- Email: [email protected]
Mercury Toxicity
Mercury Toxicity L Rani, B Basnet, and A Kumar, All India Institute of Medical Sciences, New Delhi, India & 2011 Elsevier B.V. All rights reserved.
Abbreviations CE CH3HgCl CH3-THF CoA CODH 5-DI DNA ERK FDA GABA GSH GSH-Px HgCl2 HPLC IL JNK MAA MAPK MIT MNA MPA PB125l PKC T3 T4 TxR WHO
capillary electrophoresis methyl mercury chloride methyl tetrahydrofolate coenzyme A carbon monoxide dehydrogenase 5-iodothyronine deiodinase deoxyribonucleic acid extracellular-regulated kinase Food and Drug Administration gamma-aminobutyric acid glutathione glutathione peroxidase mercuric chloride high-performance liquid chromatography interleukin c-jun-N terminal kinase mercaptoacetic acid mitogen-activated protein kinase monoiodotyrosine 2-mercaptonicotinic acid mercaptopropionic acid protein-bound radioiodine protein kinase C triiodothyronine thyroxine thioredoxin reductase World Health Organization
Man lives in close proximity to mercury; it is in mirrors, dry cell batteries, thermometers, sphygmomanometer, and barometers. It is present in the fluorescent light bulbs and switches in children’s shoes. It is released from leaking gas meters. It is there in tooth fillings. It is present in antiseptics and indigenous drugs. It does affect human health. Hatters used to lace hats with mercuric compounds in the ancient times. They often used to die of insanity. Remember, Lewis Carroll’s Mad Hatter in Alice in Wonderland? In the 1950s, huge amounts of mercuric chloride in the industrial waste found its way to the bays of Minamata and Niigata in Japan, killing 600 men and women. Later, in the years 1956 and 1960, the outbreaks of mercury toxicity occurred in Iraq. These outbreaks triggered investigations in the toxicity of this metal.
Forms of Mercury Mercury occurs primarily in three forms: (1) elemental or metallic mercury (Hg0) is a liquid form that releases vapor at room temperature, has relatively low solubility in water, and is primarily present in the atmosphere; (2) inorganic mercury in the form of Hgþ or Hg2þ cations as well as their compounds such as mercuric chloride, mercurous chloride, mercuric acetate, mercuric sulfide, phosphate, etc. (this form of mercury has strong affinity for sulfur-containing ligands); (3) organic compounds such as ethyl and methylmercury and phenyl mercury acetate.
Mercury in Environment Natural sources of environmental mercury are volcanoes, mines, forest fires, fossil fuels such as coal and petroleum, and volatilization from the oceanic depots. Almost 80% of the mercury in the environment comes from human activities such as fuel combustion, mining, and solid waste incineration, whereas 20% comes from agricultural use and municipal and industrial waste. A typical 100 MW thermal power plant can release over 10 kg of mercury each year. Approximately 200 metric tons of mercury escapes from industrial chimneys and effluents in a developing country such as India. Coal-fired power plants in the United States account for 40% of domestic (5% of global) human-caused mercury emission. Mercury releases from coal-fired power plants in Asia are major contributors of the global atmospheric pool (450%) and are increasingly significant source of atmospheric mercury deposition. The uncharged metallic mercury form (Hg0) is highly volatile and enters the atmosphere where it can reside for extended periods. Atmospheric Hg0 is eventually transformed to various inorganic and organic compounds. Most of the transformed mercury compounds are removed or get precipitated on the ground and water surfaces. The common mercury transformation is shown as follows: Hg 0
Oxidation
$
Reduction
Hg þ or Hg 2þ
Methylation
$
Demethylation
HgCH3 =HgCH3 Hg
The major transformation process for sedimental and aquatic mercury compounds is methylation. Over 95% of mercury biomethyltion is carried out by sulfate-reducing
705
706
Mercury Toxicity
microorganisms, mainly sulfur-reducing forms of anaerobic bacteria such as Desulfobacter, Desulfovibrio desulfuricans, Desulfobulbus propionicus, Desulfococcus multivorans, etc. Research on the biological mechanism of mercury methylation has been conducted mainly on one strain of sulfate-reducing bacteria, D. desulfuricans LS. It was originally proposed that a methylcorrinoid derivative is the methylating agent for inorganic mercury. The corrinoidcontaining protein is responsible for mercury methylation in D. desulfuricans LS and this methylation process involves acetyl-coenzyme A (CoA) pathway. The acetylCoA pathway is a carbon metabolism pathway that converts acetate into carbon dioxide (and vice versa), through the breakdown of acetate into carbon monoxide and a methyl moiety by carbon monoxide dehydrogenase (CODH), and subsequent oxidation of both to CO2. In the acetyl-CoA pathway, the methyl group may originate from serine or from formate. It has been shown that in the synthesis of methylmercury by D. desulfuricans LS, the methyl group is transferred from CH3-tetrahydrofolate (CH3-THF) to the cobalt of corrinoid protein via methyltransferase. The actual synthesis of methylmercury from CH3-THF and mercuric chloride (HgCl2) can be divided into two steps: CH3 -THFþ Corrinoid protein Methyl transferase I
-
CH3 -corrinoid protein
CH3 -þ Corrinoid protein þ Hg2þ Methyl transferase II
-
CH3 Hgþ þ Corrinoid protein
Apart from sulfur-reducing bacteria, iron-reducing bacteria in the genera Geobacter and Desulfuromonas were also found to methylate mercury. Various strains of Geobacter and Desulfuromonas had been shown to methylate mercury while reducing ferric, nitrate, or fumarate. Another biochemical pathway that has been suggested for mercury methylation is the methionine synthase pathway in Neurospora crassa ; however, N. crassa has not been considered to be an important producer of methylmercury in nature. Factors Affecting Methylation Methylation depends on the level of dissolved organic carbon in the environment. Low pH also increases the methylation of mercury and reduces ionic mercury to elemental form. Acid rain is an important cause of reduction of pH in soil and water bodies. Methylation of mercury is decreased by increasing chloride-ion concentration. Influence on methylation process in relation to various physical and chemical conditions is given in Table 1.
Table 1 Influence of different physical or chemical conditions on methylation of mercury Physical or chemical condition
Qualitative influence on methylation
Low dissolved oxygen Decreased pH
Enhanced methylation Enhanced methylation in water column Decreased methylation in sediment Enhanced methylation in sediment
Decreased pH Increased dissolved organic carbon Increased dissolved organic carbon Increased salinity Increased nutrient concentrations Increased selenium concentrations Increased temperature Increased sulfate concentrations Increased sulfide concentrations
Decreased methylation in water column Decreased methylation Enhanced methylation Decreased methylation Enhanced methylation Enhanced methylation Enhanced methylation
Source: www.weblakes.com/Mercury/mercury_methylation.html.
Speciation of Mercury Metal speciation is important in a variety of environmental, biological, geological, and medical applications. An accurate determination of each species is important to evaluate the potential risk of some metals as the chemical and physical properties of a metal species depend very much on its oxidation state. As there are three different species of mercury, the need for analytical methods able to differentiate between these mercury species is necessary. Such determinations are termed ‘speciation.’ Various methods have been introduced for the speciation of metals with different oxidation states. Highperformance liquid chromatography (HPLC) is the premier technique for metal speciation. For routine analysis, a simple, conventional detection based on UV–vis absorption is also in use. Among the separation methods, capillary electrophoresis (CE) is a relatively new and still developing technique. CE proves useful in elucidation of binding mechanisms and interaction of separated species with various biomolecules. Direct separation of mercury cations is not very much in use due to the aqueous solubility of most of the separated species. The separation method is carried out with or without organic modification of the sample. The separated species can be detected by UV–vis absorption. However, due to weak absorbance of mercury species in UV–vis region, the use of this process is limited. The most commonly used separation mode is separation of mercury species as anionic complexes after modification with a suitable complexation agent. The complexing agents generally used are L-cysteine and other mercapto compounds such
Mercury Toxicity
as 2-mercaptonicotinic acid (MNA), mercaptoacetic acid (MAA), mercaptopropionic acid (MPA), glutathione (GSH), etc. Sensitive amperometric detection can be used since the mercury species can be reduced, or hyphenation to element specific detection is also possible.
