Assessment of metal extract toxicity on human lymphocytes cultured in vitro

In this study the toxic effects of chromium, nickel, and cobalt extracts on in vitro cultured lymphocytes were evaluated. Graphite furnace atomic absorption spectrometry was used to measure the ion concentration. After serial dilution of the extracts, the viability of lymphocytes at 24, 48, and 72 h was estimated by flow cytometry, including propidium iodide staining and light scatter property assessment, and by MTT reduction test. The results of the investigation allowed us to conclude that 1) standardization of the procedure for preparing extracts is fundamental to obtaining repeatability of results; 2) the toxicity of an extract cannot be evaluated with a single viability assay; a combination of functional and structural tests is required; 3) when methods based on enzymatic reactions are performed, e.g. MTT test, it is advisable to replace the extract containing metal ions with fresh medium in order to avoid any interference with viability testing; 4) the amount of Co and Ni in the extract is similar, but the Cr release is very poor; 5) the lower toxicity of Cr extract probably is due to the lower ion concentration; 6) the assessment of 50% cytotoxic concentration (TC50) allows quantification of materials toxicity and comparison of various metals; and 7) the determination of a noncytotoxic concentration, i.e., a concentration lower than TC10, is required for subsequent investigation of cell functions because such studies can be carried out only on viable cell population.