Exploration on the action mechanism of Qufeng Mingmu Pills on autoimmune uveitis rats based on the AMPK/SIRT1 pathway
Objective:To study the mechanism of action of Qufeng Mingmu Pills on the AMPK/SIRT1 pathway and Th1/Th17 cells in experimental autoimmune uveitis(EAU)rats.Methods:Seventy SPF grade male Lewis rats were randomly selected,with 19 rats as the blank group,and the remaining 51 rats were used to establish an EAU model.After modeling,inflammation scores were performed.After successful modeling,they were randomly divided into model group,Chinese medicine constant group,and Chinese medicine high magnification group.The Chinese medicine constant group and Chinese medicine high magnification group were orally administered with 2.4,4.8 g/kg Qufeng Mingmu Pills solution every day,while the blank group and model group were orally administered with equal amounts of distilled water every day.ELISA detection of serum IFN-y,IL-6,IL-17A,IL-22 levels after 14 days,flow cytometry detection of peripheral Th1/Th17 cell ratio,qRT-PCR quantitative detection of CCL20,CXCL9,CXCL10,CCR6,and CXCR3 expression in the retina,Western Blot detection of retinal AMPK,P-AMPK,and SIRTI protein expression.Results:Caspi score showed that from the third day after self modeling,the inflammation score of the anterior segment of EAU rats in the model group was significantly higher than that in the blank group(P＜0.01).ELISA showed that compared with the blank group,the levels of IFN-γ,IL-6,IL-17A,and IL-22 of the model group,significantly increased(P＜0.05);Compared with the model group,the expression of inflammatory factors in the constant and high magnification groups of traditional Chinese medicine decreased significantly(P＜0.05).qRT-PCR showed that compared with the blank group,the levels of CXCL9,CXCL10,CCL20,CCR6,and CXCR3 in the model group were significantly increased(P＜0.05).Compared with the model group,the expression of chemokines and receptors in the high magnification group of traditional Chinese medicine was significantly downregulated(P＜0.05),while the expression of CXCL10,CCL20,CCR6,and CXCR3 in the constant Chinese medicine group was significantly reduced(P＜0.05).Flow cytometry analysis showed that compared with the blank group,the proportion of Th1 and Th17 cell subsets in the model group significantly increased(P＜0.05).Compared with the model group,the proportion of Th1 and Th17 cells in the high magnification group of traditional Chinese medicine was significantly down regulated(P＜0.05),while the proportion of Th1 cells in the constant dosage group of traditional Chinese medicine was significantly down regulated(P＜0.05).Western Blot showed that compared with the blank group,the expression of P-AMPK/AMPK and SIRTI in the model group was significantly reduced(P＜0.05).Compared with the model group,the expression of traditional Chinese medicine constant group and high magnification group significantly increased(P＜0.05).Conclusion:The mechanism of Qufeng Mingmu Pills in reducing inflammation and inhibiting Th1/Th17 cells in EAU rats may be related to the activation of the AMPK/SIRT1 pathway.
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