Calcitonin gene-related peptide protects type II alveolar epithelial cells from hyperoxia-induced DNA damage and cell death.
Calcitonin gene-related peptide protects type II alveolar epithelial cells from hyperoxia-induced DNA damage and cell death.
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DOI：
10.3892/etm.2017.4132
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发表时间：
2017-04
影响因子：
2.7
通讯作者：
Xu F
中科院分区：
文献类型：
--
作者：
Fu H;Zhang T;Huang R;Yang Z;Liu C;Li M;Fang F;Xu F
急性肺损伤 (ALI) 的高氧治疗可能会意外地导致活性氧 (ROS) 产生并导致额外的 ALI。降钙素基因相关肽 (CGRP) 是一种 37 个氨基酸的神经肽，可调节炎症小体的激活。然而，CGRP 在高氧期间 DNA 损伤中的作用尚不清楚。因此，本研究的目的是探讨 CGRP 对暴露于 60% 氧气的 II 型肺泡上皮细胞 (AEC II) 的 DNA 损伤和细胞死亡的影响。 AEC II 从妊娠 19-20 天的胎鼠肺中分离出来，并在 CGRP 或特异性 CGRP 受体拮抗剂 CGRP8-37 治疗期间暴露于空气或 60% 的氧气中。使用免疫荧光评估细胞以检查表面活性剂蛋白-C，并通过用 2',7'-二氯荧光素二乙酸酯探测来测量 ROS 水平。通过流式细胞仪分析AEC II的凋亡率和细胞周期，通过活化的caspase 3的蛋白质印迹分析测定细胞凋亡。通过高内涵分析通过H2AX免疫荧光证实DNA损伤。与空气组相比，高氧组ROS水平、凋亡细胞数和γH2AX表达显着升高。与之相一致的是，与高氧组或 CGRP8-37/O2 组相比，CGRP/O2 组的 ROS 水平、凋亡细胞数和 γH2AX 的表达显着降低，并且 S 期和 G2/M 期显着停滞。结论是 CGRP 可以保护肺上皮细胞免受高氧损伤，并且 CGRP 的上调可能是治疗高氧肺损伤的新治疗靶点。
Hyperoxia therapy for acute lung injury (ALI) may unexpectedly lead to reactive oxygen species (ROS) production and cause additional ALI. Calcitonin gene-related peptide (CGRP) is a 37 amino acid neuropeptide that regulates inflammasome activation. However, the role of CGRP in DNA damage during hyperoxia is unclear. Therefore, the aim of the present study was to investigate the effects of CGRP on DNA damage and the cell death of alveolar epithelial type II cells (AEC II) exposed to 60% oxygen. AEC II were isolated from 19–20 gestational day fetal rat lungs and were exposed to air or to 60% oxygen during treatment with CGRP or the specific CGRP receptor antagonist CGRP8–37. The cells were evaluated using immunofluorescence to examine surfactant protein-C and ROS levels were measured by probing with 2′,7′-dichlorofluorescin diacetate. The apoptosis rate and cell cycle of AEC II were analyzed by flow cytometry, and apoptosis was determined by western blotting analysis of activated caspase 3. The DNA damage was confirmed with immunofluorescence of H2AX via high-content analysis. The ROS levels, apoptotic cell number and the expression of γH2AX were markedly increased in the hyperoxia group compared with those in the air group. Concordantly, ROS levels, apoptotic cell number and the expression of γH2AX were significantly lower with a significant arrest of S and G2/M phases in the CGRP/O2 group than in the hyperoxia or CGRP8–37/O2 groups. CGRP was concluded to protect lung epithelium cells against hyperoxic insult, and upregulation of CGRP may be a possible novel therapeutic target to treat hyperoxic lung injury.