Lee Won-Woo, Nam Ki-Hoan, Terao Keiji, Yoshikawa Yasuhiro
Tsukuba Primate Centre, National Institute of Infectious Diseases, 1 Hachimandai, Tsukuba, Ibaraki 305-0843, Japan.
Immunology. 2002 Apr;105(4):458-65. doi: 10.1046/j.1365-2567.2002.01386.x.
Telomere length is a good biomarker to study the cellular senescence as well as aging of an organism, because it regulates the replicative capacity of vertebrate somatic cells. To demonstrate age-related telomere length dynamics in the peripheral blood mononuclear cells (PBMC) of the cynomolgus monkey, we introduced a novel method of measuring telomere length by fluorescence in situ hybridization with a Peptide Nucleic Acid (PNA) labelled probe and flow cytometry (Flow FISH). A highly significant correlation was observed between the intensity of telomere-specific fluorescence by Flow FISH and telomere length by Southern blot analysis (R = 0.923, n = 22). The intensity of telomere fluorescence in PBMC significantly decreased with age in 55 monkeys aged from 0 to 34 years and this decrease corresponded to the loss of 62.7 base pairs per year (R = - 0.52, P < 0.00004). We also analysed the expression of naive cell-associated markers, CD28, CD62L and CD45RA/CD62L in T lymphocytes of 47 cynomolgus monkeys. An age-related increase in the CD28- subset was observed in CD8+ T lymphocytes in monkeys less than 11 years old and in CD4+ T lymphocytes in monkeys over 23 years old, respectively. The percentage of CD62L+ subsets was significantly decreased with age in both CD4+ (R = - 0.55) and CD8+ T lymphocytes (R = - 0.73). From the comparison of telomere length among PBMC, CD62L+ and CD62L- T lymphocytes, it was clearly evident that loss of naive subsets results in the shortening of telomere length in vivo. These results show that this method can be applicable to studying the turnover and precursor-progeny of PBMC in cynomolgus monkeys as an animal model of aging.
端粒长度是研究细胞衰老以及生物体衰老的良好生物标志物，因为它调节脊椎动物体细胞的复制能力。为了证明食蟹猴外周血单核细胞（PBMC）中端粒长度与年龄相关的动态变化，我们引入了一种新方法，即使用肽核酸（PNA）标记探针进行荧光原位杂交和流式细胞术（流式荧光原位杂交）来测量端粒长度。通过流式荧光原位杂交检测到的端粒特异性荧光强度与Southern印迹分析测得的端粒长度之间存在高度显著的相关性（R = 0.923，n = 22）。在55只年龄从0到34岁的猴子中，PBMC中端粒荧光强度随年龄显著降低，这种降低相当于每年损失62.7个碱基对（R = -0.52，P < 0.00004）。我们还分析了47只食蟹猴T淋巴细胞中与幼稚细胞相关的标志物CD28、CD62L和CD45RA/CD62L的表达。分别在小于11岁的猴子的CD8 + T淋巴细胞和大于23岁的猴子的CD4 + T淋巴细胞中观察到CD28 - 亚群随年龄增加。CD62L + 亚群的百分比在CD4 + （R = -0.55）和CD8 + T淋巴细胞（R = -0.73）中均随年龄显著降低。通过比较PBMC、CD62L + 和CD62L - T淋巴细胞之间的端粒长度，明显可以看出幼稚亚群的丢失导致体内端粒长度缩短。这些结果表明，该方法可用于研究作为衰老动物模型的食蟹猴PBMC的更新和前体 - 子代关系。
