Figure 2. The in vitro AID response of B cells to influenza vaccination decreases with age.

A. B cells (10⁶ cells/ml), isolated from the PBMC of subjects of different ages, before (t0) or after vaccination (t28), were cultured with the vaccine, for 7 days. At the end of this time, cells were processed as described in Materials and Methods. Thirty-seven subjects were evaluated, 29 young and 8 old. Two young subjects were not included because they were considered outliers (AID mRNA values >2 SDs above the mean). The dashed horizontal line indicates the threshold of a positive response in AID mRNA expression which was arbitrarily defined as 2-fold increase. Spearman’s rho =−0.527, p=0.001 (two-tailed). B. B cells (10⁶ cells/ml), isolated from the PBMC of 10 subjects of different ages (6 young and 4 elderly), stimulated in vitro with the influenza vaccine for 7 days to induce optimal AID mRNA expression. At the end of this time, cells were harvested, RNA extracted and qPCR reactions performed to evaluate AID normalized to GAPDH transcripts. Results are expressed as fold-increase in AID mRNA expression after vaccination, calculated as follows: AID values after vaccination/AID values before vaccination. The difference between young and old is significant at both t7 (p=0.008) t28 (p=0.032), as evaluated by the Wilcoxon test. White columns: young. Black columns: elderly.