On-line studies of activation events in primary human T lymphocytes

In this paper, we review our NMR studies of human peripheral blood T lymphocytes. These studies focus on the physiological and biochemical alterations accompanying cell cycle progression. In particular, we have characterized phosphorus metabolism, glucose utilization and lactate production, and pH regulation using 31P, 13C, and 19F NMR, respectively. These studies required developing new methods for monitoring on-line stimulation of quiescent T cells under sterile, physiological conditions (i.e., CO2/HCO3- buffer, 37 degrees C) for prolonged periods of time. A perfusion system optimized for T lymphocytes inside agarose beads is described. In addition, custom-designed 19F NMR pH indicators were synthesized, characterized, and used to determine intracellular pH in quiescent lymphocytes, stimulated lymphocytes, and lymphocytes undergoing the G0-G1 transition. These unique molecular probes are described in detail. Finally, the physiological relevance of our findings regarding carbon metabolism and pH regulation is considered in the context of mitogenesis.