miR-34a 通过调控 Cdc42-WASP-Arp2/3 通路介导的 F-actin 重塑和 ROS 产生诱导中性粒细胞凋亡。
miR-34a induces neutrophil apoptosis by regulating Cdc42-WASP-Arp2/3 pathway-mediated F-actin remodeling and ROS production.
机构信息
Department of Gastroenterology, Guangzhou Women and Children's Medical Center, Guangzhou Medical University, Guangzhou, People's Republic of China.
Center for child health and mental health, Shenzhen Childen's Hospital, Shenzhen, People's Republic of China.
出版信息
Redox Rep. 2022 Dec;27(1):167-175. doi: 10.1080/13510002.2022.2102843.
BACKGROUND
The number of neutrophils is significantly reduced in myelodysplastic syndrome (MDS), but the molecular basis remains unclear. We recently found that miR-34a was significantly increased in MDS neutrophils. Therefore, this study aims to clarify the effects of aberrant miR-34a expression on neutrophil counts.
METHODS
miR-34a mimics/inhibitor transfection were performed in neutrophil-like differentiated HL60 (dHL60) cells, and a FACSCalibur flow cytometer was used to measure ROS production and apoptosis. In addition, the Cdc42-WASP-Arp2/3 pathway inhibitor (ML141) and activator (CN02) treated the dHL60 cells, and then ROS production, apoptosis and related proteins expression were detected. And, luciferase reporter assay to verify the relationship of miR-34a and the Cdc42-WASP-Arp2/3 pathway.
RESULTS
overexpression of miR-34a could induce ROS production and apoptosis, decrease the expression levels of DOCK8, p-WASP, WASP, Arp2, Arp3, and increase F-actin's expression. Meanwhile, knockdown of miR-34a could decrease ROS production and apoptosis, increase the expression of DOCK8, p-WASP, WASP, Arp2, Arp3, and decrease F-actin's expression. Immunofluorescence staining showed aberrant miR-34a and Cdc42-WASP-Arp2/3 pathway could induce F-actin membrane transfer. Luciferase reporter assay indicated that DOCK8 was a direct target gene of miR-34a.
CONCLUSION
These data indicates miR-34a may induce neutrophil apoptosis by regulating Cdc42-WASP-Arp2/3 pathway-mediated F-actin remodeling and ROS production.
背景
骨髓增生异常综合征（MDS）患者中性粒细胞数量显著减少，但分子机制尚不清楚。我们最近发现 miR-34a 在 MDS 中性粒细胞中显著上调。因此，本研究旨在阐明异常 miR-34a 表达对中性粒细胞计数的影响。
方法
在 HL60 细胞（dHL60）中进行 miR-34a 模拟物/抑制剂转染，使用 FACSCalibur 流式细胞仪测量 ROS 产生和细胞凋亡。此外，用 Cdc42-WASP-Arp2/3 通路抑制剂（ML141）和激活剂（CN02）处理 dHL60 细胞，然后检测 ROS 产生、细胞凋亡和相关蛋白表达。并通过荧光素酶报告基因实验验证 miR-34a 与 Cdc42-WASP-Arp2/3 通路的关系。
结果
miR-34a 的过表达可诱导 ROS 产生和细胞凋亡，降低 DOCK8、p-WASP、WASP、Arp2、Arp3 的表达水平，并增加 F-actin 的表达。相反，miR-34a 的下调可减少 ROS 产生和细胞凋亡，增加 DOCK8、p-WASP、WASP、Arp2、Arp3 的表达，并降低 F-actin 的表达。免疫荧光染色显示异常 miR-34a 和 Cdc42-WASP-Arp2/3 通路可诱导 F-actin 膜转移。荧光素酶报告基因实验表明 DOCK8 是 miR-34a 的直接靶基因。
结论
这些数据表明，miR-34a 可能通过调节 Cdc42-WASP-Arp2/3 通路介导的 F-actin 重塑和 ROS 产生诱导中性粒细胞凋亡。
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