外源性甲状腺素对胎儿酒精效果出生后大鼠小脑发育的改善作用

Effect of exogenous thyroxine on postnatal development of cerebellar cortex in rats with fetal alcohol exposure

目的研究胎儿酒精效果对大鼠小脑皮质中脑源性神经营养因子(BDNF)和酪氨酸激酶B(TrkB)的影响及外源性甲状腺素(T4)的效应。方法选择怀孕第6天的SD孕鼠随机分为酒精组、正常组、酒精+T4组以及代理母鼠组。酒精组孕鼠摄取1 475 J/d的酒精;正常组孕鼠摄取与酒精组同热卡的奶粉;酒精+T4组孕鼠摄取与酒精组同量的酒精,同时皮下注射5μg/(kg.d)甲状腺素;代理母鼠组摄取正常鼠食。酒精组、正常组、酒精+T4组母鼠分娩6 h后处死采血,测试血中酒精浓度和甲状腺素含量。三组仔鼠由代理母鼠养育,并分别于生后第7、14、21、28、60、90天麻醉处死,采用免疫组织化学染色法,观察小脑皮质中BDNF和TrkB阳性浦肯野细胞的分布及形态,并测算小脑4/5小叶中单位面积所含的BDNF或TrkB阳性浦肯野细胞数。结果三组18只母鼠(每组6只)及其所生仔鼠中144只(每组均选择48只)纳入分析。酒精组、酒精+T4组母鼠的血液酒精浓度高于正常组,差异有统计学意义(P<0.05);酒精+T4组母鼠血液甲状腺素含量高于酒精组,差异有统计学意义(P<0.05)。酒精+T4组仔鼠从第14天开始,小脑皮质中出现分布及形态与正常组类似的BDNF和TrkB阳性浦肯野细胞,酒精组仔鼠小脑皮质中始终未出现BDNF和TrkB阳性浦肯野细胞。酒精+T4组仔鼠小脑中BDNF和TrkB阳性浦肯野细胞数,从第14天起与正常组无差异(P>0.05);酒精组仔鼠小脑中BDNF和TrkB阳性浦肯野细胞数,从第28天起较正常组及酒精+T4组显著减少(P<0.01),并持续到第90天。结论外源性甲状腺素能促进小脑皮质中BDNF和其特异性受体TrkB的合成,促进BDNF-及TrkB-阳性浦肯野细胞的发育,进而改善胎儿酒精效果致低甲状腺素所引起的小脑发育障碍。

Objective To investigate the influence of fetal alcohol effect on the change of brain-derived neurotrophic factor（BDNF）and tyrosine kinase receptor B（TrkB）in the rat cerebellar cortex,and the effect of exogenous thyroxine on the postnatal development of cerebellar cortex in rats with fetal alcohol effect.Methods The rats with 6 pregnant days were divided into four groups,and consisting of the alcohol group with receiving 1 475 J liquid alcohol per day,the control group with receiving 35-calories milk per day,the alcohol ＋ T4 group with receiving 1 475 J liquid alcohol and exogenous thyroxine（5 μg/kg·d）subcutaneously,and the surrogate group with receiving normal diet,respectively.The plasma concentrations of alcohol and thyroxine were determined in maternal rats 6 hours after delivery.All pups in the alcohol,the control and the alcohol ＋ T4 groups were fostered by surrogate mothers,and executed under anesthesia at postnatal 7,14,21,28,60,90 days for measuring the distributions and forms of BDNF and TrkB-immunoreactive（IR）Purkinje cells with the use of immunohistochemistry.The numbers of BDNF-and TrkB-IR Purkinje cells were counted in the lobule 4/5 of cerebellum.Results A total of 18 mother rats and 144 their pups were involved in the result analysis.The maternal alcohol concentrations in the alcohol and the alcohol ＋ thyroxine groups were significantly higher than that in the control group（P 〈0.05）,and the maternal thyroxine concentration was significantly higher in the alcohol ＋ T4 group than that in the alcohol group（P 〈0.05）.A similar developmental pattern of mature BDNF-and TrkB-IR Purkinje cells were observed in the alcohol ＋ T4 and the control groups at P14 and after that,while their immature features were seen in the alcohol group.The single-layer arrangement of these Purkinje cells in the alcohol group was not completely achieved throughout postnatal life.No significant difference in the number of BDNF-and trkB-IR Purkinje cells was observed between the alcohol ＋ T4 group and the control group at P14 and after that（P 〈0.05）.There was a significant reduction of BDNF-and trkB-IR Purkinje cells that lasted till P90 in the alcohol group compared to that in the control group at P28（P 〈0.01）.Conclusions The maternal administration of exogenous thyroxine may promote the synthesis of BDNF and TrkB in the cerebellar cortex,and ameliorate the dysthyriod state induced by fetal alcohol effects during early postnatal life.