Expression of p16(INK4a) in peripheral blood T-cells is a biomarker of human aging

Abstract

Expression of the p16(INK4a) tumor suppressor sharply increases with age in most mammalian tissues, and contributes to an age-induced functional decline of certain self-renewing compartments. These observations have suggested that p16(INK4a) expression could be a biomarker of mammalian aging. To translate this notion to human use, we determined p16(INK4a) expression in cellular fractions of human whole blood, and found highest expression in peripheral blood T-lymphocytes (PBTL). We then measured INK4/ARF transcript expression in PBTL from two independent cohorts of healthy humans (170 donors total), and analyzed their relationship with donor characteristics. Expression of p16(INK4a), but not other INK4/ARF transcripts, appeared to exponentially increase with donor chronologic age. Importantly, p16(INK4a) expression did not independently correlate with gender or body-mass index, but was significantly associated with tobacco use and physical inactivity. In addition, p16(INK4a) expression was associated with plasma interleukin-6 concentration, a marker of human frailty. These data suggest that p16(INK4a) expression in PBTL is an easily measured, peripheral blood biomarker of molecular age.

Figure 1. Expression of p16^INK4a in PBTL is associated with chronologic age

(A) Comparison of relative p16^INK4a expression in different cell types of the peripheral blood. Error bars indicate standard error of the mean (SEM). (B) Linear relationship between log₂-transformed p16^INK4a mRNA expression and chronological age. Aggregate data from the exploratory and validation cohorts are shown. The correlation coefficients and p-values of individual cohort are shown in Table 1. Dotted lines indicate the 95% confidence intervals (CI) of the fitted line. (C) Expression of p16^INK4a protein increases with chronologic age in a representative subset of patients.

Figure 2. Expression of other INK4/ARF transcripts is associated with p16^INK4a expression, but not age

Dotted lines indicate the 95% confidence intervals (CI). The correlations between the variables shown in the left panels are significant (p<0.0001) while those in the right panels are not significant (p>0.4). Data shown are from the exploratory cohort.

Figure 3. Expression of p16^INK4a is associated with smoking, exercise and plasma IL-6 concentrations

(A) The comparison of p16^INK4a -age linear regression among never smokers, former smokers and current smokers is shown for the combined cohort. Slope comparison: Current, 7.5 ± 0.1 × 10^-2; Never, 3.9 ± 0.7 × 10^-2. *p<0.05, adjusted for multiple comparisons. (B) Expression of p16^INK4a is associated with tobacco exposure. Categorized pack-years: Mild (>0, ≤5); Moderate (5-10); Heavy (11-30); Severe (31-50); Extreme (51 and up). (C) Decreased p16^INK4a mRNA expression is associated with increased exercise intensity. (D) Plasma IL-6 levels are associated with increased p16^INK4a expression. Aggregate data from the exploratory and the validation cohorts are shown in a, b, and c; whereas d represents data only from the validation cohort. The Pearson correlations and p-values in each cohort are shown in Table 1. Dotted lines in c and d indicate 95% CI.