摘要
目的:观察菟丝子对恐伤孕鼠所产子代学习记忆能力及海马N6-甲基腺苷(m^(6)A)相关修饰酶和m^(6)A总水平的影响。方法:将45只孕鼠随机分为空白组、模型组和菟丝子组。在孕1 d~孕19 d,模型组和菟丝子组采用旁观电击法造模,菟丝子组5 g·kg^(-1)·d^(-1)菟丝子水煎剂灌胃,其余2组等量纯净水灌胃处理,所产仔鼠延续母代分组。仔鼠于21 d龄进行水迷宫实验观察行为学改变;采用透射电镜观察仔鼠海马神经元发育情况;采用实时荧光定量聚合酶链式反应(Real-time PCR)、蛋白免疫印迹法(Western blot)、免疫组化法(IHC)分别检测仔鼠海马甲基转移酶样蛋白3(METTL3)、甲基转移酶样蛋白14(METTL14)、Wilms瘤1相关蛋白(WTAP)、脂肪量和肥胖相关蛋白(FTO)、Alk B同源蛋白5(ALKBH5)mRNA及蛋白表达水平;采用高效液相色谱串联质谱法(LC-MS/MS)检测仔鼠海马m^(6)A总含量。结果:与空白组比较,模型组仔鼠平均潜伏期延长,进入目标象限次数、目标象限停留时间及穿越平台次数均显著减少(P<0.01);海马CA1、CA3神经元结构损伤严重,线粒体肿胀,内质网扩张,部分突触结构模糊,数量较少;METTL3、METTL14、FTO、ALKBH5表达水平下降,m^(6)A总水平明显下降(P<0.05,P<0.01)。与模型组比较,菟丝子组仔鼠平均潜伏期缩短,进入目标象限次数、目标象限停留时间及穿越平台次数均显著增加(P<0.01);海马CA1、CA3神经元损伤程度减轻,线粒体、内质网结构较好,突触结构清晰、完整,分布密集;METTL3、METTL14、FTO表达水平上升,m^(6)A总水平明显上升(P<0.05,P<0.01)。结论:菟丝子能够有效改善孕期恐伤肾所致的子代学习记忆能力下降,保护海马神经元,上调海马METTL3、METTL14、FTO表达水平及m^(6)A总水平。
Objective:To observe the effects of Cuscutae Semen on the learning and memory ability,N6-methyladenosine(m^(6)A)-related modification enzymes and total m^(6)A level in hippocampus of the offspring of fear-damaged pregnant rats.Method:Forty-five pregnant rats were randomly divided into blank group,model group and Cuscutae Semen group.From the 1^(st)day to the 19^(th) day of pregnancy,rats in the model group and the Cuscutae Semen group were induced by observing electric shock of other rats.The Cuscutae Semen group was treated with 5 g·kg^(-1)·d^(-1)Cuscutae Semen decoction(ig),while the other two groups were treated with the same amount of purified water.The offspring were assigned following the grouping method of their maternal generation.The behavioral changes of the offspring were tested by Morris water maze on 21^(st)day after birth,and the development of hippocampal neurons was observed by transmission electron microscopy.The mRNA and protein expression levels of methyltransferase-like 3(METTL3),METTL14,Wilms tumor 1 associated protein(WTAP),fat mass and obesity-associated protein(FTO)and Alk B homologue 5(ALKBH5)were detected by Real-time polymerase chain reaction(Real-time PCR),Western blot and immunohistochemistry(IHC).The total content of m^(6)A in hippocampus was determined by high performance liquid chromatography tandem mass spectrometry(LC-MS/MS).Result:Compared with the conditions in the blank group,the average latency duration in the model group was prolonged,and the number of entries in the target quadrant,the target quadrant duration and the number of crossing the platform were decreased(P<0.01).Additionally,the model group had seriously damaged structure of hippocampal CA1 and CA3 neurons,swollen mitochondria,expanded endoplasmic reticulum,and small number of synapses with some having blurred structure,and the expression levels of METTL3,METTL14,FTO,ALKBH5 as well as the total m^(6)A level were lower than those in the blank group(P<0.05,P<0.01).Compared with the model group,the Cuscutae Semen g
作者
张桐
侯俊林
江煜
杨丽萍
杜志欣
刘岳轩
丁申奥
杨雪明
闫毅楠
ZHANG Tong;HOU Junlin;JIANG Yu;YANG Liping;DU Zhixin;LIU Yuexuan;DING Shenao;YANG Xueming;YAN Yinan(Henan Province Engineering Technology Research Center for Chinese and Western Integrative Prevention and Treatment of Brain Cognitive Disease,Zhengzhou Key Laboratory of Chinese&Western Integrative Prevention and Treatment of Brain Cognitive Disease,Henan University of Chinese Medicine,Zhengzhou 450046,China;Third Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450008,China)
出处
《中国实验方剂学杂志》
CAS
CSCD
北大核心
2023年第2期104-112,共9页
Chinese Journal of Experimental Traditional Medical Formulae
基金
国家自然科学基金项目(81973596)
河南省科技攻关项目(212102310341)。