Mercury in Food and Drugs The exposure of mercury via food normally occurs when seafood containing high level of mercury or when the grain treated with fungicides, such as ethyl and methyl mercurials, is consumed. It has been found that between 80–95% of the total mercury found in fish is present as methylmercury. Methylmercury is readily absorbed by marine life, and gets accumulated and magnified as it ascends the aquatic food chain. As a result, the highest tissue concentrations are found in animals at the highest trophic levels, including long-lived species such as sharks, swordfish, and tuna. Sea mammals and shellfish carry a variable concentration of methylmercury in their muscle tissues. Methylmercury remains bound to free amino acids on the muscle tissues despite cooking or processing. As little as 0.9 g of mercury, which is 1/70th of a teaspoon, is enough to contaminate a 25 acre lake and rendering its fish unsafe. The dental amalgam used to fill in the cavities contains tin, silver, or gold in 50% of mercury. These amalgams have been found to result in galvanic currents in the metals that drive the metals into the saliva and tissues of the oral cavity. Mercury salts such as mercuric chloride, mercuric oxide, mercuric iodide, mercurous acetate, and mercurous chloride, which are used as topical disinfectants and preservatives such as thimerosal and merbromin, are another source of poisoning. Thimerosal is an organic mercurial compound that has been in use since 1930s as a preservative in over 30 USlicensed vaccines in concentrations of 0.003–0.01%. After analysis by the Food and Drug Administration (FDA), a statement was released stating that thimerosal should be reduced or eliminated from vaccines.
Absorption of Mercury The primary entry route for the elemental mercury is inhalation, whereas that for the inorganic mercury is through oral exposure. Elemental mercury can also pass through the skin. Approximately 80% of inhaled elemental mercury is absorbed by rapid diffusion through the lungs, whereas only 0.01% of elemental mercury is absorbed through the gastrointestinal tract. The absorbed mercury undergoes conversion to divalent mercury following its binding to sulfhydryl groups in the gastrointestinal tract. Therefore, the ingested elemental mercury usually passes out of the body without much harm.
707
Mercury vapors released from amalgam fillings are inhaled into the lungs and only a small portion of the compound is swallowed. The absorption of inorganic mercury via lungs is low because of the mucociliary escalator in upper respiratory tract. It clears the mercury particles trapped in the mucus by moving the latter up and out of the lungs. Solubility is the prime factor that determines the extent of transport of inorganic mercury across the intestinal tract. It determines the extent of dissociation of the compound in the lumen, and the availability of mercury for the absorption. The absorption of Hg2þ in the gastrointestinal tract is approximately 7%. In the gastrointestinal tract, methylmercury is absorbed to approximately 95%. In in vitro studies, it has been shown that the rate of absorption of mercuric compounds in the laboratory rodents increases with the increase in the intestinal pH. The rate of absorption of mercuric compounds also depends on age. Early neonatal age is a critical period for metal accumulation. At this period of life, the ability to absorb and retain the metal is high and therefore the metal toxicity is more damaging.
Distribution of Mercury in the Body Elemental mercury is lipophilic and distributed throughout the body. It readily crosses the blood–brain and placental barriers. As an uncharged monatomic gas, it is highly diffusible and lipid soluble. It is well absorbed in the lungs and easily crosses cell membranes. In red blood cells, it is oxidized to its divalent form and these divalent cations exist in both diffusible and nondiffusible forms. In the plasma, these ions exist in nondiffusible forms and bind to albumin and globulins. Except in the brain, where peak level of mercury distribution takes 23 days, mercury distributes to all other tissues and reaches peak in them within 24 h. The highest level of elemental mercury deposition following inhalation has been found to be in the kidney. Inorganic divalent mercury has poor lipid solubility. A low amount of inorganic divalent mercury crosses the blood–brain and placental barriers. Inorganic mercury is concentrated in liver and kidney more readily than elemental mercury. Methyl mercurials are lipophilic and absorbed in the gastrointestinal tract. They are distributed in all the tissues, including brain and fetus. After ingestion, the concentration of methylmercury is 13–21 times higher in erythrocytes than in plasma. Methylmercury mainly concentrates in the brain and forms methylmercury L-cysteine complex that mimics the molecular structure of methionine. In this form, the methylmercury crosses the blood–brain barrier by the L-type amino acid
708
Mercury Toxicity
transport system disguised as amino acid. However, the uptake of methylmercury by the brain is inhibited by the presence of other amino acids such as leucine, methionine, phenylalanine, and other large, neutral amino acids. Within tissues, methylmercury is slowly demethylated to Hg2þ. Mercury in hair may be increased as a result of direct absorption of mercury vapor to the hair strands. The concentration of methylmercury in hair is proportional to the concentration in blood at the time of formation of the hair strands. Once incorporated into hair strand, its concentration remains unchanged.
Metabolism and Excretion of Mercury In humans and the laboratory animals studied so far, the metabolism of all forms of inorganic mercury is similar. Once absorbed, both elemental and inorganic mercury enter an oxidation–reduction cycle. Elemental mercury is oxidized to divalent inorganic cation in the red blood cells. The mercury vapors are oxidized in red blood cells by hydrogen peroxide–catalase pathway into divalent cations. These divalents are in turn reduced to the metallic or monovalent form and released as exhaled elemental mercury vapor. Elimination and excretion of mercury occurs mainly through urine and feces. In the case of elemental mercury, fecal excretion accounting for 50% of total elimination has been found to be dominant after the first week of exposure. In long-term occupational exposures, urinary excretion becomes dominant. Owing to the rapid oxidation of the vapor, the mercury excreted in feces is probably in the form of mercuric mercury. The exhaled air, sweat, and saliva also contribute to the excretion of the metal to some extent. Inorganic mercury is also excreted in breast milk. The rate of excretion of methylmercury in human and laboratory animals is directly proportional to the simultaneous body burden. Approximately 90% of methylmercury is excreted via the fecal route. Urinary excretion of methylmercury is negligible. It accounts for r10% of the total elimination from the body. Methylmercury is also excreted in the breast milk that constitutes approximately 5% of the methylmercury in the maternal blood. It has been shown in rats that antibiotic results in a total loss of the ability to excrete mercury. The normal gut anaerobes convert methylmercury into inorganic mercury. Methylmercury is highly absorbable in the gastrointestinal tract, whereas inorganic mercury is excreted out. In contrast to anaerobes, yeast and E. coli methylate inorganic mercury to methylmercury. The usage of antibiotics could alter normal gut ecology and increase yeast and E. coli and enhance mercury toxicity.
Tests for Mercury Exposure Blood test is useful in measuring all the three forms of mercury. In the fetus, cord blood is used for testing purposes. Mercury remains in the bloodstream for only a few days; therefore, the test must be done soon after the exposure. According to Oklahoma State University, New Jersey State Department of Health, the blood level of mercury in unexposed individuals ranges from 0 to 2 mg dl1. The levels above 2.8 mg dl1 are considered to be toxic. In the blood, more than 90% of methylmercury is bound to hemoglobin in the red blood cells. Total mercury in red blood cells is also sometimes used as a proxy measure of methylmercury exposure and total mercury in plasma is used as a proxy measure of inorganic mercury exposure (Hg2þ and Hg0). The concentration of methylmercury in the brain is about five times the corresponding concentration in blood. Methylmercury also accumulates in the growing hair of scalp. Concentration of methylmercury in hair closely follows blood levels. In scalp hair, concentration of methylmercury is approximately 250 times the corresponding concentration in blood. The concentration of total mercury in hair is often used as a measure of methylmercury exposure, assuming that 480% of mercury in hair is in the form of methylmercury. Therefore, hair can also be used as biological indicators of methylmercury exposure in adult and fetal brain. In the latter case, maternal hair is used. Urine mercury test measures elemental and inorganic mercury since organic mercury is not appreciably excreted in urine. The total mercury concentration in urine is used as a measure of inorganic mercury exposure as methylmercury is excreted primarily via the bile (as GSH complex) and feces (approximately 90%) and only to a limited extent (approximately 10%) in urine. Urinary levels range from 0 to 20 mg l1 in unexposed individuals. According to WHO, the mean reference values for the total mercury in commonly used indicator media are whole blood 8 mg l1, hair 2 mg g1, urine 4 mg l1, and placenta 10 mg kg1 wet weights.
