Department of Orthopedics, Dongzhimen Hospital of Beijing University of Chinese Medicine, Beijing, P.R. China.
Eur Rev Med Pharmacol Sci. 2020 Jan;24(2):517-525. doi: 10.26355/eurrev_202001_20027.
This study aimed to research the effect of miR-202-5p-mediated ATG7 on autophagy and apoptosis of degenerative nucleus pulposus cells.
The intervertebral disc nucleus pulposus (NP) tissue of patients with intervertebral disc degenerative disease and normal intervertebral disc nucleus pulposus (NP) tissue of patients with spinal fractures was collected as the research object. Normal NP cells and degenerative NP cells were isolated. Low expression of miR-202-5p and overexpression of ATG7 were carried out in degenerative NP cells. The expression of miR-202-5p and ATG7 mRNA was detected by RT-PCR. The expression of ATG7, LC3-II, Bax, and Bcl-2 proteins was detected by Western blot. The autophagy of cells was detected by MDC staining. The apoptosis of NP cells was detected by flow cytometry. The targeting relationship between miR-202-5p and ATG7 was detected by Dual-Luciferase reporter.
In the degenerative NP tissues, miR-202-5p was highly expressed and ATG7 was low expressed. The inhibition of miR-202-5p expression can effectively promote autophagy of NP cells, increase the expression of ATG7 and LC3-II, inhibit the apoptosis of NP cells, inhibit the expression of pro-apoptotic proteins Bax, and promote the expression of pro-apoptotic proteins Bcl-2 proteins. The upregulation of ATG7 expression in degenerative NP cells alone had the same effect as the downregulation of miR-202-5p. The assay of the Dual-Luciferase reporter confirmed the targeting relationship between miR-202-5p and ATG7.
MiR-202-5p can affect the autophagy and apoptosis of degenerative nucleus pulposus cells through targeted adjustment of ATG7, which may be a new therapeutic target for intervertebral disc degenerative diseases.
本研究旨在探讨 miR-202-5p 介导的 ATG7 对退变髓核细胞自噬和凋亡的影响。
收集椎间盘退行性疾病患者的椎间盘髓核组织和脊柱骨折患者的正常椎间盘髓核组织作为研究对象。分离正常 NP 细胞和退变 NP 细胞。在退变 NP 细胞中低表达 miR-202-5p 并过表达 ATG7。通过 RT-PCR 检测 miR-202-5p 和 ATG7mRNA 的表达。通过 Western blot 检测 ATG7、LC3-II、Bax 和 Bcl-2 蛋白的表达。通过 MDC 染色检测细胞自噬。通过流式细胞术检测 NP 细胞凋亡。通过双荧光素酶报告检测 miR-202-5p 与 ATG7 的靶向关系。
在退变的 NP 组织中，miR-202-5p 高表达，ATG7 低表达。抑制 miR-202-5p 的表达能有效促进 NP 细胞的自噬，增加 ATG7 和 LC3-II 的表达，抑制 NP 细胞的凋亡，抑制促凋亡蛋白 Bax 的表达，促进促凋亡蛋白 Bcl-2 蛋白的表达。单独上调退变 NP 细胞中的 ATG7 表达具有与下调 miR-202-5p 相同的作用。双荧光素酶报告检测证实了 miR-202-5p 与 ATG7 之间的靶向关系。
miR-202-5p 可通过靶向调节 ATG7 影响退变髓核细胞的自噬和凋亡，可能成为治疗椎间盘退行性疾病的新靶点。
