茯苓多糖对LPS诱导猫肾细胞炎症的保护作用研究

doi:10.16431/j.cnki.1671-7236.2023.11.041

摘要/Abstract

摘要： 【目的】研究茯苓多糖（Poria cocos polysaccharide，PCP）对脂多糖（lipopolysaccharide，LPS）诱导的猫肾细胞（crandell rees feline kidney，CRFK）炎症反应的保护作用及抗炎机制。【方法】通过MTT法检测不同浓度PCP（5、10、15、25、35、45 μg/mL）和LPS（20、40、60、80、100、125、150 μg/mL）对CRFK细胞活力的影响；用不同浓度PCP处理LPS诱导的CRFK炎症模型，观察PCP干预后的细胞形态变化，通过硝酸还原酶法和流式细胞术检测CRFK中一氧化氮（NO）释放和细胞凋亡情况；利用实时荧光定量PCR和Western blotting检测细胞炎症和凋亡相关基因以及蛋白表达水平。【结果】5~25 μg/mL PCP对CRFK无毒性，且可逆转LPS刺激后CRFK细胞形态变化。25 μg/mL PCP可显著降低LPS刺激后CRFK内NO释放和细胞凋亡率（P<0.05）。实时荧光定量PCR结果表明，不同浓度PCP能够显著降低LPS诱导CRFK中IL-6、TNF-α、Caspase3、Caspase8、Caspase9、Bid和Bax mRNA表达量，显著上调Bcl-2 mRNA表达量（P<0.05）。Western blotting结果显示，LPS诱导的CRFK中TLR4、NF-κB、Caspase3和Caspase9蛋白表达水平显著升高（P<0.05），PCP处理能显著降低TLR4、NF-κB、Caspase3和Caspase9蛋白表达量（P<0.05）。【结论】PCP能够通过调控细胞凋亡抑制LPS诱导的CRFK炎症损伤，其作用机制可能与抑制TLR4-NF-κB信号通路有关。研究结果为PCP治疗CRFK炎症提供了新的靶点和试验依据。

关键词: 茯苓多糖; 猫肾细胞; 脂多糖; 炎症; 细胞凋亡

Abstract: 【Objective】 This study was aimed to investigate the protective effect of Poria cocos polysaccharide (PCP)on lipopolysaccharide (LPS) induced inflammation in crandell rees feline kidney (CRFK) and its anti-inflammatory mechanism.【Method】 MTT method was used to detect the changes of cell activity by different concentrations of PCP (5, 10, 15, 25, 35 and 45 μg/mL) and LPS (20, 40, 60, 80, 100, 125 and 150 μg/mL) on CRFK.The LPS-induced CRFK inflammation model was treated with different concentrations of PCP to observe the morphological changes of cells.Nitric oxide (NO) release and cell apoptosis in CRFK were detected by nitric reductase method and flow cytometry, respectively.Real-time quantitative PCR and Western blotting were used to detect the expression levels of genes and proteins related to cell inflammation and apoptosis.【Result】 5-25 μg/mL PCP was non-toxic to CRFK and could reverse the cell morphological changes of CRFK after LPS stimulation.25 μg/mL PCP significantly decreased NO release and apoptosis rate in CRFK after LPS stimulation (P<0.05).Real-time quantitative PCR results showed that different concentrations of PCP could significantly reduce the mRNA expressions of IL-6, TNF-α, Caspase3, Caspase8, Caspase9, Bid and Bax in LPS-induced CRFK, and significantly up-regulate the mRNA expression of Bcl-2 (P<0.05).Western blotting results showed that the expression levels of TLR4, NF-κB, Caspase3 and Caspase9 proteins in LPS-induced CRFK were significantly increased (P<0.05), but after PCP treatment, the expression levels of TLR4, NF-κB, Caspase3 and Caspase9 proteins were significantly decreased (P<0.05).【Conclusion】 PCP could inhibit LPS-induced cell inflammatory damage of CRFK by regulating apoptosis, and its mechanism might be related to the inhibition of TLR4-NF-κB signaling pathway.The results provided a new target and experimental basis for the treatment of CRFK inflammation by PCP.

Key words: Poria cocos polysaccharide; crandell reese feline kidney; lipopolysaccharide; inflammatory; apoptosis

中图分类号:

S859.7

刘佳丽, 林冰, 刘欣, 闫普普, 夏瑾瑾, 黄永熙, 白若男, 郭利伟, 刘国平. 茯苓多糖对LPS诱导猫肾细胞炎症的保护作用研究[J]. 中国畜牧兽医, 2023, 50(11): 4747-4758.

LIU Jiali, LIN Bing, LIU Xin, YAN Pupu, XIA Jinjin, HUANG Yongxi, BAI Ruonan, GUO Liwei, LIU Guoping. Protective Effect of Poria cocos Polysaccharides on LPS-induced CRFK Cell Inflammation[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(11): 4747-4758.

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