Studies being conducted as part of an interagency agreement between NIEHS and FDA as part of the NIEHS AIDS effort are determining the relationship between decrements in immune cell phenotypes and susceptibility to infection or tumors. Studies conducted in the laboratory of Dr. James Weaver at FDA, using monoclonal antibodies (Moabs) against CD4 and CD8, to establish titration curves for the depletion of these cell subpopulations in B6C3F1 mice have been completed. Pilot studies with antibodies to CD45R/B220 and TCRb are in progress. A method for serial phenotypic analysis of mouse peripheral blood has been validated and a manuscript reporting these results has been submitted for publication. Data are comparable for both single time point and time course experiments. Time course experiments with the anti-CD8 monoclonal show that acceptable levels of depletion extend out to at least 24 days following a single depletion injection. Depletion of CD8 cells does not have significant effects on levels of CD4, or B-Cells (CD45R+). Acceptable levels of depletion can also be achieved with the anti-CD4 monoclonal, however, multiple depletion injections of 5-10 micrograms of antibody are required. Depletion of CD4+ cells has no obvious effects on levels of circulating CD8+ cells. At NIEHS, we have begun to examine the effects of cell depletion in specific host resistance models selected for these studies. These models include infection using bacteria (Listeria monocytogenes or Streptococcus pnuemoniae), an intracellular parasite (Plasmodium yoelii) or virus (Influenza) and a tumor challenge (PYB6 or B16F10). Three host resistance models will be evaluated for each cell subpopulation, with the selected model dependent on the specific target cell. Two PYB6 tumor challenges have been completed in mice depleted of CD8+ cells. As expected, no significant differences were observed in frequency or latency of PYB6 tumors following CD8 depletion. A PYB6 tumor study in CD4-depleted mice is in progress. The identification of chemicals that have the potential to cause injury to the immune system is of considerable public health significance, as alterations in immune function can lead to increased incidence of hypersensitivity disorders, autoimmune or infectious diseases or neoplasias. Experimental animal data collected over the past 15 years using standardized testing panels has provided a database from which the sensitivity and predictability of a variety of tests commonly used for the screening of chemicals for immunotoxicity has been evaluated. These results have been used as guidelines for risk assessment in immunotoxicity and have been the basis for a number of regulatory activities. In April 1998 a Workshop was held at NIEHS to establish study design to determine the sensitivity and predictability of extended histopathology as an indicator of immunotoxicity as compared with the National Toxicology Program?s functional testing battery. Standardized slide sets were generated for 10 chemicals, which had previously been evaluated for their immunotoxicity using functional tests. The histological evaluation of the slides has been completed and the data has been incorporated into a database. Dr. Russell Helms and Dr. Chris Portier, a collaborator on the original risk assessment studies, have completed the data analysis of the pathology scoring. The data indicate that for a majority of the outcomes, there was good agreement between the pathologists. However, even for variables with excellent agreement there was some evidence of inconsistency between pathologists. A direct comparison of the ratings for each pathologist indicates that, even where there was good agreement, certain individuals tended to be more conservative while others were more able to discern subtle changes than others. Additional analyses examined the consistency of a pathologist's ratings in a single tissue by investigating the correlation among all the measures in the same tissue type. When data from all pathologists were combined, measures in each of the four tissues seemed highly correlated with the other measures from that tissue. However, when correlations were examined for each pathologist independently, only the thymus evaluations maintained the same high degree of correlation. For spleen, bone marrow and lymph node measures, the outcomes for each pathologist were less well correlated, raising concern about the consistency of a pathologist's ratings across different measures in each of these tissues. A manuscript reporting these findings is in preparation.
