In Vitro Cellular Aging in T-Lymphocyte Cultures: Analysis of DNA Content and Cell Size

Abstract

Like other normal diploid mammalian cells, human T-lymphocytes display a limited in vitro lifespan. Long-term cultures of normal adult peripheral blood T cells, activated in vitro and passaged in the presence of interleukin-2 undergo 23 ± 7 cumulative population doublings. Cell cycle analysis revealed that in senescent T cell cultures, restimulation with the original antigen resulted in 16-22% of the cells entering S phase, compared to 60% entering cycle in young cultures. In addition, within 1 week of restimulation, the senescent cultures do not increase in cell number, but 93% of the cells return to the G1/G0 DNA content, a proportion typically seen in quiescent (nonrestimulated) cultures. Coculture of early- and late-passage cells at two different ratios excluded a putative inhibitory factor produced by the old cells and similarly eliminated a possible stimulatory product in early cultures. Flow cytometry measure of forward angle light scatter revealed no difference in cell size between early- and late-passage cells, in contrast to the findings with senescent fibroblasts. Thus, while increasing cell size may contribute to the senescent phenotype of fibroblast cultures, it is not a factor in the senescence of human T-lymphocytes, and it is therefore doubtful that alterations in cell size are fundamental to in vitro cellular aging.