The limitations and validities of senescence associated-beta-galactosidase activity as an aging marker for human foreskin fibroblast Hs68 cells.
The senescence associated-beta-galactosidase (SA-betaG) assay has become one of the most commonly used markers of cell-aging. However, the reliability of the assay is questionable because the enzyme is a non-specific marker for cell-aging. In this study, we found that the SA-betaG activity increased with cell age as well as in confluent quiescent cells or cells under serum starvation, and in cells treated with H2O2. Importantly, we found that SA-betaG activity was irreversibly increased in the senescent cells or H2O2-teated cells, but was reversible in quiescent cells induced by serum starvation or confluence. Using fluorescein di-beta-d-galactopyranoside (FDG) method for SA-betaG detection, we showed that senescent human foreskin fibroblast Hs68 cells did not express a specific enzyme that has a maximal activity at pH 6.0. In the pH profile of the cellular betaG activity in senescent Hs68 cells, only a single peak was found (with maximum at pH 4.6), and no addition peak was found at or around pH 6.0 that could be attributed to the SA-betaG activity. These results support the contention that SA-betaG is the lysosomal betaG that is detectable at suboptimal pH (i.e. pH 6.0) and demonstrate that cell-aging is not the only factor that can increase SA-betaG activity, rendering SA-betaG activity unspecific for cell-aging. Thus, the assay for cell-aging is only reliable when these confounding factors are controlled or excluded.
Yang
Nae-Cherng
NC
Department of Biotechnology, Chungchou Institute of Technology, Changhua, Taiwan, ROC.
Hu
Miao-Lin
ML
D000368:Aged; D015415:Biomarkers / Q000032:analysis; D002460:Cell Line; D016922:Cellular Senescence / Q000502:physiology*; D005347:Fibroblasts / Q000201:enzymology*; D006651:Histocytochemistry / Q000379:methods; D006801:Humans; D008297:Male; D008954:Models, Biological; D012867:Skin; D001616:beta-Galactosidase / Q000032:analysis*
D015415:Biomarkers; D001616:beta-Galactosidase