Pharmacology & Toxicology Poster Session
Discussion and Conclusion
It was shown by these studies with RRBC that EDTA had no direct toxic effect in the absence of extra-cellular calcium and magnesium ions. EDTA has been shown to be capable of penetrating biological membranes and tissue stroma (Grass et al. 1985, Grass and Robinson 1988). If the toxicity of EDTA is assumed to be related to its ability to penetrate the cell, this might suggest that EDTA requires the cations as a transport medium, and probably uses the same specific membrane channels of the cations to penetrated the cell (Rosenblatt et al. 1978, Brownie et al. 1986). The cations may therefore be required in assisting the intracellular transport of EDTA to effect its toxic effects. On the other hand, it might be postulated that the cytotoxicity of EDTA is related to its ability to bind to intracellular metal ligands, thereby resulting in the formation a more active toxic-inducing form.
Calcium demonstrated a bimodal action on the toxicity of EDTA. While low concentrations up to 5mM enhanced EDTA toxicity, higher concentrations inhibited the same. Calcium has been shown to form a thin film on cell membrane (Naccache et al. 1979), which might block the proposed model "pores" on the cell membrane through which certain drug molecules, especially polar compounds, penetrate the cell. The ability of any agent such as EDTA to bind to calcium would then be an advantage, by allowing it to bind closely to the surface of the cell membrane, and thereby more readily gain entrance into the cell. Excess calcium probably exerts an inhibitory effect on EDTA toxicity by forming complexes with EDTA in the extra-cellular fluid with a reduced capacity for being transported across the cell membrane, as a result of changes in molecular size, ionic charges or both. In this respect, Ca-EDTA has also been shown to be a less toxic compound than EDTA, and to be able to protect the cell membrane lipid against peroxidation (Beales et al. 1985, Kane et al. 1989).
Intracellular accumulation of calcium on the other hand, is associated with enhanced toxicity of certain chemicals, such as galactosamine- and 1,2-dichlorvinyl-cysteine (DCVC)-induced hepatic necrosis, 7,12-dimethylbenz(a)anthracene (DMBA)-induced lymphocyte toxicity and membrane attack complement cytolysis (MacDonald et al. 1985, van-de-Water et al. 1993, Papadimitriou et al. 1994). This effect of intracellular calcium may be related to the activation of phospholipase A2 activity. The effect of intracellular calcium on the toxicity of EDTA is unknown.
Magnesium caused a concentration-dependent enhancement of EDTA toxicity above 1mM concentration. It is probable that Mg-EDTA complexes are of a smaller molecular weight and enhance the transport of EDTA through the specific Mg2+ channel on the cell membrane. This observation may also be due to physicochemical differences between the calcium and magnesium channels. The Mg2+ membrane recognition site is said to possess a very high anionic field strength (Grubbs et al. 1985). The toxicity of EDTA might also be independently enhanced by intracellular levels of magnesium cations, as for some other cytotoxins (Patel et al. 1994). Though the actual level of interaction between calcium and magnesium cations has not been specifically examined in this study, it may be both intra- and extra-cellular.
A practical conclusion from this study would include the fact that EDTA levels of at least 10mM concentration might cause toxic hemolysis when used as an anticoagulant in blood samples, in the presence of serum levels of divalent cations. It may therefore not be suitable for an assay which depends on measurements of hemoglobin, methhemoglobin or cellular components.
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|Ogundele, M.O.; (1998). the Role of Divalent Cations in the Mechanisms of EDTA Cytotoxicity. Presented at INABIS '98 - 5th Internet World Congress on Biomedical Sciences at McMaster University, Canada, Dec 7-16th. Available at URL http://www.mcmaster.ca/inabis98/pharmtox/ogundele0173/index.html|
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