精神分裂症易感基因在人类与小鼠脑组织单细胞中的表达差异分析

目的·探究精神分裂症易感基因在人类和小鼠脑组织各类细胞中的表达模式。方法·采用4类遗传研究方法(全基因组关联研究、连锁和关联研究、拷贝数变异研究和多模态功能基因组学研究)发现精神分裂症易感基因。利用人类和小鼠脑组织单细胞RNA测序(RNA-seq)数据,分析精神分裂症易感基因在神经元、星形胶质细胞、小胶质细胞、少突胶质细胞和少突胶质前体细胞的表达模式,并采用DAVID数据库对人类脑组织单细胞特异表达的易感基因进行功能富集分析。结果·精神分裂症易感基因在人类和小鼠脑组织单细胞的表达模式存在差异。其中,在人类和小鼠的神经元、星形胶质细胞及少突胶质细胞中,两者共同表达的易感基因数目较多;而在小胶质细胞和少突胶质前体细胞中,两者共同表达的易感基因数目少。此外,在人类脑组织单细胞特异表达的精神分裂症易感基因参与突触可塑性调控和钙离子信号转导等生物学过程。结论·精神分裂症易感基因在人类和小鼠脑组织单细胞水平的表达模式具有细胞类型的差异性,可为采用小鼠模型来研究精神分裂症病因学机制提供线索和依据。

Deletion of ephrin-B2 from vGAT neurons in mouse causes social anxiety disorder like phenotypes

OBJECTIVE Social anxiety disorder,also known as social phobia,is the most common of all anxiety disorders.Despite the seriousness of this disorder,it has rarely been studied and as a consequence,little is known about its underlying neurobiology.Growing evidence suggests that inhibitory neurons dysfunction could affect the balance of neuronal activity in psychotic disorders including anxiety,SAD,etc.Ephrin-B(EB) proteins were previously demonstrated to ensure normal distribution and function of inhibitory interneurons in the brain.We thus proposed that deletion of EB2 in inhibitory neurons might destroy the homeostasis of excitatory neurons and inhibitory neurons,and induce behavioral deficits associated with psychotic diseases.METHODS(1)Mice.We generated conditional EB2 knockout mice as a model of SAD and characterized the behaviors and the biochemical changes in the brains of the knockout mice.We also generated a mouse line with a selective deletion of EB2 from vGAT neurons by breeding a v GAT-Cre line with a loxP-flanked alleles in EB2.For EB2 detection,we crossed vGAT-Cre;EphrinB2 loxp/loxp mice with Rosa26-STOP-tdTomato Cre indicator(Ai9) mice to label the vGAT neurons with tdTomato.Mice were maintained in a mixed CD1/129 genetic background.Al experiments involving mice were carried out in accordance with the Shanghai Jiaotong University.Genotypes were unlocked only after completion of analysis.(2) General procedures for behavioral testing.Open field test:Locomotor activity was measured using an open field test.The size of the open field box was 44 cm×44 cm×44 cm.Each mouse was placed in the corner of the open-field apparatus at the beginning of the test,and allowed to explore it for 15 min.The total distance traveled(in cm) and counts for stereotyped behaviors were recorded and analyzed.The total distance traveled was used as an index of locomotor activity.The percentage of center zone entries was considered as anxiety indexes.EPM test:The elevated plus maze apparatus was made of dark gray plastic and comprised two open arms(30 cm×7 cm×0.25 cm) opposed to two enclosed arms(30 cm × 7 cm × 15 cm) elevated60 cm from the floor.Animals were placed in the central area of the apparatus with their head facing an enclosed arm(test duration,5 min).Digitized video recordings with EthoVision software were used for behavioral analysis.The percentage of time spent in open arms and the percentage of open-arm entries were calculated.Social behaviors:Briefly,a top open acrylic box was divided into three compartments by two clear acrylic glass partitions with an opening in the bottom.Wire mesh cages that are either empty or have social stimulus mouse in it were put in the center of each outer compartment.Unfamiliar naive CD1/129 mice of the same age were used as stimulus mice.Prior to the test,the mouse was introduced into the center chamber for 5 min,with both gates closed.Gates were then open and the test mouse was acclimated to explore the empty three-chambered apparatus freely for another 10 min.After the initial 15 min habituation session,mice were placed in the center compartment and allowed to explore freely all the three compartments.To examine the social contact,the following activities were recorded via a video surveillance for 10 min.The number of entries to each compartment,the time spent in each compartment as well as the time and frequency each mouse spent in sniffing were analyzed with EthoVision XT 8.5(Noldus,Wageningen,Netherlands).RESULTS(1)Social preference assays were performed in WT-vGATCre,EB2-flox,and EB2-vGATCre mice respectively.To evaluate social approach behavior,we used a three-chamber social arena to evaluate animals for their social interaction.The control groups of WTvGATCre,EB2-flox mice showed a significant preference for spending time in the social chamber(P<0.05);whereas the EB2-vGATCre mice did not show this preference and spent roughly equal time in the social and nonsocial chambers.(2)To determine if the decrease in social preference could result from changes in locomotion,we compared locomotor activity in EB2-vG ATCre mice and control mice.We did not observe a significant lower locomotive activity in mice.Hence,EB2 deletion caused a change in social preference but not in locomotion.Taken together,EB2 deletion exerts abnormal psychotogenic activity of social deficits which was often considered as an important feature of SAD.(3) The time spent in the open arms is widely used to measure anxiety in mice,with greater time spent in the open arm being considered as less anxious behavior.To normalize for individual differences in overall activity level,the time spent in the open arms was divided by the total time.The ratio was higher for WT-vGATCre mice and EB2-loxp mice as compared to the EB2-vG ATCre mice(P=0.0112,P=0.0197).Thus,results showed that there was a change in anxiety levels from in EB2-vGATCre mice as compared with control mice.CONCLUSION EB2 may be a candidate risk gene for SAD and that the EB2 conditional knockout model could be a tool for studying the underlying mechanisms in SAD.

