黄芪百合颗粒通过干预炎症反应缓解大鼠低氧性肺动脉高压的作用机制

Mechanism of Huangqi Baihe granules on hypoxic pulmonary hypertension in rats by modulating the inflammatory response

关键词:  低氧性肺动脉高压  转录组学  PI3K/AKT/NF-κB  炎症反应  黄芪百合颗粒

Key words:  hypoxic pulmonary hypertension  transcriptomics  PI3K/AKT/NF-κB  inflammatory effects  Huangqi Baihe Granules

摘要:

目的 探讨黄芪百合颗粒通过干预炎症反应缓解大鼠低氧性肺动脉高压的作用机制。方法 将60只雄性SD大鼠分为6组（n = 10）：空白组、模型组、地塞米松组（5 mg·kg-1）、黄芪百合颗粒高、中、低剂量组（4.1、2.05、1.025 g·kg-1）。适应饲养3 d后给药，空白组和模型组灌胃给予等量生理盐水，中药组连续灌胃14 d（1次/d），地塞米松组作为阳性药物组于造模前腹腔注射给药3 d（1次/d）；第15 d时除空白组外，其余各组置于实验动物低压模拟舱持续低氧暴露14 d。造模结束后检测右心室收缩压（RVSP）、右心室肥厚指数（RVHI）、HE染色法观察肺组织病理变化、并计算肺中小动脉的血管壁面积/血管总面积（WA%）、血管直径/血管总直径（WT%）；检测肺泡灌洗液（BALF）中肿瘤坏死因子-α（TNF-α）和白细胞介素-1β（IL-1β）的含量。转录组学检测肺组织中差异表达基因，并对差异表达基因进行KEGG、GO富集分析，通过RT-qPCR法验证差异基因表达。结果 与空白组相比，模型组大鼠的RVSP、RVHI明显升高（P ＜ 0.01），HE染色结果显示肺组织出现明显病理损伤，肺血管壁增厚并伴随着管腔狭窄，肺血管重构相关指标WT%、WA%具有明显的上升趋势（P ＜ 0.01），BALF中TNF-α、IL-1β炎症因子的水平显著上升（P ＜ 0.01）。与模型组相比，地塞米松组以及黄芪百合颗粒高、中、低剂量组RVSP、RVHI均显著降低（P ＜ 0.01）；HE染色结果显示各组大鼠肺中小动脉壁增厚及管腔狭窄程度改善，肺血管病理损伤明显减轻；肺血管重构相关指标WT%、WA%具有明显的下降趋势（P ＜ 0.01）；BALF中TNF-α、IL-1β炎症因子的水平显著降低（P ＜ 0.01）。转录组学空白组和模型组的差异表达基因，模型组和高剂量组的差异表达基因取交集，可得771个交集基因，这些交集基因KEGG富集通路为磷脂酰肌醇3-激酶/蛋白激酶B（PI3K-AKT）、核转录因子-κB（NF-κB）等信号通路，RT-qPCR法结果显示模型组差异表达基因PI3K、AKT、NF-κB、白介素-6（IL-6）mRNA的表达水平显著升高（P ＜ 0.05，P ＜ 0.01）。而给予黄芪百合颗粒处理后，上述基因的表达水平降低（P ＜ 0.05，P ＜ 0.01）。结论 黄芪百合颗粒可通过调控PI3K/AKT/NF-κB炎症通路减轻HPH大鼠的病理进程，提示其作为HPH防治策略的潜在价值。

Abstract:

ABSTRACT: OBJECTIVE Investigation of how Huangqi Baihe granules work to reduce hypoxic pulmonary hypertension in rats by affecting the inflammatory response. METHODS Sixty male SD rats were divided into six groups (n=10): the blank group, model group, dexamethasone group (5 mg·kg?1), and Huangqi Baihe granules high, medium, and low dose groups (4.1, 2.05, and 1.025 g·kg?1). After 3 days of acclimatization, the blank and model groups received equal amounts of saline by gavage. The traditional Chinese medicine groups received continuous gavage for 14 days (once daily). The dexamethasone group received intraperitoneal injections for 3 days as a positive drug group before modeling (once daily). On the 15th day, all groups except the blank group were placed in a low-pressure simulation chamber for 14 days of continuous hypoxic exposure. At the end of the modeling period, right ventricular systolic pressure (RVSP) and right ventricular hypertrophy index (RVHI) were measured. Pathological changes in lung tissue were observed with HE staining. The vessel wall area/total vessel area (WA%) and vessel diameter/total vessel diameter (WT%) of small arteries in the lungs were calculated. Levels of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in alveolar lavage fluid (BALF) were detected. Transcriptomics was used to detect differentially expressed genes in lung tissues. KEGG and GO enrichment analyses were performed for these genes. RT-qPCR verified differential gene expression. RESULTS Compared with the blank group, rats in the model group showed significantly elevated RVSP and RVHI (P < 0.01). The staining showed obvious pathological damage in the lung tissue. There was thickening of the pulmonary vascular wall and narrowing of the lumen. Pulmonary vascular remodeling indicators, WT% and WA%, showed a significant upward trend (P < 0.01). The levels of TNF-α and IL-1β in the BALF were also significantly increased (P < 0.01). Compared with the model group, the dexamethasone group and the high-, medium-, and low-dose groups of Huangqi Baihe granules showed significantly reduced RVSP and RVHI (P < 0.01). HE staining revealed improvements in the thickening of the small arterial walls and a reduction in lumen narrowing, resulting in reduced pathological damage to the pulmonary vasculature. WT% and WA% showed a significant downward trend (P < 0.01). Levels of TNF-α and IL-1β in BALF were also significantly reduced (P < 0.01). Differentially expressed genes were compared between the transcriptomic blank and model groups, as well as the model and high-dose groups. There were 771 intersected genes. These genes were enriched in signaling pathways, such as the PI3K-AKT and NF-κB pathways, according to KEGG analysis. RT-qPCR results showed that the model group had significantly higher expression of PI3K, AKT, NF-κB, and IL-6 mRNA (P < 0.05, P < 0.01). Administration of Huangqi Baihe granules reduced the expression of these genes (P < 0.05, P < 0.01). CONCLUSION Huangqi Baihe granules may alleviate pathological progression in HPH rats by regulating the PI3K/AKT/NF-κB inflammatory pathway, suggesting their potential value as a therapeutic strategy for HPH.