hazard. Nevertheless, there has been some debate regarding the utility of this method for the detection of metal contact allergens. OBJECTIVE: In these investigations, we have used the LLNA to determine the skin sensitization potential of 13 metal salts, 8 of which were considered to possess a significant ability to sensitize man, whereas the remaining 5 were judged to lack such potential. RESULTS: The predictions from the LLNA were correct for 7 of the 8 (88%) sensitizing metals and for 4 of the 5 (80%) nonsensitizers when considered against the experience of these metals as human skin sensitizers. Thus, the overall predictive accuracy of the LLNA in relation to metals was 11/13 (85%), which is very similar to the accuracy of approximately 88% in relation to a much larger number of low-molecular-weight organic chemicals, as reported previously. CONCLUSION: These data provide support for the potential utility of the LLNA in hazard identification of metal contact allergens.

Allergens↗
PubMed1999

The impact of vehicle on assessment of relative skin sensitization potency of 1,4-dihydroquinone in the local lymph node assay.

BACKGROUND: The murine local lymph node assay (LLNA) has been validated as an alternative method for the identification of skin sensitization hazards. Contact allergens are identified as a function of proliferative responses induced in draining lymph nodes. The quantitative nature of the LLNA data also provides the opportunity of assessing relative potency by reference to dose response analyses. OBJECTIVE: In the current investigations, the influence of vehicle on the skin sensitization potency of a known skin sensitizer (1,4-dihydroquinone) was assessed. METHODS: 1, 4-dihydroquinone was tested in the LLNA using 7 different vehicle systems in each of 2 independent laboratories. RESULTS: Results from the 2 laboratories were almost identical. LLNA dose response data were interpolated to derive the estimated concentration (EC) of 1, 4-dihydroquinone necessary to cause a three-fold stimulation of proliferation compared with controls, the EC3 value. The vehicles used and mean EC3 values obtained were: methyl ethyl ketone 0.07%, acetone 0.08%, acetone/olive oil (80/20 v/v) 0.15%, dimethyl formamide 0.22%, dimethyl sulfoxide 0.4%, and propylene glycol and acetone/saline (50/50 v/v) vehicles gave negative results. However, when tested at higher concentrations, positive results were obtained in these vehicles. CONCLUSION: These data reveal that the vehicle in which a chemical is encountered in the skin can have a significant impact on a quantitative measure of skin sensitization potency. The implication is that accurate assessment of risk to humans will require an understanding of the matrix in which skin exposure is likely to occur.

Animals↗
PubMed1999

Toxicology of protein allergenicity: prediction and characterization.

The ability of exogenous proteins to cause respiratory and gastrointestinal allergy, and sometimes systemic anaphylactic reactions, is well known. What is not clear however, are the properties that confer on proteins the ability to induce allergic sensitization. With an expansion in the use of enzymes for industrial applications and consumer products, and a substantial and growing investment in the development of transgenic crop plants that express novel proteins introduced from other sources, the issue of protein allergenicity has assumed considerable toxicological significance. There is a need now for methods that will allow the accurate identification and characterization of potential protein allergens and for estimation of relative potency as a first step towards risk assessment. To address some of these issues, and to review progress that has been made in the toxicological investigation of respiratory and gastrointestinal allergy induced by proteins, a workshop, entitled the Toxicology of Protein Allergenicity: Prediction and Characterization, was convened at the 37th Annual Conference of the Society of Toxicology in Seattle, Washington (1998). The subject of protein allergenicity is considered here in the context of presentations made at that workshop.

Allergens↗
PubMed1999

Influence of application vehicle on skin sensitization to methylchloroisothiazolinone/methylisothiazolinone: an analysis using the local lymph node assay.

The murine local lymph node assay (LLNA) is a method for the identification of skin sensitizing chemicals in which activity is measured as a function of proliferative responses induced in draining lymph nodes following topical exposure of mice to the test material. More recently, the LLNA has also been used for the determination of relative skin sensitizing potency based upon the mathematical derivation of an EC3 value, this being the estimated concentration of test chemical necessary to provoke a 3-fold increase in lymph-node cell-proliferative activity compared with concurrent vehicle-treated controls. Here we describe the use of the LLNA to determine the influence of vehicle on the skin-sensitizing potency of methylchloroisothiazolinone/methylisothiazolinone (MCI/MI), the active ingredient of preservatives such as Kathon CG. To this end, LLNA responses to MCI/ MI were measured using the vehicles 4:1 acetone:olive oil (AOO), methyl ethyl ketone, dimethylsulfoxide, dimethylformamide, propylene glycol (PG) and acetone. It was found that the vehicle in which MCI/MI was applied had a substantial impact on activity, with derived EC3 values varying from 0.0049% with AOO to 0.048% with PG. With the other vehicles, EC3 values ranged from 0.0068 to 0.0076%. The skin sensitizing potency of MCI/MI as judged from LLNA responses is consistent with what is known of the requirements for sensitization in humans. It is proposed that the LLNA not only provides a method for determination of relative skin sensitizing potency, but is also appropriate for assessing the influence of vehicle matrix on sensitizing activity.

Allergens↗
PubMed1998

Assessment of the skin sensitization potential of topical medicaments using the local lymph node assay: an interlaboratory evaluation.

