Mercury exposure, serum antinuclear antibodies, and serum cytokine levels in the Long Island Study of Seafood Consumption: A cross-sectional study in NY, USA.

Mercury (Hg) is a well-known neurotoxin, and has been more recently studied specifically as an immunotoxin. In experimental and a few epidemiologic studies, Hg has been associated with distinct cytokine profiles and antinuclear antibody (ANA) positivity, though patterns at lower levels of exposure, typical of seafood consumers with a western diet, are not well characterized. Seafood consumers (n=287) recruited on Long Island, NY completed food frequency and health questionnaires and provided blood for analysis of Hg, poly-unsaturated fatty acids (omega-3 and omega-6 fatty acids), selenium (Se), ANA, and several cytokines (IL-1ß, IL-4, IL-10, TNF-α, IL-17, IFN-γ, and IL-1ra). Logistic and linear regression analyses were conducted to evaluate associations between serum Hg and cytokines and ANA. Adjusted models accounted for gender, age, ethnicity, income, education, smoking, BMI, selenium, omega-3 fatty acids, omega-6 fatty acids, omega-6/omega-3 ratio, and fish intake. Sex-stratified models were also generated with the expectation that immune profiles would differ between women and men. Median blood Hg was 4.58µg/L with 90th %ile =19.8µg/L. Nine individuals displayed ANA positivity at serum titers above 1:80; many of the cytokines were below detection limits, and the ability to detect was used in the logistic regression analyses. In linear and logistic regression analyses, Hg was not significantly associated with any of the seven investigated cytokines or with ANA-positivity. Therefore, Hg was not associated with altered immune profiles in this population of seafood consumers.

Low-dose mercury heightens early innate response to coxsackievirus infection in female mice.

OBJECTIVE: Mercury is a ubiquitous environmental contaminant with toxic outcomes over a range of exposures. In this study, we investigated the effects of mercury exposure on early immune responses to coxsackievirus B3 (CVB3) infection in a murine model of autoimmune heart disease. MATERIALS AND METHODS: Female BALB/c mice, susceptible to CVB3-induced autoimmune myocarditis, were treated with mercuric chloride (200 µg/kg body weight every other day for 2 weeks) prior to infection with CVB3. Six hours post-infection, immune cells were isolated from the spleen and peritoneum for flow cytometry, gene expression, and cytokine profiling. Thirty-five days post-infection, hearts were collected for histological examination of immune cell infiltration. RESULTS: As for male mice, mercury exposure significantly increased autoimmune myocarditis and immune infiltration into the heart. During the innate response 6 h post-infection, mercury increased expression of co-stimulatory molecules and innate immune receptors on peritoneal macrophages. At the same time point, the alternatively activated macrophage gene, arginase, was increased while the classically activated macrophage gene, inducible nitric oxide synthase, was unaffected. Expression of activation markers were decreased on peritoneal B cells with mercury exposure while T cells were unaffected. Mercury increased production of pro-inflammatory mediators in the spleen. Macrophage-recruiting chemokines and activating cytokines, such as CCL2, CCL4, and IL-6, were increased with mercury following CVB3 infection. CONCLUSIONS: Thus, mercury treatment exacerbates autoimmune myocarditis in female mice and alters early innate signaling on peritoneal macrophages. Mercury also modulates the cytokine profile in the spleen toward a macrophage-activating milieu, and upregulates alternatively activated macrophage genes, providing evidence that mercury exposure promotes inflammation in the context of infection.

Novel biomarkers of mercury-induced autoimmune dysfunction: a cross-sectional study in Amazonian Brazil.

