MicroRNA-326通过靶向Klk7调控MAPK信号通路对抑郁模型大鼠的神经保护作用

Neuroprotective role of microRNA-326 in depression model rats by targeting KLK7 to regulate MAPK signal pathway

目的探究MicroRNA-326通过靶向激肽释放酶7(Kallikrein 7,Klk7)调控有丝分裂原激活的蛋白激酶(mitogen activated protein kinase,MAPK)信号通路对抑郁模型大鼠的作用。方法20只SD大鼠随机分为空白对照组和慢性不可预测轻度应激(chronic unpredictable mild stress,CUMS)组,每组各10只。CUMS诱导制备抑郁症大鼠模型。蔗糖偏好实验、旷场实验和强迫游泳实验检测大鼠行为学变化;免疫组织化学检测Klk7表达,TUNEL染色检测细胞凋亡;荧光素酶报告基因实验验证miR-326与Klk7之间的靶向结合作用;CCK-8检测海马神经元细胞活力;实时荧光定量PCR和Western blot检测miR-326表达和Klk7 mRNA表达;Western blot检测Klk7和MAPK信号通路相关基因蛋白表达。结果与空白对照组相比,CUMS组大鼠蔗糖偏爱值、穿越次数均显著下调(P<0.05),静止时间、Klk7表达和细胞凋亡均显著上调(P<0.05);Klk7为miR-326的靶基因;与阴性对照组(negative control,NC)相比,miR-326通过靶向抑制Klk7,从而抑制MAPK信号通路活化,促进CUMS大鼠海马神经元细胞增殖,抑制细胞凋亡(P<0.05);与miR-326 inhibitor组相比,同时下调miR-326和Klk7抑制MAPK信号通路活化,促进CUMS大鼠海马神经元细胞增殖,抑制细胞凋亡(P<0.05)。结论MicroRNA-326通过靶向Klk7调控MAPK信号通路对抑郁模型大鼠发挥神经保护作用。

Objective To explore the effect of microRNA-326 by targeting Kallikrein 7(Klk7)to regulate the mitogen activated protein kinase(MAPK)signaling pathway on depression model rats.Methods A total of 20 SD rats were randomly divided into blank control group and chronic unpredictable mild stress(CUMS)group,with 10 rats in each group.CUMS was used to induce and prepare a rat model of depression.Sucrose preference test,open field test and forced swimming test were used to detect the behavioral changes of rats.Immunohistochemical was employed to detect Klk7 expression and TUNEL staining was used to detect apoptosis.Luciferase reporter gene experiment was used to verify the targeted binding between miR-326 and Klk7,and CCK-8 was used to detect the cell viability of hippocampal neurons.Real-time quantitative PCR and Western blot were used to detect the expression of miR-326 and Klk7,and Western blot was used to detect protein expression of Klk7 and MAPK signaling pathway related gene.Results Compared with the blank control group,the sucrose preference value and the number of crossings of the CUMS group were significantly down-regulated(P<0.05),while the resting time,Klk7 expression and apoptosis were significantly up-regulated(P<0.05).Klk7 was a target gene of miR-326.Compared with the negative control(NC)group,miR-326 inhibited the activation of MAPK signaling pathway by targeting Klk7,thereby promoting the proliferation of hippocampal neurons in CUMS rats and inhibiting apoptosis(P<0.05).Compared with miR-326 inhibitor group,miR-326 and Klk7 were simultaneously down-regulated to inhibit the activation of MAPK signaling pathway,promote the proliferation of hippocampal neurons in CUMS rats,and inhibit apoptosis(P<0.05).Conclusion MicroRNA-326 exerts a neuroprotective effect on depression model rats by targeting Klk7 to regulate MAPK signaling pathway.