ASSESSMENT OF PERIPHERAL-BLOOD MONONUCLEAR CELL-PROLIFERATION BY [2-H-3]ADENINE UPTAKE IN THE WOODCHUCK MODEL

The levels of incorporated exogenous [H-3]thymidine of peripheral bloo d mononuclear cells (PBMC) of the woodchuck were low after stimulation with mitogens concanavalin A (ConA), phytohemagglutinin (PHA), and po keweed mitogen (PWM) when compared with other cell systems. The use of EDTA as an anticoagulant for blood sampling and AIM-V medium for cult uring of PBMC improved the [H-3]thymidine uptake of PBMC. A pronounced uptake is observed after use of [H-3]adenine instead of [H-3]thymidin e for PBMC proliferation measurement. One likely explanation for the d ifference in [H-3]adenine versus [H-3]thymidine uptake is that the alt ernative pathway for thymidine monophosphate synthesis is important: t he conversion of uridine to uridine monophosphate and, thereafter, to thymidine monophosphate. The optimal conditions for mitogen-induced pr oliferation of PBMC of the woodchuck were 2 mu g/ml ConA and PHA at da y 4 and 0.14 mu g of PWM/ml at day 5. No consistent differences of [H- 3]adenine uptake were observed between PBMC from four woodchuck hepati tis virus-infected woodchucks and five uninfected animals. (C) 1996 Ac ademic Press, Inc.