Mercury is a ubiquitous environmental contaminant, causing both neurotoxicity and immunotoxicity. Given its ability to amalgamate gold, mercury is frequently used in small-scale artisanal gold mining. We have previously reported that elevated serum titers of antinuclear autoantibodies (ANA) are associated with mercury exposures of miners in gold mining. The goal of this project was to identify novel serum biomarkers of mercury-induced immunotoxicity and autoimmune dysregulation. We conducted an analysis of serum samples from a cross-sectional epidemiological study on miners working in Amazonian Brazil. In proteomic screening analyses, samples were stratified based on mercury concentrations and ANA titer and a subset of serum samples (N=12) were profiled using Immune Response Biomarker Profiling ProtoArray protein microarray for elevated autoantibodies. Of the up-regulated autoantibodies in the mercury-exposed cohort, potential target autoantibodies were selected based on relevance to pro-inflammatory and macrophage activation pathways. ELISAs were developed to test the entire sample cohort (N=371) for serum titers to the highest of these autoantibodies (anti-glutathione S-transferase alpha, GSTA1) identified in the high mercury/high ANA group. We found positive associations between elevated mercury exposure and up-regulated serum titers of 3760 autoantibodies as identified by ProtoArray. Autoantibodies identified as potential novel biomarkers of mercury-induced immunotoxicity include antibodies to the following proteins: GSTA1, tumor necrosis factor ligand superfamily member 13, linker for activation of T cells, signal peptide peptidase like 2B, stimulated by retinoic acid 13, and interferon induced transmembrane protein. ELISA analyses confirmed that mercury-exposed gold miners had significantly higher serum titers of anti-GSTA1 autoantibody [unadjusted odds ratio=89.6; 95% confidence interval: 27.2, 294.6] compared to emerald miners (referent population). Mercury exposure was associated with increased titers of several autoantibodies in serum including anti-GSTA1. These proteins play a wide variety of roles, including as antioxidants, in the regulation of pro- and anti-inflammatory cytokines, as well as danger and oxidative stress signaling. Dysregulation of these proteins and pathways is believed to play a role in autoimmune diseases such as rheumatoid arthritis, Sjögren׳s syndrome, and multiple sclerosis. Taken together, these results suggest that mercury exposure can induce complex autoimmune dysfunction and the immunotoxic effects of this dysfunction may be measured by serum titers to autoantibodies such as anti-GSTA1.

Mechanisms of environmental influence on human autoimmunity: a National Institute of Environmental Health Sciences expert panel workshop.

The mechanisms leading to autoimmune diseases remain largely unknown despite numerous lines of experimental inquiry and epidemiological evidence. The growing number of genome-wide association studies and the largely incomplete concordance for autoimmune diseases in monozygotic twins support the role of the environment (including infectious agents and chemicals) in the breakdown of tolerance leading to autoimmunity via numerous mechanisms. The present article reviews the major theories on the mechanisms of the environmental influence on autoimmunity by addressing the different degrees of confidence that characterize our knowledge. The theories discussed herein include (i) the role of innate immunity mediated by toll-like receptors in triggering the autoimmune adaptive response characterizing the observed pathology; (ii) changes in spleen marginal zone B cells in autoantibody production with particular focus on the B10 subpopulation; (iii) Th17 cell differentiation and T regulatory cells in the aryl hydrocarbon receptor model; (iv) self antigen changes induced by chemical and infectious agents which could break tolerance by post-translational modifications and molecular mimicry; and finally (v) epigenetic changes, particularly DNA methylation, that are induced by environmental stimuli and may contribute to autoimmunity initiation. We are convinced that these working hypotheses, in most cases supported by solid evidence, should be viewed in parallel with animal models and epidemiological observations to provide a comprehensive picture of the environmental causes of autoimmune diseases.

Fetal and maternal immune responses to methylmercury exposure: a cross-sectional study.

