内皮素受体A抑制剂BQ-123对骨癌痛小鼠脊髓星形胶质细胞活化及大电导钙激活钾离子通道的作用  

Effect of endothelin-A receptor antagonism BQ-123 on spinal large conductance Ca2+-activated K+channel and astrocyte activation in a murine model of bone cancer pain

作　　者：张宇[1,2] 刘际童 黄晓玲[1,2] 胡霞 廖潜[1,2] 李莉 唐轶珣 Zhang Yu;Liu Jitong;Huang Xiaoling;Hu Xia;Liao Qian;Li Li;Tang Yixun(Department of Anesthesiology,Hunan Provincial People′s Hospital(the First Affiliated Hospital of Hunan Normal University),Changsha 410005,China;Clinical Research Center For Anesthesiology of ERAS in Hunan Province,Changsha 410005,China;Hunan Provincial Key Laboratory of Emergency and Critical Care Metabonomics,Changsha 410005,China)

机构地区：[1]湖南省人民医院(湖南师范大学附属第一医院)麻醉医学中心,长沙410005 [2]湖南省围术期加速康复麻醉临床医学研究中心,长沙410005 [3]湖南省急救医学研究所急危重症代谢组学湖南省重点实验室,长沙410005

出　　处：《中华实验外科杂志》2021年第7期1238-1242,共5页Chinese Journal of Experimental Surgery

基　　金：长沙市科技局项目(kq1901056);湖南省自然科学基金青年基金项目(2018JJ3296);湖南省卫生健康委项目(20200428);湖南省肝胆胰肠诊疗能力提升项目(湘卫[2019]118号)。

摘　　要：目的:观察内皮素受体A(ETA)抑制剂BQ-123对C57BL/6小鼠骨癌痛的镇痛效果,并探讨脊髓星形胶质细胞活化及大电导钙激活钾离子通道(BK Ca)通道的作用。方法:选用C56BL/6雄性小鼠30只,体重18~20 g,随机数字表法分为3组(n=10),假手术组(S组)、癌痛组(BCP组)、BQ-123组(BQ组)。BCP组和BQ组于左下肢股骨骨髓腔内接种Lewis肺癌细胞(2×106个)制备骨癌痛模型。S组则注射等量D-Hanks’液。于接种后第7~21天,BQ组腹腔内注射BQ-123(10 nmol/d),BCP组则注射等量双蒸水(10μl/d)。于术前1 d(T0)、术后第7天(T1)、第10天(T2)、第13天(T3)、第15天(T4)、第18天(T5)、第21天(T6)测定机械痛阈值(MWT)和后足使用评分。术后第21天,取L4~L6脊髓,分别采用蛋白质印迹法(Western blot)法检测BK Ca通道蛋白、星形胶质细胞活化标志物胶质纤维状酸性蛋白(GFAP)蛋白表达,酶联免疫吸附剂测定(ELISA)检测肿瘤坏死因子-α(TNF-α)和白细胞介素(IL)-1β的表达。正态分布资料组间比较采用单因素方差分析或重复测量资料方法分析,两两比较采用Tukey检验。结果:BQ组于T2~T6时时间点MWT高于BCP组[(3.97±0.37)g比(3.31±0.56)g、(3.89±0.45)g比(3.22±0.88)g、(3.48±0.56)g比(2.78±0.51)g、(3.13±0.49)g比(2.20±0.33)g、(3.28±0.52)g比(1.95±0.54)g],差异有统计学意义(q=3.367、3.418、3.571、4.744、6.734,P<0.05);于T3~T6时行走评分高于BCP组[(2.70±0.72)g比(2.20±0.49)g、(2.63±0.59)g比(1.77±0.40)g、(2.02±0.65)g比(1.30±0.51)g、(1.83±0.61)g比(1.27±0.52)g],差异有统计学意义(q=3.406、5.859、4.905、3.815,P<0.05);脊髓BKCa通道表达明显高于BCP组(0.27±0.03比0.18±0.02),GFAP表达低于BCP组(0.31±0.02比0.41±0.03),差异有统计学意义(q=3.406、7.276,P<0.05);TNF-α和IL-1β表达低于BCP组[(286.45±15.94)pg/mg蛋白比(217.62±15.48)pg/mg蛋白、(178.28±18.26)pg/mg蛋白比(148.15±12.87)pg/mg蛋白],差异有统计学意义(q=8.152、4.795,P<0.05)。结论:腹腔内注射ETA�Objective To observe the analgesic effect of endothelin-A receptor antagonism BQ-123 on bone cancer pain(BCP)in mice and the role of spinal large conductance Ca2+-activated K+channel(BKCa)and astrocyte activation.Methods Totally,30 male C57BL/6 mice,weighing 18-20 g,were randomized into three groups(n=10 each):Sham group(S group),BCP group,BQ-123 group(BQ group).BCP was induced by Lewis′s lung cells suspension(2×106 cells)in 10 l D-Hank′s injected into the left femur bone marrow in the BCP group and BQ group.The equal volume of D-Hank′s was injected into the left femur bone marrow in the S group.On the day 7-21 after inoculation,BQ-123(10 nmol/per day)was intraperitoneally injected in the BQ group,and the equal volume of DD water was injected in the BCP group.The mechanical withdrawal threshold(MTW)and limb use score were measured on 1 day before establishment of BCP model(T0)and on the 7th day(T1),10th day(T2),13th day(T3),15th day(T4),18th day(T5),21st day(T6)after establishment of BCP model.On the 21st day after establishment of BCP model,the L4-L6 fragments of spinal cord were removed for determination of BKCa,glial fibrillary cidic protein(GFAP)(the marker of astrocyte activation)by Western blotting.The enzyme-linked immunosorbent assay(ELISA)was used for detecting the level of tumor necrosis factor-α(TNF-α)and interleukin(IL)-1β.comparisons between groups were statistically evaluated by a one-way analysis of variance(ANOVA)or repeated measurement data analysis of variance with a Tukey post hoc test.Results As compared with the BCP group,the BQ group showed:The MWT at T2-T6[(3.31±0.56)g vs.(3.97±0.37)g,(3.22±0.88)g vs.(3.89±0.45)g,(2.78±0.51)g vs.(3.48±0.56)g,(2.20±0.33)g vs.(3.13±0.49)g,(1.95±0.54)g vs.(3.28±0.52)g,q=3.367,3.418,3.571,4.744,6.734,P<0.05]and limb use score at T3-T6[(2.20±0.49)g vs.(2.70±0.72)g,(1.77±0.40)g vs.(2.63±0.59)g,(1.30±0.51)g vs.(2.02±0.65)g,(1.27±0.52)g vs.(1.83±0.61)g,q=3.406,5.859,4.905,3.815,P<0.05)]were significantly increased;The expression of BKCa

关 键 词：骨癌痛 内皮素受体 钾离子通道 脊髓 星形胶质细胞 

分 类 号：R738.1[医药卫生—肿瘤]