TGR5 激活通过 Pellino3 抑制 caspase-8/NLRP3 减轻大鼠大脑中动脉闭塞后的神经炎症。

TGR5 activation attenuates neuroinflammation via Pellino3 inhibition of caspase-8/NLRP3 after middle cerebral artery occlusion in rats.

机构信息

Department of Neurology, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

Department of Physiology and Pharmacology and Department of Anesthesiology, Loma Linda University, 11041 Campus St, Risley Hall, Room 219, Loma Linda, CA, 92354, USA.

出版信息

J Neuroinflammation. 2021 Feb 2;18(1):40. doi: 10.1186/s12974-021-02087-1.

BACKGROUND

Nucleotide-binding oligomerization domain-like receptor pyrin domain-containing protein 3 (NLRP3) plays an important role in mediating inflammatory responses during ischemic stroke. Bile acid receptor Takeda-G-protein-receptor-5 (TGR5) has been identified as an important component in regulating brain inflammatory responses. In this study, we investigated the mechanism of TGR5 in alleviating neuroinflammation after middle cerebral artery occlusion (MCAO).

METHODS

Sprague-Dawley rats were subjected to MCAO and TGR5 agonist INT777 was administered intranasally 1 h after MCAO. Small interfering RNAs (siRNA) targeting TGR5 and Pellino3 were administered through intracerebroventricular injection 48 h before MCAO. Infarct volumes and neurologic scores were evaluated, and ELISA, flow cytometry, immunofluorescence staining, immunoblotting, and co-immunoprecipitation were used for the evaluations.

RESULTS

Endogenous TGR5 and Pellino3 levels increased after MCAO. TGR5 activation by INT777 significantly decreased pro-inflammatory cytokine, cleaved caspase-8, and NLRP3 levels, thereby reducing brain infarctions; both short- and long-term neurobehavioral assessments showed improvements. Ischemic damage induced the interaction of TGR5 with Pellino3. Knockdown of either TGR5 or Pellino3 increased the accumulation of cleaved caspase-8 and NLRP3, aggravated cerebral impairments, and abolished the anti-inflammatory effects of INT777 after MCAO.

CONCLUSIONS

TGR5 activation attenuated brain injury by inhibiting neuroinflammation after MCAO, which could be mediated by Pellino3 inhibition of caspase-8/NLRP3.

背景

核苷酸结合寡聚化结构域样受体含pyrin 结构域蛋白 3（NLRP3）在介导缺血性脑卒中期间的炎症反应中发挥重要作用。胆酸受体 Takeda-G 蛋白受体 5（TGR5）已被确定为调节大脑炎症反应的重要组成部分。在这项研究中，我们研究了 TGR5 在减轻大脑中动脉闭塞（MCAO）后神经炎症中的作用机制。

方法

Sprague-Dawley 大鼠接受 MCAO，MCAO 后 1 小时经鼻内给予 TGR5 激动剂 INT777。MCAO 前 48 小时通过脑室内注射小干扰 RNA（siRNA）靶向 TGR5 和 Pellino3。通过酶联免疫吸附试验、流式细胞术、免疫荧光染色、免疫印迹和免疫共沉淀进行评估。

结果

MCAO 后内源性 TGR5 和 Pellino3 水平增加。INT777 激活 TGR5 可显著降低促炎细胞因子、裂解的半胱天冬酶-8 和 NLRP3 水平，从而减少脑梗死；短期和长期神经行为评估均显示出改善。缺血性损伤诱导 TGR5 与 Pellino3 的相互作用。TGR5 或 Pellino3 的敲低均增加了裂解的半胱天冬酶-8 和 NLRP3 的积累，加重了脑损伤，并消除了 MCAO 后 INT777 的抗炎作用。