Effect of Mercury Molecular and Cellular Effects Mercury binds to sulfhydryl (SH), hydroxyl (OH), amino (NH2), and chloride (Cl) groups in proteins, enzymes, coenzymes, and cell membranes, and thereby disrupts many cellular enzymatic and coenzymatic processes. The binding of the metal with these molecules alters cell membrane charge, resulting in a change in cell permeability and cell death. Mercury ions bound to plasma proteins bind with –SH group of the water-soluble
Mercury Toxicity
component of lipoproteins on the cell membrane. As a result, there is a free exchange of these metals between the serum protein and the red blood cells. The –SH-rich hemoglobin molecule is an important site for the metal binding. The binding of mercury compounds to –SH groups of GSH blocks its function as a free radical scavenger. Thus, free radicals become available to cause DNA damage. These mechanisms can lead to ‘double-strand breaks’ that can cause chromatid gaps or chromosome rearrangements. It has been shown in human lymphocyte cultures that exposure of methylmercury chloride (CH3HgCl) alone or in combination with mercury chloride (HgCl2) resulted in a significant increase in the relative frequency of chromosome aberrations. The frequency of polyploid cells was also significantly increased. CH3HgCl exposure resulted in a significant decrease in the mitotic index showing a cytotoxic effect. The decrease in mitotic index was followed by an increase in the generation of reactive oxygen species. The strong affinity of mercury for –SH groups available in the spindle impairs its functions, leading to mistakes in chromosome segregation during cell division and consequently to polyploidy or aneuploidy. It has been reported that CH3HgCl induced translocation of cytochrome c from mitochondria to the cytosol of T cells. It also activates caspase cascade, leading to apoptosis of T cells. It has been reported that mercury has a strong inhibitory effect on the activity of dipeptidyl peptidase, the enzyme involved in the digestion of the milk protein casein and also xanthine oxidase, which catalyzes the oxidation of hypoxanthine to xanthine and can further catalyze the oxidation of xanthine to uric acid. It has been demonstrated that mercury inhibits the activities of alkaline phosphatase, lipase, aminotripeptidase, and glycylglycine dipeptidase in the liver and digestive tract of the fish, Channa punctatus. Addition of a chelating agent such as EDTA restored the mercury-inhibited enzyme activity. Mercury has a great chemical propensity toward thiols and selenols. Methylmercury binding to the thiol group is a rapid and reversible process. Moreover, because of its free-moving nature from one thiol protein group to the other, it can damage proteins present in the cellular machinery. The alteration in intracellular thiol can depolarize the inner mitochondrial membrane along with an increase in the formation of hydrogen peroxide. These events along with depletion of GSH lead to oxidative stress characterized by an increase in the susceptibility of the mitochondrial membrane to irondependent lipid peroxidation. Mercury compounds induce a general collapse of antioxidant mechanisms in the cell by binding to the sulfhydryl groups of GSH, a radical scavenger. Such a collapse results in cell degeneration, loss of membrane integrity, and finally cell necrosis.
709
High level of mercury has been found to inhibit the peptide-elongation step of the protein synthesis. Methylmercury also interferes with lipids, myelin, and mitochondrial DNA synthesis. Both organic and inorganic mercurial compounds inhibit the mammalian thioredoxin system, particularly thioredoxin reductase (TxR), which is responsible for cellular stress response, protein repair, and protection against oxidative damage. The thioredoxin system is an electron donor for peroxiredoxins involved in eliminating peroxides and its inhibition by mercury increases reactive oxygen species, which, in turn, causes depletion of GSH. Organic mercury has a higher bioavailability than its inorganic form. It has also been reported that organic mercury is more potent in inhibiting the growth of HeLa and HEK 293 cells in culture than the inorganic compounds. Inorganic ions get chelated and thereby restrained from action. Systemic Effects Mercury is a systemic poison. Depending on the dose and length of exposure period, it can affect various organ systems and functions that are discussed in the following text. Immune system
Mercury inhibits T-cell proliferation in the presence of monocytes. It inhibits the ability of these cells to synthesize and secrete interleukin-1 (IL-1). Mercury also impairs T-cell expression of IL-2 receptor and the transferrin receptor, thereby decreasing the T-cell function. In both in vitro and in vivo studies, lead and mercury have been found to enhance the production of IL-4 by the Th2 clone and inhibit the proliferation of Th1. This imbalance between the Th1 and Th2 results in increased production of antibodies to self-antigen and thereby generation of autoimmunity. Heavy metals cause aberrant expression of major histocompatibility complex II (MHC II) molecules on the target cells, inhibit the T-suppressor cells, alter the idiotype–anti-idiotype network, and induce heat-shock proteins. These factors may play a very important role in metal-induced autoimmunity such as multiple sclerosis, rheumatoid arthritis, and amyotrophic lateral sclerosis. Heavy metals, as discussed earlier, bind to SH and other groups and thereby modify other proteins. These modified proteins might activate B cell via the T cell and cause alteration of self-protein target for autoantibodies. The metals can bind to the MHC II without prior processing by the antigen-presenting cells or directly to the T-cell receptors. All these mechanisms have been proposed to be involved in metal-induced autoimmunity. In genetically susceptible mice and rats, the immunological effects such as hyperimmunoglobulinemia, development
710
Mercury Toxicity
of antinucleolar antibodies and antifibrillar antibodies, potentiation of mitogen-induced lymphocyte proliferation, and increased expression of class II molecules are caused by dental amalgam. Endocrine system
Mercury disrupts the function of the pituitary, thyroid, and adrenal glands even at low-level exposure. It has been reported that levels of mercury in the thyroid and pituitary glands were higher than those observed in kidney, brain, and liver tissue in people working in mercury mines in Solvenia. It reduces the level of posterior pituitary hormone, oxytocin. The bouts of depression and suicidal thoughts are mainly associated with the reduction of oxytocin. Mercury amalgam in dental fillings is a contributing factor in reducing pituitary functions. Administration of oxytocin alleviates many of these mood problems. Intramuscular administration of mercury chloride has been shown to induce hyperthyroidism in rabbits within 24 h of its administration. Mercury increased thyroid peroxidase activity, and induced a rise in the triiodothyronine (T3) and a remarkable fall in the thyroxine (T4) level. The T3/T4 ratio became higher than the controls. T3-toxicosis may be produced by a preferential synthesis of T3 or preferential deiodination of T4 to T3. On the other hand, it has also been reported that mercury causes hypothyroidism. Mercury has also been shown to cause autoimmune thyroiditis. The thyroid imbalance caused by exposure to mercury could play a major role in chronic heart conditions such as clogged arteries, myocardial infarction, and chronic heart failure. In an experiment, administration of 2.5 mg of mercuric chloride daily for 40 days to rats enhanced the thyroidal weight, thyroidal 125I uptake, and serum protein-bound radio-iodine (PB125I). Further investigation with the chromatographic analysis of thyroid pronase hydrolysates of the mercury-treated rats displayed an increase in percentage of labeled monoiodotyrosine (MIT) without any alteration in the percentage of labeled T3, indicating a coupling defect in the synthesis of T3. Prenatal exposure to methylmercury severely inhibits the activity of 5-iodothyronine deiodinases (5-DI) in the fetal brain, and increases the activity of 50 -DI. This manifests as hypothyroidism. Mercury also accumulates in the lipid-rich cortex of the adrenal gland and disrupts the function of the gland by blocking various enzymatic pathways. Heavy metals have been found to cause hyperandrogenemia or partial hypoadrenalism and polycystic ovarian syndrome in women. Respiratory system
A survey conducted on the workers exposed to mercury vapors accidentally at an estimated concentration of
approximately 44.3 mg m3 for 4–8 h showed that they exhibited chest pains, dyspnea, cough, hemoptysis, reduction in vital capacity leading to failure of pulmonary functions, and also interstitial pneumonitis. Inorganic mercuric compounds also contribute toward respiratory problems such as pulmonary edema and shortness of breathing.
Nervous system
A variety of disturbances primarily in the sensory and motor nerves have been reported. The cardinal neurological sign of mercury vapor exposure is tremor. Other neuropsychological signs and symptoms are emotional lability, insomnia, memory loss, weakness, muscle atrophy, muscle twitching, headaches, paresthesias, and polyneuropathy. Long-term exposure is associated with unsteady walk, poor concentration, tremulous speech, blurred vision, impaired hearing, reduced finger to eye coordination, etc. One of the major mechanisms of the toxicity of methylmercury is the inhibition of tubulin polymerization. Microtubular fragmentation has been found in cultured rat cerebellar granular neurons treated with 0.5–1 mM of methylmercury. Fragmentation of microtubule by methylmercury results in degeneration of neurons and inhibition of neuronal migration. Methylmercury depolarizes the presynaptic membrane that increases the Na2þ ion concentration and decreases Kþ ion concentration, causing disruption of Ca2þ homeostasis leading to increased intracellular Ca2þ concentration. Increased calcium damages the cell membrane and disrupts neurotransmitter signaling. Methylmercury stimulates the release of neurotransmitters such as dopamine, glutamate, gamma-aminobutyric acid (GABA), glycine, choline, and acetylcholine. It inhibits the uptake of excitatory amino acids such as glutamate, and aspartate in the astrocytes. Mercury has been shown to accumulate in the lysosomes of the efferent neurons of spinal cord by a retrograde axonal transport. Methylmercury exposure is associated with increased mitochondrial membrane permeability in cultured astrocytes. This leads to a collapse of the mitochondrial inner membrane potential (Cm). Loss of the Cm results in colloid osmotic swelling of the mitochondrial matrix, movement of metabolites across the inner membrane, defective oxidative phosphorylation, cessation of ATP synthesis, generation of reactive oxygen species and consequent oxidative injury. Methylmercury has been shown to induce generation of reactive oxygen species in synaptosomes by activating protein kinase C (PKC), and mitogen-activated protein kinase (MAPK) pathway kinases. The generated reactive oxygen species cause the downstream activation of a variety of kinases (p38MAPK, ERK, and JNK), leading to a cytotoxic response.