Methylmercury (MeHg) is a ubiquitous environmental contaminant with known neurodevelopmental effects. In humans, prenatal exposures primarily occur through maternal consumption of contaminated fish. In this study, we evaluated the association between prenatal exposure to MeHg and titers of total immunoglobulins (Ig) and specific autoantibodies in both mothers and fetuses by analyzing maternal and cord blood serum samples. We examined multiple immunoglobulin isotypes to determine if these biomarkers could inform as to fetal or maternal responses since IgG but not IgM can cross the placenta. Finally, we evaluated serum cytokine levels to further characterize the immune response to mercury exposure. The study was conducted using a subset of serum samples (N=61 pairs) collected from individuals enrolled in a population surveillance of MeHg exposures in the Brazilian Amazon during 2000/2001. Serum titers of antinuclear and antinucleolar autoantibodies were measured by indirect immunofluorescence. Serum immunoglobulins were measured by enzyme-linked immunosorbent assay (ELISA) and BioPlex multiplex assay. Serum cytokines were measured by BioPlex multiplex assay. In this population, the geometric mean mercury level was within the 95th percentile for US populations of women of childbearing age but the upper level of the range was significantly higher. Fetal blood mercury levels were higher (1.35 times) than those in their mothers, but highly correlated (correlation coefficient [r]=0.71; 95% CI: 0.54, 0.89). Total IgG (r=0.40; 95% CI: 0.19, 0.62) and antinuclear autoantibody (odds ratio [OR]=1.05; 95% CI: 1.02, 1.08) levels in paired maternal and fetal samples were also associated; in contrast, other immunoglobulin (IgM, IgE, and IgA) levels were not associated between pairs. Total IgG levels were significantly correlated with both maternal (r=0.60; 95% CI: 0.25, 0.96) and cord blood mercury levels (r=0.61; 95% CI: 0.25, 0.97), but individual isotypes were not. Serum cytokines, interleukin-1ß (r=0.37; 95% CI: 0.01, 0.73), interleukin-6 (r=0.34; 95% CI: 0.03, 0.65), and tumor necrosis factor-α (r=0.24; 95% CI: 0.015, 0.47), were positively correlated between maternal and fetal samples. Antinuclear and antinucleolar autoantibody titer and serum cytokine levels, in either maternal or cord blood, were not significantly associated with either maternal or cord blood mercury levels. These data provide further evidence that there are likely IgG biomarkers of mercury-induced immunotoxicity in this population since IgG levels were elevated with increased, and associated with, mercury exposure. However, unlike previous data from adult males and non-pregnant females, we found no evidence that antinuclear and antinucleolar autoantibody titer is a reliable biomarker of mercury immunotoxicity in this population.

Mercury exposure, serum antinuclear/antinucleolar antibodies, and serum cytokine levels in mining populations in Amazonian Brazil: a cross-sectional study.

Mercury is an immunotoxic substance that has been shown to induce autoimmune disease in rodent models, characterized by lymphoproliferation, overproduction of immunoglobulin (IgG and IgE), and high circulating levels of auto-antibodies directed at antigens located in the nucleus (antinuclear auto-antibodies, or ANA) or the nucleolus (antinucleolar auto-antibodies, or ANoA). We have reported elevated levels of ANA and ANoA in human populations exposed to mercury in artisanal gold mining, though other confounding variables that may also modulate ANA/ANoA levels were not well controlled. The goal of this study is to specifically test whether occupational and environmental conditions (other than mercury exposure) that are associated with artisanal gold mining affect the prevalence of markers of autoimmune dysfunction. We measured ANA, ANoA, and cytokine concentrations in serum and compared results from mercury-exposed artisanal gold miners to those from diamond and emerald miners working under similar conditions and with similar socio-economic status and risks of infectious disease. Mercury-exposed gold miners had higher prevalence of detectable ANA and ANoA and higher titers of ANA and ANoA as compared to diamond and emerald miners with no occupational mercury exposure. Also, mercury-exposed gold miners with detectable ANA or ANoA in serum had significantly higher concentrations of pro-inflammatory cytokines IL-1beta, TNF-alpha, and IFN-gamma in serum as compared to the diamond and emerald miners. This study provides further evidence that mercury exposure may lead to autoimmune dysfunction and systemic inflammation in affected populations.

In vitro interactions between splenocytes and dansylamide dye-embedded nanoparticles detected by flow cytometry.

