ADAM10抑制剂对孤独症样行为大鼠突触结构和血脑屏障通透性的影响

Effects of ADAM10 inhibitor on synaptic structure and blood-brain barrier permeability in rats with autism-like behavior

目的探讨解聚素金属蛋白酶10(a disintegrin and metalloprotease domain 10,ADAM10)抑制剂对孤独症样行为大鼠突触结构和血脑屏障(blood-brain barrier,BBB)通透性的影响。方法按照随机数字表法将50只孕期丙戊酸(valproic acid,VPA)诱导的子代雄性孤独症样行为大鼠分为2组(n=25):丙戊酸组(VPA组)、ADAM10抑制剂TAT-Pro-ADAM10(709-729)治疗组(TAT-Pro组),另取25只孕期生理盐水诱导的子代雄性大鼠作为对照组(CON组)。TAT-Pro组在生后第20天腹腔注射TAT-Pro-ADAM10(709-729)(3 nmol/g),CON组和VPA组给予等体积的生理盐水。记录造模期间各组子代大鼠体质量及尾长变化。Western blot检测给药24 h后子代大鼠海马组织中的ADAM10、血管内皮钙黏蛋白(vascular endothelial cadherin,VE-Cad)、咬合蛋白(Occludin)、带状闭合蛋白-1(zonula occludens-1,ZO-1)及突触后致密物-95(post-synaptic density,PSD95)、突触素(synaptophysin,SYN)的蛋白表达量。免疫荧光进一步验证ZO-1的蛋白表达。生后24 d进行旷场、梳毛、三箱社交及水迷宫行为学检测。高尔基染色检测生后28 d子代大鼠海马组织中树突棘密度。结果相较于VPA组,TAT-Pro组大鼠体质量及尾长无显著改变,但重复刻板行为、社交能力及学习记忆功能均显著改善(P<0.05),海马组织中ADAM10及PSD95蛋白表达水平显著降低(P<0.05),VE-Cad、Occludin蛋白表达水平均显著升高(P<0.05),且树突棘的密度显著降低(P<0.05),未成熟的树突棘也减少。结论突触发育关键期,抑制ADAM10过表达可减轻VPA诱导的孤独症样行为大鼠的学习记忆功能,且对孤独症样行为大鼠的异常突触结构及BBB通透性有一定的修复作用。

Objective To investigate the effects of a disintegrin and metalloprotease domain 10(ADAM10)inhibitor on synaptic structure and blood-brain barrier(BBB)permeability in rats with autism-like behavior.Methods A total of 50 male offspring rats with prenatal exposure to valproic acid(VPA)to induce autism-like behavior were randomly divided into VPA group and ADAM10 inhibitor,TAT-Pro-ADAM10(709-729)group(TAT-Pro group),with 25 rats in each group.And another 25 offspring male rats induced by normal saline during pregnancy served as control group(CON group).The TAT-Pro group was given intraperitoneal injection on postnatal day 20 with 3 nmol/g TAT-Pro-ADAM10(709-729),and equal volumes of normal saline were given to the CON and VPA groups.The changes of body mass and tail length of offspring rats were measured and recorded during modeling.Western blotting was performed to detect the protein levels of ADAM10,vascular endothelial cadherin(VE-Cad),Occludin,zonula occludens-1(ZO-1),post-synaptic density protein-95(PSD95),and synaptophysin(SYN)in the hippocampal tissue of the offspring rats in 24 h after administration.The expression of ZO-1 protein was further verified by immunofluorescence assay.Behavioral tests were conducted from postnatal day 24,including open field,grooming,three-chamber social interaction,and water maze tests.The density of dendritic spines in hippocampal tissue in the offspring rats on postnatal day 28 was detected by Golgi staining.Results Compared with the VPA group,the TAT-Pro group showed no significant changes in body weight and tail length,but significantly improved repetitive stereotyped behaviors,social skills and learning memory ability(P<0.05),reduced protein levels of ADAM10 and PSD95 in hippocampal tissues(P<0.05),and elevated levels of VE-cad and Occludin(P<0.05).The density of dendritic spines was significantly reduced(P<0.05),and the number of immature dendritic spines also decreased in the TAT-Pro group than the VPA group.Conclusion Inhibition of ADAM10 overexpression during the critical period of synaptic development can alleviate learning and memory dysfunctions in VPA-induced rats with autism-like behavior,and has a certain restorative effect on abnormal synaptic structures and blood-brain barrier permeability.