微RNA-24过表达对缺血性脑梗死大鼠的脑保护作用及对神经细胞凋亡的影响

Cerebroprotective effect of microRNA-24 overexpression on ischemic cerebral infarction in rats and its effect on neuronal cell apoptosis

目的研究微RNA(miR)-24过表达对缺血性脑梗死(ICI)大鼠的脑保护作用及对神经细胞凋亡的影响,并探讨相关机制。方法2020年1月至2022年1月将45只大鼠以随机数字表法分为健康组(注射生理盐水)、脑梗死组(注射生理盐水)、脑梗死过表达组(注射miR-24 agomir)、脑梗死低表达组(注射miR-24 antagomir)、脑梗死阴性对照(NC)组(注射miR-24-NC)。注射后24 h建立ICI模型,最终各组均纳入8只。建模成功后1、3 d评价大鼠神经功能;实时荧光定量逆转录聚合酶链反应(qRTPCR)检测脑组织miR-24 mRNA表达;TTC染色法检测脑梗死体积;免疫组织化学染色法检测脑皮质神经元核抗原(NeuN)表达;蛋白质印迹法检测脑组织B细胞淋巴瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)蛋白表达。结果与脑梗死组(2.06±0.25)分、(1.72±0.19)分比较,建模后1、3 d脑梗死过表达组Zea-longa神经功能评分(0.98±0.11)分、(0.82±0.09)分降低,脑梗死低表达组(2.71±0.24)分、(2.36±0.21)分升高(P<0.05)。与健康组0.56±0.12比较,脑梗死组脑组织miR-24 mRNA表达0.21±0.03降低(P<0.05);与脑梗死组比较,脑梗死过表达组脑组织miR-24 mRNA表达1.38±0.26升高(P<0.05),脑梗死低表达组0.06±0.01降低(P<0.05)。与脑梗死组(40.31±5.30)mm^(3)、(292.20±35.60)个/视野、0.13±0.02、0.52±0.06比较,脑梗死过表达组脑梗死体积(21.25±4.93)mm^(3)缩小,脑梗死皮质区NeuN阳性表达神经元数目(385.20±45.13)个/视野增加,脑组织Bcl-2蛋白表达0.54±0.07升高,Bax蛋白表达0.36±0.04降低,脑梗死低表达组脑梗死体积(50.56±6.14)mm^(3)增大,脑梗死皮质区NeuN阳性表达神经元数目(154.40±17.91)个/视野减少,Bcl-2蛋白表达0.05±0.01降低,Bax蛋白表达0.69±0.07升高(P<0.05)。结论miR-24过表达可改善ICI大鼠神经功能,缩小脑梗死体积,减少神经元凋亡,可能通过调节脑组织Bcl-2、Bax蛋白表达来实现。

Objective To investigate the cerebroprotective effect of microRNA(miR)-24 overexpression on ischemic cerebral infarction(ICI)rats and the influence on neuronal apoptosis and to explore the related mechanisms.Methods From January 2020 to January 2022,45 rats were randomly divided into a healthy group(injection with saline),a cerebral infarction group(injection with saline),a cerebral infarction overexpression group(injection with miR-24 agomir),a cerebral infarction low-expression group(injection with miR24 antagomir),and a cerebral infarction negative control(NC)group(injection with negative sequence)using a random number table method.The ICI model was established 24 h after injection,and finally 8 rats were included in each group.The neurological function of the rats was evaluated 1 and 3 d after successful modeling.Real-time fluorescence quantitative reverse transcription polymerase chain reaction(qRT‒PCR)was used to detect miR-24 mRNA expression in brain tissue;TTC staining was used to detect cerebral infarction volume;immunohistochemical staining was used to detect neuronal nuclear antigen(NeuN)expression in the cerebral cortex;and protein blotting was used to detect B-cell lymphoma-2(Bcl-2),and Bcl-2-related X protein(Bax)protein expression in brain tissue.Re⁃sults Compared with the cerebral infarction group(2.06±0.25 and 1.72±0.19),the Zea-Longa neurological function scores were lower in the cerebral infarction overexpression group(0.98±0.11 and 0.82±0.09)at 1 and 3 d after modeling and higher in the cerebral infarction overexpression group(2.71±0.24 and 2.36±0.21)(P<0.05).Compared with the healthy group(0.56±0.12),brain tissue miR-24 mRNA expression in the cerebral infarction group(0.21±0.03)was decreased(P<0.05);compared with the cerebral infarction group,brain tissue miR-24 mRNA expression in the cerebral infarction overexpression group(1.38±0.26)was elevated(P<0.05),and that in the cerebral infarction low-expression group(0.06±0.01)decreased(P<0.05).Compared with the cerebral infarction group(40.31±5.30)mm^(3),(292.20±35.60)neurons/field of view,0.13±0.02,0.52±0.06,cerebral infarction volume in the cerebral infarction overexpression group(21.25±4.93)mm^(3) was reduced,the number of neurons positively expressed in the cortical area of cerebral infarction(385.20±45.13)neurons/field of view increased,brain tissue Bcl-2 protein expression(0.54±0.07)increased,Bax protein expression(0.36±0.04)decreased,and cerebral infarction volume(50.56±6.14)mm^(3) increased in the cerebral infarction low-expression group,and the number of NeuN-positively expressing neurons in the cerebral infarction cortical area(154.40±17.91)decreased/field of view,and Bcl-2 protein expression(0.05±0.01)decreased and Bax protein expression(0.69±0.07)increased(P<0.05).Conclusion The overexpression of miR24 can improve neurological function,reduce the volume of cerebral infarction and decrease neuronal apoptosis in ICI rats,possibly by regulating the expression of Bcl-2 and Bax proteins in brain tissue.