Brain, Behavior, and Immunity ( IF 15.1 ) Pub Date : 2021-08-26 , DOI: 10.1016/j.bbi.2021.08.230 Xiaojin Feng 1 , Fenfang Zhan 2 , Deqiang Luo 3 , Jialing Hu 4 , Gen Wei 4 , Fuzhou Hua 5 , Guohai Xu 5
Objective
Cognitive impairment is a common neurological disease of which NLRP3-related neuroinflammation has been demonstrated to be an essential mediator. Previous studies have indicated that long non-coding RNAs (lncRNAs) are critical for the development of neurological disorders. However, the roles and functions of lncRNA 4344 in neuroinflammation during cognitive impairment are unknown and need to be further elucidated.
Methods
Lipopolysaccharide (LPS)-induced rat cognitive impairment and rat microglia (RM) cell inflammation models were established in vitro and in vivo. The Morris water maze test was used to evaluate the cognitive behavior of the rats. Gene expression was assessed using real-time quantitative polymerase chain reaction, and protein levels using enzyme-linked immunosorbent assay, or western blot analysis. The targeting relationship between lncRNA 4344, miR-138-5p, and NLRP3 was identified using bioinformatics analysis and a dual-luciferase reporter gene assay. Hematoxylin-Eosin and Nissl stainings, terminal deoxynucleotidyl transferase dUTP nick end labeling, or immunofluorescence staining assays were performed to detect pathological changes, neuronal apoptosis, or positive cells in hippocampal tissues, respectively.
Results
The expression levels of lncRNA 4344 and NLRP3 were upregulated in the hippocampal tissues of LPS-treated rats and RM cells, and showed a strong positive correlation between each other. LncRNA 4344 overexpression further enhanced the expression of NLRP3 and its downstream genes (caspase-1, IL-1β, and IL-18), as well as neuronal apoptosis in LPS-stimulated RM cells, whereas lncRNA 4344 silencing attenuated the inflammatory injuries. Moreover, miR-138-5p was the direct target of lncRNA 4344 and was downregulated in the RM cell inflammation model. We also found that miR-138-5p directly reduced the expression of NLRP3 and its downstream genes. Subsequently, the results of the animal experiments showed that the lncRNA 4344/miR-138-5p/NLRP3 axis plays an essential role in regulating the cognitive behavior, pathological changes and apoptosis of hippocampal neurons, expression of inflammation-related factors (NLRP3, caspase-1, IL-1β, and IL-18), and microglial activation in LPS-induced cognitive impairment rats.
Conclusion
Our results demonstrated for the first time that lncRNA 4344 regulates NLRP3-related neuroinflammation and cognitive impairment by targeting miR-138-5p, providing a possible target for the treatment of diseases characterized by a cognitive deficit.
中文翻译:
LncRNA 4344 通过靶向 miR-138-5p 促进 NLRP3 相关的神经炎症和认知障碍
客观的
认知障碍是一种常见的神经系统疾病,NLRP3 相关的神经炎症已被证明是一种重要的介质。先前的研究表明,长链非编码 RNA (lncRNA) 对神经系统疾病的发展至关重要。然而,lncRNA 4344在认知障碍期间神经炎症中的作用和功能尚不清楚,需要进一步阐明。
方法
在体外和体内建立脂多糖(LPS)诱导的大鼠认知障碍和大鼠小胶质细胞(RM)细胞炎症模型. Morris水迷宫测试用于评估大鼠的认知行为。使用实时定量聚合酶链反应评估基因表达,使用酶联免疫吸附测定或蛋白质印迹分析评估蛋白质水平。使用生物信息学分析和双荧光素酶报告基因分析确定了 lncRNA 4344、miR-138-5p 和 NLRP3 之间的靶向关系。进行苏木精-伊红和尼氏染色、末端脱氧核苷酸转移酶 dUTP 缺口末端标记或免疫荧光染色测定,分别检测海马组织中的病理变化、神经元凋亡或阳性细胞。
结果
lncRNA 4344和NLRP3的表达水平在LPS处理的大鼠和RM细胞的海马组织中上调,并且彼此之间呈强正相关。LncRNA 4344 过表达进一步增强了 NLRP3 及其下游基因(caspase-1、IL-1β 和 IL-18)的表达,以及 LPS 刺激的 RM 细胞中的神经元凋亡,而 lncRNA 4344 沉默减轻了炎症损伤。此外,miR-138-5p 是 lncRNA 4344 的直接靶标,在 RM 细胞炎症模型中下调。我们还发现 miR-138-5p 直接降低了 NLRP3 及其下游基因的表达。随后,动物实验结果表明,lncRNA 4344/miR-138-5p/NLRP3轴在调节认知行为中起着至关重要的作用,
结论
我们的研究结果首次证明,lncRNA 4344 通过靶向 miR-138-5p 调节 NLRP3 相关的神经炎症和认知障碍,为治疗以认知缺陷为特征的疾病提供了可能的靶点。