中药复方治疗抗精神病药物所致代谢综合征的用药规律和潜在作用机制研究

目的解析近10年临床治疗抗精神病药物所致代谢综合征(antipsychotic-induced metabolic syndromes,AIMS)的中药复方的用药规律及预测这些中药的潜在作用机制。方法采用数据挖掘的方法对中文数据库和英文数据库中关于中药复方治疗AIMS的临床研究进行归纳总结,对其用药规律进行分析;再利用网络药理学方法预测高频中药的潜在作用靶点和作用通路。结果从44篇文献中整理出复方50个(最高频复方为六郁汤),涉及86味中药(最高频君药为半夏);通过频数统计和关联分析发现这些中药多味甘,性温,归脾、肺经。从中药数据库与分析平台(Traditional Chinese Medicine Database and Analysis Platform,TCMSP)数据库筛选出排名前5的高频中药已验证的靶点285个(经过合并、去重),从GeneCards数据库筛选出抗精神病药物奥氮平、氯氮平和利培酮的作用靶点675个(经过合并、去重),从DisGeNET数据库中筛选出代谢综合征相关的靶点1027个,将3部分靶点取交集,发现共有靶点33个。最后,将这33个靶点在String数据库构建蛋白质-蛋白质互作网络,将结果在Cytoscape软件中进行可视化并进行拓扑分析,找到核心靶点8个。采用Metascape数据库进行京都基因与基因组百科全书(Kyoto encyclopedia of genes and genomes,KEGG)分析,发现这些靶点主要富集在磷酸腺苷活化蛋白激酶(adenosine monophosphate activated protein kinase,AMPK)信号通路上。结论治疗AIMS的中药属性及作用机制具备一定的分布规律,可为中药治疗AIMS提供理论依据,为进一步发掘其潜在作用机制提供方向。