银杏叶提取物对脑卒中后抑郁大鼠的影响

Effect of Ginkgo biloba extract in post-stroke depression model rats

目的观察银杏叶提取物(GBE)对脑卒中后抑郁(PSD)模型大鼠抑郁样行为的干预作用,并研究其通过调控Toll样受体4/核因子-κB(TLR4/NF-κB)通路抑制神经炎症的作用机制。方法大鼠随机分为假手术组、脑缺血组、PSD组、帕罗西汀组、银杏叶提取物低剂量(GBE-L)组及银杏叶提取物高剂量(GBE-H)组,每组10只。除假手术组外,其余各组大鼠均进行大脑中动脉栓塞术(MCAO)制备左侧脑缺血再灌注模型。除假手术组和脑缺血组外,其余各组大鼠予慢性不可预知温和刺激(CUMS)建立PSD大鼠模型,持续刺激8周。刺激4周后,帕罗西汀组、GBE-L及GBE-H组分别给予帕罗西汀5 mg·kg^(-1)、GBE50 mg·kg^(-1)、GBE100 mg·kg^(-1)治疗,假手术组、脑缺血及PSD组给予等体积0.9%NaCl,连续灌胃给药28 d。在造模第4周和第8周,分别测定大鼠体质量和糖水偏好率。用酶联免疫吸附测定(ELISA)法检测血清白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)水平及大脑皮质中去甲肾上腺素(NE)、5-羟色胺(5-HT)、多巴胺(DA)水平;用聚合酶链反应法检测大鼠海马Tlr4、Nfkb1、核因子κB激酶亚基β抑制因子(Ikbkb)mRNA水平;用蛋白质印迹法检测大鼠海马组织NF-κB、核因子κB抑制蛋白α(IKBα)以及磷酸化核因子κB抑制蛋白α(p-IKBα)蛋白表达水平。结果假手术组、脑缺血组、PSD组、帕罗西汀组、GBE-L及GBE-H组给药治疗后大鼠体质量分别为(427.10±6.36)、(403.10±7.37)、(310.10±9.71)、(355.00±4.03)、(347.90±9.88)和(391.90±5.07)g;给药治疗后大鼠糖水消耗率分别为(93.93±1.78)%、(91.57±1.03)%、(54.72±7.34)%、(88.35±4.36)%、(63.55±12.73)%和(81.04±4.31)%;大脑皮质NE含量分别为(1951.14±52.86)、(1827.27±23.63)、(1662.12±35.92)、(2033.58±72.28)、(1887.31±33.07)和(2175.00±42.54)pg·mL^(-1);大脑皮质5-HT含量分别为(237.07±8.86)、(226.15±10.27)、(214.51±3.46)、(297.13±5.79)、(274.14±7.63)和(285.34±8.72)ng·mL^(-1);大脑皮质DA含量分别为(1531.11±47.26)、(1209.89±58.09)、(1143.15±36.31)、(1812.67±51.28)、(1651.56±31.82)和(1853.33±20.42)pg·mL^(-1)。与PSD组相比,GBE能显著增加大鼠体质量(P<0.01),提高大鼠糖水消耗率,表现出抗抑郁样行为作用。GBE显著降低血清中TNF-α、IL-1β水平(均P<0.01),增加大脑皮质中NE、5-HT、DA水平(均P<0.01);下调Tlr4、Nfkb1、Ikbkb mRNA水平(P<0.05,P<0.01);减少NF-κB蛋白表达(P<0.01),降低IKBα磷酸化水平(P<0.01)。结论银杏叶提取物可减轻PSD大鼠抑郁样行为,具有抗抑郁作用,其机制与抑制TLR4/NF-κB通路减轻神经炎症有关。

Objective To observe the effect of Ginkgo biloba extract(GBE)on depression like behavior in post stroke depression(PSD)model rats,and explore the mechanism of regulating Toll like receptor 4/nuclear factor-κB(TLR4/NF-κB)pathway to inhibit neuroinflammation.Methods Rats were randomly divided into 6 groups,sham,cerebral ischemia,PSD,paroxetine,low-dose Ginkgo biloba extract(GBE-L)and high-dose Ginkgo biloba extract(GBE-H)groups,10 rats in each group.Except for the sham group,middle cerebral artery occlusion(MCAO)was performed to prepare a left focal cerebral ischemia model.Except for the sham group and cerebral ischemia group,other groups were subjected to chronic unpredictable mild stress(CUMS)to establish PSD rat model for 8 weeks.After 4 weeks of CUMS,the paroxetine group,GBE-L,and GBE-H were treated with paroxetine 5 mg·kg^(-1),GBE 50 mg·kg^(-1),and GBE 100 mg·kg^(-1),respectively.The sham group,cerebral ischemia group,and PSD group were treated with the same volume of 0.9%NaCl and continuously administered by gavage for 28 d.After 4 weeks and 8 weeks of CUMS,the body weight and sugar preference test were measured.Levels of serum tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β)and levels of norepinephrine(NE),serotonin(5-HT),and dopamine(DA)in the cerebral cortex were measured by enzyme-linked immunosorbent assay(ELISA).The mRNA levels of Tlr4,Nfkb1,and nuclear factorκB-kinase subunitβinhibitory factor(Ikbkb)in the hippocampus of rats were detected by polymerase chain reaction.The protein levels of NF-κB,nuclear factorκB inhibitory proteinα(IKBα)and phosphorylation nuclear factorκB inhibitory proteinα(p-IKB)in hippocampal tissue were detected by Western blot.Results The body weights of rats in the sham group,cerebral ischemia group,PSD group,paroxetine group,GBE-L group and GBE-H group were(427.10±6.36),(403.10±7.37),(310.10±9.71),(355.00±4.03),(347.90±9.88)and(391.90±5.07)g;sugar preference rate were(93.93±1.78)%,(91.57±1.03)%,(54.72±7.34)%,(88.35±4.36)%,(63.55±12.73)%and(81.04±4.31)%;the levels of NE in the cerebral cortex were(1951.14±52.86),(1827.27±23.63),(1662.12±35.92),(2033.58±72.28),(1887.31±33.07)and(2175.00±42.54)pg·mL^(-1);the levels of 5-HT in the cerebral cortex were(237.07±8.86),(226.15±10.27),(214.51±3.46),(297.13±5.79),(274.14±7.63)and(285.34±8.72)ng·mL^(-1);the levels of DA in the cerebral cortex were(1531.11±47.26),(1209.89±58.09),(1143.15±36.31),(1812.67±51.28),(1651.56±31.82)and(1853.33±20.42)pg·mL^(-1).Compared with the PSD group,GBE significantly increased the body weight of rats(P<0.01)and increased the preference rate of sugar water in rats,showing the antidepressant like behavioral.GBE significantly reduced the levels of serum TNF-α,IL-1β(all P<0.01),increased the levels of NE,5-HT,and DA in the cerebral cortex(all P<0.01),down regulate the mRNA levels of Tlr4,Nfkb1 and Ikbkb(P<0.05,P<0.01),reduced the expression of NF-κB(P<0.01),and reduced the phosphorylation of IKBα(P<0.01).Conclusion Ginkgo biloba extract can improve depression-like behavior in PSD model rats,and has antidepressant effect.Its mechanism is related to the inhibition of TLR4/NF-κB pathway,thus reducing neuroinflammation.