通鼻消涕颗粒治疗大鼠慢性鼻窦炎的作用机制研究

Study on the mechanism of Tongbixiaoti granules in the treatment of chronic rhinosinusitis in rats

目的通过动物实验研究为通鼻消涕颗粒治疗慢性鼻窦炎提供客观实验数据支持。方法SPF级SD大鼠60只,取10只大鼠作为正常组,其余随机分为模型组、克拉霉素片组、通鼻消涕颗粒低剂量组、通鼻消涕颗粒中剂量组、通鼻消涕颗粒高剂量组,每组各10只。造模成功后分别应用克拉霉素和通鼻消涕颗粒灌胃治疗2周后,HE染色检测鼻黏膜炎症情况。免疫组化染色及RT-PCR实验检测各组鼻黏膜组织的PLUNC、Cu/ZnSOD、PACAP、DJ-1、谷胱苷肽S转移酶-pi、NKEF-B的蛋白和mRNA;ELISA法检测大鼠血清的核转录因子(NF-κB)表达水平;流式细胞术检测大鼠外周血Th淋巴细胞亚群Th1/Th2水平。采用t检验进行统计学分析,对比各组大鼠服药前后各个阶段的免疫指标含量变化有无显著差异。结果HE染色与免疫组化染色发现PLUNC等鼻黏膜保护性蛋白均表达在鼻黏膜纤毛柱状上皮及黏膜下腺体中。RT-PCR检测结果表明经通鼻消涕颗粒治疗后,大鼠鼻黏膜上皮及黏膜下腺体反应性增生,产生PLUNC等鼻黏膜保护性蛋白增多来抵抗慢性炎症,克拉霉素治疗后主要是促进炎症消退,而鼻黏膜保护性蛋白产生不多。ELISA检测大鼠血清的NF-κB表达水平提示:通鼻消涕颗粒及克拉霉素片均能减轻慢性鼻窦炎大鼠血清中的NF-κB水平达到抗炎效果。流式检测结果表明中药通鼻消涕颗粒和克拉霉素片均能提高Th1细胞比例,降低Th2细胞比例,从而有助于促进炎症消退。结论中药通鼻消涕颗粒可通过提高鼻黏膜组织中PLUNC等鼻黏膜保护性蛋白含量来改善鼻黏膜局部免疫功能并降低血清NF-κB和Th2细胞比例促进炎症消退从而达到“扶正祛邪”的治疗效果。

Objective To provide objective experimental data support for the treatment of chronic rhinosinusitis(CRS)with Tongbixiaoti granules through animal experiments.Methods A total of 60 SPF SD rats were selected,10 rats were selected as the normal control group,and the others were randomly divided into model group,clarithromycin tablet group,Tongbixiaoti granule low-dose,medium-dose and high-dose groups,with 10 rats in each group.After successful modeling,clarithromycin tablets and Tongbixiaoti granules were administered by gavage for 2 weeks,and the nasal mucosal inflammation was detected by HE staining.The protein and mRNA of PLUNC,Cu/ZnSOD,PACAP,DJ-1,glutathione S-transferase-pi and NKEF-B in nasal mucosa of each group were detected by immunohistochemical staining and reverse transcription polymerase chain reaction(RT-PCR).The expression level of NF-κB in serum was detected by enzyme-linked immunosorbent assay(ELISA).Flow cytometry was used to detect Th1/Th2 levels of Th lymphocyte subsets in peripheral blood of rats.t-test was used for statistical analysis to compare the changes of immune indexes before and after medication.Results HE staining and immunohistochemical staining revealed that protective proteins such as PLUNC were expressed in the ciliated columnar epithelium and submucosal glands of the nasal mucosa.RT-PCR test results showed that the epithelia and submucosal glands of the rat nasal mucosa were reactive proliferation after treatment with Tongbixiaoti granules,and the nasal mucosal protective proteins such as PLUNC were increased to resist chronic inflammation.Clarithromycin treatment mainly promoted inflammation resolution,while the production of protective proteins in the nasal mucosa was not significant.ELISA detection of NF-κB expression levels in rat serum suggested that both Tongbixiaoti granules and Clarithromycin tablets could reduce NF-κB levels in the serum of rats with CRS and achieve anti-inflammatory effect.The flow cytometry results showed that both Tongbixiaoti granules and clarithromycin tablets could increase the proportion of Th1 cells and decrease the proportion of Th2 cells,thereby helping to promote inflammation resolution.Conclusion Traditional Chinese medicine Tongbixiaoti granules can improve the local immune function of nasal mucosa by increasing the content of PLUNC and other protective proteins in nasal mucosa tissue,and reduce the ratio of serum NF-κB and Th2 cells to promote the remission of inflammation,so as to achieve the therapeutic effect of“strengthening the body resistance to eliminate pathogenic factors”.