Jaime M. Hatcher-Martin, Marla Gearing, […], and Kurt D. Pennell
Polychlorinated biphenyls (PCBs) are synthetic chemicals primarily used as coolants and insulators in electrical equipment. Although banned for several decades, PCBs continue to exist in the environment because of their long half-life, continued presence in items produced before the ban, and poor disposal practices. Epidemiological and experimental studies have identified exposure to PCBs as a potential risk factor for Parkinson’s disease, perhaps more so in females. The objective of this work was to examine the association between PCB levels in post-mortem human brain tissue and the diagnosis of Parkinson’s disease, as well as the degree of nigral depigmentation. We also sought to determine if this association was more significant when patients were stratified by sex. Post-mortem brain samples from control patients and those diagnosed with Parkinson’s disease were obtained from the Emory University Brain Bank and from the Nun Study. Concentrations of eight prevalent PCB congeners were extracted from post-mortem brain tissue and analyzed using gas chromatography-mass spectrometry. PCB congeners 153 and 180 were significantly elevated in the brains of Parkinson’s disease patients. When stratified by sex, the female Parkinson’s disease group demonstrated significantly elevated concentrations of total PCBs and specifically congeners 138, 153, and 180 compared to controls, whereas PCB concentrations in males were not significantly different between control and Parkinson’s disease groups. In a separate population of women (Nun Study) who had no clinical signs or symptoms of PD, elevated concentrations total PCB and congeners 138, 153 and 180 were also observed in post-mortem brain tissue exhibiting moderate nigral depigmentation compared to subjects with mild or no depigmentation. These quantitative data demonstrate an association between brain PCB levels and Parkinson’s disease-related pathology. Furthermore, these data support epidemiological and laboratory studies reporting a link between PCB exposure and an increased risk for Parkinson’s disease, including greater susceptibility of females.
Keywords: Parkinson’s disease, polychlorinated biphenyl, PCB, sex, exposure, neurodegenerative disease, mass spectrometry
Polychlorinated biphenyls (PCBs) are anthropogenic compounds primarily used as dielectric fluids in capacitors and transformers due to their chemical stability and physical properties. Although the manufacture, processing, and distribution of PCBs was discontinued in the United States (U.S.) in 1977 (ATSDR, 2000), total worldwide production through 1980 was estimated to be approximately 1.1 million metric tons (2.4 billion pounds), with the majority of production (60 %) occurring in the U.S. (Bletchly, 1983). Despite stringent regulatory measures, the use and servicing of “totally enclosed” PCBs, including electrical and railroad transformers, circuit breakers and liquid-filled cables, are still authorized in the U.S. (CFR, 2011). Due to the legacy of allowable uses, releases from hazardous waste sites, and improper disposal practices, combined with their persistence in the environment, PCBs continue to pose long-term threats to human health and the environment.
