Mechanism of liver-soothing and stomach-harmonizing recipe alleviating airway inflammation in rats with gastroesophageal reflux disease by regulating PPAR-γ/RXR signaling pathway
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摘要:目的
探讨疏肝和胃方通过调控过氧化物酶体增殖物激活受体γ/类视黄醇X受体(PPAR-γ/RXR)信号通路缓解胃食管反流病(GERD)大鼠气道炎症的机制。
方法采用食管下端盐酸灌注法构建GERD合并气道炎症大鼠模型。将50只SD雄性大鼠随机分为对照组、模型组、疏肝和胃方低剂量组(10.49 g/kg生药量)、疏肝和胃方高剂量组(20.98 g/kg生药量)和奥美拉唑组(3.67 mg/kg), 每组10只, 灌胃14 d。采用苏木精-伊红(HE)染色观察气管组织的病理学变化; 采用RT-qPCR检测支气管肺泡灌洗液中炎症因子[白细胞介素(IL)-17、IL-33、诱导型一氧化氮合酶(iNOS)]以及抗炎因子[IL-10、克拉拉细胞蛋白16(CC16)、表面活性蛋白-D(SP-D)]的mRNA表达水平; 采用免疫印迹法检测PPAR-γ、RXR-α、核因子-κB(NF-κB)、活化蛋白-1(AP-1)的蛋白相对表达量。
结果HE染色结果显示，模型组可见大量炎症细胞浸润，疏肝和胃方低剂量组、高剂量组和奥美拉唑组炎症细胞浸润明显减少。与对照组比较，模型组支气管肺泡灌洗液中IL-17、IL-33、iNOS的mRNA表达水平升高, IL-10、CC16、SP-D的mRNA表达水平降低, PPAR-γ、RXR-α的蛋白相对表达量升高, NF-κB、AP-1的蛋白相对表达量降低，差异均有统计学意义(P < 0.05)。与模型组比较，疏肝和胃方高剂量组和奥美拉唑组上述指标均有改善，差异有统计学意义(P < 0.05)。
结论疏肝和胃方可以有效缓解GERD大鼠气道炎症，其作用机制可能与激活PPARγ/RXR信号通路有关。
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关键词:
- 疏肝和胃方 /
- 胃食管反流病 /
- 过氧化物酶体增殖物激活受体 /
- 类视黄醇X受体 /
- 信号通路
Abstract:ObjectiveTo investigate the mechanism of liver-soothing and stomach-harmonizing recipe alleviating airway inflammation in rats with gastroesophageal reflux disease (GERD) by regulating peroxisome proliferator-activated receptor γ/retinoid X receptor (PPAR-γ/RXR) signaling pathway.
MethodsA rat model of GERD with airway inflammation was established by perfusion of hydrochloric acid into the lower esophagus. A total of 50 SD male rats were randomly divided into control group, model group, low-dose liver-soothing and stomach-harmonizing recipe group (10.49 g/kg crude drug dosage), high-dose liver-soothing and stomach-harmonizing recipe group (20.98 g/kg crude drug dosage), and omeprazole group (3.67 mg/kg), with 10 rats in each group. The rats were gavaged for 14 days. Hematoxylin and eosin (HE) staining was used to observe the pathological change in tracheal tissue. RT-qPCR was used to detect the mRNA expression levels of inflammatory cytokines[interleukin (IL)-17, IL-33, inducible nitric oxide synthase (iNOS)]and anti-inflammatory cytokines[IL-10, Clara cell 16-kDa protein (CC16), surfactant protein-D (SP-D)]in bronchoalveolar lavage fluid; the Western blot was used to detect the relative protein expression levels of PPAR-γ, RXR-α, nuclear factor-κB (NF-κB), and activator protein-1 (AP-1).
ResultsThe HE staining results showed that a large number of inflammatory cell infiltrations were observed in the model group, while the inflammatory cell infiltrations were significantly reduced in the low-dose liver-soothing and stomach-harmonizing recipe group, high-dose liver-soothing and stomach-harmonizing recipe group, and omeprazole group. Compared with the control group, the mRNA expression levels of IL-17, IL-33 and iNOS in bronchoalveolar lavage fluid were significantly increased while the mRNA expression levels of IL-10, CC16 and SP-D were significantly decreased in the model group, and the relative protein expression levels of PPAR-γ and RXR-α were significantly increased, while the relative protein expression levels of NF-κB and AP-1 were significantly decreased in the model group (P < 0.05). Compared with the model group, the above indicators were significantly improved in the high-dose liver-soothing and stomach-harmonizing recipe group and omeprazole group (P < 0.05).
ConclusionLiver-soothing and stomach-harmonizing recipe can effectively alleviate airway inflammation in rats with GERD, and its mechanism may be related to the activation of the PPAR-γ/RXR signaling pathway.
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表 1 各组大鼠支气管肺泡灌洗液中促炎症介质表达水平比较(x±s)
组别 n IL-17 mRNA IL-33 mRNA iNOS mRNA 对照组 10 0.86±0.03 0.63±0.10 0.98±0.12 模型组 10 2.04±0.32* 0.91±0.05* 2.21±0.30* 疏肝和胃方低剂量组 10 1.88±0.14 0.82±0.13 1.78±0.24# 疏肝和胃方高剂量组 10 0.87±0.09# 0.61±0.02# 1.42±0.11# 奥美拉唑组 10 0.94±0.07# 0.65±0.10# 1.57±0.10# IL-17 : 白细胞介素-17; IL-33 : 白细胞介素-33; iNOS: 诱导型一氧化氮合酶。与对照组比较, * P < 0.05; 与模型组比较, #P < 0.05。 
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表 2 各组大鼠支气管肺泡灌洗液中抗炎症介质表达水平比较(x±s)
组别 n IL-10 mRNA CC16 mRNA SP-D mRNA 对照组 10 0.94±0.19 1.07±0.16 0.96±0.08 模型组 10 0.38±0.04* 0.44±0.06* 0.43±0.07* 疏肝和胃方低剂量组 10 0.60±0.10# 0.53±0.07 0.46±0.05 疏肝和胃方高剂量组 10 0.79±0.20# 0.87±0.13# 0.82±0.05# 奥美拉唑组 10 0.71±0.23# 0.84±0.10# 0.75±0.09# IL-10 : 白细胞介素-10; CC16 : 克拉拉细胞蛋白16; SP-D: 表面活性蛋白-D。与对照组比较, * P < 0.05; 与模型组比较, #P < 0.05。
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表 3 各组大鼠支气管肺泡灌洗液中PPAR-γ、RXR-α、NF-κB、AP-1蛋白相对表达量比较(x±s)
组别 n PPAR-γ RXR-α NF-κB AP-1 对照组 10 0.20±0.02 0.57±0.06 0.38±0.02 0.22±0.03 模型组 10 0.05±0.01* 0.14±0.02* 0.89±0.13* 1.02±0.25* 疏肝和胃方低剂量组 10 0.13±0.01# 0.35±0.07# 0.65±0.06# 0.71±0.12# 疏肝和胃方高剂量组 10 0.20±0.05# 0.60±0.07# 0.53±0.09# 0.43±0.08# 奥美拉唑组 10 0.19±0.06# 0.56±0.10# 0.58±0.05# 0.33±0.06# PPAR-γ: 过氧化物酶体增殖物激活受体γ; RXR-α: 类视黄醇X受体α; NF-κB: 核因子-κB; AP-1: 活化蛋白-1。与对照组比较, * P < 0.05; 与模型组比较, #P < 0.05。
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