Cytometric analysis on cytotoxicity of 4,4′-methylenediphenyl diisocyanate, a chemical allergen, in rat thymocytes

4,4’-Methylenediphenyl diisocyanate (MDI) is a cross-linking agent. Chemical reactivity of MDI with endogenous substances such as albumin and glutathione (GSH) is assumed to be responsible for MDI toxicity. We examined the cytotoxic effect of MDI on rat thymocytes, under the condition that endogenous biological substances except for cells were nominally absent, in order to study chemico-biological interactions between MDI and cells. The treatment of 10-50 µM MDI for 3 hr significantly increased the side scatter signal intensity of cytograms, without affecting forward scatter intensity. The increase in side scatter signal intensity by MDI was associated with an increase in cell lethality. The treatment of cells with 50 µM MDI for 3 hr increased cell lethality without increasing the population of preapoptotic annexin V-positive living cells. In contrast, H2O2 at 100 µM significantly increased the population of annexin V positive living cells prior to cell death. MDI at 30-50 µM did not affect the increase in cell lethality induced by H2O2 or A23187. Simultaneous application of 50 µM GSH did not affect the cytotoxicity of 50 µM MDI. It was therefore concluded that the process of cell death induced by MDI could not be attributed to oxidative stress and intracellular Ca2+ overload, and that MDI possesses cytotoxic actions that are not significantly related to its chemical reactivity with GSH.