Isolation and identification of senescent renal tubular epithelial cells using immunomagnetic beads based on DcR2
Isolation and identification of senescent renal tubular epithelial cells using immunomagnetic beads based on DcR2
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基于DcR2的免疫磁珠分离鉴定衰老肾小管上皮细胞
DOI：
10.1016/j.exger.2017.04.008
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发表时间：
2017-09-01
影响因子：
3.9
通讯作者：
He, Ya-Ni
中科院分区：
文献类型：
--
作者：
Chen, Jia;Chen, Ke-Hong;He, Ya-Ni
细胞衰老在肿瘤和慢性疾病（如糖尿病和慢性肾脏疾病）的进展中起着重要作用。衰老细胞是研究衰老相关疾病的重要模型，目前尚无有效的方法对衰老细胞进行分选。诱饵受体2 （Decoy receptor 2, DcR2）是肿瘤坏死因子超家族的跨膜受体，在衰老细胞中特异性表达。在这项研究中，我们使用磁激活细胞分选（MACS）分离高纯度的dcr2阳性肾小管上皮细胞（RTECs），基于三种衰老细胞模型，包括第五代细胞、晚期糖基化终产物（AGEs）和h2o2诱导细胞。与预先分选的细胞相比，G1期和S期DcR2阳性RTECs的百分比分别增加了20%和4%。在dcr2阳性rtec中，SA-13-gal、p16和衰老相关异染色质病灶（SAHF）的阳性率分别比细胞分选前高40%、30%和44%。与分选前相比，上清液中IL-6和tgf - β 1的水平分别增加了1.7倍和1.5倍。连续培养5天后，未见明显细胞死亡。MACS后，DcR2阴性RTECs中Ki-67阳性表达率显著高于DcR2阳性RTECs。我们证明了使用DcR2对活的、衰老的rtec进行分类，具有高特异性和稳定性。我们的发现为进一步研究衰老rtec在慢性肾脏疾病进展中的作用奠定了基础。(C) 2017 Elsevier Inc版权所有
Cell senescence plays a major role in the progression of tumors and chronic conditions such as diabetes and chronic kidney disease. Senescent cells are an important model for the study of aging-related diseases, and there is currently no efficient method for sorting out senescent cells. Decoy receptor 2 (DcR2) is a transmembrane receptor of the tumor necrosis factor superfamily, which is specifically expressed in senescent cells. In this study, we used magnetic activated cell sorting (MACS) isolation of a highly-pure populations DcR2-positive renal tubular epithelial cells (RTECs) based on three senescent cell models including the fifth passage cells, advanced glycation end-products (AGEs)-and H2O2-induced cells. The percentages of DcR2 positive RTECs in G1 and S phases increased by 20% and 4%, respectively, as compared to that in the pre-sorted cells. The positivity rates of SA-13-gal, p16, and senescence-associated heterochromatin foci (SAHF) in DcR2-positive RTECs were about 40%, 30%, and 44% higher than that prior to cell sorting. The levels of IL-6 and TGF-beta 1 in the supernatant were increased by 1.7 and 1.5 folds, respectively, as compared to that observed prior to sorting. No significant cell death was observed after 5 days of continuous culture. Ki-67 positive expression rate in DcR2 negative RTECs was significantly higher than that in DcR2 positive RTECs after MACS. We demonstrated the use of DcR2 to classify live, senescent RTECs with a high specificity and stability. Our findings lay the foundation for further study of senescent RTECs in the progression of chronic kidney disease. (C) 2017 Elsevier Inc All rights reserved.