Developments of immunotoxicology methods in the rat and applications to the study of environmental pollutants

Immunotoxicity can either be expressed as immunosuppression, or as allergy or autoimmunity. This paper deals with immunodeficiency investigation in the rat, and emphasis is given to the role of histopathology. As current guidelines for toxicity testing pay only marginal attention to the immune system, it is evident that improvements have been proposed for this type of study. For evaluating the immune system as a possible target, we chose a tiered testing approach, in which basis testing was done in a subacute toxicity study in the rat following the OECD guideline 407. General parameters of the immune system included careful histopathological evaluation of lymphoid organs and tissues combined with data on the weights of lymphoid organs, white blood cell parameters and serum immunoglobulin measurements. No parameters were included in this screening that would compromise these toxicity experiments (e.g. immunization). The outcome of this first tier determined the need for additional tier-two immune function studies that were aimed at confirming and characterizing further immunotoxicity, and that comprised non-specific and specific immune responses as well as host-resistance models to infectious diseases. The latter models are especially of great importance for human risk assessment. This tiered system has been validated by the known immunosuppressants azathioprine and cyclosporine A. Although conventional histopathology has shown its great value in identifying immunotoxic agents, morphological characterization can be improved by immunohistochemistry, hybrido-histochemistry (in situ hybridization) and morphometric analyses; these technologies are also important for the characterization of the mechanism of toxicant-induced immune alterations. Insight into the mechanism of action can also be provided by using specific animal models such as the athymic rat and the so-called severe combined immune deficient (scid) mouse, in which lymphoid cells of human and animal origin (e.g. thymus tissue) can be grafted. By comparing the sensitivity of the grafted tisues with immunotoxicants, data can be obtained that are useful for the assessment of the immunotoxicological risk for humans. Examples of chemicals that have been identified to be immunotoxic in the tiered test system in the rat are the environmental contaminants hexachlorobenzene and tributyltin oxide, and the results of these studies were discussed. Based on these data it was recommended that additional test parameters be incorporated into the existing OECD guideline.