Mercury Toxicity
Cardiovascular system
Mercury in both organic and ionic forms accumulates in the heart and causes hypertension, tachycardia, and ventricular arrhythmias. Increased blood pressure and palpitation is observed on the inhalation of elemental mercury vapor. As the duration of mercury exposure increases, there is also an increase in heart rate. It has been reported that children treated with mercurous chloride tablets for worms or with ammoniated mercury ointment for diaper rash suffered from tachycardia and elevated blood pressure. Gastrointestinal system
Exposure to mercury results in the inflammation of oral mucosa, excessive salivation, and difficulty in swallowing. It also leads to diarrhea, sore gums, ulceration of the lips and tongue, and vomiting. Renal system
The target organ for both ionic elemental or mercuric mercury is the kidney where it has a half-life of 2 months due to its biotransformation to ionic forms. It accumulates in the kidney. The binding of mercury to glomerular basement proteins causes severe damage to the kidney and results in proteinuria, hematuria, and oliguria. An increase in tubular antigens and enzymes has been reported in workers with chronic low-level exposure to elemental mercury vapor. It has been reported that 62 out of 86 cases of ethyl mercury poisoning from Iraqi grain seed exhibited clinical symptoms of kidney damage. It has been reported that when mice were fed on a diet containing methyl mercury for approximately 1 year, they developed renal carcinoma. Reproductive system Female
Maternal exposure to mercury before and during pregnancy may lead to spontaneous abortion, preterm birth, and low birth weight. It has been observed that abnormalities in menstrual cycle and miscarriage are common among the women exposed to mercury vapor. Female dental assistants exposed to elemental mercury have a lower fertility. The mercury vapor from toothfilling amalgam may interfere with corpus luteum function. Mercury inhibited the production of progesterone by the human granulosa cells in vitro. An early rise of estradiol level in the follicular phase in women showing high levels of mercury has been observed. This too could cause luteal phase defect. Injection of mercuric chloride into the female hamster resulted in prolongation of estrous cycle and morphological changes in the corpora lutea along with the inhibition of follicular maturation and elevation of pituitary FSH. The injection of mercuric chloride in female mice before mating resulted in dose-related
711
increase in preimplantation and early postimplantation fetal loss. Male
In males, mercury mainly affects sperm production, motility, and morphology. Mercury decreases sperm motility, and induces a higher percentage of abnormal sperm tail forms marked by bent, kinked, and coiled tails in Macaca fasicularis. However, no changes in serum testosterone were observed. Methylmercury has been suggested to inhibit mitotic divisions in the germ cells to decrease the sperm production and also inhibit microtubule assembly during spermiogenesis. In men as well, methylmercury decreases sperm count, increases abnormal morphology and impotency, and decreases libido. Antifertility effects of mercury in mice can be attributed to the direct inhibition of the DNA synthesis in the spermatogonial cells since the mercury-bound DNA can affect the unwinding and rewinding of DNA essential for replication and transcription. Intercalation of DNA in turn affects the RNA transcription and protein synthesis. Mercury also has an inhibitory effect on essential spermatogenic enzymes. Fetal and child health
Mercury vapor can pass through the feto-placenta membrane and reach the brain of the fetus and endocrine glands. The blood level of mercury in fetus reaches a level higher than the level in maternal circulation. It has also been reported that mercuric ions accumulate in the placenta and thereby limit the exchange of nutrients and gases to the fetus. At peak maternal hair mercury levels above 70 mg g1, there is a high risk, more than 30%, of neurological disorder in the offspring. It has been reported that even at a low dose, mercuric ions may affect embryonic development and cause fetal malformation. The administration of large doses of mercury to pregnant rodents resulted in cleft palate. Prenatal exposure of rats produced abnormalities in renal function that were detected in offspring at 42 days of age. Thimerosol used as preservative in vaccines and anti-RhoD-immunoglobulins administered to Rh-negative pregnant women and methylmercury in seafood could poison the fetus. One out of six women in the United States has mercury levels high enough to increase the risk of neurotoxicity to fetus. The report of ingestion of flour from grain treated with alkyl mercury by pregnant women in Sweden showed normal birth of the offspring but later there were signs of brain damage manifested as mental retardation and inability to move. In a case study, a 3-month pregnant woman consumed methylmercurycontaminated meat, and delivered a male infant. The infant showed elevated urinary level of mercury. At 3 months of age, the infant was hypotonic and irritable, and exhibited myoclonic seizures, and at 6 years of age the
712
Mercury Toxicity
child displayed severe neurological impairment such as blindness, neuromuscular weakness, and inability to speak. Children with regressive autism, chronic constipation, and diarrhea have 10 000 times the normal level of E. coli in their stool. They have a higher intake of antibodies in the first 36 months of their life and therefore a higher possibility of further change in gut ecology. These children have a higher level of mercury in their infantile teeth in comparison with normal controls. Their reduced ability to excrete mercury may further exacerbate their neurological deficit. Children exposed to mercury vapors show acrodynia, which is a hypersensitivity reaction to mercury and expressed by irritability, mood changes, erythema of the hands, feet, and sometimes nose and other parts of the body, leg pains or cramps, peeling of the palms of the hands and soles of the feet, and insomnia.
Several thiol-containing complexing agents have also been successfully used to remove methylmercury from the body. The most promising complexing agent is N-acetylcysteine, which when given by mouth, traps the methylmercury secreted in bile and carries it into the feces. It enhances methylmercury excretion when given orally, has a low toxicity, and is widely available in the clinical setting.
Conclusion Mercury toxicity is a major health risk particularly to the developing economies that depend on thermal power generation. The risk to these countries is also heightened by the disposal of mercury waste in their shores and soil by the developed countries. The world needs to take cognizance of this toxin and develop regulatory measures and health policies.
Modification of Mercury Toxicity Selenium (Se) has been known to bind toxic metals. Owing to this property, it plays a role in potentially reducing their toxicity. It has been shown that accumulation or retention of selenium is increased in response to accumulation of mercury in tissues of marine mammals or mineworkers following exposure or ingestion. When inorganic mercury and sodium selenite are administered together, sodium selenite forms a high-molecular-weight complex with mercury and plasma protein, thereby reducing the accumulation of mercury in kidney; it has also been shown to form an inert high-molecular-weight complex, which stays in the red blood cells without doing any harm to other tissues. The plasma protein to which Hg–Se complex binds has been identified as plasma selenoprotein (selenoprotein P). Various studies demonstrated the binding of complexes of mercury–selenium, silver–selenium, and cadmium–selenium by plasma selenoprotein P, suggesting that this protein may function to chelate heavy metals, reducing their toxicity. Mercury exposure leads to significant increase in the expression of both selenoprotein P and glutathione peroxidase (GSHPx) in mercury miners. These increases were accompanied by elevated selenium concentrations in serum. Selenoproteins play two important roles in protecting against mercury toxicity. First, they may bind more mercury through their highly reactive selenol group, and second, their antioxidative properties help eliminate the reactive oxygen species induced by mercury in vivo.
Further Reading Ballatori N (2002) Transport of toxic metals by molecular mimicry. Environmental Health Perspectives 110: 689--694. Berry MJ and Ralston NVC (2009) Mercury toxicity and the mitigating role of selenium. Ecohealth 5: 456--459. Carvalho CML, Chew EH, Hashemy SI, Lu J, and Holmgren A (2008) Inhibition of the human thioredoxin system. A molecular mechanism of mercury toxicity. Journal of Biological Chemistry 283: 11913--11923. Clarkson TW (1995) Environmental contaminants in the food chain. American Journal of Clinical Nutrition 61: 682s--686s. Clarkson TW (2002) The three modern faces of mercury. Environmental Health Perspectives 110(supplement 1): 11--23. Clarkson TW and Magos L (2006) The toxicology of mercury and its chemical compounds. Critical Reviews in Toxicology 36: 609--662. Davidson PW, Myers GJ, and Weiss B (2004) Mercury exposure and child development outcomes. Paediatrics 113: 1023--1029. De Flora S, Benniceli C, and Bagnasco M (1994) Genotoxicity of mercury compounds. A review. Mutation Research 317: 57--79. Kuban P, Pelcova P, Margetınova J, and Kuban V (2009) Mercury speciation by CE: An update. Electrophoresis 30: 92--99. Magos L and Clarkson TW (2006) Overview of the clinical toxicity of mercury. Annals of Clinical Biochemistry 43: 257--268. Oklahoma State University, New Jersey State Department of Health, Division of Occupational and Environmental Health. The Health Effects of Mercury, Environmental Health and Safety. www.ehs. okstate.edu/training/mercury.htm (accessed January 2010). Risher JF (2003) Elemental Mercury and Inorganic Mercury Compounds: Human Health Aspects Agency for Toxic Substances and Disease Registry (ATSDR). Atlanta, GA: WHO. (Concise International Chemical Assessment Document 50). Risher JF, Murray HE, and Prince GR (2002) Organic mercury compounds: Human exposure and its relevance to public health. Toxicology and Industrial Health 18: 109--160. Who (1990) Environment Health Criteria: Methyl Mercury. Geneva: World Health Organization. pp. 1–144.