Engineered nanoparticles (NPs) possess a range of biological activity. In vitro methods for assessing toxicity and efficacy would be enhanced by simultaneous quantitative information on the behavior of NPs in culture systems and signals of cell response. We have developed a method for visualizing NPs within cells using standard flow-cytometric techniques and uniquely designed spherical siloxane NPs with an embedded (covalently bound) dansylamide dye. This method allowed NP visualization without obscuring detection of relevant biomarkers of cell subtype, activation state, and other events relevant to assessing bioactivity. We determined that NPs penetrated cells and induced a range of biological signals consistent with activation and costimulation. These results indicate that NPs may affect cell function at concentrations below those inducing cytotoxicity or apoptosis and demonstrate a novel method to image both localization of NPs and cell-level effects. FROM THE CLINICAL EDITOR: A method for visualizing NPs within cells using standard flow-cytometric techniques is reported in this paper. The novel method allowed NP visualization without obscuring detection of relevant biomarkers of cell subtype, activation state, and other events relevant to assessing bioactivity. NPs also induced a range of biological signals consistent with activation and costimulation.

Modulators of mercury risk to wildlife and humans in the context of rapid global change.

Environmental mercury (Hg) contamination is an urgent global health threat. The complexity of Hg in the environment can hinder accurate determination of ecological and human health risks, particularly within the context of the rapid global changes that are altering many ecological processes, socioeconomic patterns, and other factors like infectious disease incidence, which can affect Hg exposures and health outcomes. However, the success of global Hg-reduction efforts depends on accurate assessments of their effectiveness in reducing health risks. In this paper, we examine the role that key extrinsic and intrinsic drivers play on several aspects of Hg risk to humans and organisms in the environment. We do so within three key domains of ecological and human health risk. First, we examine how extrinsic global change drivers influence pathways of Hg bioaccumulation and biomagnification through food webs. Next, we describe how extrinsic socioeconomic drivers at a global scale, and intrinsic individual-level drivers, influence human Hg exposure. Finally, we address how the adverse health effects of Hg in humans and wildlife are modulated by a range of extrinsic and intrinsic drivers within the context of rapid global change. Incorporating components of these three domains into research and monitoring will facilitate a more holistic understanding of how ecological and societal drivers interact to influence Hg health risks.

Inflammatory response following in vitro exposure to methylmercury with and without n-3 long chain polyunsaturated fatty acids in peripheral blood mononuclear cells from systemic lupus erythematosus patients compared to healthy controls.

Methylmercury (MeHg) is a proposed environmental stimulus in systemic lupus erythematosus (SLE). Humans are primarily exposed to MeHg through fish consumption. Fish are also important sources of n-3 long chain polyunsaturated fatty acids (n-3 LCPUFA). This in vitro study investigated the inflammatory response of isolated peripheral blood mononuclear cells (PBMCs), when exposed to either MeHg alone or with added n-3 LCPUFA, from SLE patients (N = 12) compared to healthy sex matched controls (N = 12). The PBMCs were isolated and exposed to 200 nM of MeHg for 24 h with or without pre-exposure to eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA) at a concentration of 100 µM each. Supernatants were analyzed for the inflammatory markers. Following exposure to MeHg, mean TNF-α concentrations were significantly higher in SLE patients (2226.01 ±â€¯348.98pg/ml) compared to controls (701.40 ±â€¯680.65 pg/ml) (P = .008). Pre-exposure of cells with MeHg and EPA resulted in a significantly higher concentration of IL-8 in supernatants from SLE patients (2137.83 ±â€¯1559.01 pg/ml) compared to that of the controls (879.26 ±â€¯979.49 pg/ml) (P = .030). EPA and DHA attenuated the pro-inflammatory inducing effects of MeHg in SLE and control cells. In summary, exposure to MeHg stimulated a higher TNF-α response in SLE patients compared with healthy controls; nevertheless the presence of n-3 LCPUFA reduced the overall inflammatory response, albeit to a lesser degree in SLE patients.

Sex differences in coxsackievirus B3-induced myocarditis: IL-12Rbeta1 signaling and IFN-gamma increase inflammation in males independent from STAT4.

