Dental amalgam has been used extensively for over one hundred years as a restorative material. Dental amalgam is composed of approximately 50% mercury, now known to be released over the lifetime of the restoration. There has been considerable discussion as to whether the amount of mercury released is significant to patient health, in particular the immune system.
For more than 20 years, there have been studies in murine and human models of the effects of mercury on immune function. Murine studies have demonstrated genetic susceptibility to mercury-induced autoimmune disease, with factors such as level and duration of exposure being of significance. Studies of human lymphocytes have demonstrated conflicting results with mercury being reported to depress and activate immune responses.
The aims of this study therefore were to examine Immune responses by analysing proliferation and cytokine production for a group of 19 healthy patients undergoing amalgam replacement with composite resin and to determine whether the number of amalgam surfaces removed or blood group influenced the results of these assays.
Peripheral blood mononuclear cells (PBMC) function was investigated using spontaneous proliferation assays before and after amalgam replacement. The plant mitogens phytohaemmagglutinin (PHA) and concanavalin A (Con A) stimulate T lymphocytes preferentially, and poke weed mitogen (PWM) stimulates both T and B cells and therefore proliferation studies using these mitogens are a general measure of cellular immunocompetence. There were no differences in responses to PHA and PWM. However there was a significant decrease in the proliferative response to Con A after amalgam replacement (p=0.027). The metal salts mercuric chloride (HgCl2) and aluminium chloride (AICI3) were used in the proliferation assays to investigate the effects of these metals on PBMC proliferation. There was a significant increase in the proliferative response to HgCl2 (p=0.001) but not to AlCl3 after amalgam replacement.
Cytokines are small proteins involved in communication between cells during immune responses. Interleukin (IL-2) is produced by T cells and large granular lymphocytes and interferon gamma (IFN-y) is produced by antigen-specific T cells and by natural killer cells recruited by IL-2. Commercial ELISAs were used to measure IL-2 and IFN-γ levels in the culture supernatants collected from the PBMC cultures containing PHA and HgCl2 before and after amalgam replacement. There was a significant decrease in IL-2 levels in the PHA- and HgCl2-stimulated culture supernatants between visits. IFN-y levels were found to be significantly lower in the PHA but not in the HgCl2 culture supernatants after amalgam replacement.
Analysis of the proliferation and cytokine levels in relation to the number of amalgam surfaces replaced demonstrated that the number of amalgam surfaces influenced the proliferative responses to both the metal salts, HgCl2 and AICI3 and to PHA in combination with HgCl2 but not to the plant mitogens. There was also a correlation with the number of amalgam surfaces and IFN-γ and IL-2 production in response to HgCl2. Patients with less than 13 amalgam surfaces had a significantly higher proliferative response to HgCl2 and AICI3 after amalgam replacement compared with patients with 13-48 amalgams. IFN-γ and IL-2 production in response to HgCl2 was greater for patients with greater than 36 amalgam surfaces after amalgam replacement.
The results of this study also demonstrated associations between blood groups and proliferative responses to the plant mitogens PWM and Con A and to the metal salt AICI3, Proliferative responses to mitogens and metal salts were higher for blood group O patients. Patients with blood group A demonstrated the highest proliferative response to PWM. There was no correlation between blood groups and the levels of IFN-γ or IL-2 produced in response to HgCl2 or to PHA after amalgam replacement. Cytokine production for blood group A patients tended to decrease after amalgam replacement, especially IL-2 production in response to both PHA and HgCl2. Blood group B and AB patients similarly experienced either no change or a decrease in cytokine levels. In contrast, most patients with blood group O experienced an increase in IFN-γ levels in response to HgCl2 after amalgam replacement.
In conclusion, this study has demonstrated that the functional activity of PBMC in some patients undergoing amalgam replacement is altered as reflected in changes in proliferative responses to plant mitogens, especially Con A and to HgCl2 and in changes in production of IL-2 and IFN-γ before and after amalgam replacement. The influence of blood group was established with O group patients having higher proliferative responses to Con A and HgCl2 and an altered pattern of cytokine production after amalgam replacement. Further investigation into the role of genetic susceptibility may allow at-risk individuals to be identified.