Of the 209 possible PCB congeners, 84 were present in commercial mixtures (e.g., Aroclors) at concentrations of greater than 0.5 weight percent (Frame et al., 1996). Of these congeners, six are widely considered to be “indicators” of PCB exposure (PCB 28, 52, 101, 138, 153, 180) based on their persistence in the environment and accumulation in animal and human tissue (Bachour et al., 2000). Of these congeners, PCBs 138, 153, and 180 were also elevated in mouse brain following exposure to Aroclor 1254:1260 mixture (Caudle et al., 2006). Due to their lipophilic (hydrophobic) properties and low rates of metabolism, the more highly chlorinated PCB congeners preferentially accumulate in fatty tissues and readily pass the blood-brain barrier. Dewailly and colleagues (1999)measured the concentrations of 14 PCB congeners in subcutaneous fat, omental fat, brain, and liver autopsy tissue from an Inuit population living in Greenland and found that congeners 138, 153 and 180 accounted for 63-68 % of the PCB burden. In brain tissue samples, concentrations of PCB congeners 138, 153 and 180 ranged from approximately 30 to 400 ppb (ug/kg lipid) (Dewailly et al., 1999)
Exposure to PCBs has been associated with a range of neurological effects, including neurobehavioral abnormalities in newborns and young children (e.g., Branchi et al., 2005; Jacobson et al., 1990; Schantz et al., 2003) and neurochemical alterations in laboratory animals (Bemis and Seegal, 2004; Malkiewicz et al., 2006; Mariussen and Fonnum, 2001; Seegal et al., 2002). A consistent finding of both in vitro and in vivo studies is the significant reduction in dopamine concentrations in the striatal tissue following acute exposures to Aroclor 1016, 1254, or 1260 (Chishti et al., 1996; Richardson and Miller, 2004; Seegal et al., 1991). Previous work in our laboratory demonstrates that exposure of mice to moderate levels of PCB mixtures (7.5 or 15 mg/kg/day of 1:1 Aroclor 1254:1260) results in dose-dependent reductions in striatal dopamine transporter (DAT) expression and function, although no observable changes in dopamine levels or tyrosine hydroxylase (TH) expression were detected (Caudle et al., 2006). Several studies have identified the importance of the DAT and the vesicular monoamine transporter 2 (VMAT2) in the regulation of dopamine levels in the striatum (Fon et al., 1997; Gainetdinov et al., 1998; Miller et al., 1999a,b). The primary function of DAT is to remove dopamine from the synapse following exocytosis, while VMAT2 sequesters cytosolic dopamine into vesicles for release. Altered transporter (i.e., DAT and VMAT2) function can impede dopamine sequestration and storage, resulting in elevated levels of dopamine in the cytoplasm, where it is available to react to form oxidized byproducts (e.g., 3,4-dihydroxyphenylacetic acid) and reactive oxygen species (ROS). Such PCB-induced oxidative stress can lead to dopaminergic cell damage, and ultimately death, as demonstrated by Lee and colleagues (2006).
Epidemiological studies have shown an increased incidence of Parkinson’s disease (PD) in humans, especially females (Standardized Mortality Ratio [SMR] = 2.95) who were occupationally-exposed to PCBs (Steenland et al., 2006). In a study of 8 Parkinson’s disease patients (3 female, 5 male) and 7 controls, elevated concentrations of PCB congeners 153 (p < 0.05) and 180 (p < 0.01) were detected in the caudate nucleus of Parkinson’s disease patients (Corrigan et al., 1998). Since these data were published, few, if any, studies have attempted to relate concentrations of PCB measured in human tissue to the incidence of Parkinson’s disease, despite the apparent association. Over this same time period, advancements in analytical instrumentation, and in particular mass spectrometry, have resulted in greatly improved mass resolution and lower detection limits. This is particularly relevant to PCBs, for which gas-chromatography (GC) combined with electron capture detection (ECD) has been the accepted analytical technique for nearly 30 years (e.g. EPA, 2000; Erickson, 1997). However, PCB quantification by GC-ECD suffers from two important limitations, (a) poor resolution that can result in co-elution of congeners that cannot be distinguished without extended runs times (e.g., > 2 hr) and/or sophisticated separation techniques (e.g., two-dimensional analysis) and (b) nonspecific response of ECDs, especially to oxygenated species, which is particularly problematic for tissue and plasma samples (Hardin et al., 1990).
The objective of this research was to quantify selected PCB congeners in post-mortem brain tissue from patients with Parkinson’s disease or Alzheimer’s disease (AD)-related neuropathology and age-matched controls using advanced gas-chromatography-mass spectrometry (GC-MS) techniques. Correlations between individual PCB congeners and summed concentrations, sex, and degree of neuropathological features are evaluated in two independent cohorts obtained from the Emory Brain Bank and the Nun Study. Confirmation of elevated PCB concentrations in the brain tissue of patients from an additional cohort exhibiting Parkinson’s disease neuropathology provides further insight into PD etiopathogenesis and supports the potential link between PCB exposures and neurodegeneration.