Cardiovascular disease is the number one killer of men and women in North America. Male BALB/c mice infected with coxsackievirus B3 (CVB3) develop more severe inflammatory heart disease compared to female mice, similar to the increased heart disease that occurs in men. We show here that increased inflammation in male mice is not due to increased viral replication in the heart, but associated with increased proinflammatory cytokines IL-1beta, IL-18 and IFN-gamma. We have previously reported that IL-12Rbeta1 signaling increases CVB3-induced myocarditis and IL-1beta/IL-18 levels in males, while IL-12(p35)/STAT4-induced IFN-gamma does not alter the severity of acute disease. However, whether differences exist between males and females in these two cytokine signaling pathways is unknown. In this study, we examined sex differences in 1) IL-12Rbeta1 signaling or 2) STAT4/IFN-gamma pathways following CVB3 infection in BALB/c mice. We found that male and female mice deficient in IL-12Rbeta1 had decreased inflammation and viral replication in the heart, indicating that IL-12Rbeta1 signaling increases myocarditis in both sexes. In contrast, STAT4 deficiency did not alter the sex difference in myocarditis, with males maintaining increased inflammation over females. IFN-gamma deficient males, however, had decreased myocarditis and viral replication compared to females. Thus, IFN-gamma increases inflammation in males independent from STAT4. These results demonstrate that sex differences greatly influence viral replication and the severity of acute CVB3-induced myocarditis.

Mercury, selenium and fish oils in marine food webs and implications for human health.

Humans who eat fish are exposed to mixtures of healthful nutrients and harmful contaminants that are influenced by environmental and ecological factors. Marine fisheries are composed of a multitude of species with varying life histories, and harvested in oceans, coastal waters and estuaries where environmental and ecological conditions determine fish exposure to both nutrients and contaminants. Many of these nutrients and contaminants are thought to influence similar health outcomes (i.e., neurological, cardiovascular, immunological systems). Therefore, our understanding of the risks and benefits of consuming seafood require balanced assessments of contaminants and nutrients found in fish and shellfish. In this paper, we review some of the reported benefits of fish consumption with a focus on the potential hazards of mercury exposure, and compare the environmental variability of fish oils, selenium and mercury in fish. A major scientific gap identified is that fish tissue concentrations are rarely measured for both contaminants and nutrients across a range of species and geographic regions. Interpreting the implications of seafood for human health will require a better understanding of these multiple exposures, particularly as environmental conditions in the oceans change.

Mercury and autoimmunity: implications for occupational and environmental health.

Mercury (Hg) has long been recognized as a neurotoxicant; however, recent work in animal models has implicated Hg as an immunotoxicant. In particular, Hg has been shown to induce autoimmune disease in susceptible animals with effects including overproduction of specific autoantibodies and pathophysiologic signs of lupus-like disease. However, these effects are only observed at high doses of Hg that are above the levels to which humans would be exposed through contaminated fish consumption. While there is presently no evidence to suggest that Hg induces frank autoimmune disease in humans, a recent epidemiological study has demonstrated a link between occupational Hg exposure and lupus. In our studies, we have tested the hypothesis that Hg does not cause autoimmune disease directly, but rather that it may interact with triggering events, such as genetic predisposition, exposure to antigens, or infection, to exacerbate disease. Treatment of mice that are not susceptible to Hg-induced autoimmune disease with very low doses and short term exposures of inorganic Hg (20-200 microg/kg) exacerbates disease and accelerates mortality in the graft versus host disease model of chronic lupus in C57Bl/6 x DBA/2 mice. Furthermore, low dose Hg exposure increases the severity and prevalence of experimental autoimmune myocarditis (induced by immunization with cardiac myosin peptide in adjuvant) in A/J mice. To test our hypothesis further, we examined sera from Amazonian populations exposed to Hg through small-scale gold mining, with and without current or past malaria infection. We found significantly increased prevalence of antinuclear and antinucleolar antibodies and a positive interaction between Hg and malaria. These results suggest a new model for Hg immunotoxicity, as a co-factor in autoimmune disease, increasing the risks and severity of clinical disease in the presence of other triggering events, either genetic or acquired.

