小儿肺咳颗粒对脂多糖致大鼠慢性支气管炎的防治作用及机制研究

Preventive and therapeutic effects and mechanism of Xiaoer Feike Granules on chronic bronchitis induced by lipopolysaccharide in rats

探讨小儿肺咳颗粒(XEFK)对大鼠慢性支气管炎(chronic bronchitis, CB)的防治作用及其机制。将60只SD大鼠,除空白组10只外,剩余50只建立脂多糖诱导大鼠慢性支气管炎模型,第22天将造模动物按照体质量随机分为5组,并按组别分别给予纯化水、咳特灵胶囊0.11 g·kg^(-1)以及XEFK 3.2,1.6,0.8 g·kg^(-1)(给药浓度分别为0.32,0.16,0.08 g·mL^(-1)),给药体积均为10 mL·kg^(-1),每日1次,连续给药21 d。末次给药后1 h,麻醉动物,灌洗支气管及肺泡,涂片固定,镜下计数中性粒细胞,比色法测定支气管灌洗液中谷胱甘肽过氧化物酶(GSH-Px)、超氧化物歧化酶(SOD)、丙二醛(MDA)的含量;流式细胞仪检测血清中T细胞亚群CD4^(+),CD8^(+),CD4^(+)/CD8^(+)的含量变化;全自动血液流变仪检测血液流变学相关指标;酶联免疫吸附测定(ELISA)法检测血清中肿瘤坏死因子-α(TNF-α),白细胞介素IL-1β,IL-2,IL-6,IL-10的含量;实时荧光定量PCR(RT-qPCR)法检测肺组织中TNF-α和IL-10 mRNA的表达情况;光镜下进行支气管组织形态学病理改变观察。与模型组比较,XEFK高、中剂量组能显著降低支气管灌洗液中性粒细胞和MDA的含量,显著升高GSH-Px和SOD活性,改善和修复支气管黏膜层及黏膜下层慢性炎细胞浸润和淋巴组织增生;高剂量组能降低大鼠的血浆黏度,但对其余血液流变学指标未见统计学差异;各剂量组血清CD4^(+),CD8^(+),CD4^(+)/CD8^(+),IL-2和IL-10的含量明显升高,血清TNF-α,IL-1β和IL-6的含量显著降低;各剂量组能显著下调肺组织TNF-α mRNA的表达水平,IL-10 mRNA的表达明显增加。XEFK能够降低脂质过氧化反应,升高外周血T细胞亚群的含量,调节炎症因子的释放分泌,修复支气管组织形态学病理改变,其作用机制可能与改善炎症反应和增强机体免疫功能有关。

The aim of this paper was to investigate the preventive and therapeutic effects of Xiaoer Feike Granules(XEFK) on chronic bronchitis in rats and its mechanism. Except for 10 rats in the blank group, the remaining 50 of the 60 SD rats were used to establish a model of chronic bronchitis induced by LPS. On the 22 nd day, the model rats were randomly divided into 5 groups according to their body weight, and administrated with purified water, Keteling Capsules 0.11 g·kg^(-1), XEFK 3.2, 1.6 and 0.8 g·kg^(-1)(the dosing concentrations were 0.32, 0.16, 0.08 g·mL^(-1), respectively). These rats took the corresponding drug orally once a day, for consecutive 21 days. The rats were anesthetized 1 hour after the last administration, and the lavage bronchus and alveoli were collected. Then, after the fixation of the smear, neutrophils were counted microscopically, and the contents of glutathione peroxidase(GSH-Px), superoxide dismutase(SOD) and malondialdehyde(MDA) in the bronchoalveolar lavage fluid(BALF) were detected by colorimetric method. Flow cytometry was used to detect the content changes of T cell subsets CD4^(+), CD8^(+), CD4^(+)/CD8^(+) in serum. Hemorheology related indexes were detected by automatic hemorheology. Enzyme-linked immunosorbent assay(ELISA) was used to detect the contents of tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), IL-2, IL-6 and IL-10 in serum. The expression of TNF-α and IL-10 mRNA in lung was detected by Real-time quantitative PCR(RT-qPCR). HE staining was used to observe the pathological changes in the bronchitis tissues. Compared with the model group, XEFK high and medium dose groups could significantly reduce the contents of neutrophils and MDA in bronchial lavage fluid, and increase the activities of GSH-Px and SOD in BALF, and repair the chronic inflammatory cell infiltration and lymphoid tissue hyperplasia in the bronchial mucosal layer and submucosal layer. The high-dose group could reduce the plasma viscosity of rats, but there was no statistical difference in other hemorheological indexes. CD4^(+), CD8^(+), CD4^(+)/CD8^(+), IL-2 and IL-10 contents in each dose group were significantly increased, and TNF-α, IL-1β and IL-6 contents were significantly decreased in serum. Each dose group could significantly down-regulate the expression level of TNF-α mRNA in the lung and increase the expression of IL-10 mRNA. XEFK could reduce lipid peroxidation, increase the content of peripheral blood T cell subsets, regulate the release and secretion of inflammatory factors, and repair the morphological and pathological changes of bronchial tissue. Its mechanism might be related to the improvement of inflammatory response and the enhancement of immune function.