Further evaluation of the local lymph node assay in the final phase of an international collaborative trial.

E.I. du Pont de Nemours, Inc. Haskell Laboratory, Newark, DE 19711-0050, USA.
Toxicology (Impact Factor: 3.75). 05/1996; 108(1-2):141-52. DOI: 10.1016/0300-483X(95)03279-O
Source: PubMed

ABSTRACT The local lymph node assay (LLNA) is a method used for the prospective identification in mice of chemicals that have the potential to cause skin sensitization. We report here the results of the second and final phase of an international trial in which the performance of the assay has been evaluated using seven test materials in five independent laboratories. The additional chemicals examined here included compounds which are considered less potent allergens than some of those tested in the first phase of the investigation, and includes hexylcinnamic aldehyde (HCA), a chemical recommended by the Organization for Economic Cooperation and Development (OECD) as a positive control for skin sensitization studies. In each laboratory all skin sensitizing chemicals examined (2,4-dinitrochlorobenzene {DNCB}, HCA, oxazolone, isoeugenal and eugenol) elicited positive responses of comparable magnitude as judged by the derived lowest concentration of test chemical required to elicit a 3-fold or greater increase in the proliferative activity of draining lymph node cells compared with vehicle-treated controls. We observed that sodium lauryl sulphate, considered to be a non-sensitizing skin irritant, also induced a positive response in the assay. Para-aminobenzoic acid (pABA), a nonsensitizing chemical, was negative at all test concentrations in each laboratory. Some laboratories incorporated minor modifications into the standard assay procedure, including the evaluation of lymph nodes pooled from individual mice rather than treatment groups and the use of statistical analyses. The use of statistics did not markedly change the determination of the lowest concentration yielding a positive response. These data confirm that the local lymph node assay is robust and yields equivalent results when performed independently.

1 Follower
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: 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.
    Toxicology 05/2012; 299(1):20-4. DOI:10.1016/j.tox.2012.05.002 · 3.75 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The skin-sensitizing potential of chemicals is an important concern for public health and thus a significant end point in the hazard identification process. To determine skin-sensitizing capacity, large research efforts focus on the development of assays, which do not require animals. As such, an in vitro test has previously been developed based on the differential expression of CREM and CCR2 transcripts in CD34(+) progenitor-derived dendritic cells (CD34-DC), which allows to classify chemicals as skin (non-)sensitizing. However, skin sensitization is not an all-or-none phenomenon, and up to now, the assessment of relative potency can only be derived using the in vivo local lymph node assay (LLNA). In our study, we analyzed the feasibility to predict the sensitizing potency, i.e., the LLNA EC3 values, of 15 skin sensitizers using in vitro data from the CD34-DC-based assay. Hereto, we extended the in vitro-generated gene expression data set by an additional source of information, the concentration of the compound that causes 20% cell damage (IC20) in CD34-DC. We statistically confirmed that this IC20 is linearly independent from the gene expression changes but that it does correlate with LLNA EC3 values. In a further analysis, we applied a robust linear regression with both IC20 and expression changes of CREM and CCR2 as explanatory variables. For 13 out of 15 compounds, a high linear correlation was established between the in vitro model and the LLNA EC3 values over a range of four orders of magnitude, i.e., from weak to extreme sensitizers.
    Toxicological Sciences 04/2010; 116(1):122-9. DOI:10.1093/toxsci/kfq108 · 4.48 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The local lymph node assay (LLNA) is used to test the potential of low molecular weight (LMW) compounds to induce sensitization via the skin. In the present study, a respiratory LLNA was developed. Male BALB/c mice were exposed head/nose-only during three consecutive days for 45, 90, 180, or 360 min/day to various LMW allergens. Ear application (skin LLNA) was used as a positive control. Negative controls were exposed to the vehicle. Three days after the last exposure, proliferation was determined in the draining mandibular lymph nodes, and the respiratory tract was examined microscopically. Upon inhalation, the allergens trimellitic anhydride, phthalic anhydride, hexamethylene diisocyanate, toluene diisocyanate, isophorone diisocyanate (IPDI), dinitrochlorobenzene, and oxazolone were positive and showed stimulation indices (SIs) up to 11, whereas trimeric IPDI, formaldehyde, and methyl salicylate were negative (viz. SI < 3). All compounds, except trimeric IPDI, induced histopathological lesions predominantly in the upper respiratory tract. Exposure by inhalation is a realistic approach to test respiratory allergens. However, based on the local toxicity, the dose that can be applied is (generally) much lower than can be achieved by skin application. It is concluded that strong LMW allergens, regardless their immunological nature, besides the skin can also sensitize the body via the respiratory tract. In addition, the contact allergens were as potent as the respiratory allergens, although the potency ranking differed from that in a skin LLNA.
    Toxicological Sciences 10/2008; 106(2):423-34. DOI:10.1093/toxsci/kfn199 · 4.48 Impact Factor