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In an oral environment, minute amounts of constituent metals dissolve from the surfaces of restorations. The uptake of the metals through the mucosa may cause systemic or local effects. The purpose of this study was to investigate these effects and mechanisms, focusing on the nickel as a dental metal. A nickel-chromium dental casting alloy was stored in 3 immersion solutions for 4 weeks and the amount of nickel dissolved was measured. NiCl2 was administrated to the cultured medium to examine the cytotoxicity for HeLa cells and periodontal ligament-derived cells as well as the nickel distribution in the HeLa cells. The results were as follows: 1. The amounts of nickel released from the alloy were 1.2 micrograms/cm2 (human saliva), 2.0 micrograms/cm2 (PBS (-)) and 2.5 micrograms/cm2 (MILLI-Q water). 2. 3.0 mM NiCl2 showed a large cytotoxicity resulting in the cell growth rate and morphological changes. 3. Incubated for 72 hours in 0.15 mM NiCl2, the amount of nickel in the 10(6) cells was 20 ng. After 24 hours of incubation with NiCl2, and the following 72 hours without NiCl2, the amount of nickel had decreased, but 30% still remained. 4. Incubated for 72 hours in 0.15 mM NiCl2, 65% of the nickel in the cells was bound by the soluble fraction. These results suggest that nickel released from the oral prosthesis may be sequestered for a long term in the cells and possibly causes some effects.

To elucidate the possible toxicity of heavy metals in a renal tubular epithelial cell line derived from a normal cynomolgus female monkey (JTC-12, P3 (F], the effects of HgCl2, Na2CrO4 and NiCl2 on the dome formation and the release of intrinsic enzymes from the cells were studied. The results were as follows: 1. The JTC-12. P3(F) cells showed an evident dome formation when an inducer of differentiation (hexamethylene bisacetamide or N, N-dimethylformamide) was added to the medium. 2. Ouabain inhibited the dome formation of the JTC-12. P3(F) cells, suggesting that the dome formation is dependent on the active transepithelial transport of Na+. 3. The addition of 40 microM HgCl2, 10 microM Na2CrO4 or 20 microM NiCl2 inhibited the dome formation. 40 microM HgCl2, 10 microM Na2CrO4 or 150 microM NiCl2 caused significant cell death. 4. The addition of 5 microM HgCl2, 1 microM Na2CrO4 or 20 microM NiCl2 resulted in an increased release of lactate dehydrogenase into the medium for 6 hours. 20 microM HgCl2, 1 microM Na2CrO4 or 20 microM NiCl2 increased the medium gamma-glutamyl transpeptidase concentration for 6 hours, but 40 microM HgCl2, 5 microM Na2CrO4 or 100 microM NiCl2 did not cause a significant increase in the medium alkaline phosphatase concentration. The results suggest that the JTC-12. P3(F) cells possess, at least in part, functional characteristics similar to the other kidney proximal tubular cells and that the inhibition of dome formation and enzyme release from the cells may be the early indicators that predict metal toxicity to the cells.

The effects of metal ions and mechanical stress on cells were examined in vitro, to shed light on tissue response to metal prosthesis. Seven commercial base alloy plates, four Co-Cr alloys, one pure Ti, one Ni-Cu-Ga alloy and one Ni-Cr alloy for solder were used. Two kinds of static load, 5 gf/cm2 and 10 gf/cm2, were applied to three kinds of tissue culture cells, L-929, HEp-2 and Gin-1 cells, for 24 hours, and then the cells were incubated for 7 days under normal culture conditions. Cell recovery was evaluated by cell number and protein content. The amount of dissolved metal ions was measured by atomic absorption spectroscopy (Experiment 1). A concomitant study of cell culture with known amounts of Co, Cr, Ni was carried out, and cell recovery was examined (Experiment 2). Atomic absorption spectroscopy showed that dissolution of either Co or Cr from the alloys tested was extremely small. However, the maximum amount of Ni and Cu dissolved from Ni-Cu-Ga alloy and Ni-Cr alloy for solder was over 10 ppm. Ti was not dissolved from pure Ti. Four Co-Cr alloys and pure Ti did not inhibit cell recovery or protein content with loading. However, Ni-Cu-Ga alloy and Ni-Cr alloy significantly inhibited cell recovery and protein content, and the effects were enhanced by loading. The cytotoxicity of Co, Cr or Ni ion at a static load of 10 gf/cm2 was higher than that at 5 gf/cm2. These results suggest that mechanical stress increases the cytotoxicity of dissolved metal ions.

The effects of cobalt (Co) and nickel (Ni) on the humoral immune response were studied by two indexes of specific IgM antibody production against sheep red blood cells (SRBC) and polyclonal IgG antibody production in the spleens of mice intraperitoneally injected with cobalt chloride or nickel chloride. An experiment for the effect of both metals on specific IgM production was carried out by measuring IgM plaque-forming cells in the spleens of mice intraperitoneally injected with both metal salts using 1/10, 1/100 or 1/200 of LD50 for i.p. injection three times every other day and were immunized with SRBC on the day of the last injection of each metal salt. The other experiment for the effect of both metals on polyclonal IgG production was done by measuring, on days +3 or +6 in relation to the last injection of metal salts, polyclonal IgG-forming cells in the spleens of mice injected with both metal salts using 1/10 or 1/100 of LD50 for i.p. injection three times every other day by the reverse plaque-forming method. The following results may be drawn from this study: 1. Co may cause changes in the homeostasis of humoral immune response even more than affecting the immune system with immunotoxicity as antigenicity. 2. On the other hand, Ni may have antigenicity even more than an acting as immunomodulator influencing the immune system.

In order to investigate biocompatibility of pure metals, which are widely used, the gingival tissues were brought into contact with pure metals. Nine kinds of pure metals, such as gold, platinum, silver, palladium, copper, nickel, zinc, indium and tin were used in the analysis of biocompatibility. With a No. 1/2 round bar, a standardized small cavity was prepared in the mesiolingual cervical portion of the upper first molar. Nine kinds of pure metals powder were inserted into the small cavity of the maxillary root surface in rats. Post operative changes of the gingival tissue were studied histopathologically. The results were as follows: 1. Gold, palladium and indium powder were observed in the lamina propria of the gingiva. Infiltration of cells was not observed around these pure metal powders. Gold, palladium and indium powders showed little cytotoxicity. 2. Zinc powders were not associated with chronic inflammatory charges. Zinc powders were surrounded by an abundance of connective tissue substances. The fragments were surrounded by collagen fibers, but no inflammatory cells were present. Zinc powders showed little cytotoxicity. 3. Platinum and tin powders were not associated with chronic inflammatory changes. These pure metal powders were surrounded by macrophages, but no other mononuclear inflammatory cells were present. 4. Clumps and granules of silver powder were not associated with chronic inflammatory changes. Multinucleate giant cells were shown surrounding silver powders. Silver powders were observed to have been taken into the multinucleate giant cells. Many of these cells also contained fine dark granules in their cytoplasm, and a few contained small pieces of silver. The tissue reaction to silver showed little cytotoxicity. 5. Copper and nickel powders were associated with chronic inflammatory cells. Moderate chronic inflammation and occasional neutrophils leucocytes and lymphocytes were shown to be associated with these pure metal powders. Copper and nickel caused extensive damage to the gingival tissues.