ATP depletion, purine riboside triphosphate accumulation and rat thymocytedeath induced by purine riboside

Purine riboside (purine-1-D-ribofuranoside, nebularine), an adenosine analo g, exerts cytotoxic effect both in vivo and in vitro. However, exact bioche mical mechanism for its toxicity and sensitivity of lymphoid cells remains unknown. The present experiments have examined the sequential metabolic cha nges leading to cell death, induced in cultured rat thymocytes during incub ation with purine riboside. Among 22 analogs tested, purine-riboside and tu bercidin were most toxic as determined by trypan blue exclusion and lactate dehydrogenase leakage from the cells. 2-Chloroadenosine and 2'-deoxyadenos ine were only moderately toxic, whereas other analogs tested were without e ffect on cell viability. In the presence of purine riboside, more than 90% of ATP was lost after 2 h of incubation. Hypoxanthine accumulated in the me dium and the formation of purine-riboside triphosphate exceeded 4-fold the physiological concentration of ATP in the cell. Inhibition of adenosine kin ase by 5-iodotubercidin reversed the cytotoxic effect of purine riboside. I nterestingly, cells virtually deprived of ATP after 2 h of incubation with purine riboside maintained high nucleotide energy charge value and high via bility. Purine riboside triphosphate was capable to replace ATP in stimulat ion of glycolysis in cell-free thymus extract. We conclude that for a short time (a few hours) purine riboside triphosphate formed in the cell may ser ve in the absence of ATP as an intermediate of cellular energy metabolism i n rat thymocytes. However, possibly due to toxic effects of purine-riboside triphosphate, cells were finally dying. Thus, ATP depletion and adenosine kinase mediated purine riboside phosphates formation are the principle caus es of rat thymocytes death exposed to purine riboside. (C) 1999 Published b y Elsevier Science Ireland Ltd. All rights reserved.