Mercury exposure, malaria, and serum antinuclear/antinucleolar antibodies in Amazon populations in Brazil: a cross-sectional study.

BACKGROUND: Mercury is an immunotoxic metal that induces autoimmune disease in rodents. Highly susceptible mouse strains such as SJL/N, A.SW, B10.S (H-2s) develop multiple autoimmune manifestations after exposure to inorganic mercury, including lymphoproliferation, elevated levels of autoantibodies, overproduction of IgG and IgE, and circulating immune complexes in kidney and vasculature. A few studies have examined relationships between mercury exposures and adverse immunological reactions in humans, but there is little evidence of mercury-associated autoimmunity in humans. METHODS: To test the immunotoxic effects of mercury in humans, we studied communities in Amazonian Brazil with well-characterized exposures to mercury. Information was collected on diet, mercury exposures, demographic data, and medical history. Antinuclear and antinucleolar autoantibodies (ANA and ANoA) were measured by indirect immunofluorescence. Anti-fibrillarin autoantibodies (AFA) were measured by immunoblotting. RESULTS: In a gold mining site, there was a high prevalence of ANA and ANoA: 40.8% with detectable ANoA at > or =1:10 serum dilution, and 54.1% with detectable ANA (of which 15% had also detectable ANoA). In a riverine town, where the population is exposed to methylmercury by fish consumption, both prevalence and levels of autoantibodies were lower: 18% with detectable ANoA and 10.7% with detectable ANA. In a reference site with lower mercury exposures, both prevalence and levels of autoantibodies were much lower: only 2.0% detectable ANoA, and only 7.1% with detectable ANA. In the gold mining population, we also examined serum for AFA in those subjects with detectable ANoA (> or =1:10). There was no evidence for mercury induction of this autoantibody. CONCLUSIONS: This is the first study to report immunologic changes, indicative of autoimmune dysfunction in persons exposed to mercury, which may also reflect interactions with infectious disease and other factors.

Determining biological activity of nanoparticles as measured by flow cytometry.

Flow cytometry is a powerful tool to evaluate cellular responses at the single cell level. Applicability to evaluating biological activity of in vitro and in vivo exposure to compounds is limited only by the number of fluorochrome emission spectra a particular instrument can detect and the availability of antibodies specific for a particular cellular protein. Here, I describe the general method considerations and provide an example experimental design for utilizing flow cytometry to evaluate the biological activity of nanoparticles on primary murine immune cells.

Low-dose inorganic mercury increases severity and frequency of chronic coxsackievirus-induced autoimmune myocarditis in mice.

Mercury is a widespread environmental contaminant with neurotoxic impacts that have been observed over a range of exposures. In addition, there is increasing evidence that inorganic mercury (iHg) and organic mercury (including methyl mercury) have a range of immunotoxic effects, including immune suppression and induction of autoimmunity. In this study, we investigated the effect of iHg on a model of autoimmune heart disease in mice induced by infection with coxsackievirus B3 (CVB3). We examined the role of timing of iHg exposure on disease; in some experiments, mice were pretreated with iHg (200 µg/kg, every other day for 15 days) before disease induction with virus inoculation, and in others, they were treated with iHg after the acute (viral) phase of disease but before the development of dilated cardiomyopathy (DCM). iHg alone had no effect on heart pathology. Pretreatment with iHg before CVB3 infection significantly increased the severity of chronic myocarditis and DCM compared with control animals receiving vehicle alone. In contrast, treatment with iHg after acute myocarditis did not affect the severity of chronic disease. The increased chronic myocarditis, fibrosis, and DCM induced by iHg pretreatment were not due to increased viral replication in the heart, which was unaltered by iHg treatment. iHg pretreatment induced a macrophage infiltrate and mixed cytokine response in the heart during acute myocarditis, including significantly increased interleukin (IL)-12, IL-17, interferon-γ, and tumor necrosis factor-α levels. IL-17 levels were also significantly increased in the spleen during chronic disease. Thus, we show for the first time that low-dose Hg exposure increases chronic myocarditis and DCM in a murine model.

