(B) Percentages of Annexin V+ cells in the indicated groups; Statistical significance was assessed using the Mann-Whitney 0.05 were considered as statistically significant, unless otherwise indicated as NS. progressed into adulthood (20C40 years old) but then decreased during further progression toward old age ( 60 years old). The lowered numbers of circulating MAIT cells in the elderly was correlated with a gradual increase of apoptosis. A majority of circulating MAIT cells expressed the chemokine receptors CCR5 and CCR6, and most also expressed CD8 and CD45RO. Few expressed CD69 in cord blood, but the frequency increased with age. Upon activation with PMA plus ionomycin or IL12 plus IL18, fewer MAIT cells isolated from the young adult group expressed IFN-, IL17A and Granzyme B then cells from other age groups while the proportion of cells that expressed TNF- was comparable. Taken together, our data provide information for guiding the assessment of normal levels and phenotypes of MAIT cells at different ages in healthy individuals and patients. 0.05 are considered as statistically significant (* 0.05, ** 0.01, *** 0.001, **** 0.0001). Results Increased Circulating MAIT Cell Frequency From CB to Young Subjects, but Decreased From Young to Elderly Subjects Firstly, we defined human blood circulating MAIT cells as CD3+TCRV7.2+TCR?CD161hi cells by flow cytometry (Supplemental Physique 1) as suggested by a previous report (8). To determine how age may influence the frequency of circulating MAIT cells in humans, we examined MAIT cells in blood samples from D2PM hydrochloride 379 healthy individuals, which included 13 cord blood, 100 children (under 14 years old), 90 youths (20C40 years old), 88 middle-age persons (41C60 years old), 88 elderly (above 60 years old) (Table 1). The frequencies of V7.2+CD161hi MAIT cells in the CD3+TCR? population progressively increased when comparison is made from groups of CB to youth, at a respective average frequency of 0.09, 1.17, and 2.88% in the CB, Children and Youth groups. However, MAIT cell frequencies progressively decreased from groups of youth to elderly, at a respective average frequency of 2.88, 2.18, and 1.42% in the youth, middle-age, and elderly groups (Figures 1A,B). A similar trend was observed in the MAIT cell frequencies as relative to whole PBMCs (CB, mean SEM: 0.01 0.003%; Children, 0.75 0.08%; Youth, 1.51 0.13%; Middle-age, 1.09 0.12%; and Elderly, 0.56 0.07%) (Physique 1C). Corresponding to the changes in frequency, the numbers of MAIT cells increased from CB to youth, and then decreased from youth to elderly (CB, 0.076 0.017; Children, 2.78 0.31; Youth, 3.92 0.34; Middle-age, 2.6 0.29; and Elderly, 1.53 0.19 104/ml) (Figure 1D). Therefore, both the percentage and number of MAIT cells are very low in cord blood, increase during childhood, peak during youth, and then progressively decreased from middle to old age. Open in a separate window Physique 1 Circulating MAIT cell and CD3+ T cell frequencies and numbers in different cohorts. Freshly isolated PBMCs from 379 healthy individuals (grouped as shown in Table 1) were analyzed by flow cytometry. MAIT cells were gated as 7-AAD-TCR? CD3+TCRV7.2+CD161hi. (A) Representative FACS plots showing TCRV7.2 and CD161 expression in live gated TCR?CD3+ Mouse monoclonal to IHOG cells. Numbers adjacent to the rectangles are percentages within D2PM hydrochloride live gated TCR?CD3+ cells. (B) MAIT cell percentages in CD3+ TCR? T cells. (C) MAIT cell percentages in viable D2PM hydrochloride PBMCs. (D) Absolute MAIT cell numbers in PBMCs per milliliter of blood. (E) CD3+ cell percentages in viable PBMCs. (F) CD3+ cells absolute Number. Each symbol represents an individual subject. Statistical significance was assessed using the Mann-Whitney 0.05 were considered as statistically significant, unless otherwise indicated as NS (NS, Not significant). To determine whether age-associated changes.