Abbreviations CE CH3HgCl CH3-THF CoA CODH 5-DI DNA ERK FDA GABA GSH GSH-Px HgCl2 HPLC IL JNK MAA MAPK MIT MNA MPA PB125l PKC T3 T4 TxR WHO
capillary electrophoresis methyl mercury chloride methyl tetrahydrofolate coenzyme A carbon monoxide dehydrogenase 5-iodothyronine deiodinase deoxyribonucleic acid extracellular-regulated kinase Food and Drug Administration gamma-aminobutyric acid glutathione glutathione peroxidase mercuric chloride high-performance liquid chromatography interleukin c-jun-N terminal kinase mercaptoacetic acid mitogen-activated protein kinase monoiodotyrosine 2-mercaptonicotinic acid mercaptopropionic acid protein-bound radioiodine protein kinase C triiodothyronine thyroxine thioredoxin reductase World Health Organization
Man lives in close proximity to mercury; it is in mirrors, dry cell batteries, thermometers, sphygmomanometer, and barometers. It is present in the fluorescent light bulbs and switches in children’s shoes. It is released from leaking gas meters. It is there in tooth fillings. It is present in antiseptics and indigenous drugs. It does affect human health. Hatters used to lace hats with mercuric compounds in the ancient times. They often used to die of insanity. Remember, Lewis Carroll’s Mad Hatter in Alice in Wonderland? In the 1950s, huge amounts of mercuric chloride in the industrial waste found its way to the bays of Minamata and Niigata in Japan, killing 600 men and women. Later, in the years 1956 and 1960, the outbreaks of mercury toxicity occurred in Iraq. These outbreaks triggered investigations in the toxicity of this metal.
Forms of Mercury Mercury occurs primarily in three forms: (1) elemental or metallic mercury (Hg0) is a liquid form that releases vapor at room temperature, has relatively low solubility in water, and is primarily present in the atmosphere; (2) inorganic mercury in the form of Hgþ or Hg2þ cations as well as their compounds such as mercuric chloride, mercurous chloride, mercuric acetate, mercuric sulfide, phosphate, etc. (this form of mercury has strong affinity for sulfur-containing ligands); (3) organic compounds such as ethyl and methylmercury and phenyl mercury acetate.
Mercury in Environment Natural sources of environmental mercury are volcanoes, mines, forest fires, fossil fuels such as coal and petroleum, and volatilization from the oceanic depots. Almost 80% of the mercury in the environment comes from human activities such as fuel combustion, mining, and solid waste incineration, whereas 20% comes from agricultural use and municipal and industrial waste. A typical 100 MW thermal power plant can release over 10 kg of mercury each year. Approximately 200 metric tons of mercury escapes from industrial chimneys and effluents in a developing country such as India. Coal-fired power plants in the United States account for 40% of domestic (5% of global) human-caused mercury emission. Mercury releases from coal-fired power plants in Asia are major contributors of the global atmospheric pool (450%) and are increasingly significant source of atmospheric mercury deposition. The uncharged metallic mercury form (Hg0) is highly volatile and enters the atmosphere where it can reside for extended periods. Atmospheric Hg0 is eventually transformed to various inorganic and organic compounds. Most of the transformed mercury compounds are removed or get precipitated on the ground and water surfaces. The common mercury transformation is shown as follows: Hg 0
Oxidation
$
Reduction
Hg þ or Hg 2þ
Methylation
$
Demethylation
HgCH3 =HgCH3 Hg
The major transformation process for sedimental and aquatic mercury compounds is methylation. Over 95% of mercury biomethyltion is carried out by sulfate-reducing
705
706
Mercury Toxicity
microorganisms, mainly sulfur-reducing forms of anaerobic bacteria such as Desulfobacter, Desulfovibrio desulfuricans, Desulfobulbus propionicus, Desulfococcus multivorans, etc. Research on the biological mechanism of mercury methylation has been conducted mainly on one strain of sulfate-reducing bacteria, D. desulfuricans LS. It was originally proposed that a methylcorrinoid derivative is the methylating agent for inorganic mercury. The corrinoidcontaining protein is responsible for mercury methylation in D. desulfuricans LS and this methylation process involves acetyl-coenzyme A (CoA) pathway. The acetylCoA pathway is a carbon metabolism pathway that converts acetate into carbon dioxide (and vice versa), through the breakdown of acetate into carbon monoxide and a methyl moiety by carbon monoxide dehydrogenase (CODH), and subsequent oxidation of both to CO2. In the acetyl-CoA pathway, the methyl group may originate from serine or from formate. It has been shown that in the synthesis of methylmercury by D. desulfuricans LS, the methyl group is transferred from CH3-tetrahydrofolate (CH3-THF) to the cobalt of corrinoid protein via methyltransferase. The actual synthesis of methylmercury from CH3-THF and mercuric chloride (HgCl2) can be divided into two steps: CH3 -THFþ Corrinoid protein Methyl transferase I
-
CH3 -corrinoid protein
CH3 -þ Corrinoid protein þ Hg2þ Methyl transferase II
-
CH3 Hgþ þ Corrinoid protein
Apart from sulfur-reducing bacteria, iron-reducing bacteria in the genera Geobacter and Desulfuromonas were also found to methylate mercury. Various strains of Geobacter and Desulfuromonas had been shown to methylate mercury while reducing ferric, nitrate, or fumarate. Another biochemical pathway that has been suggested for mercury methylation is the methionine synthase pathway in Neurospora crassa ; however, N. crassa has not been considered to be an important producer of methylmercury in nature. Factors Affecting Methylation Methylation depends on the level of dissolved organic carbon in the environment. Low pH also increases the methylation of mercury and reduces ionic mercury to elemental form. Acid rain is an important cause of reduction of pH in soil and water bodies. Methylation of mercury is decreased by increasing chloride-ion concentration. Influence on methylation process in relation to various physical and chemical conditions is given in Table 1.
Table 1 Influence of different physical or chemical conditions on methylation of mercury Physical or chemical condition
Qualitative influence on methylation
Low dissolved oxygen Decreased pH
Enhanced methylation Enhanced methylation in water column Decreased methylation in sediment Enhanced methylation in sediment
Decreased pH Increased dissolved organic carbon Increased dissolved organic carbon Increased salinity Increased nutrient concentrations Increased selenium concentrations Increased temperature Increased sulfate concentrations Increased sulfide concentrations
Decreased methylation in water column Decreased methylation Enhanced methylation Decreased methylation Enhanced methylation Enhanced methylation Enhanced methylation
Source: www.weblakes.com/Mercury/mercury_methylation.html.
Speciation of Mercury Metal speciation is important in a variety of environmental, biological, geological, and medical applications. An accurate determination of each species is important to evaluate the potential risk of some metals as the chemical and physical properties of a metal species depend very much on its oxidation state. As there are three different species of mercury, the need for analytical methods able to differentiate between these mercury species is necessary. Such determinations are termed ‘speciation.’ Various methods have been introduced for the speciation of metals with different oxidation states. Highperformance liquid chromatography (HPLC) is the premier technique for metal speciation. For routine analysis, a simple, conventional detection based on UV–vis absorption is also in use. Among the separation methods, capillary electrophoresis (CE) is a relatively new and still developing technique. CE proves useful in elucidation of binding mechanisms and interaction of separated species with various biomolecules. Direct separation of mercury cations is not very much in use due to the aqueous solubility of most of the separated species. The separation method is carried out with or without organic modification of the sample. The separated species can be detected by UV–vis absorption. However, due to weak absorbance of mercury species in UV–vis region, the use of this process is limited. The most commonly used separation mode is separation of mercury species as anionic complexes after modification with a suitable complexation agent. The complexing agents generally used are L-cysteine and other mercapto compounds such
Mercury Toxicity
as 2-mercaptonicotinic acid (MNA), mercaptoacetic acid (MAA), mercaptopropionic acid (MPA), glutathione (GSH), etc. Sensitive amperometric detection can be used since the mercury species can be reduced, or hyphenation to element specific detection is also possible.
Mercury in Food and Drugs The exposure of mercury via food normally occurs when seafood containing high level of mercury or when the grain treated with fungicides, such as ethyl and methyl mercurials, is consumed. It has been found that between 80–95% of the total mercury found in fish is present as methylmercury. Methylmercury is readily absorbed by marine life, and gets accumulated and magnified as it ascends the aquatic food chain. As a result, the highest tissue concentrations are found in animals at the highest trophic levels, including long-lived species such as sharks, swordfish, and tuna. Sea mammals and shellfish carry a variable concentration of methylmercury in their muscle tissues. Methylmercury remains bound to free amino acids on the muscle tissues despite cooking or processing. As little as 0.9 g of mercury, which is 1/70th of a teaspoon, is enough to contaminate a 25 acre lake and rendering its fish unsafe. The dental amalgam used to fill in the cavities contains tin, silver, or gold in 50% of mercury. These amalgams have been found to result in galvanic currents in the metals that drive the metals into the saliva and tissues of the oral cavity. Mercury salts such as mercuric chloride, mercuric oxide, mercuric iodide, mercurous acetate, and mercurous chloride, which are used as topical disinfectants and preservatives such as thimerosal and merbromin, are another source of poisoning. Thimerosal is an organic mercurial compound that has been in use since 1930s as a preservative in over 30 USlicensed vaccines in concentrations of 0.003–0.01%. After analysis by the Food and Drug Administration (FDA), a statement was released stating that thimerosal should be reduced or eliminated from vaccines.
