β-galactosidase activity as a biomarker of replicative senescence during the course of human fibroblast cultures

It has been called into question whether the commonly used beta-galactosidase staining is a reliable biomarker of cellular senescence because induction of beta-galactosidase activity also occurs independently of senescence. Here, we tested whether cytochemically detectable beta-galactosidase activity is reproducible and reflects the rate of cellular aging in vitro. Therefore, we serially cultured fibroblasts from 12 different donors and stained the cells for beta-galactosidase at pH 6 until the onset of the permanent growth arrest of the individual cultures. All fibroblast strains displayed a high replicative capacity with a similar growth pattern during the exponential growth phase and a very high interbiopsy variability in the onset of decreased mitotic activity and in the onset of growth arrest. Correspondingly, beta-galactosidase activity was low during the exponential growth phase, with an individually defined significant increase in activity when the growth speed of the culture decreased. The increase in beta-galactosidase activity was a better predictor for the onset of the decreased growth speed than the chronological life span of the culture expressed in population doublings. Within the phase of decreased mitotic activity, we observed a high fluctuation in the percentage of positively stained fibroblasts. Thus, our results support beta-galactosidase activity as a reliable biomarker for the course of replicative senescence, if used under defined standardized conditions.