Possible involvement of p21 but not of p16 or p53 in keratinocyte senescence

Abstract

It has been reported that p21, p53, and p16 affect the cell cycle and cell senescence. However, their roles in keratinocyte senescence are not clear. We established primary keratinocyte strains from 15 donors and maintained them until replicative senescence; their population doublings ranged from 5.7-45.2. These strains were classified based on their population doublings as short (5.7-10.4), intermediate (13.9-17.4), and long (21.5-45.2). To investigate the roles of p21, p53, and p16 in the cellular senescence of the cultured keratinocytes, we quantitatively analyzed p21, p53, and p16 levels of keratinocyte strains with different life spans by Western blot with Fluorol mager. p21 levels increased in the senescent phase but not in the nonsenescent phase in all of the short, intermediate, and long life-span strains. Northern blot analysis also revealed induction of p21 mRNA was similar to that of p21 protein levels. There were no apparent differences in p53 levels between senescent and nonsenescent cells. The short life-span strains exhibited a significant increase in p16 levels in the senescent phase (eighth or tenth passage). However, in two long life-span strains, p16 levels were increased in the nonsenescent phase (eighth passage) but then declined as the cells reached senescence (twenty-seventh passage). Therefore, induction of p16 appeared not to be associated with senescence in long life-span strains. In conclusion, p21 but not p16 or p53 may play roles in keratinocyte senescence.

Apoptosis Signal-regulating Kinase 1 (ASK1) Is an Intracellular Inducer of Keratinocyte Differentiation

“…Keratinocyte Culture-Normal human keratinocytes were cultured with MCDB153 medium supplemented with insulin (5 g/ml), hydrocortisone (5 ϫ 10 Ϫ7 M), ethanolamine (0.1 mM), phosphoethanolamine (0.1 mM), bovine hypothalamic extract (100 g/ml), and Ca 2ϩ (0.1 mM) as described previously (9).…”

p16INK4a can initiate an autonomous senescence program

“…INK4a accumulates as ®broblasts, T-lymphocytes, keratinocytes, mammary epithelial cells, uroepithelial cells, and prostate cells senesce (Alcorta et al, 1996;Brenner et al, 1998;Erickson et al, 1998;Jarrard et al, 1999;Kiyono et al, 1998;Rezniko et al, 1996;Sayama et al, 1999).…”

The ink4a/arf Tumor Suppressors Cooperate with p21 in the Processes of Mouse Epidermal Differentiation, Senescence, and Carcinogenesis

“…This is in agreement with the data obtained in human head and neck primary keratinocytes (51). However, it seems to be at variance with the recently reported involvement of p21 (but not of p16 or p53) in human skin keratinocyte senescence (52). It is important to note that the two proteins inactivated in the ink4a ⌬2,3 mutation, p16 ink4a and p19 arf , act in overlapping pathways in cellular immortalization (53).…”