Department of anesthesiology, Tongde hospital of Zhejiang Province, Hangzhou, 310012, Zhejiang, China.
Cancer Institute of Integrated traditional Chinese and Western Medicine, Zhejiang Academy of Traditional Chinese Medicine, Tongde hospital of Zhejiang Province, NO.234, Gucui Road, Hangzhou, 310012, Zhejiang, China.
J Biomed Sci. 2020 Mar 6;27(1):40. doi: 10.1186/s12929-020-00635-0.
The present study aimed to verify whether long noncoding RNA (lncRNA) MALAT1 is involved in brain tissue damage induced by ischemia-reperfusion injury, and to explore the mechanism by which MALAT1 regulates aquaporin 4 (AQP4).
In this study, we established glucose deprivation (OGD)/reoxygenation (RX) astrocyte cell model and middle cerebral artery occlusion (MCAO)/reperfusion mouse model in vitro and in vivo. Then cell counting kit-8 assay, flow cytometry analysis, Triphenyltetrazolium chloride (TTC) staining, and western blotting were used to determine cell viability, cell apoptosis, cerebral infarction volume, and the abundance of AQP4, respectively.
We found that the level of MALAT1 was significantly upregulated in both the MCAO/reperfusion model and OGD/RX model. Knockdown of MALAT1 increased cell viability and reduced cell apoptosis in MA-C cells, while an AQP4 siRNA combined with a siRNA targeting MALAT1 could not enhance this effect. Further experiments showed that MALAT1 positively regulated AQP4 expression via miR-145. The MALAT1 siRNA did not alleviate the exacerbation of damage after miR-145 inhibitor action. However, an miR-145 inhibitor reversed the protection effects of MALAT1, indicating that MALAT1 silencing protects against cerebral ischemia-reperfusion injury through miR-145. TTC staining showed that the infracted area of whole brain was significantly attenuated in treated with sh-MALAT1 group in vivo.
Taken together, our study confirmed that MALAT1 promotes cerebral ischemia-reperfusion injury by affecting AQP4 expression through competitively binding miR-145, indicating that MALAT1 might be a new therapeutic target for treatment cerebral ischemic stroke.
本研究旨在验证长链非编码 RNA（lncRNA）MALAT1 是否参与缺血再灌注损伤引起的脑组织损伤，并探讨 MALAT1 调节水通道蛋白 4（AQP4）的机制。
本研究采用体外葡萄糖剥夺（OGD）/复氧（RX）星形胶质细胞模型和体内大脑中动脉闭塞（MCAO）/再灌注小鼠模型。然后，通过细胞计数试剂盒-8 检测、流式细胞术分析、三苯基四唑氯化物（TTC）染色和 Western blot 检测，分别测定细胞活力、细胞凋亡、脑梗死体积以及 AQP4 的丰度。
我们发现，在 MCAO/再灌注模型和 OGD/RX 模型中，MALAT1 的水平均显著上调。在 MA-C 细胞中，敲低 MALAT1 可增加细胞活力并减少细胞凋亡，而 AQP4 siRNA 与 MALAT1 siRNA 联合使用则不能增强这种作用。进一步的实验表明，MALAT1 通过 miR-145 正向调节 AQP4 的表达。MALAT1 siRNA 并不能减轻 miR-145 抑制剂作用后损伤的加剧。然而，miR-145 抑制剂逆转了 MALAT1 的保护作用，表明 MALAT1 沉默通过 miR-145 来保护脑缺血再灌注损伤。TTC 染色显示，体内给予 sh-MALAT1 后，全脑梗死面积明显减轻。
综上所述，我们的研究证实 MALAT1 通过竞争性结合 miR-145 影响 AQP4 的表达，从而促进脑缺血再灌注损伤，表明 MALAT1 可能成为治疗缺血性脑卒中的新治疗靶点。
