[1]张擎天,黄泽灵,陈华,等.枳壳甘草汤调控炎性小体改善腰椎间盘突出症痛敏的实验研究[J].中国中医骨伤科杂志,2024,32(01):13-19.[doi:10.20085/j.cnki.issn1005-0205.240103]
　ZHANG Qingtian,HUANG Zeling,CHEN Hua,et al.Experimental Study of Zhiqiao Gancao Decoction in Regulating Inflammatory Bodies to Improve Pain Sensitivity of Lumbar Intervertebral Disc Herniation[J].Chinese Journal of Traditional Medical Traumatology & Orthopedics,2024,32(01):13-19.[doi:10.20085/j.cnki.issn1005-0205.240103]
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枳壳甘草汤调控炎性小体改善腰椎间盘突出症痛敏的实验研究()
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《中国中医骨伤科杂志》[ISSN:1005-0205/CN:42-1340/R]

卷:
第32卷
期数:
2024年01期
页码:
13-19
栏目:
实验研究
出版日期:
2024-01-15

文章信息/Info

Title:
Experimental Study of Zhiqiao Gancao Decoction in Regulating Inflammatory Bodies to Improve Pain Sensitivity of Lumbar Intervertebral Disc Herniation
文章编号:
1005-0205(2024)01-0013-07
作者:
张擎天1; 黄泽灵1; 陈华1; 段星星1; 陆斌杰1; 沈晓峰1; 李宇卫1; 徐波1△
1南京中医药大学附属苏州中医医院(江苏 苏州,215100)
Author(s):
ZHANG Qingtian1; HUANG Zeling1; CHEN Hua1; DUAN Xingxing1; LU Binjie1SHEN Xiaofeng1; LI Yuwei1; XU Bo1△
1Suzhou Hospital of Traditional Chinese Medicine Affiliated to Nanjing University of Chinese Medicine,Suzhou 215100,Jiangsu China.
关键词:
枳壳甘草汤;  腰椎间盘突出症;  炎性小体;  酸敏感离子通道
Keywords:
Zhiqiao Gancao decoction;  lumbar disc herniation;  NLRP3 inflammatory body;  acid-sensing ion channels(ASICs)
分类号:
R681.5
DOI:
10.20085/j.cnki.issn1005-0205.240103
文献标志码:
A
摘要:
目的:探讨髓核组织Nod样受体蛋白3(NLRP3)炎性小体活化对酸敏感离子通道3(ASIC3)所介导痛敏环节的激活作用,研究枳壳甘草汤改善腰椎间盘突出症(Lumbar Disc Herniation,LDH)疼痛的干预机制。方法:将60只SD大鼠随机分为空白组、模型组、Caspase-1抑制剂组、中药低剂量组(生药3 g/kg)、中药中剂量组(生药6 g/kg)、中药高剂量组(生药12 g/kg)每组10只。除空白组外,各组大鼠进行腰椎间盘突出症造模5 d后,进行相应干预及灌胃给药,持续20 d。于造模前及造模后5,12,19 d测定各组大鼠机械与冷刺激缩足阈值。取材后用苏木精-伊红(HE)染色检测椎间盘退变程度,免疫组化检测髓核组织Caspase-1p10的表达,ELISA法检测血清中IL-6、TNF-α、IL-1β的含量,蛋白免疫印迹法(Western Blot)检测椎间盘组织NLRP3、Caspase-1p10与背根神经节(Dorsal Root Ganglion,DRG)中ASIC3的表达,实时聚合酶链式反应(RT-PCR)检测背根神经节中ASIC3的mRNA表达。结果:机械与冷刺激缩足阈值结果:腰椎间盘突出症模型组大鼠表现出典型的冷痛敏和机械痛敏状态,抑制剂组、枳壳甘草汤治疗各组中大鼠痛敏状态均获缓解,冷刺激与机械刺激缩足阈值均较模型组有明显改善,差异有统计学意义(P<0.05)。HE染色结果:腰椎间盘突出症模型组大鼠纤维环排列紊乱,部分肿胀断裂; 抑制剂组及枳壳甘草汤治疗各组与模型组相比,纤维环排列均较为有序,且纤维环无明显断裂。免疫组化结果:与模型组相比,枳壳甘草汤治疗组免疫组化Caspase-1p10平均光密度更弱,差异有统计学意义(P<0.05)。ELISA检测结果:治疗各组较模型组炎症因子IL-6、IL-1β、TNF-α表达均显著下降,差异有统计学意义(P<0.05)。PCR结果:抑制剂组及枳壳甘草汤治疗各组与模型组相比,ASIC3的mRNA表达均显著下降,差异有统计学意义(P<0.05)。Western Blot结果:抑制剂组及枳壳甘草汤治疗各组与模型组相比,NLRP3、Caspase-1p10、ASIC3蛋白表达水平均出现显著下降,差异有统计学意义(P<0.05)。结论:NLRP3炎性小体的活化能够调控ASIC3的转录及功能,进而构建并维持腰椎间盘突出症的痛敏状态。枳壳甘草汤改善腰椎间盘突出症痛敏的机制与其对NLRP3炎性小体/ASIC3信号通路的调控有关。
Abstract:
Objective:To elucidate the activation of Nod-like receptor thermal protein domain associated protein 3(NLRP3)inflammatory corpuscles in nucleus pulposus tissue on the activation of pain sensitive links mediated by acid-sensing ion channel 3(ASIC3),and to explore the intervention mechanism of Zhiqiao Gancao decoction in improving the pain of lumbar disc herniation(LDH).Methods:60 SD rats were randomly divided into blank group,model group,Caspase-1 inhibitor group,low dose group of Zhiqiao Gancao decoction(crude drug 3 g/kg),medium dose group of Zhiqiao Gancao decoction(crude drug 6 g/kg),and high dose group of Zhiqiao Gancao decoction(crude drug 12 g/kg),with 10 rats in each