电针对慢性不可预见性温和应激致抑郁小鼠海马氧化应激-calpain-炎症信号轴的影响

Effect of Electroacupuncture on Oxidative Stress-Calpain-Inflammatory Signaling Axis in the Hippocampus of Mice with Depression Due to Chronic Unpredicted Mild Stress

目的 基于海马氧化应激-calpain-炎症信号轴探讨电针对小鼠抑郁样行为的干预作用及机制。方法 采用慢性不可预见性温和应激方法复制小鼠抑郁模型,将24只小鼠随机分为正常组、模型组、电针组和维生素C(vitamin C,VC)组,每组6只;正常组、模型组、电针组小鼠给予0.5 mL生理盐水灌胃3周,VC组小鼠给予10 mg/kg VC注射液灌胃3周,电针组小鼠电针“百会-神门-三阴交”组穴,每日1次,每次20 min,连续干预3周,正常组、模型组和VC组小鼠不予电针干预。另有24只小鼠给予腹腔注射脂多糖复制急性抑郁模型,将24只小鼠分为正常组、模型组、钙蛋白酶抑制剂(calpeptin inhibitor, CI)组和N-乙酰半胱氨酸(N-acetylcysteine, NAC)组,每组6只;NAC组小鼠经脑定位注射3μL NAC到侧脑室,CI组小鼠按2 mg/kg经腹腔注射CI。观察并记录小鼠体质量变化;采用蔗糖水偏好实验、旷场实验、强迫游泳实验、悬尾实验评价电针对小鼠抑郁样行为的干预效果;ELISA法检测小鼠海马组织超氧化物歧化酶(superoxide dismutase, SOD)活性,丙二醛(malondialdehyde, MDA)、肿瘤坏死因子α(tumor necrosis factor-α,TNF-α)、白细胞介素(interleukin, IL)-1β、5-羟色胺(5-hydroxytryptamine, 5-HT)水平及血清皮质酮(corticosterone, CORT)水平;Western blot法检测小鼠海马组织calpain-1、超视交叉核生物钟振荡蛋白(suprachiamatic nucleus circadian oscillatory protein, SCOP)和IL-1β表达水平。结果 模型组小鼠表现出典型的抑郁样行为,小鼠体质量、糖水偏好指数显著下降(P<0.05),强迫游泳实验和悬尾实验中小鼠静止不动时间显著增加(P<0.05),海马组织5-HT水平显著降低(P<0.05),血清CORT水平显著升高(P<0.05);模型组小鼠海马组织SOD活性显著下降(P<0.05),MDA水平显著上升(P<0.05),calpain-1、TNF-α、IL-1β水平显著升高(P<0.05),SCOP水平显著下降(P<0.05)。而经电针干预后,小鼠抑郁样行为得到显著改善(P<0.05);且电针干预能显著改善慢性不可预见性温和刺激诱导的小鼠海马组织的氧化损伤,下调calpain-1及炎症因子的水平(P<0.05),并能抑制慢性不可预见性温和刺激诱导的小鼠海马组织SCOP表达水平的降低(P<0.05)。结论 电针“百会-神门-三阴交”组穴能改善慢性不可预见性温和应激诱导的小鼠抑郁样行为,其作用机制可能与调控小鼠海马氧化应激-calpain-炎症信号轴有关。

Objective To investigate the intervention effect of electroacupuncture on depression-like behavior in rats and its mechanism based on the hippocampal oxidative stress-calpain-inflammatory signaling axis.Methods The chronic unpredicted mild stress(CUMS)method was used to establish a mouse model of depression,and 24 mice were randomly divided into normal group,model group,electroacupuncture group,and vitamin C(VC)group,with 6 mice in each group.The mice in the normal group,the model group,and the electroacupuncture group were given 0.5 mL of normal saline by gavage for 3 weeks,those in the VC group were given 10 mg/kg VC injection by gavage for 3 weeks,and those in the electroacupuncture group were given electroacupuncture at Baihui-Shenmen-Sanyinjiao acupoints for 20 minutes each time,once a day for 3 consecutive weeks;the mice in the normal group,model group,and VC group were not given electroacupuncture.Another 24 mice were given intraperitoneally injected lipopolysaccharide to establish a mouse model of acute depression,and then they were divided into normal group,model group,calpeptin inhibitor(CI)group,and N-acetylcysteine(NAC)group,with 6 mice in each group;the mice in the NAC group were given stereotactic brain injection of 3μL of NAC into the lateral ventricle,and those in the CI group were given intraperitoneal injection of CI at a dose of 2 mg/kg.The change in body weight was observed and recorded;the sucrose preference test,open field test,forced swimming test,and tail suspension test were used to evaluate the intervention effect of electroacupuncture on the depression-like behavior of mice;ELISA was used to measure the activity of superoxide dismutase(SOD)and the levels of malondialdehyde(MDA),tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),and 5-hydroxytryptamine(5-HT)in the hippocampus of mice,as well as the serum level of corticosterone(CORT);Western blot was used to measure the protein expression levels of calpain-1,suprachiasmatic nucleus circadian oscillatory protein(SCOP),and IL-1βin the hippocampus of mice.Results The mice in the model group showed typical depression-like behavior,with significant reductions in body weight and sucrose preference index(P<0.05),a significant increase in immobility time in forced swimming test and tail suspension test(P<0.05),a significant reduction in the level of 5-HT in the hippocampus(P<0.05),and a significant increase in the serum level of CORT(P<0.05);in addition,the model group had a significant reduction in the activity of SOD(P<0.05),significant increases in the levels of MDA,calpain-1,TNF-α,and IL-1β(P<0.05),and a significant reduction in the level of SCOP in the hippocampus(P<0.05).After electroacupuncture intervention,there was a significant improvement in the depression-like behavior of mice(P<0.05),and electroacupuncture intervention significantly improved CUMS-induced oxidative injury in the hippocampus,downregulated the levels of calpain-1 and in flammatory factors(P<0.05),and inhibited the reduction in the expression level of SCOP in the hippocampus induced by CUMS(P<0.05).Conclusion Electroacupuncture at Baihui-Shenmen-Sanyinjiao acupoints can improve the depression-like behavior of mice induced by CUMS,possibly by regulating the oxidative stress-calpain-inflammatory signaling axis in the hippocampus of mice.