p16INK4A is a robust in vivo biomarker of cellular aging in human skin

The cell-cycle regulating gene, p16(INK4A), encoding an inhibitor of cyclin-dependent kinases 4 and 6, is considered to play an important role in cellular aging and in premature senescence. Although there is an age-dependent increase of p16(INK4A) expression in human fibroblast senescence in vitro, no data are available regarding the age dependency of p16(INK4A) in vivo. To determine whether p16(INK4A) expression in human skin correlates with donor age, p16(INK4A) expression was analyzed by immunohistochemistry as well as the expression of the p16(INK4A) repressor BMI1. Samples from the age groups 0-20, 21-70, and 71-95 years were selected from a bank of healthy human skin. We show that the number of p16(INK4A) positive cells is significantly higher in elderly individuals compared to the younger age groups. The number of p16(INK4A) positive cells was found to be increased in both epidermis and dermis, compartments with strictly different proliferative activities. BMI1 gene expression was significantly down-regulated with increasing donor age, whereas no striking age differences were observed for Ki67. In immunofluorescence co-expression studies, Ki67-positive cells were negative for p16(INK4A) and BMI1-expressing cells also stained negatively for Ki67. In conclusion, we provide for the first time evidence that p16(INK4A) expression directly correlates with chronological aging of human skin in vivo. p16(INK4A) therefore is a biomarker for human aging in vivo. The data reported here suggest a model for changes in regulatory gene expression that drive aging in human skin.