Grantee Research Project Results
Final Report: Comparative In Vitro Immunotoxicology of Organochlorine Mixtures: Validation for Use in Risk Assessment
EPA Grant Number: R829361Title: Comparative In Vitro Immunotoxicology of Organochlorine Mixtures: Validation for Use in Risk Assessment
Investigators: DeGuise, Sylvain
Institution: University of Connecticut
EPA Project Officer: Hahn, Intaek
Project Period: October 1, 2001 through September 30, 2004 (Extended to September 30, 2005)
Project Amount: $666,649
RFA: Complex Chemical Mixtures (2000) RFA Text | Recipients Lists
Research Category: Hazardous Waste/Remediation , Land and Waste Management , Safer Chemicals
Objective:
The objectives of this research project were to: (1) determine the effects of in vitro exposure to mixtures of organochlorines on peripheral blood neutrophils phagocytosis and splenocytes proliferation and natural killer (NK) cell activity using B6C3F1 mice, the common model in immunotoxicology; (2) compare the effects of in vitro exposure to mixtures of organochlorines on immune functions of different species of concern, including humans and marine mammals using B6C3F1 mice as a reference species; (3) validate in mice the significance of in vitro exposure to determine immunotoxicity compared to traditional in vivo exposures; and (4) investigate the use of monoclonal and polyclonal antibodies to rapidly assess the exposure directly at the level of the individual target cell using flow cytometry, which will allow the association between exposure and effects at the cell level.
Summary/Accomplishments (Outputs/Outcomes):
Objective 1
The immunomodulatory effects of polychlorinated biphenyls (PCBs), individually and in vitro, were assessed upon in vitro exposure in humans, mice, and several (up to nine) species of marine mammals. The results are reported in several peer-reviewed publications (see below). Immune functions studied include B and T lymphocyte proliferation, phagocytosis, and respiratory burst. NK cell activity was abandoned for quality control concerns when it was realized that PCBs were directly toxic to the target cells used, which would have biased the results.
Objective 2
One of the most important findings of this project is that immunotoxicity in humans and marine mammals could not be predicted from that in mice. Dendrograms demonstrated that phylogeny could not help predict toxicity (i.e., closely related species were not more useful than mice to predict toxicity). This remained true for all immune functions tested.
Another important finding is that the toxicity of mixtures is complex. In most species and assays tested, there were abundant examples that the toxicity of a mixture was not equal to the sum of the toxicity of the components of the mixture, as evidenced with discrete examples and simple mathematical models. Several examples of synergistic and antagonistic interactions were discovered where the toxicity of a mixture was greater or lesser, respectively, than that of the sum of toxicity of the components of the mixture.
Also of concern is the fact that for most species and assays tested, the immunotoxicity was mostly associated with the noncoplanar congeners, which are generally considered relatively nontoxic, rather than with dioxin and the dioxin-like coplanar congeners, the toxicity of which is much better understood. Not surprisingly, the traditional toxicity equivalency (TEQ) approach, which measures the dioxin-like toxicity of a mixture by adding the dioxin-like toxicity of its components, consistently failed to predict the immunotoxicity of mixtures in this study. Although the mechanisms by which noncoplanar PCBs are immunotoxic are poorly understood, preliminary results in our laboratory suggest the involvement of calcium mobilization.
Objective 3
The validation of the in vitro model was performed by exposing mice (orally) with PCBs doses that resulted in tissue (blood and spleen) concentrations within the range of those used in vitro. The effects in mice were then compared with those at a similar concentration in an in vitro dose-response curve. There was a good agreement between the effects detected upon in vitro and in vivo exposure.
Objective 4
Attempts to label single cell suspensions of leukocytes with commercially available monoclonal and polyclonal antibodies to PCBs were consistently unsuccessful. This remained true when cells were experimentally exposed to individual PCBs and laboratory or commercial mixtures of PCBs. Labeling remained unsuccessful when cell membranes were permeabilized. This suggests that either: (1) the antibodies were not specific for PCBs, (2) labeling was consistently lower than background, or (3) the flow cytometric method was not sensitive enough to detect PCB exposure using this method.
Conclusions
Our study clearly demonstrated that the immunotoxicity of PCBs differs from conventional dogmas in that: (1) there are very significant differences in immunotoxicity between species and immunotoxicity in a target species cannot be predicted from that in the mouse model; (2) the toxicity of mixtures is different from that of the sum of its components; and (3) the immunotoxic effects of PCBs are largely associated with the noncoplanar congeners, which are generally considered relatively nontoxic, and through largely unknown mechanisms. The outcome of these findings is very serious, with the conventional models (the traditional mouse model and the well-accepted TEQ approach) failing to predict immunotoxicity in target species, leading to inaccurate and irrelevant risk assessment. Our results will provide the basis for the development of new, species-specific approaches to risk assessment.
Journal Articles on this Report : 6 Displayed | Download in RIS Format
Other project views: | All 56 publications | 12 publications in selected types | All 8 journal articles |
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Type | Citation | ||
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Levin M, Morsey B, Mori C, DeGuise S. Specific non-coplanar PCB-mediated modulation of bottlenose dolphin and beluga whale phagocytosis upon in vitro exposure. Journal of Toxicology and Environmental Health-Part A-Current Issues 2004;67(19):1517-1535. |
R829361 (2004) R829361 (Final) |
not available |
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Levin M, Morsey B, Mori C, Nambiar PR, DeGuise S. Non-coplanar PCB-mediated modulation of human leukocyte phagocytosis: A new mechanism for immunotoxicity. Journal of Toxicology and Environmental Health-Part A-Current Issues 2005;68(22):1977-1993. |
R829361 (2004) R829361 (Final) |
not available |
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Levin M, Morsey B, Mori C, Nambiar PR, De Guise S. PCBs and TCDD, alone and in mixtures, modulate marine mammal but not B6C3F1 mouse leukocyte phagocytosis. Journal of Toxicology and Environmental Health-Part A 2005;68(8):635-656. |
R829361 (2004) R829361 (Final) |
not available |
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Levin M, Morsey B, De Guise S. Modulation of the respiratory burst by organochlorine mixtures in marine mammals, humans and mice. Journal of Toxicology and Environmental Health-Part A 2007;70(1):73-83. |
R829361 (Final) |
not available |
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Levin M, Morsey B, De Guise S. Non-coplanar PCBs induce calcium mobilization in bottlenose dolphin and beluga whale, but not in mouse leukocytes. Journal of Toxicology and Environmental Health Part A 2007;70(14):1220-31. |
R829361 (Final) |
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Mori C, Morsey B, Levin M, Nambiar PR, De Guise S. Immunomodulatory effects of in vitro exposure to organochlorines on T-cell proliferation in marine mammals and mice. Journal of Toxicology and Environmental Health-Part A 2006;69(4):283-302. |
R829361 (2004) R829361 (Final) |
not available |
Supplemental Keywords:
PCBs, dioxin, in vitro immunotoxicity, marine mammals, human, comparative, coplanar, non-coplanar, mechanisms, risk assessment,, RFA, Scientific Discipline, Health, Waste, Environmental Chemistry, Health Risk Assessment, Chemistry, chemical mixtures, Risk Assessments, Hazardous Waste, Ecology and Ecosystems, Hazardous, complex mixtures, organochlorine mixtures, chemical exposure, risk assessment, fate and transport , biodegradation, in vitro immunotoxicology, PCBs, exposure, environmental transport and fate, PCB, human exposure, hazardous chemicals, marine mammals, immunotoxicology, animal bioassays, contaminated soils, environmental chemicals, exposure assessmentProgress and Final Reports:
Original AbstractThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.