nNOS-NOS1AP复合体在瑞芬太尼诱发大鼠痛觉过敏中的作用  

Role of nNOS-NOS1AP coupling in remifentanil-induced hyperalgesia in rats

作　　者：舒瑞辰 李媛 张璇 张增利 宋振国 尹毅青 Shu Ruichen;Li Yuan;Zhang Xuan;Zhang Zengli;Song Zhenguo;Yin Yiqing(Department of Anesthesiology,Tianjin Medical University Cancer Institute and Hospital National Clinical Research Center for Cancer Tianjin′s Clinical Research Center for Cancer Key Laboratory of Cancer Prevention and Therapy,Tianjin 300060,China)

机构地区：[1]天津医科大学肿瘤医院麻醉科、国家恶性肿瘤临床医学研究中心、天津市恶性肿瘤临床医学研究中心、天津市肿瘤防治重点实验室,天津300060

出　　处：《中华麻醉学杂志》2024年第1期66-70,共5页Chinese Journal of Anesthesiology

基　　金：国家自然科学基金(82101351,82001479,81801889);天津市科技计划项目(22JCQNJC01050);天津市医学重点学科(专科)建设项目(TJYXZDXK-009A)。

摘　　要：目的评价神经元型一氧化氮合酶(nNOS)-一氧化氮合酶1衔接蛋白(NOS1AP)复合体在瑞芬太尼诱发大鼠痛觉过敏中的作用。方法清洁级健康成年雄性SD大鼠40只,体质量240~260 g,2~3月龄,采用随机数字表法分为4组(n=10):对照组(C组)静脉输注生理盐水0.1 ml·kg^(-1)·min^(-1)60 min;瑞芬太尼组(R组)静脉输注瑞芬太尼1.0μg·kg^(-1)·min^(-1)60 min;nNOS-NOS1AP抑制剂ZLc002组(C+Z组)和瑞芬太尼+ZLc002组(R+Z组)每天腹腔注射ZLc00210 mg/kg,连续3 d,然后分别静脉输注生理盐水0.1 ml·kg^(-1)·min^(-1)或瑞芬太尼1.0μg·kg^(-1)·min^(-1)60 min。分别于静脉输注前24 h和输注结束后6、24、48 h(T0-3)时测定机械缩足反应阈(MWT)和热缩足潜伏期(TWL)。最后一次痛阈测定结束后处死大鼠,取L4-6脊髓组织,采用RT-PCR法检测脊髓nNOS、NOS1AP、G蛋白信号传导激活蛋白1(Dexras1)的mRNA表达;生物素转化法提取亚硝基化蛋白,Western blot法测定nNOS、NOS1AP、总体和亚硝基化Dexras1的表达;免疫共沉淀法检测nNOS-NOS1AP共表达;测定脊髓NO含量。结果与C组比较,R组T1-3时MWT降低,TWL缩短,脊髓nNOS、NOS1AP及其mRNA表达上调,nNOS-NOS1AP共表达和NO生成增加,亚硝基化Dexras1表达上调(P<0.05),C+Z组上述各指标差异无统计学意义(P>0.05);与R组比较,R+Z组T1-3时MWT升高,TWL延长,脊髓nNOS-NOS1AP共表达和NO生成减少,亚硝基化Dexras1表达下调(P<0.05),nNOS、NOS1AP及其mRNA表达差异无统计学意义(P>0.05);4组间脊髓Dexras1及其mRNA表达差异无统计学意义(P>0.05)。结论瑞芬太尼诱发大鼠痛觉过敏的机制可能与上调脊髓nNOS和NOS1AP的表达,促进二者相互作用,介导NO生成和Dexras1亚硝基化修饰有关。Objective To evaluate the role of neuronal nitric oxide synthase(nNOS)-nitric oxide synthase 1 adaptor protein(NOS1AP)coupling in remifentanil-induced hyperalgesia in rats.Methods Forty clean-grade healthy adult male Sprague-Dawley rats,weighing 240-260 g,aged 2-3 months,were divided into 4 groups(n=10 each)using a random number table method:control group(group C),remifentanil group(group R),nNOS-NOS1AP inhibitor ZLc002 group(group C+Z)and remifentanil+ZLc002 group(group R+Z).Normal saline was intravenously infused at a rate of 0.1 ml·kg^(-1)·min^(-1) for 60 min in C group.Remifentanil was intravenously infused at a rate of 1.0μg·kg^(-1)·min^(-1) for 60 min in R group.ZLc00210 mg/kg was intraperitoneally injected for 3 consecutive days,and then normal saline 0.1 ml·kg^(-1)·min^(-1) and remifentanil 1.0μg·kg^(-1)·min^(-1) were intravenously infused for 60 min in C+Z group and R+Z group.The mechanical paw withdrawal threshold(MWT)and thermal paw withdrawal latency(TWL)were measured at 24 h before intravenous infusion and 6,24 and 48 h after intravenous infusion(T0-3).All the rats were sacrificed after the last measurement of pain thresholds,and the L4-6 segments of the spinal cord were removed for determination of the expression of nNOS,NOS1AP and Dexamethasone-induced Ras-related protein 1(Dexras1)protein and mRNA using the real-time polymerase chain reaction.Nitrosylated proteins were extracted by biotin conversion for determination of the expression of nNOS,NOS1AP and total and nitrosylated Dexras1(by Western blot)and co-expression of nNOS-NOS1AP(by co-immunoprecipitation).The content of NO in the spinal cord was measured.Results Compared with group C,the MWT was significantly decreased,and the TWL was shortened at T1-3,the expression of nNOS and NOS1AP protein and mRNA was up-regulated,the co-expression of nNOS-NOS1AP and NO production were increased,and the expression of nitrosylated Dexras1 was up-regulated in group R(P<0.05),and no significant change was found in each aforementioned parameter in gr

关 键 词：瑞芬太尼 痛觉过敏 一氧化氮合酶Ⅰ型 一氧化氮合酶1衔接蛋白 

分 类 号：R614[医药卫生—麻醉学]