group.Except for blank group,the LDH model was established in all other groups of rats.5 d after modeling,the rats in each group were given corresponding intervention and intragastric administration for 20 d.The threshold of mechanical and cold stimulation foot shrinkage contraction of rats in each group was measured before modeling and 5,12 and 19 d after modeling.After extraction,hematoxylin-eosin(HE)staining was used to detect the degree of intervertebral disc degeneration; immunofluorescence was used to detect the expression of Caspase-1p10 in nucleus pulposus tissue; ELISA was used to detect IL-6,TNF-α and IL-1β content; Western Blot was used to detect the expression of NLRP3,Caspase-1p10 in intervertebral disc tissue and ASIC3 in dorsal root ganglia(DRG).The real-time polymerase chain reaction(RT-PCR)was used to detect the mRNA expression of ASIC3 in DRG.Results:Mechanical and cold stimulation foot shrinkage threshold results:The LDH model group rats showed typical cold and mechanical pain sensitivity states,and the pain sensitivity state of the rats in the inhibitor group and Zhiqiao Gancao decoction treatment groups were relieved,and the cold stimulation and mechanical stimulation foot shrinkage threshold were significantly improved compared with the model group(P<0.05).HE staining results:In the LDH model group,the fibrous rings were arranged in disorder,and some of them were swollen and broken.Compared with the model group,the arrangement of fibrous rings in the inhibitor group and the Zhiqiao Gancao decoction treatment groups were more orderly,and the fibrous rings were not significantly broken.Immunohistochemistry:Compared with the model group,the average optical density of Caspase-1p10 in the treatment group of Zhiqiao Gancao decoction was weaker,with a significant difference(P<0.05).ELISA results:The inflammatory factors IL-6,IL-1β and TNF-α in the treatment groups decreased significantly compared with model group,with statistical differences(P<0.05).PCR results:Compared with the model group,the expression of ASIC3 mRNA in the inhibitor group and the Zhiqiao Gancao decoction treatment groups decreased significantly,with statistical difference(P<0.05).Western Blot results:Compared with the model group,the expression levels of NLRP3,Caspase-1p10 and ASIC3 protein in the inhibitor group and Zhiqiao Gancao decoction treatment groups were significantly decreased,with statistical difference(P<0.05).Conclusion:The activation of NLRP3 inflammatory corpuscles can regulate the transcription and function of ASIC3,and thereby construct and maintain the pain sensitivity state of LDH.The mechanism of Zhiqiao Gancao decoction in improving LDH pain is related to its regulation of NLRP3 inflammatory body/ASIC3 signal pathway.

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备注/Memo

备注/Memo:
基金项目:国家自然科学基金项目(82174399)
江苏省自然科学基金项目(BK20211084)
江苏省中医药科技发展计划青年项目(QN202007)
江苏省科教能力提升工程:江苏省医学重点学科/实验室建设单位项目(苏卫科教〔2022〕17号)
苏州市中西医结合学会科研项目(SKJYD2021227)
苏州市科技发展计划项目(SKYXD2022053)
第七批全国老中医药专家学术经验继承工作项目(国中医药人教函〔2022〕76号)
中医骨伤重点实验室建设项目(JSDW202253,SZS2022019)
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更新日期/Last Update: 2024-01-15