Biomarkers of methylmercury exposure immunotoxicity among fish consumers in Amazonian Brazil.

BACKGROUND: Mercury (Hg) is a ubiquitous environmental contaminant with neurodevelopmental and immune system effects. An informative biomarker of Hg-induced immunotoxicity could aid studies on the potential contribution to immune-related health effects. OBJECTIVES: Our objectives were to test the hypothesis that methylmercury (MeHg) exposures affect levels of serum biomarkers and to examine interactions between Hg and selenium (Se) in terms of these responses. METHODS: This cross-sectional epidemiological study assessed adults living along the Tapajós River, a system long affected by MeHg. We measured antinuclear (ANA) and antinucleolar (ANoA) autoantibody levels and eight cytokines in serum samples (n = 232). Total Hg (including MeHg) and Se were measured in blood, plasma, hair, and urine. RESULTS: The median (range) total Hg concentrations were 14.1 µg/g (1.1-62.4), 53.5 µg/L (4.3-288.9), 8.8 µg/L (0.2-40), and 3.0 µg/L (0.2-16.1) for hair, blood, plasma, and urine, respectively. Elevated titers of ANA (but not ANoA) were positively associated with MeHg exposure (log-transformed, for blood and plasma), unadjusted [odds ratio (OR) = 2.6; 95% confidence interval (CI): 1.1, 6.2] and adjusted for sex and age (OR = 2.9; 95% CI: 1.1, 7.5). Proinflammatory [interleukin (IL)-6 and interferon (IFN)-γ], anti-inflammatory (IL-4), and IL-17 cytokine levels were increased with MeHg exposure; however, in the subset of the population with elevated ANA, proinflammatory IL-1ß, IL-6, IFN-γ, and tumor necrosis factor (TNF)-α and anti-inflammatory (IL-4) cytokine levels were decreased with MeHg exposure. Although Se status was associated with MeHg level (correlation coefficient = 0.86; 95% CI: 0.29, 1.43), Se status was not associated with any changes in ANA and did not modify associations between Hg and ANA titers. CONCLUSIONS: MeHg exposure was associated with an increased ANA and changes in serum cytokine profile. Moreover, alterations in serum cytokine profiles differed based on ANA response, suggesting a specific phenotype of MeHg susceptibility. Further research on the potential health implications of these observed immunological changes is warranted.

Differential immunotoxic effects of inorganic and organic mercury species in vitro.

Despite the fact that humans are exposed to multiple forms of mercury (elemental, inorganic, and organic), most research on mercury toxicity has focused on methylmercury (MeHg) and on neurotoxic outcomes and mechanisms. Recent work has indicated that the immunotoxic effects of mercury compounds may be significant contributors to human disease as well as mechanistically relevant to other target organ toxicities. In this study, we compared the effects of inorganic Hg (iHg) to organic Hg species (MeHg and ethylmercury, EtHg) in human peripheral blood mononuclear cells (PBMCs) in vitro at sub-cytotoxic concentrations, using methods developed to characterize response of human PBMCs to iHg in vitro. PBMCs were isolated from six volunteer blood donors (three males and three females) and cultured in the presence and absence of lipopolysaccharide (LPS) and low levels (up to 200nM of each Hg species, separately) for 24h in culture. Cell culture supernatants were analyzed for cytokine concentrations with a bead-based multiplex assay. We report that iHg and MeHg both increase pro-inflammatory cytokine release in LPS-stimulated PBMCs, while EtHg decreases IFN-gamma release as well pro-inflammatory cytokine release. IL-17 release is significantly increased only in response to iHg treatment. Levels of anti-inflammatory cytokines (IL-1Ra and IL-10) were not significantly altered by any Hg treatment. These results indicate that both organic and inorganic species of Hg can affect the human immune system, but that they may exert different effects on immune function.