There is compelling epidemiological evidence to indicate that environmental factors, including PCBs, are associated with an increased risk of Parkinson’s disease (Corrigan et al., 1998; Hatcher et al., 2008; Steenland et al., 2006). However, the specific compounds and related mechanisms responsible for this association are still unclear. PCBs are a class of synthetic compounds that are persistent in the environment. From a toxicological standpoint, these compounds exhibit many features that make them ideal candidates for being involved in the risk of developing Parkinson’s disease. These compounds exhibit low volatility, chemical stability, and lipophilic properties, making them extremely persistent in the environment. In addition, the strong tendency of PCBs to bioaccumulate and biomagnify increases the risk of human exposure and accumulation at toxic levels.
In this study, we report that concentrations of selected PCB congeners are elevated in post-mortem brain tissue from patients with Parkinson’s disease. These congeners represent the most abundant PCB congeners found in the mouse brain after exposure to a mixtures of Aroclor 1254:1260, the most common form in which PCBs where used (Caudle et al., 2006). These findings corroborate other studies reporting associations between PCB exposure and an increased risk of Parkinson’s disease. These results are also consistent with previous animal studies, in which exposure of mice to moderate doses of a 1:1 mixture of Aroclor 1254 and 1260 resulted in decreased expression of dopaminergic markers, including DAT and TH (Caudle et al., 2006). When stratified by sex, we found that the association between brain PCB concentrations was primarily driven by females. This is especially interesting given that women typically have a lower incidence of Parkinson’s disease in the general population. Steenland and colleagues reported a similar increased risk in Parkinson’s disease and other neurodegenerative diseases in a cohort of PCB-exposed female workers (Steenland et al., 2006).
This study is not a mere replication of previous studies showing elevated PCBs in post-mortem human brain (Corrigan et al., 1998). The fact that these samples were collected over a decade later than previous studies demonstrate a continued association between PCBs and Parkinson’s disease. Advances in mass spectrometry over the past decade now allow for precise identification and quantification of PCBs in human samples at ppb levels. Given the long prodromal period in the disease, exposures that occurred in the last decades of the past century are very relevant to patients with recent diagnosis. Exposures from birth to age 50’s could have a significant impact on the development and progression of the disease. Therefore, even with the continued decline in environmental levels of PCBs, they remain relevant for individuals at the average age of diagnosis of Parkinson’s disease.
The Nun Study represents a unique collection of postmortem brain samples. In contrast to the participants from the Emory Cohort of post-mortem tissues who spent at least some time in their life in the southeast United States, participants in the Nun Study represent a Midwestern population with different chemical exposure profiles. While none of the 40 subjects had a diagnosis of Parkinson’s disease, which with an average age of 95 is striking, many did have neuropathological evidence of nigrostriatal degeneration. The fact that the same three PCB congeners (i.e., 138, 153, and 180) were elevated in samples with moderate nigrostriatal degeneration is remarkable as these patients represent a completely different geographical cohort from the Emory population.
The methods presented in this study provide useful means to identify chemical exposures without the obstacles of recall bias and inability to identify specific exposures. Patients with neurodegenerative diseases could have been exposed to these compounds as early as their teens, and continuing up through the point of diagnosis. Persistence of these compounds in the brain provides the cumulative and toxicokinetic features indicative of chronically-acting neurotoxicants. In light of the long prodromal period in Parkinson’s disease, such persistent neurotoxicants are in prime position to be lead candidates for the link between environmental exposure and Parkinson’s disease. It is important to note that the elevated levels of the various PCB congeners do not necessarily mean that they cause disease. Rather, the unbiased identification of these compounds, when combined with previous epidemiological and toxicological studies linking this class of compounds to the disease, provides additional evidence to support the inclusion of PCB exposure as a risk factor for Parkinson’s disease. Expanded use of the analytical techniques presented here could help further elucidate the role of persistent environmental pollutants in Parkinson’s disease, and potentially, other idiopathic neurodegenerative diseases.
Concentrations of PCB congeners in post-mortem brain tissue from Nun Study subjects exhibiting no, mild, and moderate nigral depigmentation.