Absorption of Mercury The primary entry route for the elemental mercury is inhalation, whereas that for the inorganic mercury is through oral exposure. Elemental mercury can also pass through the skin. Approximately 80% of inhaled elemental mercury is absorbed by rapid diffusion through the lungs, whereas only 0.01% of elemental mercury is absorbed through the gastrointestinal tract. The absorbed mercury undergoes conversion to divalent mercury following its binding to sulfhydryl groups in the gastrointestinal tract. Therefore, the ingested elemental mercury usually passes out of the body without much harm.
707
Mercury vapors released from amalgam fillings are inhaled into the lungs and only a small portion of the compound is swallowed. The absorption of inorganic mercury via lungs is low because of the mucociliary escalator in upper respiratory tract. It clears the mercury particles trapped in the mucus by moving the latter up and out of the lungs. Solubility is the prime factor that determines the extent of transport of inorganic mercury across the intestinal tract. It determines the extent of dissociation of the compound in the lumen, and the availability of mercury for the absorption. The absorption of Hg2þ in the gastrointestinal tract is approximately 7%. In the gastrointestinal tract, methylmercury is absorbed to approximately 95%. In in vitro studies, it has been shown that the rate of absorption of mercuric compounds in the laboratory rodents increases with the increase in the intestinal pH. The rate of absorption of mercuric compounds also depends on age. Early neonatal age is a critical period for metal accumulation. At this period of life, the ability to absorb and retain the metal is high and therefore the metal toxicity is more damaging.
Distribution of Mercury in the Body Elemental mercury is lipophilic and distributed throughout the body. It readily crosses the blood–brain and placental barriers. As an uncharged monatomic gas, it is highly diffusible and lipid soluble. It is well absorbed in the lungs and easily crosses cell membranes. In red blood cells, it is oxidized to its divalent form and these divalent cations exist in both diffusible and nondiffusible forms. In the plasma, these ions exist in nondiffusible forms and bind to albumin and globulins. Except in the brain, where peak level of mercury distribution takes 23 days, mercury distributes to all other tissues and reaches peak in them within 24 h. The highest level of elemental mercury deposition following inhalation has been found to be in the kidney. Inorganic divalent mercury has poor lipid solubility. A low amount of inorganic divalent mercury crosses the blood–brain and placental barriers. Inorganic mercury is concentrated in liver and kidney more readily than elemental mercury. Methyl mercurials are lipophilic and absorbed in the gastrointestinal tract. They are distributed in all the tissues, including brain and fetus. After ingestion, the concentration of methylmercury is 13–21 times higher in erythrocytes than in plasma. Methylmercury mainly concentrates in the brain and forms methylmercury L-cysteine complex that mimics the molecular structure of methionine. In this form, the methylmercury crosses the blood–brain barrier by the L-type amino acid
708
Mercury Toxicity
transport system disguised as amino acid. However, the uptake of methylmercury by the brain is inhibited by the presence of other amino acids such as leucine, methionine, phenylalanine, and other large, neutral amino acids. Within tissues, methylmercury is slowly demethylated to Hg2þ. Mercury in hair may be increased as a result of direct absorption of mercury vapor to the hair strands. The concentration of methylmercury in hair is proportional to the concentration in blood at the time of formation of the hair strands. Once incorporated into hair strand, its concentration remains unchanged.
Metabolism and Excretion of Mercury In humans and the laboratory animals studied so far, the metabolism of all forms of inorganic mercury is similar. Once absorbed, both elemental and inorganic mercury enter an oxidation–reduction cycle. Elemental mercury is oxidized to divalent inorganic cation in the red blood cells. The mercury vapors are oxidized in red blood cells by hydrogen peroxide–catalase pathway into divalent cations. These divalents are in turn reduced to the metallic or monovalent form and released as exhaled elemental mercury vapor. Elimination and excretion of mercury occurs mainly through urine and feces. In the case of elemental mercury, fecal excretion accounting for 50% of total elimination has been found to be dominant after the first week of exposure. In long-term occupational exposures, urinary excretion becomes dominant. Owing to the rapid oxidation of the vapor, the mercury excreted in feces is probably in the form of mercuric mercury. The exhaled air, sweat, and saliva also contribute to the excretion of the metal to some extent. Inorganic mercury is also excreted in breast milk. The rate of excretion of methylmercury in human and laboratory animals is directly proportional to the simultaneous body burden. Approximately 90% of methylmercury is excreted via the fecal route. Urinary excretion of methylmercury is negligible. It accounts for r10% of the total elimination from the body. Methylmercury is also excreted in the breast milk that constitutes approximately 5% of the methylmercury in the maternal blood. It has been shown in rats that antibiotic results in a total loss of the ability to excrete mercury. The normal gut anaerobes convert methylmercury into inorganic mercury. Methylmercury is highly absorbable in the gastrointestinal tract, whereas inorganic mercury is excreted out. In contrast to anaerobes, yeast and E. coli methylate inorganic mercury to methylmercury. The usage of antibiotics could alter normal gut ecology and increase yeast and E. coli and enhance mercury toxicity.
Tests for Mercury Exposure Blood test is useful in measuring all the three forms of mercury. In the fetus, cord blood is used for testing purposes. Mercury remains in the bloodstream for only a few days; therefore, the test must be done soon after the exposure. According to Oklahoma State University, New Jersey State Department of Health, the blood level of mercury in unexposed individuals ranges from 0 to 2 mg dl1. The levels above 2.8 mg dl1 are considered to be toxic. In the blood, more than 90% of methylmercury is bound to hemoglobin in the red blood cells. Total mercury in red blood cells is also sometimes used as a proxy measure of methylmercury exposure and total mercury in plasma is used as a proxy measure of inorganic mercury exposure (Hg2þ and Hg0). The concentration of methylmercury in the brain is about five times the corresponding concentration in blood. Methylmercury also accumulates in the growing hair of scalp. Concentration of methylmercury in hair closely follows blood levels. In scalp hair, concentration of methylmercury is approximately 250 times the corresponding concentration in blood. The concentration of total mercury in hair is often used as a measure of methylmercury exposure, assuming that 480% of mercury in hair is in the form of methylmercury. Therefore, hair can also be used as biological indicators of methylmercury exposure in adult and fetal brain. In the latter case, maternal hair is used. Urine mercury test measures elemental and inorganic mercury since organic mercury is not appreciably excreted in urine. The total mercury concentration in urine is used as a measure of inorganic mercury exposure as methylmercury is excreted primarily via the bile (as GSH complex) and feces (approximately 90%) and only to a limited extent (approximately 10%) in urine. Urinary levels range from 0 to 20 mg l1 in unexposed individuals. According to WHO, the mean reference values for the total mercury in commonly used indicator media are whole blood 8 mg l1, hair 2 mg g1, urine 4 mg l1, and placenta 10 mg kg1 wet weights.
