In the absence of BDE-47, cell cultures stimulated with LPS elicit an innate immune response with the production of cytokines primarily from the monocyte/macrophage cell lineage. Previous reports have shown innate immune responses are elevated in postmortem brain specimens and CSF samples from ASD subjects compared with controls (Vargas et al.,2005
). In the current study, we demonstrate for the first time, that BDE-47 pretreatment of LPS-stimulated cell cultures results in divergent innate cytokine responses in ASD children compared with TD general population pediatric controls. In cell cultures from TD controls, LPS stimulated cytokine/chemokine production was significantly reduced in the presence of 100nM BDE-47 for a number of cytokines including, GM-CSF, IL-12p40, TNFα, IL-6 MIP-1α and MIP-1β whereas only IL-6 was decreased in cell cultures from ASD children. In contrast, we demonstrate that there is increased production of IL-1β in similar cell cultures from ASD subjects, compared with no detectable change in IL-1β in cell culture from TD controls. Similarly, pre-treatment with 500nM concentrations of BDE-47 induced increased production of IL-8 in LPS stimulated cell cultures from ASD children but not TD controls. These results suggest that innate immune cytokine response may be differentially affected by BDE-47 in subjects with ASD compared with TD controls.
The innate immune response (also called natural or native immunity) is considered the first line of defense and provides a powerful and rapid response to eliminate microbes from host tissue. Innate immunity relies primarily on pattern recognition of structures that are shared by various classes of microbes but not mammalian cells, such as LPS present in gram negative bacteria and double stranded RNA in viruses. Upon encountering invading pathogens, cells of the innate immune system are responsible for their phagocytosis, processing, and the subsequent presentation of antigens in combination with MHC molecules to cells of the adaptive immune system resulting in the propagation of the immune response. Monocyte and monocyte- derived cell lineages comprise the major cell types responsible for the phagocytosis and presentation of foreign antigenic molecules from pathogens to T cells, and are an important source of immunomodulatory cytokines and chemokines. Interference in this process can have profound effects on both the innate and acquired immune systems.
Previous studies indicate that there is an over-production of pro-inflammatory cytokines, in particular IL-1β and TNFα from peripheral mononuclear cells stimulated with bacterial LPS in children with autism, findings which strongly indicate an inappropriate monocyte driven innate immune response (Singh, 1996
). However, these studies were all performed a subset of autism subjects that have gastrointestinal (GI) abnormalities. As the subjects studied herein were not evaluated for GI abnormalities by a pediatric gastroenterologist it may be that we were testing a different group of autistic individuals compared with these previous reports. In addition, the age range of our study were much younger than these previous reports and may account for the fact that although we found a slight elevation in TNFα and IL-1β levels in the ASD cases compared with controls, this did not reach statistical significance. Furthermore, our experimental conditions required that we use DMSO as a control. Although in previous experiments we found that LPS stimulated cytokine responses are the same with or without DMSO, we cannot directly compare our experiments with the previous published reports.
The presence of specific monocyte phenotypes that produce enhanced levels of cytokines has also been associated with other immune-mediated disorders such as multiple sclerosis (TNFα) and Type 1 diabetes (IFNα) (Bergh et al., 2004
). In the CNS, antigen presenting cells from the monocyte cell lineage have been found in healthy meninges, the choroid plexus, and cerebrospinal fluid (Pashenkov et al., 2003
). During inflammation in the CNS there is a recruitment of these cells where they are thought to play equal roles in the defense against infections, but may also contribute to the break-down of tolerance to CNS autoantigens (Pashenkov et al., 2003
Microglial cells share many similarities with cells of the monocyte lineage and are considered part of the CNS innate immune system. The CNS is largely populated by astroglia and microglia that not only respond to cytokines, but also produce cytokines upon activation. Cell culture studies have shown that neuropoietic cytokines such as IL-1 and IL-8 can have direct effects on neurons and glial cells, including changes in proliferation, survival, death, neurite outgrowth and gene expression (Mehler et al., 98
; Gadient et al., 1999
). Microglial cells participate in many reactive processes in the CNS and have been implicated in the exacerbation of neurological conditions such as multiple sclerosis, Alzheimer’s disease, AIDS, and viral encephalitis (Nelson et al., 2002
). Vargas et al
., recently showed the presence of innate immune activation in both brain specimens and cerebrospinal fluid (CSF) from subjects with autism (Vargas et al., 2005
), with active neuroinflammation present in the cerebral cortex and cerebellum. This inflammatory process was characterized by a marked cellular activation of microglial and astroglial cells and the presence of an altered cytokine pattern, including enhanced proinflammatory cytokine levels in the CSF. These results suggest that abnormal innate immune responses in the neuroglia of individuals with autism may influence neural function and neural development.
Organochlorine (OC) contaminants, notably polychlorinated biphenyls (PCBs), are ubiquitous in all ecosystems and found in the tissues of humans and wildlife. We have previously demonstrated this ubiquitous environmental contaminant to be associated with various adverse conditions, such as impaired immunological function in marine wildlife (Neale et al., 2002
). Although the immunotoxicity of coplanar, dioxin-like PCBs is well documented, the adverse effects exerted by non-coplanar, non-dioxinlike PCBs have received little attention. A direct causal relationship between PCB exposure and the observed detrimental effects on the immune system observed in marine mammals has yet to be fully established in humans. Levin et al (2005) demonstrated a suppression of phagocytosis by non-coplanar PCBs, suggesting a previously unrecognized aryl hydrocarbon receptor (AhR)-independent pathway. Similarly, brominated flame retardants are a novel class of environmental contaminants; within this group, the polybrominated diphenyl ethers are currently used in large quantities, disperse similarly to PCBs and DDT, and bioaccumulate and biomagnify (Herzke et al., 2005
; Meerts et al., 2001
). Little work has been done on the effects of PBDEs on immune function in humans. In one study on harbor porpoises, thymic atrophy and splenic depletion were significantly correlated with an increase in both PCB and PBDE levels (Beineke et al., 2005
). Moreover, the structural similarities between PBDEs and immunotoxic halogenated aromatic compounds suggest that the PBDEs may exert an affect on the immune system (Fernlof et al., 1997
). However, the choice of which cogener to use may be critical to realize any effect. There is in vitro
evidence that BDE-47 and its major metabolites inhibit aromatase activity (CYP19) and can be cytotoxic (Canton et al., 2005
). In addition, many of the biochemical effects of PBDEs, with BDE-47 being the most potent, on protein kinase activity and calcium homeostatis in rat neuronal cultures have been shown to be similar to those seen following exposure to structurally related PCBs (Kodavanti et al., 2005
Little is known about the effects of BDE-47 and immune function. However, we demonstrate herein that BDE-47 can affect innate immune responses previously shown to be elevated in subjects with ASD. This may be due to the lipophilic nature of the BDEs, which would allow more efficient incorporation into the lipid bilayer of (Rahman, et al
2001). We further hypothesize that patients with autism may have an altered sensitivity to the immunomodulatory effects of BDEs. It may be this increased susceptibility that is responsible for some of the immune anomalies that we have previously noted (Ashwood et al., 2006
). The precise mechanism by which PDE effects PBMC function in the pediatric populations described herein is under further investigation.