Corrosion and wear of dental amalgam may be associated with unexpectedly high levels of endogenous exposure to heavy metals. According to WHO, the resulting uptake of mercury exceeds that from all other sources in persons not occupationally exposed (WHO 1991) and a daily uptake level of 100 μg has been reported (Barregard et al. 1995). Due to the distribution patterns of mercury, standard blood and urine analyses give meager information on the response of the organism to this exposure. We here present data from nuclear microscopy analysis of neutrophil granulocytes (short-lived cells in the immune system cascade) in peripheral blood. Blood samples were drawn from patients suffering from possible side effects from dental amalgam. Their symptoms resembled those of the Chronic Fatigue Syndrome (Fukuda 1994), and the onset or intensity of the symptoms were related to occasional increased exposure to dental amalgam e.g., unprotected placing/drilling of the material. Data showed profound derangement of several cellular trace elements in the patient group and in some cases the substitution of mercury for zinc in the nuclear area. This supports the contention that systemic side effects from dental amalgam may occur. Venous blood samples were drawn from a cohort of Caucasian patients (n = 25) with a chronic debilitating illness, possibly related to the exposure from dental amalgam, and cell preparations were done as previously described (Johansson 1984, Lindh 1997). The same procedure was performed on blood samples from an age- and sex-matched healthy control group (n = 22) with similar numbers of amalgam fillings. A freeze-dried monolayer preparation of neutrophil granulocytes from each subject was investigated by means of nuclear microscopy (Zidenberg-Cherr). Thirty cells from each subject were analyzed in a subsequent manner and means and variances of the elemental concentrations of calcium (Ca), manganese (Mn), iron (Fe), zinc (Zn), and mercury (Hg) were calculated. In addition, the intracellular distribution of zinc and mercury was investigated in a few cells from both patients and controls by means of a nuclear microscopy elemental mapping technique. Cells with mercury levels above the detection limit (0.5 μg/kg dry weight) were investigated as well as cells with no detectable mercury.

Mercury is an element with unique physical and chemical properties whose deleterious effects on various organ systems have been known for centuries. The metal (Hg°) mercury is the only element liquid at ambient temperatures and has an extremely high vapor pressure. Natural degassing of the earth’s crust by volcanoes and emissions from soils and waters are estimated to contribute on the order of 2700 to 30,000 tons per year (Nriagu 1989, Lindqvist 1991). A second source of mercury is anthropogenic from burning of coal or petroleum. The total input into the atmosphere may be up to 150,000 tons per year, with natural emissions accounting for the major input (Berlin 1986). However, estimations of contributions from different sources vary. Aristotle wrote about mercury as liquid silver (hydrargyrum) with the metallic mercury extracted in ancient times, as today, from the sulphide mineral cinnabar (HgS). Although technical developments have brought about more sophisticated methods of distilling mercury, all processes create mercury vapor, which is a potential hazard. Mercury mines pose environmental concern, due to mine tailings and waste rock contributing mercury-enriched sediment to watersheds (Rytuba 2000) such as in the California Coast Ranges (Rytuba 2000), the Idria mine in Slovenia (Hines et al. 2000), in Slovakia (Svoboda et al. 2000), and, perhaps most conspicuously, the mine tailings in Aznacollar, Spain, that caused a recent accident (Grimalt et al. 1999). Any industrial sites that utilize mercury during production may also produce contamination of the environment (Sunderland and Chmura 2000). The possible sources of mercury exposure are presented in Table 10.1. Amalgamation with mercury has been used as a method for beneficiation of gold and silver since Roman times. The total global release of mercury into the environment from these activities before 1930 was estimated as over 260,000 tons. Thereafter, with the introduction of cyanidation processing technology, the emissions declined (Lacerda and Solomons 1998). However, small-scale artisanal gold mining continues and is a serious hazard to largely unskilled persons in rural areas over the world.

OBJECTIVES The purpose of this study was to evaluate treatment of patients suffering from chronic ill health with a multitude of symptoms associated with metal exposure from dental amalgam and other metal alloys. SETTING AND DESIGN We included 796 patients in a retrospective study using a questionnaire about symptom changes, changes in quality of life as a consequence of treatment and assessment of care taking. METHODS Treatment of the patients by removal of offending dental metals and concomitant antioxidant therapy was implemented according to the Uppsala model based on a close co-operation between physicians and dentists. RESULTS More than 70% of the responders, remaining after exclusion of those who had not begun or completed removal, reported substantial recovery and increased quality of life. Comparison with similar studies showed accordance of the main results. Plasma concentrations of mercury before and after treatment supported the metal exposure to be causative for the ill health. MAIN FINDINGS Treatment according to the Uppsala model proved to be adequate for more than 70% of the patients. Patients with a high probability to respond successfully to current therapy might be detected by symptom profiles before treatment. CONCLUSIONS The hypothesis that metal exposure from dental amalgam can cause ill health in a susceptible part of the exposed population was supported. Further research is warranted to develop laboratory tests to support identification of the group of patients responding to current therapy as well as to find out causes of problems in the group with no or negative results.

