Kinetics of cell death and disintegration in human lymphocyte cultures

In order to quantitate lymphocyte proliferative responses the role of cell death in the kinetics of phytohemagglutinin-stimulated cultures was explored. Unless the disintegration time (tDIS) of nonviable lymphocytes in culture is known, the rate of cell death cannot be calculated. To obtain t(DIS) the time interval was between total and viable cell population decay after various killing events. Two subpopulations of lymphocytes were observed, the major (80%) with a mean (±SEM) t(DIS) of 16 ± 2 hr and the minor (20%) with a t(DIS) of 45 ± 7 hr. Kinetic balance sheets were constructed predicting total culture. DNA content (cells plus medium), was calculated both from proliferation rates and from observed death and disintegration rates. In an experiment characterized by extensive cell death, the 2 tallies were well-matched when the above data were utilized. The large discrepancy between predicted and observed DNA contents of the medium indicates that the DNA of disintegrated lymphocytes is extensive degraded. It is concluded that cell death explains proliferation deficits in stimulated lymphocyte cultures. This approach to quantitation of cell death may have general applicability to kinetic studies of cultured cells.