电针对急性术后痛大鼠背根神经节PD-L1/PD-1-SHP-1信号通路的影响
Effect of electroacupuncture on PD-L1/PD-1-SHP-1 signaling pathway in dorsal root ganglia of rats with acute postoperative pain
摘要目的:评价电针对急性术后痛大鼠背根神经节(DRG)程序性细胞死亡配体1(PD-L1)/程序性细胞死亡受体(PD-1)/含Src-同源区2蛋白酪氨酸磷酸酶-1(SHP-1)信号通路的影响。方法:SPF级雄性SD大鼠39只,体质量220~250 g,6~8周龄,采用随机数字表法分为3组( n=13):对照组(C组)、腹部手术组(S组)和腹部手术+电针组(S+EA组)。S组和S+EA组在异氟烷麻醉下行腹部手术。S+EA组于术毕即刻和术后2 h时,选取双侧足三里和三阴交穴进行电针刺激30 min,频率为10 Hz连续波,电流为1 mA。每组取7只大鼠,分别于造模前1 d(T 0)和造模后3、5、7、9、11、24 h时(T 1~6),依次测定机械缩足反应阈(MWT)、机械缩腹反应阈(ACT)、热缩足潜伏期(TWL)和冷缩足潜伏期(CWL)。于造模后5 h每组处死6只大鼠,取出T 12~L 4 DRG,分别采用Western blot法检测IL-6、PD-L1、PD-1和SHP-1的表达,采用免疫荧光法观察PD-L1在各类神经元的表达情况。 结果:3组不同时点TWL和CWL比较差异无统计学意义( P>0.05)。与C组比较,S组T 1~5时MWT降低,T 1~6时ACT降低,DRG IL-6表达上调,PD-L1、PD-1和SHP-1表达下调( P<0.05);与S组比较,S+EA组T 1,2时MWT和ACT升高,DRG IL-6表达下调,PD-L1、PD-1和SHP-1表达上调( P<0.05)。免疫荧光结果显示,PD-L1在小直径神经元(CGRP +神经元和IB4 +神经元)和大直径神经元(NF200 +神经元)上均存在表达,且主要表达在CGRP +神经元和IB4 +神经元上。3组间PD-L1在DRG各类神经元的表达比较差异无统计学意义( P>0.05)。 结论:电针减轻大鼠急性术后痛的机制可能与激活PD-L1/PD-1/SHP-1信号通路有关。
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abstractsObjective:To evaluate the effect of electroacupuncture (EA) on programmed cell death ligand-1 (PD-L1)/programmed cell death receptor-1 (PD-1)/Src-homology region two domain-containing phosphatase-1 (SHP-1) signaling pathway in the dorsal root ganglia (DRG) of rats with acute postoperative pain.Methods:Thirty-nine SPF male Sprague-Dawley rats, weighing 220-250 g, aged 6-8 weeks, were divided into 3 groups ( n=13 each) using a random number table method: control group (group C), abdominal surgery group (group S) and abdominal surgery+ EA group (group S+ EA). Group S and group S+ EA underwent abdominal surgery under isoflurane anesthesia. In S+ EA group, both Zusanli and Sanyinjiao acupoints were selected and stimulated for 30 min with a frequency of 10 Hz continuous wave and a current of 1 mA starting from the end of operation and 2 h after operation. Seven rats were selected from each group for measurement of the mechanical paw withdrawal threshold (MWT), abdominal contraction threshold (ACT), thermal paw withdrawal latency (TWL) and cold paw withdrawal latency (CWL) at 1 day before developing the model (T 0) and 3, 5, 7, 9, 11 and 24 h after developing the model (T 1-6). Six rats in each group were sacrificed at 5 h after the model was prepared, and T 12-L 4 DRG was removed for determination of the expression of interleukin-6 (IL-6), PD-L1, PD-1 and SHP-1 (by Western blot) and expression of PD-L1 in various neurons (by immunofluorescence). Results:There was no significant difference in TWL and CWL between the three groups at different time points ( P>0.05). Compared with group C, MWT at T 1-5 and ACT at T 1-6 were significantly decreased, and the expression of IL-6 in DRG was up-regulated, and the expression of PD-L1, PD-1 and SHP-1 in DRG was down-regulated in group S ( P<0.05). Compared with group S, MWT and ACT were significantly increased at T 1, 2, the expression of IL-6 in DRG was down-regulated, and the expression of PD-L1, PD-1 and SHP-1 in DRG was up-regulated in group S+ EA ( P<0.05). The results of immunofluorescence showed that PD-L1 was expressed on both small diameter neurons (CGRP + neurons and IB4 + neurons) and large diameter neurons (NF200 + neurons) and mainly on CGRP + neurons and IB4 + neurons. There was no significant difference in the expression of PD-L1 in various DRG neurons among the three groups ( P>0.05). Conclusions:The mechanism by which EA relieves acute postoperative pain may be related to activation of the PD-L1/PD-1-SHP-1 signaling pathway in the DRG of rats.
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