The coupling of the tetrazolium salts triphenyltetrazolium chloride and nitro-blue tetrazolium to the electron transport chain in mitochondria of thymus, spleen, liver, kidney, and Ehrlich ascites carcinoma cells has been studied with several substrates. In experiments on succinate–triphenyltetrazolium chloride reductase activity it has been possible to demonstrate a radiation lesion in the electron transport chain of mitochondria from thymus and spleen, but not in those from other tissues. This lesion is evident 4 hours after 25 rad of whole-body irradiation, or earlier with higher doses. It is not prevented by the prior administration of aminoethylisothiouronium bromide, serotonin, vitamin K1, or vitamin E, but is reduced by anoxic conditions.Lower levels of cytochrome c are found in irradiated mitochondria isolated from thymus, and the radiation lesion is believed to be produced by loosening the binding of cytochrome c to the mitochondrial membrane after X-irradiation. Decreased levels of ATP occur in thymus, spleen, and ascites cells following irradiation.

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The pyridine nucleotides of thymocytes have been examined following irradiation of the cells in vivo or in vitro. NAD was found to constitute about 95% of the total oxidized pyridine nucleotides in thymocytes while the level of NADH2 was found to be very low. There is a rapid loss of NAD from thymocytes following irradiation in vitro or in vivo which is evident with as little as 25 rads. There is no concomitant rise in the NADH2 levels following irradiation. The loss of NAD occurs principally from the nucleus of the cell and is characterized by an intracellular release of NADase from the mitochondria and nucleus into the cytoplasm. Following irradiation there is a reduced capacity for thymocytes to synthesize NAD from 7-C14-nicotinamide either in vitro or in vivo. The administration or presence of nicotinamide produces an unphysiological increase in the level of NAD in whole thymus or thymocytes in suspension. The level of NAD in treated cells is higher than in untreated controls following irradiation. This effect is not produced by nicotinic acid, cysteamine, L-cysteine, or promethazine. The administration of nicotinamide does not prevent the development of pyknotic degeneration in irradiated thymocytes. There is no change in the RNA content of the thymus under the conditions which reduce the NAD content.

A simple, rapid quantitative method for biologically evaluating radioprotective compounds a t the cellular level has been developed by ineans of Ehrlich mouse ascites tumor cells. The cells were irradiated in vitro and subsequently grown in vivo and the radioprotective compound was evaluated by its ability to prevent mitotic arrest and loss of proliferative capacity in the cells. Results statistically significant a t the 1% level were available in 48 hours when groups of 15 mice per sample were compared. L-Cysteine was found to be a better protector than 2-an~inoethylisothiouronium bromide hydrobomide or 2-mercaptoethylamine and also increased the protection of cells irradiated under anaerobic conditions.

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