LncRNA Tug1 Contributes Post-stroke NLRP3 Inflammasome-Dependent Pyroptosis via miR-145a-5p/Tlr4 Axis.

Pyroptosis, a type of programmed cell death illuminated by inflammasomes and active caspases, is implicated in post-stroke inflammation. Our previous study showed that lncRNA taurine upregulated gene 1 (Tug1) sponging miR-145a-5p modulated microglial activation after oxygen-glucose deprivation (OGD). However, the role and mechanism of Tug1 on post-stroke pyroptosis is not fully clear. Photo-thrombosis stroke mice and OGD-treated BV-2 microglia were established respectively. Tug1 knockdown or overexpression was achieved by intraventricular infusion of AAV-shTug1 in vivo, or transfection of siTug1 and pcDNA3.1-Tug1 in vitro. Neurological function and infarction volume were evaluated. Meanwhile, pyroptosis-associated proteins (IL-1β, IL-18, NLRP3, ASC, cleaved-caspase-1, and GSDMD-N), TLR4, and p-p65/p65 as well as Tug1 and miR-145a-5p were detected 24 h after photo-thrombosis or 4 h after OGD by qRT-PCR, western blot, and ELISA. The correlation between Tug1/miR-145a-5p/Tlr4 axis and pyroptosis was explored by dual-luciferase reporter assay and functional gain-and-loss experiments. Photo-thrombosis or OGD caused neural injury and upregulated pyroptosis-associated proteins, Tug1, TLR4, and p-p65 as well as downregulated miR-145a-5p, which was prevented by Tug1 knockdown in vivo and in vitro. Tlr4 gene, putatively binding with miR-145a-5p by bioinformatics analysis, was found to be a direct target of miR-145a-5p with negative interactions. Furthermore, miR-145a-5p inhibitor abolished the inhibitive effects of siTug1 on TLR4 and p-p65 as well as pyroptosis-associated proteins, whereas miR-145a-5p mimics abrogated the enhanced effects of pcDNA3.1-Tug1 on that, suggesting an involvement of Tug1/miR-145a-5p/Tlr4 axis on pyroptosis. Tug1 contributes NLRP3 inflammasome-dependent pyroptosis through miR-145a-5p/Tlr4 axis post-stroke, providing a promising therapeutic strategy against inflammatory injury.

Yao M ，Luo Y ，Li H ，Liao S ，Yu J ... -  《-》

Long Non-coding RNA TUG1 Sponges Mir-145a-5p to Regulate Microglial Polarization After Oxygen-Glucose Deprivation.

Wang H ，Liao S ，Li H ，Chen Y ，Yu J ... -  《Frontiers in Molecular Neuroscience》

MiR-139 protects against oxygen-glucose deprivation/reoxygenation (OGD/R)-induced nerve injury through targeting c-Jun to inhibit NLRP3 inflammasome activation.

Cerebral ischemia/reperfusion (I/R) injury after ischemic stroke is usually accompanied with the activation of inflammasome which seriously impairs neurological function. MiR-139 has been reported to be associated with inflammatory regulation in multiple diseases. However, its effect and mechanism on inflammation regulation after cerebral I/R injury are still poorly understood. An in vitro model of cerebral I/R injury was constructed with oxygen-glucose deprivation/reoxygenation (OGD/R) treatment. TargetScan bioinformatics analysis and dual luciferase reporter assay were utilized to confirm the targeted relationship between miR-139 and c-Jun. Cell pyroptosis was verified by flow cytometry and Caspase-1 Detection Kit. qRT-PCR assay was performed to detect the expression levels of miR-139, c-Jun, NLRP3 and ASC. Western blotting was applied to measure the protein levels of c-Jun and pyroptosis-related markers NLRP3, ASC, caspase-1, GSDMDNterm. The ELISA assay was applied to measure the release of IL-1β, IL-18 and LDH. MiR-139 was significantly downregulated whereas c-Jun was obviously upregulated after OGD/R treatment. TargetScan analysis predicted that c-Jun was a potential target of miR-139, which was verified by the dual-luciferase reporter assay. Also, overexpression of miR-139 repressed c-Jun expression. Furthermore, miR-139 inhibited OGD/R-induced cell pyroptosis and the upregulation of NLRP3, caspase-1, ASC, GSDMDNterm, and the release of IL-1β, IL-18 and LDH, while miR-139 inhibition exerted the opposite effects. However, overexpression of c-Jun aggravated OGD/R-induced nerve injury and partly abolished the neuroprotective effect of miR-139. Upregulation of miR-139 exerted neuroprotection against OGD/R-induced nerve injury by negatively regulating c-Jun/NLRP3 inflammasome signaling. This study offered insights for providing potential therapeutic targets for treating cerebral I/R injury.