Mercury induces an unopposed inflammatory response in human peripheral blood mononuclear cells in vitro.

BACKGROUND: The human immune response to mercury is not well characterized despite the body of evidence that suggests that Hg can modulate immune responses, including the induction of autoimmune disease in some mouse models. Dysregulation of cytokine signaling appears to play an important role in the etiology of Hg-induced autoimmunity in animal models. OBJECTIVES: In this study, we systematically investigated the human immune response to Hg in vitro in terms of cytokine release. METHODS: Human peripheral blood mononuclear cells (PBMCs) were isolated from 20 volunteers who donated blood six separate times. PBMCs were cultured with lipopolysaccharide and concentrations of mercuric chloride (HgCl(2)) up to 200 nM. Seven cytokines representing important pathways in physiologic and pathologic immune responses were measured in supernatants. We used multilevel models to account for the intrinsic clustering in the cytokine data due to experimental design. RESULTS: We found a consistent increase in the release of the proinflammatory cytokines interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha, and concurrent decrease in release of the antiinflammatory cytokines interleukin 1-receptor antagonist (IL-1Ra) and IL-10 in human PBMCs treated with subcytotoxic concentrations of HgCl(2). IL-4, IL-17, and interferon-gamma increased in a concentration-response manner. These results were replicated in a second, independently recruited population of 20 different volunteers. CONCLUSIONS: Low concentrations of HgCl(2) affect immune function in human cells by dysregulation of cytokine signaling pathways, with the potential to influence diverse health outcomes such as susceptibility to infectious disease or risk of autoimmunity.

Cutting edge: cross-regulation by TLR4 and T cell Ig mucin-3 determines sex differences in inflammatory heart disease.

Recent clinical studies have reinforced the importance of sex-related differences in the pathogenesis of cardiovascular diseases, with an increased incidence and mortality in men. Similar to humans, male BALB/c mice infected with coxsackievirus B3 (CVB3) develop more severe inflammation in the heart even though viral replication is no greater than in females. We show that TLR4 and IFN-gamma levels are significantly elevated and regulatory T cell (Treg) populations significantly reduced in the heart of males following CVB3 infection, whereas females have significantly increased T cell Ig mucin (Tim)-3, IL-4 and Treg. Blocking Tim-3 in males significantly increases inflammation and TLR4 expression while reducing Treg. In contrast, defective TLR4 signaling significantly reduces inflammation while increasing Tim-3 expression. Cross-regulation of TLR4 and Tim-3 occurs during the innate and adaptive immune response. This novel mechanism may help explain why inflammatory heart disease is more severe in males.

Complement receptor 1 and 2 deficiency increases coxsackievirus B3-induced myocarditis, dilated cardiomyopathy, and heart failure by increasing macrophages, IL-1beta, and immune complex deposition in the heart.

Complement and complement receptors (CR) play a central role in immune defense by initiating the rapid destruction of invading microorganisms, amplifying the innate and adaptive immune responses, and mediating solubilization and clearance of immune complexes. Defects in the expression of C or CR have been associated with loss of tolerance to self proteins and the development of immune complex-mediated autoimmune diseases such as systemic lupus erythematosus. In this study, we examined the role of CR on coxsackievirus B3 (CVB3)-induced myocarditis using mice deficient in CR1/2. We found that CR1/2 deficiency significantly increased acute CVB3 myocarditis and pericardial fibrosis resulting in early progression to dilated cardiomyopathy and heart failure. The increase in inflammation was not due to increased viral replication, which was not significantly altered in the hearts of CR1/2-deficient mice, but was associated with increased numbers of macrophages, IL-1beta levels, and immune complex deposition in the heart. The complement regulatory protein, CR1-related gene/protein Y (Crry), was increased on cardiac macrophage populations, while immature B220(low) B cells were increased in the spleen of CR1/2-deficient mice during acute CVB3-induced myocarditis. These results show that expression of CR1/2 is not necessary for effective clearance of CVB3 infection, but prevents immune-mediated damage to the heart.