Effect of Mercury Molecular and Cellular Effects Mercury binds to sulfhydryl (SH), hydroxyl (OH), amino (NH2), and chloride (Cl) groups in proteins, enzymes, coenzymes, and cell membranes, and thereby disrupts many cellular enzymatic and coenzymatic processes. The binding of the metal with these molecules alters cell membrane charge, resulting in a change in cell permeability and cell death. Mercury ions bound to plasma proteins bind with –SH group of the water-soluble
Mercury Toxicity
component of lipoproteins on the cell membrane. As a result, there is a free exchange of these metals between the serum protein and the red blood cells. The –SH-rich hemoglobin molecule is an important site for the metal binding. The binding of mercury compounds to –SH groups of GSH blocks its function as a free radical scavenger. Thus, free radicals become available to cause DNA damage. These mechanisms can lead to ‘double-strand breaks’ that can cause chromatid gaps or chromosome rearrangements. It has been shown in human lymphocyte cultures that exposure of methylmercury chloride (CH3HgCl) alone or in combination with mercury chloride (HgCl2) resulted in a significant increase in the relative frequency of chromosome aberrations. The frequency of polyploid cells was also significantly increased. CH3HgCl exposure resulted in a significant decrease in the mitotic index showing a cytotoxic effect. The decrease in mitotic index was followed by an increase in the generation of reactive oxygen species. The strong affinity of mercury for –SH groups available in the spindle impairs its functions, leading to mistakes in chromosome segregation during cell division and consequently to polyploidy or aneuploidy. It has been reported that CH3HgCl induced translocation of cytochrome c from mitochondria to the cytosol of T cells. It also activates caspase cascade, leading to apoptosis of T cells. It has been reported that mercury has a strong inhibitory effect on the activity of dipeptidyl peptidase, the enzyme involved in the digestion of the milk protein casein and also xanthine oxidase, which catalyzes the oxidation of hypoxanthine to xanthine and can further catalyze the oxidation of xanthine to uric acid. It has been demonstrated that mercury inhibits the activities of alkaline phosphatase, lipase, aminotripeptidase, and glycylglycine dipeptidase in the liver and digestive tract of the fish, Channa punctatus. Addition of a chelating agent such as EDTA restored the mercury-inhibited enzyme activity. Mercury has a great chemical propensity toward thiols and selenols. Methylmercury binding to the thiol group is a rapid and reversible process. Moreover, because of its free-moving nature from one thiol protein group to the other, it can damage proteins present in the cellular machinery. The alteration in intracellular thiol can depolarize the inner mitochondrial membrane along with an increase in the formation of hydrogen peroxide. These events along with depletion of GSH lead to oxidative stress characterized by an increase in the susceptibility of the mitochondrial membrane to irondependent lipid peroxidation. Mercury compounds induce a general collapse of antioxidant mechanisms in the cell by binding to the sulfhydryl groups of GSH, a radical scavenger. Such a collapse results in cell degeneration, loss of membrane integrity, and finally cell necrosis.
709
High level of mercury has been found to inhibit the peptide-elongation step of the protein synthesis. Methylmercury also interferes with lipids, myelin, and mitochondrial DNA synthesis. Both organic and inorganic mercurial compounds inhibit the mammalian thioredoxin system, particularly thioredoxin reductase (TxR), which is responsible for cellular stress response, protein repair, and protection against oxidative damage. The thioredoxin system is an electron donor for peroxiredoxins involved in eliminating peroxides and its inhibition by mercury increases reactive oxygen species, which, in turn, causes depletion of GSH. Organic mercury has a higher bioavailability than its inorganic form. It has also been reported that organic mercury is more potent in inhibiting the growth of HeLa and HEK 293 cells in culture than the inorganic compounds. Inorganic ions get chelated and thereby restrained from action. Systemic Effects Mercury is a systemic poison. Depending on the dose and length of exposure period, it can affect various organ systems and functions that are discussed in the following text. Immune system
Mercury inhibits T-cell proliferation in the presence of monocytes. It inhibits the ability of these cells to synthesize and secrete interleukin-1 (IL-1). Mercury also impairs T-cell expression of IL-2 receptor and the transferrin receptor, thereby decreasing the T-cell function. In both in vitro and in vivo studies, lead and mercury have been found to enhance the production of IL-4 by the Th2 clone and inhibit the proliferation of Th1. This imbalance between the Th1 and Th2 results in increased production of antibodies to self-antigen and thereby generation of autoimmunity. Heavy metals cause aberrant expression of major histocompatibility complex II (MHC II) molecules on the target cells, inhibit the T-suppressor cells, alter the idiotype–anti-idiotype network, and induce heat-shock proteins. These factors may play a very important role in metal-induced autoimmunity such as multiple sclerosis, rheumatoid arthritis, and amyotrophic lateral sclerosis. Heavy metals, as discussed earlier, bind to SH and other groups and thereby modify other proteins. These modified proteins might activate B cell via the T cell and cause alteration of self-protein target for autoantibodies. The metals can bind to the MHC II without prior processing by the antigen-presenting cells or directly to the T-cell receptors. All these mechanisms have been proposed to be involved in metal-induced autoimmunity. In genetically susceptible mice and rats, the immunological effects such as hyperimmunoglobulinemia, development
710
Mercury Toxicity
of antinucleolar antibodies and antifibrillar antibodies, potentiation of mitogen-induced lymphocyte proliferation, and increased expression of class II molecules are caused by dental amalgam. Endocrine system
Mercury disrupts the function of the pituitary, thyroid, and adrenal glands even at low-level exposure. It has been reported that levels of mercury in the thyroid and pituitary glands were higher than those observed in kidney, brain, and liver tissue in people working in mercury mines in Solvenia. It reduces the level of posterior pituitary hormone, oxytocin. The bouts of depression and suicidal thoughts are mainly associated with the reduction of oxytocin. Mercury amalgam in dental fillings is a contributing factor in reducing pituitary functions. Administration of oxytocin alleviates many of these mood problems. Intramuscular administration of mercury chloride has been shown to induce hyperthyroidism in rabbits within 24 h of its administration. Mercury increased thyroid peroxidase activity, and induced a rise in the triiodothyronine (T3) and a remarkable fall in the thyroxine (T4) level. The T3/T4 ratio became higher than the controls. T3-toxicosis may be produced by a preferential synthesis of T3 or preferential deiodination of T4 to T3. On the other hand, it has also been reported that mercury causes hypothyroidism. Mercury has also been shown to cause autoimmune thyroiditis. The thyroid imbalance caused by exposure to mercury could play a major role in chronic heart conditions such as clogged arteries, myocardial infarction, and chronic heart failure. In an experiment, administration of 2.5 mg of mercuric chloride daily for 40 days to rats enhanced the thyroidal weight, thyroidal 125I uptake, and serum protein-bound radio-iodine (PB125I). Further investigation with the chromatographic analysis of thyroid pronase hydrolysates of the mercury-treated rats displayed an increase in percentage of labeled monoiodotyrosine (MIT) without any alteration in the percentage of labeled T3, indicating a coupling defect in the synthesis of T3. Prenatal exposure to methylmercury severely inhibits the activity of 5-iodothyronine deiodinases (5-DI) in the fetal brain, and increases the activity of 50 -DI. This manifests as hypothyroidism. Mercury also accumulates in the lipid-rich cortex of the adrenal gland and disrupts the function of the gland by blocking various enzymatic pathways. Heavy metals have been found to cause hyperandrogenemia or partial hypoadrenalism and polycystic ovarian syndrome in women. Respiratory system
A survey conducted on the workers exposed to mercury vapors accidentally at an estimated concentration of
approximately 44.3 mg m3 for 4–8 h showed that they exhibited chest pains, dyspnea, cough, hemoptysis, reduction in vital capacity leading to failure of pulmonary functions, and also interstitial pneumonitis. Inorganic mercuric compounds also contribute toward respiratory problems such as pulmonary edema and shortness of breathing.
Nervous system
A variety of disturbances primarily in the sensory and motor nerves have been reported. The cardinal neurological sign of mercury vapor exposure is tremor. Other neuropsychological signs and symptoms are emotional lability, insomnia, memory loss, weakness, muscle atrophy, muscle twitching, headaches, paresthesias, and polyneuropathy. Long-term exposure is associated with unsteady walk, poor concentration, tremulous speech, blurred vision, impaired hearing, reduced finger to eye coordination, etc. One of the major mechanisms of the toxicity of methylmercury is the inhibition of tubulin polymerization. Microtubular fragmentation has been found in cultured rat cerebellar granular neurons treated with 0.5–1 mM of methylmercury. Fragmentation of microtubule by methylmercury results in degeneration of neurons and inhibition of neuronal migration. Methylmercury depolarizes the presynaptic membrane that increases the Na2þ ion concentration and decreases Kþ ion concentration, causing disruption of Ca2þ homeostasis leading to increased intracellular Ca2þ concentration. Increased calcium damages the cell membrane and disrupts neurotransmitter signaling. Methylmercury stimulates the release of neurotransmitters such as dopamine, glutamate, gamma-aminobutyric acid (GABA), glycine, choline, and acetylcholine. It inhibits the uptake of excitatory amino acids such as glutamate, and aspartate in the astrocytes. Mercury has been shown to accumulate in the lysosomes of the efferent neurons of spinal cord by a retrograde axonal transport. Methylmercury exposure is associated with increased mitochondrial membrane permeability in cultured astrocytes. This leads to a collapse of the mitochondrial inner membrane potential (Cm). Loss of the Cm results in colloid osmotic swelling of the mitochondrial matrix, movement of metabolites across the inner membrane, defective oxidative phosphorylation, cessation of ATP synthesis, generation of reactive oxygen species and consequent oxidative injury. Methylmercury has been shown to induce generation of reactive oxygen species in synaptosomes by activating protein kinase C (PKC), and mitogen-activated protein kinase (MAPK) pathway kinases. The generated reactive oxygen species cause the downstream activation of a variety of kinases (p38MAPK, ERK, and JNK), leading to a cytotoxic response.