Wang QS ，Luo XY ，Fu H ，Luo Q ，Wang MQ ，Zou DY ... -  《-》

Overexpression of lncRNA TUG1 Alleviates NLRP3 Inflammasome-Mediated Cardiomyocyte Pyroptosis Through Targeting the miR-186-5p/XIAP Axis in Coronary Microembolization-Induced Myocardial Damage.

Coronary microembolization (CME) is a complicated problem that commonly arises in the context of coronary angioplasty. The lncRNA taurine-up regulated gene 1 (TUG1), significantly contributes to cardiovascular diseases; however, its contribution to CME-induced myocardial damage remains elusive. Herein, we establish the rat CME model and investigate the role of TUG1 in CME. The cell viability was evaluated via CCK-8 assay. Serum and cell culture supernatant samples were evaluated via ELISA. The dual luciferase reporter (DLR) assay, RIP, and RNA-pull down were conducted to validate the associations between TUG1 and miR-186-5p as well as miR-186-5p and XIAP. The expression of TUG1, miR-186-5p, and XIAP mRNA were determined by RT-qPCR, and proteins were evaluated via immuneblotting. As a result, TUG1 and XIAP were significantly down-regulated, and the miR-186-5p level was found to be remarkably up-regulated in CME myocardial tissues. Overexpression of TUG1 alleviated CME-induced myocardial injury and pyroptosis, whereas TUG1 knockdown showed the opposite effects. The DLR assay, RIP, and RNA-pull down results reveal that TUG1 directly targets miR-186-5p and miR-186-5p directly targets XIAP. In vitro rescue experiments show that TUG1 overexpression alleviates LPS-caused cardiomyocyte injury and pyroptosis via sponging miR-186-5p and regulating XIAP, and depression of miR-186-5p reduces LPS-induced cardiomyocyte injury and pyroptosis by targeting XIAP. Concludingly, the overexpression of TUG1 alleviates NLRP3 inflammasome-mediated cardiomyocyte pyroptosis through targeting the miR-186-5p/XIAP axis in CME-induced myocardial injury.

Zhou Y ，Li T ，Chen Z ，Huang J ，Qin Z ，Li L ... -  《Frontiers in Immunology》

LncRNA Rian reduces cardiomyocyte pyroptosis and alleviates myocardial ischemia-reperfusion injury by regulating by the miR-17-5p/CCND1 axis.

Myocardial ischemia-reperfusion injury (MIRI) is a pathological process characterized by cardiomyocyte death. Long noncoding RNAs (lncRNAs) have been shown to be dysregulated in the course of MIRI. Accordingly, the current study investigated the mechanism of lncRNA Rian in MIRI-induced cardiomyocyte pyroptosis. First, a murine model of MIRI was established by using the left anterior descending (LAD) coronary artery ligation method. Cardiac function and myocardial histopathological changes were evaluated by echocardiography and hematoxylin and eosin staining. Then, a cell model of MIRI was established by oxygen-glucose deprivation/reoxygenation (OGD/R), followed by analysis of NLRP3, cleaved caspase-1, and GSDMD-N levels by western blotting. The levels of IL-1β, IL-18, TNF-α, and IL-10 were measured using ELISA. LncRNA Rian, miR-17-5p, and CCND1 expression in myocardial tissues and OGD/R cells were examined using RT-qPCR. Finally, the binding relationships between Rian and miR-17-5p and miR-17-5p and CCND1 were validated with the help of dual-luciferase and RNA pull-down assays. Rian was poorly expressed in MIRI mice and OGD/R cells. LncRNA Rian overexpression reduced cardiomyocyte pyroptosis in vivo and in vitro, as indicated by decreased NLRP3, cleaved caspase-1, GSDMD-N, IL-1β, IL-18, and TNF-α levels and increased IL-10 levels. Furthermore, Rian bound to miR-17-5p and promoted CCND1 transcription. Notably, miR-17-5p overexpression or CCND1 silencing reversed the inhibitory effect of Rian overexpression on cardiomyocyte pyroptosis. Collectively, our findings indicate that Rian overexpression reduces cardiomyocyte pyroptosis and alleviates MIRI through the miR-17-5p/CCND1 axis.

Kang H ，Yu H ，Zeng L ，Ma H ，Cao G ... -  《-》