CB2R调控小胶质细胞功能对小鼠孤独症样行为的影响  

Effect of cannabinoid receptor 2 on autism-like behaviors inmice by regulating microglial function

作　　者：田文茹 杨璧璘 胡婧仪 刘世丹 邹明扬 武丽杰 孙彩虹 TIAN Wenru;YANG Bilin;HU Jingyi;LIU Shidan;ZOU Mingying;WU Lijie;SUN Caihong(Department of Child and Adolescent Health,Public Health College,Harbin Medical University,Harbin,Heilongjiang 150081,China;Harbin Center for Disease Control and Prevention)

机构地区：[1]哈尔滨医科大学公共卫生学院儿少卫生与妇幼保健学教研室,黑龙江哈尔滨150081 [2]哈尔滨市疾病预防控制中心

出　　处：《中国儿童保健杂志》2025年第1期62-70,共9页Chinese Journal of Child Health Care

基　　金：国家自然科学基金(82073577);黑龙江省普通本科高等学校青年创新人才计划(UNPYSCT-2020168)。

摘　　要：目的 探讨大麻素受体2(CB2R)对小胶质细胞活化和焦亡的调控作用,以及对小鼠孤独症样行为的影响,为ASD的病因学研究提供科学依据。方法 体外实验:将BV2小胶质细胞分为对照组、LPS组、AM1241组,采用RT-PCR法检测炎性因子mRNA表达水平,Western Blot法检测焦亡标记物蛋白表达。体内实验:自2023年10月—2024年7月,构建CB2R敲除鼠(CB2R^(-/-)),将8周龄的小鼠分为WT组和CB2R^(-/-)组,采用免疫荧光评估小胶质细胞活化情况,采用RT-PCR和Western Blot检测焦亡标记物mRNA和蛋白表达水平,通过旷场、自梳、埋珠、三箱实验来评估小鼠的焦虑样行为、重复刻板行为和社交功能。结果 体外实验:与对照组相比,LPS组促炎因子TNF-α、IL-6和CCL2的mRNA水平升高,抗炎因子Arg-1和IL-13的mRNA水平降低(P<0.05);与LPS组相比,AM1241组TNF-α、IL-6和CCL2的mRNA水平降低,Arg-1、IL-10和IL-13的mRNA水平增加(P<0.000 1)。与对照组相比,LPS组中NLRP3、GSDMD、GSDMD-N、caspase-1和cleaved-caspase-1的蛋白表达明显增加,AM1241干预后,均显著降低(P<0.05)。体内实验:与WT小鼠相比,CB2R^(-/-)小鼠海马CA1、CA3区的CD86+/Iba1^(+)比值升高,而CD206^(+)/Iba1^(+)比值降低(P<0.01);NLRP3、GSDMD、caspase-1、IL-1β、IL-18的mRNA水平明显升高(t=2.20、3.12、2.55、2.54、2.30,P<0.05);NLRP3、GSDMD、GSDMD-N、caspase-1和cleaved-caspase-1的蛋白表达明显增加(t=7.43、2.87、3.86、3.99、6.55,P<0.05);移动距离、自梳时间和埋珠个数均显著升高,休息时间和社交偏好指数显著降低(P<0.05)。结论 CB2R参与调控小胶质细胞活化和焦亡,影响神经炎症反应,进而导致小鼠出现孤独症样行为,提示CB2R可能是ASD潜在治疗靶点。Objective To explore the regulatory effect of cannabinoid receptor 2(CB2R)on microglial function and its impact on autism-like behaviors in mice,in order to provide scientific basis for the etiology of ASD.Methods In vitro experiment,BV2 microglia were divided into control group,LPS group,and AM1241 group.RT-PCR was used to detect mRNA expression levels of inflammatory factors.Western Blot was used to measure the proteins expression of pyroptosis marker.In vivo experiments,the 8-week-old mice were divided into WT group and CB2R knockout mice(CB2R^(-/-)group).Immunofluorescence staining was used to evaluate the activation of microglia.RT-PCR and western blot were used to detect mRNA and proteins expression levels of pyroptosis marker.Anxiety,restricted and repetitive behaviors,and social function were evaluated by open field,self-grooming,burying marble,and three-chamber test.Results In vitro experiments,compared with the control group,the mRNA levels of pro-inflammatory factors TNF-α,IL-6,and CCL2 in the LPS group were increased,while anti-inflammatory factors Arg-1 and IL-13 were decreased(P<0.05).Compared with the LPS group,the mRNA levels of TNF-α,IL-6,and CCL2 in the AM1241 group were decreased,while Arg-1,IL-10,and IL-13 were increased(P<0.0001).Compared with the control group,the protein expression of NLRP3,GSDMD,GSDMD-N,caspase-1,and cleaved caspase-1 were significantly increased in the LPS group.After intervention with AM1241,the above protein levels were significantly reduced(P<0.05).In vivo experiments,compared with the WT group,the CD86+/Iba1^(+)ratios in the hippocampal CA1 and CA3 regions were increased,while the CD206^(+)/Iba1^(+)ratios were decreased in the CB2R^(-/-)group(P<0.01).The mRNA expression of NLRP3,GSDMD,caspase-1,IL-1β,and IL-18 in CB2R^(-/-)group was significantly increased(t=2.20,3.12,2.55,2.54,2.30,P<0.05).The protein expression levels of NLRP3,GSDMD,GSDMD-N,caspase-1,and cleaved caspase-1 in CB2R^(-/-)group were significantly increased(t=7.43,2.87,3.86,3.99,6.55,P<0.05).The CB2R^

关 键 词：大麻素受体2 孤独症谱系障碍 小胶质细胞活化 小胶质细胞焦亡 神经炎性反应 

分 类 号：R749.94[医药卫生—神经病学与精神病学]