靶向沉默ANXA3对大鼠吗啡镇痛耐受及炎症反应的影响  

Effects of targeted silencing ANX A3 on morphine analgesic tolerance and inflammation in rats

作　　者：梁冰[1] 方洁[1] Liang Bing;Fang Jie(Department of Anesthesiology,the First Affiliated Hospital of Henan University of Traditional Chinese Medicine,Zhengzhou 450000,China)

机构地区：[1]河南中医药大学第一附属医院麻醉科

出　　处：《解放军医学杂志》2020年第1期68-72,共5页Medical Journal of Chinese People's Liberation Army

摘　　要：目的研究靶向沉默ANXA3对大鼠吗啡镇痛效能的作用及其可能机制。方法雄性SD大鼠连续7 d鞘内注射15μg吗啡建立慢性吗啡镇痛耐受模型。将30只大鼠随机分为吗啡组、阴性对照组和ANXA3 siRNA组。实验第1、3、5和7天,通过测定各组大鼠后爪的热缩足反射潜伏期(PWL)计算最大可能镇痛效应百分数(%MPE)。第7天行为学测试结束后麻醉并处死大鼠,收集各组大鼠脊髓腰膨大,采用Western blotting及RT-PCR测定ANXA3 mRNA及蛋白表达水平。采用RT-PCR和ELISA试剂盒测定肿瘤坏死因子(TNF)-α、白细胞介素(IL)-1β和IL-6的mRNA及蛋白表达水平。采用Western blotting评价细胞外信号调节激酶(ERK)和c-Jun氨基端激酶(JNK)的磷酸化水平。结果注射吗啡后,阴性对照组%MPE与吗啡组比较无明显差异。注射吗啡第3、5、7天,ANXA3 siRNA组%MPE均明显高于阴性对照组(78.27%±8.50%vs.71.75%±5.98%、64.37%±25.94%vs.43.56%±2.69%、39.96%±3.27%vs.13.81%±1.25%,P<0.05)。与吗啡组比较,阴性对照组ANXA3 mRNA和蛋白相对表达水平无明显差异。ANXA3 siRNA组的ANXA3 mRNA和蛋白相对表达水平均低于阴性对照组(0.47±0.09 vs.1.14±0.08、0.38±0.05 vs.1.15±0.21,P<0.05)。与吗啡组比较,阴性对照组TNF-α、IL-1β和IL-6蛋白浓度无明显差异。ANXA3 siRNA组的TNF-α、IL-1β和IL-6蛋白浓度均低于阴性对照组[(210.15±19.86)pg/g vs.(394.93±54.18)pg/g、(313.42±42.19)pg/g vs.(534.26±49.78)pg/g、(434.68±49.15)pg/g vs.(794.92±98.32)pg/g,P<0.05]。与吗啡组比较,阴性对照组p-ERK和p-JNK蛋白相对表达水平无明显差异。ANXA3 siRNA组p-ERK和p-JNK蛋白相对表达水平明显均低于阴性对照组(0.55±0.04 vs.1.07±0.12、0.68±0.07 vs.1.15±0.04,P<0.05)。结论靶向沉默ANXA3可能通过调控ERK和JNK的磷酸化水平增强吗啡的镇痛效能并减轻炎症反应。Objective To investigate the effects and possible mechanisms of targeted silencing ANXA3 on morphine analgesia and inflammation in rats. Methods Male SD rats were injected intrathecally with 15 μg morphine for 7 days to establish a chronic morphine analgesic tolerance model. Thirty rats were randomly divided into morphine group, negative control group and ANXA3 siRNA group. The maximum possible percentage of analgesic effect(%MPE) was calculated by measuring the paw withdrawal thermal latency(PWL) of rats in each group at the 1 st, 3 rd, 5 th and 7 th day of the experiment. On the 7 th day, rats were anesthetized and sacrificed after behavioral test, and the spinal cord enlargement of rats in each group were collected. The mRNA and protein levels of ANXA3 were detected by RT-PCR and Western blotting, and of tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β) and interleukin-6(IL-6) were determined by RT-PCR and ELISA kits. In addition, Western blotting was used to evaluate the phosphorylation levels of extracellular signal-regulated kinase(ERK) and c-Jun NH2-terminal kinase( JNK). Results After injection of morphine, no significant difference existed in %MPE between the negative control group and the morphine group, while the %MPE was obviously higher in ANXA3 siRNA group than in the negative control group(P<0.05) on the 3 rd day(78.27%±8.50% vs. 71.75%±5.98%), 5 th day(64.37%±25.94% vs. 43.56%±2.69%) and 7 th day(39.96%±3.27% vs. 13.81%±1.25%) after morphine injection. The relative expression levels of ANXA3 mRNA and protein showed no significant difference between morphine group and the negative control group, but was lower in the relative expression levels of ANXA3 mRNA and protein in ANXA3 siRNA group than in the negative control group(0.47±0.09 vs. 1.14±0.08, 0.38±0.05 vs. 1.15±0.21, P<0.05). Compared with morphine group, no significant difference existed in TNF-α, IL-1β and IL-6 protein concentrations in the negative control group. The protein concentrations of TNF-α, IL-1β and IL-6 wer

关 键 词：Annxin A3 吗啡 镇痛 炎症反应 

分 类 号：R614[医药卫生—麻醉学]