Host resistance to rat cytomegalovirus (RCMV) and immune function in adult PVG rats fed herring from the contaminated Baltic Sea

Abstract

The immunotoxic potential of many classes of environmental contaminants has been well established in laboratory studies, with much attention being focussed on aryl hydrocarbon (Ah)-receptor binding polychlorinated biphenyl (PCB), polychlorinated dibenzo-p-dioxin (PCDD), and polychlorinated dibenzofuran (PCDF) congeners. In a semi-field study, we previously showed that harbour seals (Phoca vitulina) fed herring from the contaminated Baltic Sea had lower natural killer cell activity, T-lymphocyte functionality and delayed-type hypersensitivity responses than seals fed herring from the relatively uncontaminated Atlantic Ocean. While ethical and practical constraints preclude in-depth studies in seals, specific reagents and a wider array of immune function tests allow such studies in laboratory rats. We therefore carried out a feeding study in rats aimed at extending our observations of contaminant-induced immunosuppression in harbour seals. The same two herring batches used in the seal study were freeze-dried, supplemented and fed to female adult PVG rats for a period of 4 1/2 months. Daily contaminant intakes of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxic equivalents (TEQ) were estimated to be 0.3 ng/kg body weight and 1.6 ng/kg in the Atlantic and Baltic groups, respectively. At the end of the feeding experiment, no contaminant-related changes in spleen CD4+/CD8+ cellularity, natural killer cell activity, or mitogen-induced proliferative responses of thymus or spleen cells could be detected. However, total thymocyte numbers and thymus CD4+/CD8+ ratios were reduced in the Baltic group. A novel model was established to assess the specific T-cell response to rat cytomegalovirus (RCMV). When applied to the feeding study, no differences between the Atlantic and Baltic groups in the RCMV-induced proliferative T-lymphocyte responses could be detected, but virus titres in salivary glands of infected rats of the Baltic Sea group were higher. These elevated RCMV titres and changes in thymus cellularity suggest that the dietary exposure to low levels of contaminants may have been immunotoxic at a level which our immune function test could not otherwise detect. While the herring diet per se appeared to have an effect on several immune function parameters, lower plasma thyroid hormone levels in the Baltic Sea group of rats confirmed that exposure to the environmental mixture of contaminants led to adverse PHAH-related health effects.

Impaired cellular immune response in rats exposed perinatally to Baltic Sea herring oil or 2,3,7,8-TCDD

“…The lower RCMV-speci®c antibody titres observed at 25 days postinfection in the two exposure groups suggest an eect of contaminants on the humoral response, possibly re-¯e cting an eect on B-cells, T-helper cells, or their precursors. In addition, the impaired RCMV-speci®c T-cell responses may indicate that activity of cytotoxic Tlymphocytes was aected in TCDD-exposed rats (Ross et al 1996c).…”

“…In the ®rst of two parallel studies established to extend our ®ndings in seals, adult PVG rats were fed a mixture of freeze-dried herring prepared from the identical two supplies used in the seal study. Despite similar intakes of contaminants between rats and seals on a body weight basis, there was no evidence of immune alterations in the rats following 4.5 months on the respective diets (Ross et al 1996c). However, higher virus titres in the salivary glands of rat cytomegalovirus (RCMV)-infected rats fed the Baltic Sea herring suggested a possible immunotoxic eect, which could not be detected using our functional assays.…”

“…Body, thymus, spleen and liver weights were recorded, and the thymus and spleen were placed aseptically in culture medium consisting of RPMI 1640 (Gibco, Grand Island, N.Y., USA), 10% heat inactivated fetal calf serum (PAA, Linz, Austria), 100 IU/ml penicillin, 100 lg/ml streptomycin and 2 mM glutamine. Cell suspensions were prepared as described elsewhere (Ross et al 1996c), counted by Coulter counter and adjusted to the required concentration.…”

“…Using a double-staining method, CD4 cells were labelled using¯uorescein isothiocyanate (FITC) labelled ER-2 (Serotec, Oxford, UK) and CD8 cells labelled with biotinylated OX8 (Serotec) monoclonal antibodies as described previously (Ross et al 1996c). A¯uorescence-activated cell scanner (FACS; Becton Dickinson, Rutherford, N.J., USA) was used to measure triplicate samples of 10 000 cells.…”

“…RCMV-speci®c stimulations consisted of a co-incubation of 3´10 3 para-formaldehyde-®xed RCMV-infected rat embryo cells (REC) and 5´10 5 spleen cells in 150 ll/well in 96-well roundbottomed cell culture plates, using methods described previously (Ross et al 1996c). Plates were incubated at 37°C in a 5% CO 2 humidi®ed incubator and [ 3 H]thymidine incorporation measured between 72 and 96 h. For the assessment of virus titres, salivary glands during both necropsies were removed aseptically and placed in Eagle basal medium (Gibco) containing 2% fetal calf serum.…”