The murine local lymph node assay (LLNA) is a method for the predictive identification of chemicals that have a potential to cause skin sensitization. Activity is measured as a function of lymph node cell (LNC) proliferative responses stimulated by topical application of test chemicals. Those chemicals that induce a threefold or greater increase in LNC proliferation compared with concurrent vehicle controls are classified as skin sensitizers. In the present investigations we have evaluated further the reliability and accuracy of the LLNA. In the context of an international interlaboratory trial the sensitization potentials of six materials with a history of use in topical medicaments have been evaluated: benzoyl peroxide, hydroquinone, penicillin G, streptomycin sulfate, ethylenediamine dihydrochloride, and methyl salicylate. Each chemical was analyzed in the LLNA by all five laboratories. Either the standard LLNA protocol or minor modifications of it were used. Benzoyl peroxide and hydroquinone, both human contact allergens, elicited strong LLNA responses in each laboratory. Penicillin G, another material shown previously to cause allergic contact dermatitis in humans, was also positive in all laboratories. Streptomycin sulfate induced equivocal responses, in that this material provoked a positive LLNA response in only one of the five laboratories, and then only at the highest concentration tested. Ethylenediamine dihydrochloride dissolved in a 3:1 mixture of acetone with water, or in 4:1 acetone:olive oil (one laboratory), was uniformly negative. However, limited further testing with the free base of ethylene diamine yielded a positive LLNA response when applied in acetone:olive oil (AOO). Finally, methyl salicylate, a nonsensitizing skin irritant, was negative at all test concentrations in each laboratory. Collectively these data serve to confirm that the local lymph node assay is sufficiently robust to yield equivalent results when performed independently in separate laboratories and indicate also that the LLNA is of value in assessing the skin sensitization potential of topical medicaments.

Administration, Topical↗
PubMed1998

Inducible interleukin 4 (IL-4) production and mRNA expression following exposure of mice to chemical allergens.

Previous investigations have indicated that different classes of chemical allergen provoke discrete immune responses in mice characterized by the development of selective cytokine secretion patterns by draining lymph node cells. It was found that, in mice exposed chronically to chemical respiratory allergens, lymph node cells produced high levels of mitogen-inducible interleukin 4 (IL-4), a cytokine necessary for the development of IgE antibody responses. In contrast, exposure of mice to contact allergens resulted in only low levels of IL-4 production. The studies described here were conducted to determine whether differentially induced expression of IL-4 messenger RNA was independent of the need for mitogen stimulation, unlike secretion of IL-4 protein. Mice were exposed to concentrations of trimellitic anhydride (TMA), a respiratory allergen, or to 2,4-dinitrochlorobenzene (DNCB), a contact allergen, under conditions of equivalent immunogenicity and where the former, but not the latter, resulted in significant production of mitogen-inducible IL-4 protein. The steady state levels of IL-4 mRNA in draining lymph nodes were measured by reverse transcription-polymerase chain reaction after a single or repeated daily application of the test chemical to the ears of chronically sensitized mice. Expression of mRNA for IL-4 was evaluated relative to the expression of a housekeeping gene, glyceraldehyde 3-phosphate dehydrogenase. Both chemicals elicited increased steady state levels of IL-4 mRNA by draining lymph node cells compared with vehicle-treated and naive controls. However, using two different treatment protocols, a chronic and an abbreviated chronic protocol and at all times examined, the elevation in IL-4 mRNA steady state levels induced by TMA was greater than that observed with DNCB. These data provide further support for the differential stimulation by contact and respiratory chemical allergens of cytokine secretion patterns in mice and demonstrate that the divergent production of IL-4 in response to these classes of allergen is at least partly transcriptionally regulated.

Allergens↗
PubMed1998

Production and in situ localization of cutaneous tumour necrosis factor alpha (TNF-alpha) and interleukin 6 (IL-6) following skin sensitization.

The induction of contact sensitization and other cutaneous immune responses is dependent upon the activity of epidermal cytokines. One such, keratinocyte-derived tumour necrosis factor alpha (TNF-alpha), is thought to provide the stimulus for the migration of Langerhans cells from the epidermis and their accumulation as immunocompetent dendritic cells in draining lymph nodes. In these investigations we have examined the stimulation by allergen of cutaneous TNF-alpha production and the induced epidermal expression of mRNA for TNF-alpha. Topical exposure of mice to oxazolone, a skin-sensitizing chemical, resulted in cutaneous TNF-alpha protein production that was maximal 2-h following treatment and then declined markedly. The same treatment resulted in highly localized and transient expression of epidermal TNF-alpha mRNA as judged by in situ hybridization. Epidermal mRNA for TNF-alpha was apparent 10 min following exposure to oxazolone, but was no longer detectable at 20 min. A similar pattern of TNF-alpha mRNA expression in the epidermis was provoked by intradermal exposure to interleukin 1 beta, a cytokine shown previously to induce TNF-alpha. Such rigorous regulation of temporal and spatial expression was shown not to be a characteristic of all epidermal cytokines induced by chemical allergen. Exposure to oxazolone under the same conditions resulted in a more widespread and more persistent expression of epidermal mRNA for interleukin 6. These data demonstrate that during skin sensitization the induced expression of epidermal TNF-alpha is finely controlled in space and time. It is proposed that such regulation facilitates the initiation of cutaneous immune responses while preventing excessive inflammation that would result from more persistent TNF-alpha production.

Adjuvants, Immunologic↗