咽立爽口含滴丸对慢性咽炎家兔免疫细胞亚群及NF-κB信号通路的影响

doi:10.12300/j.issn.1674-5817.2021.006

摘要/Abstract

摘要： 目的探究咽立爽口含滴丸对慢性咽炎家兔模型的作用效果及机制。方法健康新西兰白兔42只,随机分为7组：空白组,模型组,慢严舒柠清喉利咽颗粒组（9 g/kg）,咽立爽口含滴丸低（0.025 g/kg）、中（0.05 g/kg）、高（0.1 g/kg）剂量组和雾化组（0.025 g/kg）。除空白组外,其余家兔均采用化学刺激法建立慢性咽炎动物模型。造模成功后第2天起,5个治疗组每日分别以慢严舒柠清喉利咽颗粒、咽立爽口含滴丸相应剂量进行不同治疗,而空白组和模型组以蔗糖（0.025 g/kg）置于家兔口腔,连续给药7 d。从造模第3天开始,每天观察家兔咽喉壁局部情况,给药结束后进行采血,处死家兔后采样待检。HE染色观察咽喉壁黏膜组织病理学变化;流式细胞术检测血液CD3⁺、CD4⁺、CD8⁺,计算CD4⁺/CD8⁺水平;ELISA检测血清白细胞介素-6（IL-6）、白细胞介素-1（IL-1）、肿瘤坏死因子-α（TNF-α）、C反应蛋白（CRP）和总抗氧化能力（T-AOC）水平;蛋白质印迹法检测咽喉壁黏膜组织IL-6、IL-1、TNF-α、核转录因子κB（NF-κB p65）、核转录因子-κB抑制因子（IκB-α）蛋白表达;实时荧光定量PCR检测咽喉壁黏膜组织NF-κB p65、IκB-α mRNA表达。结果与空白组比较,慢性咽炎家兔咽喉壁黏膜病理组织损伤;CD3⁺、CD4⁺、CD4⁺/CD8⁺和T-AOC水平显著降低,CD8⁺、IL-6、IL-1、TNF-α、CRP、NF-κB p65和IκB-α水平明显升高（P<0.01）。与模型组比较,不同治疗组均可不同程度减轻咽喉壁黏膜组织病理损伤;咽立爽口含滴丸高剂量组及雾化组CD3⁺、CD4⁺、CD4⁺/CD8⁺和T-AOC水平显著升高,CD8⁺、IL-6、IL-1、TNF-α、CRP、NF-κB p65和IκB-α水平均降低（P<0.05）,效果优于低、中剂量及慢严舒柠清喉利咽颗粒组。结论咽立爽口含滴丸可显著改善慢性咽炎家兔模型的炎性指标,减轻组织病理损伤,调节NF-κB信号通路相关因子表达,且雾化用药的效果更明显。

