长链非编码 RNA MEG3 通过调控 miR-130a-5p/CXCL12/CXCR4 轴加重神经病理性疼痛和星形胶质细胞过度激活。

lncRNA MEG3 aggravated neuropathic pain and astrocyte overaction through mediating miR-130a-5p/CXCL12/CXCR4 axis.

机构信息

Department of Anesthesiology, Qianjiang Hospital Affiliated to Renmin Hospital of Wuhan University, Qianjiang 433100, Hubei, China.

Department of Anesthesiology, The First People's Hospital of Jingzhou, Jingzhou 434000, Hubei, China.

出版信息

Aging (Albany NY). 2021 Oct 5;13(19):23004-23019. doi: 10.18632/aging.203592.

DOI:10.18632/aging.203592

PMID:34609952

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8544300/

Abstract

OBJECTIVE

Long non-coding RNAs (lncRNAs) exert a critical function in mediating neuropathic pain (NP). MEG3, a novel lncRNA, contributes to astrocyte activation and inflammation. However, its role in NP remains unclear.

METHODS

The chronic constriction injury (CCI) method was employed to construct an NP rat model. Astrocyte activation was induced by lipopolysaccharide (LPS). The profiles of MEG3, microRNA (miR)-130a-5p, CXC motif chemokine receptor 12 (CXCL12)/CXC motif chemokine receptor 4 (CXCR4), and the Rac1/NF-κB pathway in CCI rats' spinal cord tissues and astrocytes were monitored by reverse transcription-quantitative PCR (RT-qPCR) and western blot (WB). Pain scores of CCI rats were assessed. Enzyme-linked immunosorbent assay (ELISA) was adopted to monitor neuroinflammation alteration. The glial fibrillary acidic protein (GFAP)-labeled astrocytes were tested by immunohistochemistry (IHC). Bioinformatics, dual-luciferase reporter assay and RNA immunoprecipitation (RIP) were utilized to verify the molecular mechanism between MEG3 and miR-130a-3p.

RESULTS

MEG3, CXCL12 and CXCR4 were overexpressed and miR-130a-5p was knocked down in CCI rats and LPS-induced astrocytes. Up-regulating MEG3 aggravated NP, enhanced inflammatory cytokines interleukin-1β (IL-1β), tumor necrosis factor (TNF)-α, and interleukin-6 (IL-6) expression and release in CCI rats and LPS-induced astrocytes. Up-regulating miR-130-5p repressed LPS-induced inflammation in astrocytes. AS verified by the dual-luciferase reporter assay and RIP assay, MEG3 sponged miR-130a-5p as a competitive endogenous RNA (ceRNA). What's more, miR-130a-5p up-regulation weakened the MEG3-induced proinflammatory effects on LPS-induced astrocytes.

CONCLUSIONS

MEG3 aggravates NP and astrocyte activation via the miR-130a-5p/CXCL12/CXCR4 axis, which is a potential therapeutic target for NP.

摘要

目的

长链非编码 RNA（lncRNA）在介导神经病理性疼痛（NP）中发挥关键作用。MEG3 是一种新的 lncRNA，有助于星形胶质细胞激活和炎症。然而，其在 NP 中的作用尚不清楚。

方法

采用慢性缩窄性损伤（CCI）方法构建 NP 大鼠模型。用脂多糖（LPS）诱导星形胶质细胞激活。通过逆转录定量 PCR（RT-qPCR）和蛋白质印迹（WB）监测 CCI 大鼠脊髓组织和星形胶质细胞中 MEG3、微小 RNA（miR）-130a-5p、CXC 基序趋化因子受体 12（CXCL12）/CXC 基序趋化因子受体 4（CXCR4）和 Rac1/NF-κB 通路的特征。评估 CCI 大鼠的疼痛评分。采用酶联免疫吸附试验（ELISA）监测神经炎症变化。免疫组织化学（IHC）检测胶质纤维酸性蛋白（GFAP）标记的星形胶质细胞。利用生物信息学、双荧光素酶报告基因检测和 RNA 免疫沉淀（RIP）验证 MEG3 与 miR-130a-3p 之间的分子机制。

结果

在 CCI 大鼠和 LPS 诱导的星形胶质细胞中，MEG3、CXCL12 和 CXCR4 表达上调，miR-130a-5p 下调。上调 MEG3 加重 NP，增强 CCI 大鼠和 LPS 诱导的星形胶质细胞中炎性细胞因子白细胞介素-1β（IL-1β）、肿瘤坏死因子（TNF）-α和白细胞介素-6（IL-6）的表达和释放。上调 miR-130-5p 抑制 LPS 诱导的星形胶质细胞炎症。双荧光素酶报告基因检测和 RIP 实验证实，MEG3 作为竞争性内源性 RNA（ceRNA）吸附 miR-130a-5p。此外，miR-130a-5p 的上调减弱了 MEG3 对 LPS 诱导的星形胶质细胞的促炎作用。