整合“转录组-基因调控网络”探讨白虎加桂枝汤君药石膏的性-效关联内涵  

Exploration on Property-efficacy Association of Sovereign Drug Gypsum Fibrosum in Baihu Guizhitang by Integrating Transcriptomics with Gene Regulatory Network Analysis

作　　者：李玮婕 毛霞[1] 刘毓东 王可欣 张彦琼 林娜[1] LI Weijie;MAO Xia;LIU Yudong;WANG Kexin;ZHANG Yanqiong;LIN Na(Institute of Chinese Materia Medica,China Academy of Chinese Medical Sciences,Beijing 100700,China)

机构地区：[1]中国中医科学院中药研究所,北京100700

出　　处：《中国实验方剂学杂志》2023年第1期52-60,共9页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金重点项目(81630107);中国中医科学院科技创新项目(CI2021A03808,CI2021A01508)。

摘　　要：目的:从“性-效”关联角度,采用“转录组-基因调控网络”整合研究策略,系统评价白虎加桂枝汤君药石膏在全方“热者寒之”药效作用中的贡献度和贡献形式。方法:20只雄性Lewis大鼠随机分为4组,每组5只:正常组、佐剂诱导型关节炎(AIA)热证模型组、AIA热证+白虎加桂枝汤组(BHGZT,21.4 g·kg^(-1))、AIA热证+白虎加桂枝汤去石膏拆方组(BHGZT-GYP,10.7 g·kg^(-1))。利用热痹经典动物模型-AIA热证大鼠来源的转录组表达谱数据,通过“病证基因-药物效应靶标”相互作用网络挖掘,预测石膏在白虎加桂枝汤干预热痹的主要作用环节。基于AIA热证大鼠模型从受累关节情况、关节临床积分、肿胀度、关节表面温度、疼痛阈值、关节损伤等疾病严重指标和病证结合指标情况,以及体内“免疫-炎症”失衡调节层面进行整体验证,并基于体外3T3-L1前脂肪细胞开展体外成脂实验验证。结果:白虎加桂枝汤君药石膏在干预热痹中的主要作用环节是调节机体能量代谢的紊乱和矫正“免疫-炎症”的失衡。动物实验观察可见,白虎加桂枝汤及其去石膏拆方均可显著缓解AIA热证大鼠受累关节红肿、畸变和疼痛等症状(P<0.01),并抑制受累膝/踝关节损伤、滑膜炎症、骨破坏等病理改变(P<0.05),且全方作用明显优于去石膏拆方(P<0.05)。机制研究表明,白虎加桂枝汤及其去石膏拆方可显著抑制AIA热证大鼠的胸腺/脾脏损伤和内脏指数(P<0.05),并降低TLR4、TNF-α、IL-6、IL^(-1)β及IL^(-1)8等炎症因子的表达(P<0.05)。体外3T3-L1前脂肪细胞实验表明,与去石膏拆方组相比,白虎加桂枝汤全方能够更加显著地抑制脂肪细胞成脂,且单纯石膏对脂滴含量的抑制作用显著增强,表明石膏在全方抑制成脂中的重要作用。结论:白虎加桂枝汤君药石膏在全方缓解热痹相关的能量代谢紊乱和矫正“免疫-炎症”失衡中发挥着重要作用Objective:To systematically explore the roles and contributions of the sovereign drug Gypsum Fibrosum contained in Baihu Guizhitang(BHGZT)against rheumatoid arthritis(RA)with heat syndrome from property-efficacy association by an approach integrating transcriptomics with gene regulatory network analysis.Method:A total of 20 male Lewis rats were randomly assigned into 4 groups:a control group(n=5),a group of adjuvant-induced arthritis rat model with heat syndrome(AIA-H,n=5),an AIA-H+BHGZT group(BHGZT,21.4 g·kg^(-1),n=5),and an AIA-H+BHGZT without Gypsum Fibrosum group(BHGZT-GYP,10.7 g·kg^(-1),n=5).We combined the gene expression profiling based on AIA-H rat model and"disease-gene-drug effective target"network analysis to predict the major function of Gypsum Fibrosum contained in BHGZT against RA with heat syndrome.Furthermore,in vivo experiments with the AIA-H rat model were performed to validate the therapeutic effects on the severity of arthritis based on the representative images of arthritis,limb diameter,infrared thermography,pain thresholds,and joint injury,as well as at the level of immunityinflammation imbalance.Oil Red O staining was employed for the differentiation of 3T3-L1 pre-adipocytes in the AIA-H rats treated by BHGZT,BHGZT-GYP,and GYP.Result:Gene expression profiling and network analysis demonstrated that Gypsum Fibrosum mainly regulated the energy metabolism disorders and the immunity-inflammation imbalance during the development and progression of RA.In vivo experiments showed that both BHGZT and BHGZT-GYP reduced the disease severity of AIA-H rats(P<0.01)by relieving joint redness and distortion,decreasing arthritis score and limb diameter,elevating pain thresholds,alleviating joint erosion,joint inflammation,and bone destruction(P<0.05).Notably,BHGZT outperformed BHGZT-GYP(P<0.05).Both BHGZT and BHGZT-GYP inhibited the pathological changes and decreased the indexes of thymus and spleen(P<0.05),and down-regulated the expression of inflammatory cytokines including Toll-like receptor 4(TLR4),tu

关 键 词：白虎加桂枝汤 石膏 热痹 “性-效”关联 能量代谢 “免疫-炎症”平衡 

分 类 号：R2-0[医药卫生—中医学] R33R289R684.3