【摘要】 目的:探讨长链非编码RNA(LncRNA)KCNQ1OT1调节微小RNA(miR)-214-3p/N-myc下游调控基因3(NDRG3)轴对脑出血(ICH)模型大鼠神经功能的影响。方法:将SD大鼠分为假手术(Sham)组、ICH组、LncRNA KCNQ1OT1干扰质粒对照(sh-NC)组、LncRNA KCNQ1OT1干扰质粒(sh-KCNQ1OT1)组、sh-KCNQ1OT1+miR-214-3p抑制剂对照(miR-In-NC)组和sh-KCNQ1OT1+miR-214-3p抑制剂(miR-214-3p-In)组，每组12只。改良神经功能缺损评分(mNSS)评估各组大鼠神经功能变化；测量各组大鼠脑部血肿体积；HE染色观察各组大鼠脑组织病理变化；TUNEL染色观察脑组织细胞凋亡；免疫组织化学染色观察NDRG3表达量变化；ELISA检测大鼠脑组织中TNF-α和IL-1β水平；qRT-PCR检测各组大鼠脑组织中LncRNA KCNQ1OT1、miR-214-3p和NDRG3mRNA水平；Western blot检测各组大鼠脑组织中NDRG3、Bax和Bcl-2蛋白表达水平；双荧光素酶报告基因实验测定miR-214-3p与LncRNA KCNQ1OT1或NDRG3的靶向关系。结果:与Sham组相比，ICH组大鼠mNSS评分、脑血肿体积和脑组织细胞凋亡率显著增加(P<0.05),脑组织结构被破坏，红细胞、炎性细胞浸润和神经胶质细胞的数量增加，脑组织中TNF-α、IL-1β、LncRNA KCNQ1OT1、NDRG3和Bax表达水平显著升高(P<0.05),miR-214-3p和Bcl-2表达水平显著降低(P<0.05);与sh-NC组相比，sh-KCNQ1OT1组大鼠mNSS评分、脑血肿体积和脑组织细胞凋亡率显著减少(P<0.05),脑组织结构恢复，红细胞、炎性细胞和神经胶质细胞的数量减少，脑组织中TNF-α、IL-1β、LncRNA KCNQ1OT1、NDRG3和Bax表达水平显著降低(P<0.05),miR-214-3p和Bcl-2表达水平显著升高(P<0.05);与sh-KCNQ1OT1+miR-In-NC组相比，sh-KCNQ1OT1+miR-214-3p-In组大鼠mNSS评分、脑血肿体积和脑组织细胞凋亡率显著增加(P<0.05),脑组织结构被破坏，红细胞、炎性细胞浸润，神经胶质细胞数量增加，脑组织中TNF-α、IL-1β、NDRG3和Bax表达水平显著升高(P<0.05),miR-214-3p和Bcl-2表达水平显著降低(P<0.05);与miR-NC组相比，miR-214-3p组转染KCNQ1OT1-WT或NDRG3-WT的细胞的相对荧光素酶活性均显著降低(P<0.05)。结论:LncRNA KCNQ1OT1下调可能通过调控miR-214-3p/NDRG3轴，缓解ICH大鼠神经功能损伤。更多还原

【Abstract】 Objective: To investigate the effect of long non-coding RNA(LncRNA) KCNQ1OT1 on neurological function of intracerebral hemorrhage(ICH) model rats by adjusting microRNA-214-3p/N-myc downstream regulated gene 3(miR-214-3p/NDRG3) axis. Methods: SD rats were assigned into sham surgery group(Sham), ICH group, LncRNA KCNQ1OT1 interference plasmid control(sh-NC) group, LncRNA KCNQ1OT1 interference plasmid(sh-KCNQ1OT1) group, sh-KCNQ1OT1+miR-214-3p inhibitor control(miR-In-NC) group, and sh-KCNQ1OT1+miR-214-3p inhibitor(miR-214-3p-In) group, each with 12 rats. The modified neurological severity score(mNSS) was used to assess changes in neurological function. The volume of cerebral hematoma in each group was measured. HE staining was performed to observe the pathological changes in brain tissue. TUNEL staining was used to observe cell apoptosis in brain tissue. The changes in the expression level of NDRG3 were observed by immunohistochemical staining. ELISA was used to detect the levels of TNF-α and IL-1β in the rat brain tissue. qRT-PCR was used to detect the levels of LncRNA KCNQ1OT1, miR-214-3p, and NDRG3 mRNA in the brain tissues. Western blot was performed to detect the expression of NDRG3, Bax, and Bcl-2 proteins in the brain tissues. Dual luciferase reporter gene assay was used to determine the targeting relationship between miR-214-3p and LncRNA KCNQ1OT1 or NDRG3. Results: For Sham group, the mNSS score, cerebral hematoma volume, and brain tissue apoptosis rate of rats in ICH group unusually increased(P<0.05), the brain tissue structure was disrupted, the numbers of red blood cells, inflammatory cell infiltration, and glial cells increased, the TNF-α, IL-1β, LncRNA KCNQ1OT1, NDRG3, and Bax in brain tissue were unusually increased(P<0.05), and the miR-214-3p and Bcl-2 were greatly reduced(P<0.05). For sh-NC group, the mNSS score cerebral hematoma volume, and brain tissue apoptosis rate of rats in sh-KCNQ1OT1 group greatly reduced(P<0.05), the brain tissue structure was restored, the numbers of red blood cells, inflammatory cell infiltration, and glial cells declined, the TNF-α, IL-1β, LncRNA KCNQ1OT1, NDRG3, and Bax in brain tissue were unusually declined(P<0.05), and the miR-214-3p and Bcl-2 were greatly raised(P<0.05). For sh-KCNQ1OT1+miR-In-NC group, the mNSS score, cerebral hematoma volume, and brain tissue apoptosis rate of rats in sh-KCNQ1OT1+miR-214-3p-In group greatly raised(P<0.05), the brain tissue structure was disrupted, the numbers of red blood cells, inflammatory cell infiltration, and glial cells raised, the TNF-α, IL-1β, NDRG3, and Bax in brain tissue were greatly raised(P<0.05), and the miR-214-3p and Bcl-2 were greatly reduced(P<0.05). Compared with the miR-NC group, the relative luciferase activity of cells transfected with KCNQ1OT1-WT or NDRG3-WT in the miR-214-3p group was significantly decreased.(P<0.05). Conclusion: Downregulation of LncRNA KCNQ1OT1 may alleviate neurological dysfunction in ICH rats by adjusting miR-214-3p/NDRG3 axis.更多还原