关键词: 慢性咽炎, 慢严舒柠清喉利咽颗粒, 咽立爽口含滴丸, 家兔

Abstract: Objective To explore the effect and mechanism of pharyngeal refreshing pills on an animal model of chronic pharyngitis. Methods A total of 42 healthy New Zealand white rabbits were randomly divided into seven groups: control, model, manyanshuning granules (9 g/kg), pharyngeal refreshing pills at low- (0.025 g/kg), medium- (0.05 g/kg), high- (0.1 g/kg) doses, and atomization (0.025 g/kg) groups. Animal models of chronic pharyngitis were established by chemical stimulation in all rabbits except the control group. From the second day after successful modeling, the five treatment groups were administered with manyanshuning granules and different doses of pharyngeal refreshing pills every day, while the control and model groups were fed with white sugar (0.025 g/kg) for 7 days. From the third day after successful modeling, the local condition of the pharyngeal wall of each rabbit was observed every day. Blood samples were collected at the end of drug administration and examined. Histopathological changes in the pharyngeal wall mucosa were examined by hematoxylin-eosin staining, and the levels of CD3⁺, CD4⁺, and CD8⁺ were detected by flow cytometry. The levels of interleukin-6 (IL-6), IL-1, tumor necrosis factor-α (TNF-α), C-reactive protein (CRP), and total antioxidant capacity (T-AOC) in serum were detected by enzyme-linked immunosorbent assay (ELISA). Western blotting was used to detect the expression of IL-6, IL-1, TNF-α, nuclear factor-κappa B (NF-κB) p65 and NF-κB inhibitor (IκB-α) at the protein level in the pharyngeal wall mucosa. qRT-PCR was used to detect NF-κB p65 and IκB-α mRNA expression in the pharyngeal wall mucosa. Results Compared with the control group, the pathological tissues of the posterior pharyngeal mucosa of rabbits with chronic pharyngitis were significantly damaged; the levels of CD3⁺, CD4⁺, CD4⁺/CD8⁺, and T-AOC were significantly decreased, while the levels of CD8⁺, IL-6, IL-1, TNF-α, CRP, NF-κB p65, and IκB-α were significantly increased (all P < 0.01). Compared with the model group, the pathological injury of pharyngitis could be alleviated to different degrees in the treatment group; the levels of CD3⁺, CD4⁺, CD4⁺/CD8⁺, and T-AOC in the pharyngeal refreshing pills at high dose and atomization groups were significantly increased, while the levels of CD8⁺, IL-6, IL-1, TNF-α, CRP, NF-κB p65, and IκB-α were significantly decreased (all P < 0.05). The effect of high-dose pharyngeal refreshing pill administration was better than that of low-dose and middle-dose pharyngeal refreshing pills, and manyanshuning granules. Conclusion The pharyngeal refreshing pills can significantly improve the inflammatory index of chronic pharyngitis, reduce pathological injury, affect the expression of NF-κB signaling pathway-related factors, and the effect of atomization dose is more obvious.

Key words: Chronic pharyngitis, Manyanshuning granules, Pharyngeal refreshing pills, Rabbit

中图分类号:

Q95-33
R-332

唐燚, 喻国冻, 何星辰, 王佳, 张田. 咽立爽口含滴丸对慢性咽炎家兔免疫细胞亚群及NF-κB信号通路的影响[J]. 实验动物与比较医学, 2021, 41(5): 426-434.

TANG Yi, YU Guodong, HE Xingchen, WANG Jia, ZHANG Tian. Effects of Pharyngeal Refreshing Pills on Immune Cell Subsets and NF-κB Signaling Pathway in Chronic Pharyngitis[J]. Laboratory Animal and Comparative Medicine, 2021, 41(5): 426-434.