Mercury Toxicity
Cardiovascular system
Mercury in both organic and ionic forms accumulates in the heart and causes hypertension, tachycardia, and ventricular arrhythmias. Increased blood pressure and palpitation is observed on the inhalation of elemental mercury vapor. As the duration of mercury exposure increases, there is also an increase in heart rate. It has been reported that children treated with mercurous chloride tablets for worms or with ammoniated mercury ointment for diaper rash suffered from tachycardia and elevated blood pressure. Gastrointestinal system
Exposure to mercury results in the inflammation of oral mucosa, excessive salivation, and difficulty in swallowing. It also leads to diarrhea, sore gums, ulceration of the lips and tongue, and vomiting. Renal system
The target organ for both ionic elemental or mercuric mercury is the kidney where it has a half-life of 2 months due to its biotransformation to ionic forms. It accumulates in the kidney. The binding of mercury to glomerular basement proteins causes severe damage to the kidney and results in proteinuria, hematuria, and oliguria. An increase in tubular antigens and enzymes has been reported in workers with chronic low-level exposure to elemental mercury vapor. It has been reported that 62 out of 86 cases of ethyl mercury poisoning from Iraqi grain seed exhibited clinical symptoms of kidney damage. It has been reported that when mice were fed on a diet containing methyl mercury for approximately 1 year, they developed renal carcinoma. Reproductive system Female
Maternal exposure to mercury before and during pregnancy may lead to spontaneous abortion, preterm birth, and low birth weight. It has been observed that abnormalities in menstrual cycle and miscarriage are common among the women exposed to mercury vapor. Female dental assistants exposed to elemental mercury have a lower fertility. The mercury vapor from toothfilling amalgam may interfere with corpus luteum function. Mercury inhibited the production of progesterone by the human granulosa cells in vitro. An early rise of estradiol level in the follicular phase in women showing high levels of mercury has been observed. This too could cause luteal phase defect. Injection of mercuric chloride into the female hamster resulted in prolongation of estrous cycle and morphological changes in the corpora lutea along with the inhibition of follicular maturation and elevation of pituitary FSH. The injection of mercuric chloride in female mice before mating resulted in dose-related
711
increase in preimplantation and early postimplantation fetal loss. Male
In males, mercury mainly affects sperm production, motility, and morphology. Mercury decreases sperm motility, and induces a higher percentage of abnormal sperm tail forms marked by bent, kinked, and coiled tails in Macaca fasicularis. However, no changes in serum testosterone were observed. Methylmercury has been suggested to inhibit mitotic divisions in the germ cells to decrease the sperm production and also inhibit microtubule assembly during spermiogenesis. In men as well, methylmercury decreases sperm count, increases abnormal morphology and impotency, and decreases libido. Antifertility effects of mercury in mice can be attributed to the direct inhibition of the DNA synthesis in the spermatogonial cells since the mercury-bound DNA can affect the unwinding and rewinding of DNA essential for replication and transcription. Intercalation of DNA in turn affects the RNA transcription and protein synthesis. Mercury also has an inhibitory effect on essential spermatogenic enzymes. Fetal and child health
Mercury vapor can pass through the feto-placenta membrane and reach the brain of the fetus and endocrine glands. The blood level of mercury in fetus reaches a level higher than the level in maternal circulation. It has also been reported that mercuric ions accumulate in the placenta and thereby limit the exchange of nutrients and gases to the fetus. At peak maternal hair mercury levels above 70 mg g1, there is a high risk, more than 30%, of neurological disorder in the offspring. It has been reported that even at a low dose, mercuric ions may affect embryonic development and cause fetal malformation. The administration of large doses of mercury to pregnant rodents resulted in cleft palate. Prenatal exposure of rats produced abnormalities in renal function that were detected in offspring at 42 days of age. Thimerosol used as preservative in vaccines and anti-RhoD-immunoglobulins administered to Rh-negative pregnant women and methylmercury in seafood could poison the fetus. One out of six women in the United States has mercury levels high enough to increase the risk of neurotoxicity to fetus. The report of ingestion of flour from grain treated with alkyl mercury by pregnant women in Sweden showed normal birth of the offspring but later there were signs of brain damage manifested as mental retardation and inability to move. In a case study, a 3-month pregnant woman consumed methylmercurycontaminated meat, and delivered a male infant. The infant showed elevated urinary level of mercury. At 3 months of age, the infant was hypotonic and irritable, and exhibited myoclonic seizures, and at 6 years of age the
712
Mercury Toxicity
child displayed severe neurological impairment such as blindness, neuromuscular weakness, and inability to speak. Children with regressive autism, chronic constipation, and diarrhea have 10 000 times the normal level of E. coli in their stool. They have a higher intake of antibodies in the first 36 months of their life and therefore a higher possibility of further change in gut ecology. These children have a higher level of mercury in their infantile teeth in comparison with normal controls. Their reduced ability to excrete mercury may further exacerbate their neurological deficit. Children exposed to mercury vapors show acrodynia, which is a hypersensitivity reaction to mercury and expressed by irritability, mood changes, erythema of the hands, feet, and sometimes nose and other parts of the body, leg pains or cramps, peeling of the palms of the hands and soles of the feet, and insomnia.
Several thiol-containing complexing agents have also been successfully used to remove methylmercury from the body. The most promising complexing agent is N-acetylcysteine, which when given by mouth, traps the methylmercury secreted in bile and carries it into the feces. It enhances methylmercury excretion when given orally, has a low toxicity, and is widely available in the clinical setting.
Conclusion Mercury toxicity is a major health risk particularly to the developing economies that depend on thermal power generation. The risk to these countries is also heightened by the disposal of mercury waste in their shores and soil by the developed countries. The world needs to take cognizance of this toxin and develop regulatory measures and health policies.
Modification of Mercury Toxicity Selenium (Se) has been known to bind toxic metals. Owing to this property, it plays a role in potentially reducing their toxicity. It has been shown that accumulation or retention of selenium is increased in response to accumulation of mercury in tissues of marine mammals or mineworkers following exposure or ingestion. When inorganic mercury and sodium selenite are administered together, sodium selenite forms a high-molecular-weight complex with mercury and plasma protein, thereby reducing the accumulation of mercury in kidney; it has also been shown to form an inert high-molecular-weight complex, which stays in the red blood cells without doing any harm to other tissues. The plasma protein to which Hg–Se complex binds has been identified as plasma selenoprotein (selenoprotein P). Various studies demonstrated the binding of complexes of mercury–selenium, silver–selenium, and cadmium–selenium by plasma selenoprotein P, suggesting that this protein may function to chelate heavy metals, reducing their toxicity. Mercury exposure leads to significant increase in the expression of both selenoprotein P and glutathione peroxidase (GSHPx) in mercury miners. These increases were accompanied by elevated selenium concentrations in serum. Selenoproteins play two important roles in protecting against mercury toxicity. First, they may bind more mercury through their highly reactive selenol group, and second, their antioxidative properties help eliminate the reactive oxygen species induced by mercury in vivo.
Further Reading Ballatori N (2002) Transport of toxic metals by molecular mimicry. Environmental Health Perspectives 110: 689--694. Berry MJ and Ralston NVC (2009) Mercury toxicity and the mitigating role of selenium. Ecohealth 5: 456--459. Carvalho CML, Chew EH, Hashemy SI, Lu J, and Holmgren A (2008) Inhibition of the human thioredoxin system. A molecular mechanism of mercury toxicity. Journal of Biological Chemistry 283: 11913--11923. Clarkson TW (1995) Environmental contaminants in the food chain. American Journal of Clinical Nutrition 61: 682s--686s. Clarkson TW (2002) The three modern faces of mercury. Environmental Health Perspectives 110(supplement 1): 11--23. Clarkson TW and Magos L (2006) The toxicology of mercury and its chemical compounds. Critical Reviews in Toxicology 36: 609--662. Davidson PW, Myers GJ, and Weiss B (2004) Mercury exposure and child development outcomes. Paediatrics 113: 1023--1029. De Flora S, Benniceli C, and Bagnasco M (1994) Genotoxicity of mercury compounds. A review. Mutation Research 317: 57--79. Kuban P, Pelcova P, Margetınova J, and Kuban V (2009) Mercury speciation by CE: An update. Electrophoresis 30: 92--99. Magos L and Clarkson TW (2006) Overview of the clinical toxicity of mercury. Annals of Clinical Biochemistry 43: 257--268. Oklahoma State University, New Jersey State Department of Health, Division of Occupational and Environmental Health. The Health Effects of Mercury, Environmental Health and Safety. www.ehs. okstate.edu/training/mercury.htm (accessed January 2010). Risher JF (2003) Elemental Mercury and Inorganic Mercury Compounds: Human Health Aspects Agency for Toxic Substances and Disease Registry (ATSDR). Atlanta, GA: WHO. (Concise International Chemical Assessment Document 50). Risher JF, Murray HE, and Prince GR (2002) Organic mercury compounds: Human exposure and its relevance to public health. Toxicology and Industrial Health 18: 109--160. Who (1990) Environment Health Criteria: Methyl Mercury. Geneva: World Health Organization. pp. 1–144.