Department of Pharmacology, Uniformed Services University of the Health Sciences, Bethesda, Maryland, United States of America.
PLoS One. 2013 Oct 24;8(10):e78589. doi: 10.1371/journal.pone.0078589. eCollection 2013.
Accelerated senescence is a primary response to cellular stresses including DNA damaging agents (e.g., ionizing radiation) and is widely believed to be caused by continuous proliferative signaling in the presence of cell cycle arrest. Studies of signal transduction pathways leading to accelerated senescence have revealed that inhibition of mammalian target of rapamycin (mTOR) by rapamycin rescues cells from senescence. However, the molecular mechanisms upstream of mTOR following ionizing radiation (IR) are not well defined. We investigated signal transduction leading to IR-induced accelerated senescence in human pulmonary artery endothelial cells (HPAEC). Exposure of HPAEC to X-rays (10 Gy, 2.4 Gy/min) upregulated senescence markers including p53, p21/waf1, and senescence-associated beta galactosidase (SA-β-gal). Ly294002 (a phosphatidylinositol-3-kinase [PI3K] inhibitor) or rapamycin (an mTOR inhibitor) blocked the induction of cellular senescence markers suggesting roles for PI3K and mTOR. Pathway-directed microarrays revealed increased transcription of insulin-like growth factor I (IGF-1), a modulator of cell growth and proliferation upstream of mTOR. qRT-PCR confirmed that both IGF-1 and IGF-2 mRNA were increased in response to X-rays, and ELISA showed increased secretion of IGF-1 protein into the medium of irradiated HPAEC. Consistent with upregulation of these ligands, we found that X-ray exposure led to hyperphosphorylation of IGF-1R, the receptor for IGF-1 and -2. Treatment with AG1024, an IGF-1R inhibitor, suppressed IR-induced upregulation of p53, p21/waf1, and SA-β-gal. Together these findings suggest that IGF-1R is a key regulator of IR-induced accelerated senescence in a pathway that requires intact mTOR activity upstream of both p53 and p21/waf1.
加速衰老（Accelerated senescence）是细胞应激（包括 DNA 损伤剂，如电离辐射）的主要反应，广泛认为是由于细胞周期停滞时持续的增殖信号导致的。对导致加速衰老的信号转导途径的研究表明，雷帕霉素（rapamycin）抑制哺乳动物雷帕霉素靶蛋白（mTOR）可使细胞免于衰老。然而，电离辐射（IR）后 mTOR 上游的分子机制尚不清楚。我们研究了导致人肺动脉内皮细胞（HPAEC）IR 诱导加速衰老的信号转导。用 X 射线（10Gy，2.4Gy/min）照射 HPAEC 可上调衰老标志物，包括 p53、p21/waf1 和衰老相关β-半乳糖苷酶（SA-β-gal）。Ly294002（一种磷酸肌醇-3-激酶 [PI3K] 抑制剂）或雷帕霉素（mTOR 抑制剂）阻断细胞衰老标志物的诱导，表明 PI3K 和 mTOR 发挥作用。定向途径微阵列显示胰岛素样生长因子 I（IGF-1）的转录增加，IGF-1 是 mTOR 上游细胞生长和增殖的调节剂。qRT-PCR 证实 X 射线照射后 IGF-1 和 IGF-2 mRNA 均增加，ELISA 显示照射的 HPAEC 培养基中 IGF-1 蛋白分泌增加。与这些配体的上调一致，我们发现 X 射线照射导致 IGF-1R 过度磷酸化，IGF-1R 是 IGF-1 和 -2 的受体。用 IGF-1R 抑制剂 AG1024 处理可抑制 IR 诱导的 p53、p21/waf1 和 SA-β-gal 上调。这些发现表明，IGF-1R 是一个关键调节因子，在 mTOR 活性完整的情况下，通过 p53 和 p21/waf1 调节 IR 诱导的加速衰老。