图/表 6

参考文献

[1] XU C Y, YUE R S, LV X, et al.The efficacy and safety of Banxia-Houpo-Tang for chronic pharyngitis: a protocol for systematic review and meta analysis[J]. Medicine, 2020, 99(30): e19922. DOI:10.1097/MD.0000000000019922.
[2] LEONE C A, CARUSO A A, ALLOCCA V, et al.Pilot study on the effects of high molecular weight sodium hyaluronate in the treatment of chronic pharyngitis[J]. Int J Immunopathol Pharmacol, 2015, 28(4):532-538. DOI:10.1177/0394632015586497.
[3] RAISH M, AHMAD A, ANSARI M A, et al.Momordica charantia polysaccharides ameliorate oxidative stress, inflammation, and apoptosis in ethanol-induced gastritis in mucosa through NF-kB signaling pathway inhibition[J]. Int J Biol Macromol, 2018, 111:193-199. DOI:10.1016/j.ijbiomac.2018. 01.008.
[4] 陈其冰, 王燕, 李芬, 等. 慢性咽炎病因和发病机制研究进展[J]. 听力学及言语疾病杂志, 2019, 27(2):224-228. DOI:10.3969/j.issn.1006-7299.2019.02.029.
[5] 吴思恩, 李清明, 曹火太. 咽立爽口含滴丸治疗慢性咽炎的效果观察[J]. 中国当代医药, 2015, 22(20):81-83.
[6] 郭裕. 咽立爽口含滴丸超声雾化治疗急性单纯性咽炎疗效观察[J]. 世界临床药物, 2006, 27(7):431-432,442. DOI:10.3969/j.issn.1672-9188.2006.07.012.
[7] 何强亮, 刘元献, 杨龙, 等. 慢性咽炎动物模型的建立及相关指标观察分析[J]. 中国中西医结合耳鼻咽喉科杂志, 2017, 25(1):17-20. DOI:10.16542/j.cnki.issn.1007-4856.2017.01.005.
[8] 苗明三. 实验动物和动物实验技术[M]. 北京: 中国中医药出版社, 1997.
[9] CHEN J J, YIN S K, LIU S X, et al.A multicenter randomized controlled study on the treatment of acute and chronic pharyngitis with Binglianqingye Spray[J]. Lin Chung Er Bi Yan Hou Tou Jing Wai Ke Za Zhi, 2018, 32(1):1-6. DOI:10.13201/j.issn.1001-1781.2018. 01.001.
[10] 温雯, 张颖, 邹桂欣, 等. 兔慢性咽炎模型实验研究[J]. 辽宁中医杂志, 2018, 45(6):1313-1315. DOI:10.13192/j.issn.1000-1719.2018.06.060.
[11] 谢雪艳, 李天珍, 王万林, 等. 不同产源艾纳香油化学成分及其抗炎活性研究[J]. 中药新药与临床药理, 2019, 30(9):1069-1076. DOI:10.19378/j.issn.1003-9783.2019.09.008.
[12] 王立英, 刘强, 曹震. 清喉利咽颗粒联合克拉霉素治疗慢性咽炎的临床研究[J]. 现代药物与临床, 2020, 35(2):300-303. DOI: 10.7501/j.issn.1674-5515. 2020. 02.022.
[13] 王艺, 张玲, 周家璇. 咽炎消合剂对慢性咽炎大鼠咽黏膜修复作用的研究[J]. 现代中西医结合杂志, 2015, 24(15):1600-1602. DOI:10.3969/j.issn.1008-8849. 2015.15.002.
[14] 贺敏. 逍遥散联合半夏厚朴汤治疗慢性咽炎的效果及对患者血清炎症标志物及T淋巴细胞亚群的影响[J]. 中医临床研究, 2020, 12(13):69-71. DOI: 10.3969/j.issn.1674-7860.2020.13.023.
[15] 曹敏, 向钦, 刘阳, 等. CBA/CaJ小鼠脏器重量、血液生理生化指标及免疫细胞的比例[J]. 中国实验动物学报, 2018, 26(4):418-423. DOI:10.3969/j.issn. 1005-4847.2018.04.002.
[16] 张礼来, 张志琴, 周凤玲. 益气通窍中药灌洗辅助治疗鼻内镜术后鼻源性慢性咽炎疗效及对炎性细胞因子、T淋巴细胞亚群、SIgA水平的影响[J]. 现代中西医结合杂志, 2018, 27(9):959-962. DOI:10.3969/j.issn.1008-8849. 2018.09.013.
[17] 雷剑波, 荣堃, 杨丽, 等. 养阴利咽饮联合玉液散吹喉治疗阴虚肺燥型慢性咽炎效果及对血清炎症因子水平的影响[J]. 现代中西医结合杂志, 2016, 25(27):3041-3044. DOI:10.3969/j.issn.1008-8849. 2016. 27.028.
[18] ZHOU Z X, MOU S F, CHEN X Q, et al.Anti-inflammatory activity of resveratrol prevents inflammation by inhibiting NFκB in animal models of acute pharyngitis[J]. Mol Med Rep, 2018, 17(1):1269-1274. DOI:10.3892/mmr.2017.7933.
[19] 徐佳莉, 王莹, 陈鹏. 高剂量PM_2.5诱导卵清蛋白致哮喘小鼠肺损伤及其机制[J]. 细胞与分子免疫学杂志, 2017, 33(10):1297-1302. DOI:10.13423/j.cnki.cjcmi.008444.
[20] 高斐宏, 李林, 卢炀, 等. 蒙药十三味红花密诀丸对过敏性鼻炎大鼠鼻黏膜ICAM-1、VCAM-1、TLR4、NF-κBp65表达的影响[J]. 中华中医药杂志, 2017, 32(9):4143-4145.
[21] ZHU C, ZHANG S, SONG C, et al.Selenium nanoparticles decorated with Ulva lactuca poly-saccharide potentially attenuate colitis by inhibiting NF-κB mediated hyper inflammation[J]. J Nano-biotechnology, 2017, 15(1):20. DOI:10.1186/s12951-017-0252-y.