Twenty-seven consecutive patients with health problems associated with dental amalgam were recruited. In spite of thorough medical examinations, there were no diagnoses available. The patient group was dominated by women. A healthy age- and sex-matched control group with dental amalgams without symptoms was also recruited. Metal level monitoring in plasma and nuclear microscopy of isolated individual blood cells were carried out. Significant increases of copper, iron, zinc and strontium were found in patient plasma. There was no significant difference in plasma selenium between the groups. Mercury was significantly increased in patient plasma, although there was overlap between the groups. In erythrocytes a significant increase in calcium and a significant decrease in magnesium, copper, manganese and zinc were found. Calcium, magnesium, manganese and copper increased in patient neutrophil granulocytes. A significant decrease was found for zinc. A conspicuous finding was the presence of measurable mercury in a few of the cells from the patient but not in the control group. Thus, nuclear microscopy of isolated individual blood cells might provide a better diagnostic tool for metal exposure than blood plasma measurements.

Nuclear microscopy based upon developments in high energy ion beam techniques is by now an accepted technique in many fields of research. The advancements into the biomedical field have, however, been slower than expected. A major factor explaining this tendency is the availability of nuclear microscopy. This paper reviews briefly the biomedical work using nuclear microscopy that has been carried out since the 4(th) International Conference on Nuclear Microprobe Technology and Applications held in Shanghai. Nuclear microscopy of isolated individual blood cells from patients adversely affected by metal exposure from dental amalgam has been performed both before and after removal of the metallic fillings. The elemental profile of blood cells was more or less normalised after treatment. Some of these results will be presented to illustrate a medical application. Results from bulk analysis by ICP-MS of erythrocytes and plasma before and after treatment will also be presented to illustrate the difference in information content between these two approaches as well as the need for complementary information in solving biomedical problems. As part of a larger study of acute porphyria, nuclear microscopy of blood cells was included among the 78 laboratory tests. The approach in this study was unbiased in the sense that no hypothesis was formulated as to which laboratory parameters would be the most explanatory for health or disease. Multivariate discriminant analysis was applied to the large amounts of data acquired. This approach led to the hypothesis that oxidative stress increased the synthesis of manganese-dependent superoxide dismutase in the mitochondria of polymorphonuclear leukocytes, explaining the increase of manganese in these cells. Antioxidant therapy was therefore applied to a couple of patients with porphyria, however, without clinical success. (C) 1997 Elsevier Science B.V.

Interaction between selenium and the heavy metals cadmium and mercury was studied in an experimental rat model (Sprague-Dawley). The rats were administered either one single trace element or combinations of selenium and cadmium as well as selenium and mercury. Salts of these trace elements were administered intraperitoneally daily during thirty days. Thereafter the animals were sacrificed and kidneys and livers excised rapidly. Thin sections were produced by a cryotome and subsequently freeze-dried. Nuclear microscopy of the sections showed that in the combination groups there was a co-localization of selenium and the heavy metals. None of the expected pathological signs of cadmium and mercury toxicity were observed. The conclusion was that selenium exerted a protective effect against the toxicity of cadmium and mercury through mechanisms still to be unveiled.

The sensitizing properties of metals widely used in medical and dental care have been studied with the help of an optimized lymphocyte proliferative assay, MELISA. MELISA (memory lymphocyte immuno-stimulation assay) was originally developed for the screening of allergenic epitopes of drugs and other chemicals of low molecular weight, but has recently been adapted for the study of metal-induced sensitization. The patients studied suffered from various oral mucosal problems which were suspected to be caused by the release of metal ions from dental restorations. They were also troubled by chronic fatigue persisting over many years. One patient was also occupationally exposed to metals while working in a dental practice. Healthy subjects without any discomfort due to metal devices served as controls. In addition to metals used in dentistry, lymphocyte responses to organic mercurials used widely as preservatives in vaccines, eye/nose drops and contact lense fluids were studied. The results indicated that mercurials, as well as other metals such as gold or palladium, induce strong lymphocyte proliferative responses in patients with oral or systemic symptoms, but not in similarly exposed unaffected subjects. The results of MELISA performed with a pair of identical twins with chronic fatigue syndrome (CFS) indicated that metal-specific responses may be dependent on the genetics of the patient. Thus, many metals that are today accepted for use in medicine and dentistry carry a definite sensitizing risk for certain genetically predisposed individuals. Therefore, the use of these metals should be limited in the future.

To diagnose infectious mononucleosis caused by Epstein-Barr virus (EBV), a peptide from the EBV nuclear antigen (EBNA) 1 (p107) and an EBNA 2 peptide (polyproline) were used as antigens in enzyme-linked immunosorbent assays for IgG and IgM. Well-characterized serum samples (360) from healthy individuals and patients with EBV or cytomegalovirus infections were examined. The p107 IgG and IgM assays were also tested with serum from 1000 patients with suspected EBV-related disorders. The p107 and polyproline IgG assays were 100% specific for EBV seropositivity. Low p107 IgG titers (less than 1000) were found in 98% of patients with EBV infectious mononucleosis but also in 18% of patients with other diseases. A p107-to-polyproline IgG ratio of less than 1 was 98% specific for EBV infectious mononucleosis; sensitivity was 86%. In EBV capsid antigen-IgG seropositive patients, a p107 IgG titer of less than 1000 together with a p107 IgG-to-IgM ratio of less than 1 was 98% sensitive and specific for EBV infectious mononucleosis. Thus, this ratio appears adequate to measure EBNA antibodies for diagnosis of EBV mononucleosis.