孕期慢性应激对子代雌性大鼠抑郁样行为及印记基因IGF-2/H19甲基化的影响  

Effects of chronic stress during pregnancy on depression-like behavior and methylation of imprinted gene IGF-2/H19 in female offspring rats

作　　者：李叶 马淑琴 陈小惠 刘红娅 王瑞 朱家姝 关素珍 Li Ye;Ma Shuqin;Chen Xiaohui;Liu Hongya;Wang Rui;Zhu Jiashu;Guan Suzhen(School of Public Health and Management,Ningxia Medical University,Yinchuan 750004,China;Key Laboratory of Environmental Factors and Chronic Disease Control,Yinchuan 750004,China;General Hospital of Ningxia Medical University,Yinchuan 750004,China)

机构地区：[1]宁夏医科大学公共卫生与管理学院,银川750004 [2]宁夏环境因素与慢性病控制重点实验室,银川750004 [3]宁夏医科大学附属总医院,银川750004

出　　处：《中华行为医学与脑科学杂志》2022年第12期1057-1065,共9页Chinese Journal of Behavioral Medicine and Brain Science

基　　金：国家自然科学基金(81960591);宁夏回族自治区自然科学基金(2022AAC02030)。

摘　　要：目的探究孕期慢性应激对子代雌性大鼠抑郁行为及海马组织印记基因胰岛素样生长因子-2(insulin-like growth factor-2,IGF-2)/长链非编码RNA(lncRNA)H19甲基化的影响。方法32只SPF级雌性SD大鼠按照随机数字表法分为模型组和对照组,模型组大鼠采用慢性不可预知性温和应激(chronic unpredictable mild stress model,CUMS)方法建立抑郁模型,对照组正常饲养。应激刺激第7天时,雌雄鼠合笼交配。在应激刺激前1 d和应激刺激第1、7、14、21、28天对两组母鼠行内眦静脉采血,测定血浆皮质酮浓度;在雌性仔鼠出生后第28天和出生后第42天时,每组随机抽取8只,行内眦静脉采血后测定血浆皮质酮浓度;在出生后第42天时,分别通过糖水偏爱实验、悬尾实验和强迫游泳实验测定两组雌性仔鼠的抑郁样行为。采用RT-PCR和Western blot法检测仔鼠海马组织中IGF-2/H19及相关转移酶表达情况。利用Methyl Target技术,对IGF-2差异性甲基化区域(differentially methylated region,DMR)片段2的19个CpG位点,H19印记控制区(imprinting control region,ICR)片段1中8个CpG位点和H19-ICR片段2的15个CpG位点,同时捕获测序,计算每个CpG位点甲基化水平。采用SPSS 26.0,采用重复测量方差分析、t检验和非参数检验对相关数据进行统计分析。结果(1)重复测量方差分析结果显示,母鼠血浆皮质酮的时间与组别的交互效应不显著(F=2.997,P=0.066),时间主效应显著(F=4.44,P=0.010),组别主效应显著(F=41.40,P=0.001)。组间因素的单独效应分析,在应激第14、21、28天,模型组母鼠血浆皮质酮浓度均高于对照组母鼠(均P<0.001)。(2)在糖水偏爱实验中,模型组雌性仔鼠的液体总消耗[11.10(10.38,11.58)mL,13.55(12.00,15.77)mL,Z=-3.055,P=0.002]、1%蔗糖水消耗[(5.50±1.30)mL,(8.56±2.04)mL,t=-3.582,P=0.003]及1%蔗糖偏爱百分比[(51.35±8.69)%,(62.11±8.05)%,t=-2.576,P=0.022]均低于对照组。(3)模型组雌性仔鼠悬尾实验中静止Objective To investigate the effects of chronic stress during pregnancy on depressive behavior and DNA methylation of insulin-like growth factor-2(IGF-2)/long non-coding RNA(lncRNA)H19 in hippocampus of female offspring rats.Methods A total of 32 SPF female SD rats were divided into model group and control group according to the random number table.The rats in the model group were treated with chronic unpredictable mild stress(CUMS)to establish the depression model,and the rats in the control group were fed normally.On the 7th day of stress stimulation,all female rats mated with male rats.One day before stress stimulation and 1,7,14,21,28 days after stress stimulation,blood samples were collected from the inner canthus vein of the rats to determine the plasma corticosterone concentration.Eight female pups were randomly selected from each group on postnatal day 28(PND28)and postnatal day 42(PND42).Plasma corticosterone concentration was measured after angular vein blood collection.At PND42,the depression-like behavior of female pups in the two groups was measured by sucrose preference test,tail suspension test and forced swimming test.The expression of IGF-2/H19 and related transferases in hippocampus of offspring rats was detected by RT-PCR and Western blot.Methyl target technology was used to capture and sequence 19 CpG sites of IGF-2 differentially methylated region(DMR)fragment 2,8 CpG sites in H19 imprinting control region(ICR)fragment 1 and 15 CpG sites in H19-ICR fragment 2,and calculate the methylation level of each CpG site.SPSS 26.0 was used for statistical analysis of relevant data by repeated measurement ANOVA,t test and non-parametric test.Results(1)The data of plasma corticosterone content of the two groups of female rats at different times were analyzed by repeated measurement variance.The results showed that the the interaction effect between time and group was not significant(F=2.997,P=0.066),and the main effect of time was significant(F=4.44,P=0.010).The main effect of group was significant(F=41.

关 键 词：慢性不可预知温和应激 孕期 雌性子代 抑郁行为 胰岛素样生长因子-2 长链非编码RNA印记基因H19 大鼠 

分 类 号：R749.4[医药卫生—神经病学与精神病学]