补中益气汤调节DNA甲基化酶影响SCH母鼠后代脑线粒体功能改善认知能力  

Effect of Buzhong Yiqi Decoction(补中益气汤)on Brain Mitochondrial Function to Improve Cognitive Ability in The Offsprings of Maternal SCH Rats through Regulating DNA Methyltransferase

作　　者：陈巍[1,2] 黄杨玲[1] 付强 李晓钰 武跃华 马贤德[1] 高天舒[3] 贾连群[1] 王佳[1] CHEN Wei;HUANG Yangling;FU Qiang;LI Xiaoyu;WU Yuehua;MA Xiande;GAO Tianshu;JIA Lianqun;WANG Jia(Liaoning University of Traditional Chinese Medicine,Shenyang 110847,Liaoning,China;The Second Affiliated Hospital of Liaoning University of Traditional Chinese Medicine,Shenyang 110034,Liaoning,China;Affiliated Hospital of Liaoning University of Traditional Chinese Medicine,Shenyang 110032,Liaoning,China)

机构地区：[1]辽宁中医药大学,辽宁沈阳110847 [2]辽宁中医药大学附属第二医院,辽宁沈阳110034 [3]辽宁中医药大学附属医院,辽宁沈阳110032

出　　处：《中华中医药学刊》2025年第3期96-102,I0025,I0026,共9页Chinese Archives of Traditional Chinese Medicine

基　　金：国家自然科学基金青年科学基金项目(82004340);辽宁省教育厅高等学校基本科研项目(面上项目)(2100221283);辽宁省教育厅科学研究经费项目(面上项目)(LJKZ0877)。

摘　　要：目的研究补中益气汤对亚临床甲状腺功能减退(subclinical hypothyroidism,SCH)母鼠后代脑神经发育的影响并探讨其可能作用机制,同时比较了不同中药剂量与西药的疗效作用。方法SD成年雌性大鼠,采用“甲状腺全切+术后左甲状腺激素(Leftothyroxine,L-T4)皮下注射”建立SCH模型。随机分为以下6组:假手术组、模型组、补中益气汤低剂量组(4.1 g/kg·d)、补中益气汤中剂量组(8.19 g/kg·d)、补中益气汤高剂量组(16.38 g/kg·d)、优甲乐组(4.5μg/kg·d),完成SCH大鼠模型后与正常的雄鼠合笼交配,交配前2周开始给药直至取材。取材为孕13 d(E13)胎盘组织、出生后7 d(P7)仔鼠脑组织,21 d(P21)仔鼠行水迷宫实验。放射免疫法(Radioimmunoassay,RIA)检测血清中T4(T4);酶联免疫吸附试验(Enzyme linked immunosorbent assay,ELISA)检测血清中促甲状腺激素(Thyroid stimulating hormone,TSH);电镜观察脑组织线粒体形态学;采用2,7-二氯荧光素二乙酸酯(DCFH-DA)探针法检测细胞内ROS;比色法检测线粒体呼吸链复合物Ⅰ、Ⅱ活性;蛋白质印迹(Western blotting,Wb)方法检测DNA甲基转移酶(DNA methyltransferase,DNMT)DNMT1、DNMT3a、DNMT3b,及DNA去甲基化酶10-11易位蛋白(ten-eleven-translocation protein,TET)TET1、TET2。结果与假手术组比较,模型组大鼠血清促甲状腺激素(thyroid-stimulating hormone,TSH)水平明显升高、P21 d仔鼠Morris水迷宫成绩显著下降、电镜下神经元线粒体形态破坏;E13 d胎盘组织和P7 d仔鼠脑组织中活性氧(ROS)水平升高、线粒体呼吸链复合物Ⅰ、Ⅱ活性降低,P7 d脑组织促甲基化DNMT1、DNMT3a及DNMT3b蛋白灰度值下降、甲基化反向调节TET1、TET2蛋白水平上升。与模型组比较,补中益气汤中剂量组和高剂量组及西药组可降低SCH大鼠升高的TSH水平、改善P21 d仔鼠Morris水迷宫成绩、降低脑组织中ROS水平及线粒体呼吸链复合物I、Ⅱ活性,改善仔鼠神经元线粒体的形态,升Objective To study the effect of Buzhong Yiqi Decoction(补中益气汤)on the neural development of the off-springs of subclinical hypothyroidism(SCH)rats and explore its possible mechanism of action.At the same time,to compare the therapeutic effects of different doses of traditional Chinese medicine and Western medicine.Methods Adult female SD rats were used to establish an SCH model using total thyroidectomy and postoperative subcutaneous injection of L-T4,and the rats were randomly divided into the following 6 groups:sham surgery group,model group,low-dose group of Buzhong Yiqi Decoction[4.1 g/(kg·d)],medium dose group of Buzhong Yiqi Decoction[8.19 g/(kg·d)],high-dose group of Buzhong Yiqi Decoction[16.38 g/(kg·d)],euthyrox group[4.5μɡ/(kg·d)].After completing the SCH model,the model rats mated with normal male rats in a cage,and started administration 2 weeks before mating until sample collection.The samples were taken out from placental tissue at day 13 of pregnancy(E13)and brain tissue from offsprings at day 7 after birth(P7).Water maze experiments were performed on offsprings at day 21.Radioimmunoassay(RIA)method was used to detect total thyroxine(T4)in serum.En-zyme linked immunosorbent assay(ELISA)was used to detect thyroid stimulating hormone(TSH)in serum.The mitochondrial morphology of brain tissue was observed through electron microscopy.Intracellular reactive oxygen species(ROS)was detected by DCFH-DA probe method.Mitochondrial DNA copy number was detected by using RT-PCR.DNA methyltransferase(DN-MT)1,DNMT3a,DNMT3b,ten-eleven-translocation protein(TET)1 and TET2 were detected by the Western blotting(Wb)method.Result Compared with those of the sham surgery group,the serum TSH levels of the model group were significantly in-creased,the Morris water maze scores of the 21-day offspring rats were significantly decreased,and the morphology of neuronal mitochondria was damaged under electron microscopy.On day 13,the levels of ROS increased and the activities of mitochondrial respiratory chain compl

关 键 词：妊娠亚临床甲减 仔鼠脑发育 补中益气汤 线粒体能量代谢 甲基化 

分 类 号：R245.97[医药卫生—针灸推拿学]