脊髓mTOR/S6K1/Gli1信号通路在小鼠慢性吗啡耐受中的作用  

Role of spinal mTOR/S6K1/Gli1 signaling pathway in chronic morphine tolerance in mice

作　　者：王龙 徐凌飞 王行何 李童 曹君利 刘苏 Wang Long;Xu Lingfei;Wang Xinghe;Li Tong;Cao Junli;Liu Su(Jiangsu Province Key Laboratory of Anesthesiology,Xuzhou Medical University,Xuzhou 221004,China)

机构地区：[1]徐州医科大学江苏省麻醉重点实验室,221004 [2]徐州医科大学附属医院麻醉科,221006

出　　处：《中华麻醉学杂志》2021年第2期172-176,共5页Chinese Journal of Anesthesiology

基　　金：江苏省自然科学基金(BK20161175);江苏省高校自然科学重大项目(17KJA3320006);江苏省"青蓝工程"资助项目(53041605)。

摘　　要：目的评价脊髓哺乳动物雷帕霉素靶蛋白(mTOR)/核糖体蛋白S6激酶1(S6K1)/Gli1信号通路在小鼠慢性吗啡耐受中的作用。方法健康雄性昆明小鼠,8~10周龄,体重23~25 g。实验Ⅰ取小鼠50只,采用随机数字表法分为2组:生理盐水组(S组,n=10)和吗啡组(M组,n=40)。M组和S组分别皮下注射吗啡和生理盐水10 mg/kg,2次/d,连续7 d。于给药前1 d和每天最后1次给药后30 min测定热痛阈,计算最大镇痛效应百分比(MPE)。M组于给药后1、3、5和7 d热痛阈测定结束后、S组于最后1次热痛阈测定结束后随机处死10只小鼠,取脊髓组织。实验Ⅱ取小鼠40只,采用随机数字表法分为4组(n=10):KU-0063794+吗啡组(KU+M组)、二甲基亚砜(DMSO)+吗啡组(DM+M组)、吗啡+KU-0063794组(M+KU组)和吗啡+二甲基亚砜组(M+DM组)。4组皮下注射吗啡10 mg/kg,2次/d,连续7 d。于第1~3天每天吗啡注射前30 min时,KU+M组腹腔注射mTOR特异性抑制剂KU-0063794200μl(1μg/μl),DM+M组腹腔注射10%DMSO 200μl,2次/d;于第5~7天每天吗啡注射前30 min时,M+KU组腹腔注射KU-0063794200μl(1μg/μl),M+DM组腹腔注射10%DMSO 200μl,2次/d。于吗啡注射前1 d和每天最后1次给药后30 min测定热痛阈,计算MPE。最后1次热痛阈测定结束后处死大鼠,取脊髓组织。采用Western blot法检测脊髓mTOR、p-mTOR、S6K1、p-S6K1和Gli1的表达水平。结果实验Ⅰ与S组比较,M组给药后各时点MPE升高,给药后第3、5和7天时脊髓p-mTOR、p-S6K1和Gli1表达下调(P<0.05),mTOR和S6K1表达差异无统计学意义(P>0.05)。实验Ⅱ与DM+M组比较,KU+M组注射吗啡后第3~7天时MPE降低,脊髓p-mTOR、p-S6K1和Gli1表达下调(P<0.05),mTOR和S6K1表达差异无统计学意义(P>0.05);与M+DM组比较,M+KU组注射吗啡后第6和7天时MPE升高,脊髓p-mTOR、p-S6K1和Gli1表达下调(P<0.05),mTOR和S6K1表达差异无统计学意义(P>0.05)。结论脊髓mTOR/S6K1/Gli1信号通路参与了小鼠慢性吗啡耐受的形成和维持。Objective To evaluate the role of spinal mammlian target of rapamycin(mTOR)/ribosomal S6 kinase 1(S6K1)/glioma associated oncogene homolog 1(Gli1)signaling pathway in chronic morphine tolerance in mice.Methods Healthy male Kunming mice,aged 8-10 weeks,weighing 23-25 g,were used in the study.The experiment was performed in two parts.Experiment I Fifty mice were randomly assigned into 2 groups:normal saline group(group S,n=10)and morphine group(group M,n=40).In M and S groups,morphine and normal saline 10 mg/kg were injected subcutaneously,respectively,twice a day for 7 consecutive days.The thermal pain threshold(TPT)was measured and the maximum analgesic effect percentage(MPE)was calculated at 1 day before administration and 30 min after the last administration every day.Ten mice in each group were randomly selected and sacrificed after measurement of TPT at 1,3,5 and 7 days after administration in group M and after the last measurement of TPT in group S,and the lumbar segment(L4-6)of the spinal cord was removed.ExperimentⅡForty mice were randomly divided into 4 groups(n=10 each):KU-0063794+morphine group(group KU+M),dimethyl sulfoxide(DMSO)+morphine group(group DM+M),morphine+KU-0063794 group(group M+KU)and morphine+DMSO group(group M+DM).Morphine 10 mg/kg was injected subcutaneously twice a day for 7 consecutive days in 4 groups.At 1-3 days of morphine injection,mTOR specific inhibitor KU-0063794200μl(1μg/μl)and 10%DMSO 200μl was injected intraperitoneally in KU+M group and DM+M group at 30 min before administration twice a day.At 5-7 days of morphine injection,KU-0063794200μl(1μg/μl)or 10%DMSO 200μl was injected intraperitoneally in group M+KU or group M+DM at 30min before administration,respectively,twice a day.TPT was measured and MPE was calculated at 1 day before morphine injection and at 30 min after the last administration every day.The animals were sacrificed after the last measurement of TPT,and the lumbar segment(L4-6)of the spinal cord was removed for determination of the expression of spina

关 键 词：受体作用蛋白丝氨酸苏氨酸激酶类 核糖体蛋白质S6激酶类 锌指蛋白GLT1 脊髓 吗啡 药物耐受性 

分 类 号：R965[医药卫生—药理学]