基于BDNF/TrkB/CREB通路探讨参苓开心颗粒的抗抑郁作用机制

Antidepressant mechanism of Shenling Kaixin Granules based on BDNF/TrkB/CREB pathway

研究参苓开心颗粒(Shenling Kaixin Granules)对慢性不可预知性温和应激(chronic unpredictable mild stress,CUMS)模型大鼠的抗抑郁作用机制。将90只雄性SD大鼠随机分为对照组、模型组、舒肝解郁胶囊(110 mg·kg^(-1))组及参苓开心颗粒低(90 mg·kg^(-1))、中(180 mg·kg^(-1))、高(360 mg·kg^(-1))剂量组,通过CUMS法复制抑郁大鼠模型,实验结束后通过糖水偏好、旷场、高架十字迷宫及强迫游泳等实验评估大鼠行为学变化;ELISA检测血清白细胞介素1β(interleukin 1β,IL-1β)、肿瘤坏死因子α(tumor necrosis factorα,TNF-α)、脑源性神经营养因子(brain-derived neurotrophic factor,BDNF)、5-羟色胺(5-hydroxytryptamine,5-HT)含量,检测海马CA1区超氧化物歧化酶(superoxide dismutase,SOD)、过氧化氢酶(catalase,CAT)活性;苏木精-伊红(hematoxylin-eosin,HE)染色检测海马CA1区病理变化;Western blot检测海马CA1区神经生长因子(nerve growth factor,NGF)、BDNF、酪氨酸激酶受体B(p-TrkB/TrkB)、环磷腺苷效应元件结合蛋白(p-CREB/CREB)、核因子E2相关因子2(nuclear factor E2 related factor 2,Nrf2)、血红素加氧酶1(heme oxygenase 1,HO-1)、B淋巴细胞瘤-2基因(B-cell lymphoma-2,Bcl-2)/Bcl2关联X蛋白(Bcl-2 associated X protein,Bax)、胱天蛋白酶3(caspase-3)表达水平的变化。结果显示,与对照组相比,模型组大鼠糖水偏好减少,旷场实验进入中央次数、中央时间及运动总距离缩短,高架十字迷宫实验进入开臂次数及时间百分比减少,强迫游泳实验不动次数及时间增加;血清IL-1β、TNF-α含量增加,BDNF、5-HT含量下降;海马CA1区SOD、CAT活性下降,NGF、BDNF、p-TrkB/TrkB、p-CREB/CREB、HO-1、Bcl-2/Bax表达下调,Nrf2核转移降低,caspase-3表达上调。与模型组相比,各给药组大鼠糖水偏好系数、进入中央次数、中央时间、运动总距离、进入开臂次数及时间百分比均增加,强迫游泳不动次数与时间均缩短;血清IL-1β、TNF-α含量下降,BDNF、5-HT含量增加;海马CA1区SOD、CAT活性增强;NGF、BDNF、p-TrkB/TrkB、p-CREB/CREB、HO-1、Bcl-2/Bax表达上调,Nrf2核转移增加,caspase-3表达下调。综上表明,参苓开心颗粒可能通过激活BDNF/TrkB/CREB通路增加Nrf2核转移,降低海马氧化应激损伤,抑制caspase-3活性,降低海马神经细胞的凋亡,发挥抗抑郁作用。

To investigate the antidepressant mechanism of Shenling Kaixin Granules(SLKX)in treating chronic unpredictable mild stress(CUMS)model rats.Ninety male SD rats were randomly divided into control group,model group,Shugan Jieyu Capsules(110 mg·kg~(-1))group and SLKX low-(90 mg·kg~(-1)),medium-(180 mg·kg~(-1)),and high-dose(360 mg·kg~(-1))groups.Depression rat model was replicated by CUMS method.After treatment,the behavioral changes of rats were evaluated by sugar preference,open field,elevated cross maze and forced swimming experiments.The contents of interleukin 1 beta(IL-1β),tumor necrosis factorα(TNF-α),brain-derived neurotrophic factor(BDNF)and 5-hydroxytryptamine(5-HT)in serum were determined by enzyme linked immunosorbent assay(ELISA),and the activities of superoxide dismutase(SOD)and catalase(CAT)in hippocampal CA1 region were also detected.Pathological changes in hippocampal CA1 region were detected by hematoxylin-eosin(HE)staining,and Western blot was used to determine the expression of nerve growth factor(NGF),BDNF,phospho-tyrosine kinase receptor(p-TrkB)/TrkB,phospho-cAMP-response element binding protein(p-CREB)/CREB,nuclear factor E2 related factor 2(Nrf2),heme oxygenase 1(HO-1),B-cell lymphoma-2(Bcl-2)/Bcl-2 associated X protein(Bax)and caspase-3 in hippocampal CA1 region.Results showed that compared with the control group,the model group had decreased sugar preference,reduced number of entries and time spent in the center of open field and shortened total distance of movement,reduced number of entries and proportion of time spent in open arm,and increased number and time of immobility in forced swimming experiment.Additionally,the serum contents of IL-1βand TNF-αand the expression of caspase-3 were higher,while the contents of BDNF and 5-HT,the activities of SOD and CAT in hippocampal CA1 region,the expressions of NGF,BDNF,p-TrkB/TrkB,p-CREB/CREB,HO-1 and Bcl-2/Bax,and the Nrf2 nuclear translocation were lower in model group than in control group.Compared with the conditions in model group,the sugar preference,the number of entries and time spent in the center of open,total distance of movement,and the number of entries and proportion of time spent in open arm in treatment groups were increased while the number and time of immobility in forced swimming experiment were decreased;the serum contents of IL-1βand TNF-αand the expression of caspase-3 were down regulated,while the contents of BDNF and 5-HT,the activities of SOD and CAT in hippocampal CA1 region,the expressions of NGF,BDNF,p-TrkB/TrkB,p-CREB/CREB,HO-1,Bcl-2/Bax,and Nrf2 nuclear translocation were enhanced.In conclusion,SLKX might regulate the Nrf2 nucleus translocation by activating BDNF/TrkB/CREB pathway,lower oxidative stress damage in hippocampus,inhibit caspase-3 activity,and reduce apoptosis of hippocampal nerve cells,thereby playing an antidepressant role.