龟鹿二仙胶对膝骨关节炎大鼠肠道菌群的影响:机器学习与16S rDNA分析  

Effects of Guilu Erxian Glue on gut microbiota in rats with knee osteoarthritis:machine learning and 16S rDNA analysis

作　　者：顾富城 杨美鑫 吴伟欣 蔡玮俊 钦洋溢 孙铭一 孙健 耿秋东 李楠 Gu Fucheng;Yang Meixin;Wu Weixin;Cai Weijun;Qin Yangyi;Sun Mingyi;Sun Jian;Geng Qiudong;Li Nan(College of Traditional Chinese Medicine,Fujian University of Traditional Chinese Medicine,Fuzhou 350122,Fujian Province,China;Key Laboratory of Traditional Chinese Medicine Orthopedics and Sports Rehabilitation,Ministry of Education,Fujian University of Traditional Chinese Medicine,Fuzhou 350122,Fujian Province,China)

机构地区：[1]福建中医药大学中医学院,福建省福州市350122 [2]福建中医药大学中医骨伤及运动康复教育部重点实验室,福建省福州市350122

出　　处：《中国组织工程研究》2026年第4期1058-1072,共15页Chinese Journal of Tissue Engineering Research

基　　金：福建省省级财政专项(X2022001-财政专项;X2022002-财政专项),项目负责人:李楠;福建省自然科学基金(2023J01865),项目负责人:李楠;福建省自然科学基金(2022J01364),项目负责人:耿秋东。

摘　　要：背景:龟鹿二仙胶由龟甲胶、鹿角胶、枸杞子和人参组成。在早期的临床观察中,发现使用龟鹿二仙胶治疗肝肾不足型膝骨关节炎患者能有效缓解膝关节疼痛、增加运动范围、提高行走能力。然而,口服龟鹿二仙胶能够对膝关节产生局部治疗作用的确切机制尚不清楚。目的:研究龟鹿二仙胶对膝骨关节炎大鼠肠道菌群的影响,并利用16SrDNA测序和机器学习分析评估其作用机制。方法:将18只雄性SD大鼠随机分为3组:空白组、模型组、龟鹿二仙胶组,每组8只。模型组和龟鹿二仙胶组采用内侧半月板失稳手术制备膝骨关节炎模型。造模成功后,龟鹿二仙胶组灌胃龟鹿二仙胶水煎剂,空白组和模型组则给予等量的蒸馏水。持续干预28 d后,使用高效液相色谱技术检测龟鹿二仙胶有效成分;MRI成像观察大鼠膝关节软骨情况采集各组大鼠粪便样本,使用试剂盒提取DNA,通过PCR扩增及纯化,构建Illumina测序文库,利用Illumina MiSeq平台进行高通量测序,生成原始序列数据。得到原始数据后,使用QIIME2软件对数据进行处理,使用线性判别分析效应值和随机森林算法对微生物数据进行差异性物种筛选,使用KEGG和MetaCyc功能通路分析探索关键菌群与实验组间的关联,使用线性判别分析效应值和随机森林算法筛选差异性物种,使用关联网络分析微生物群落间的相互作用,结合机器学习方法分析肠道菌群组成和变化。结果与结论:(1)龟鹿二仙胶中药复方进行LC-MS成分鉴定,共指出7种有效成分。(2)MRI成像显示,龟鹿二仙胶组大鼠高密度影的滑膜炎症范围减少,且股骨头内侧髁软骨较模型组退化较少。(3)16SrDNA测序显示模型组大鼠表现出明显的菌群失衡,在门水平上厚壁菌门和拟杆菌门的丰度显著降低,而变形菌门的比例显著增加(P<0.05)。龟鹿二仙胶组大鼠的肠道菌群结构有了显著改善,其厚壁菌门和拟杆BACKGROUND:The Guilu Erxian Glue consists of Testudinis Plastrum,Cornu Cervi,Lycii Fructus,and Ginseng Radix.In earlier clinical observations,it is discovered that using Guilu Erxian Glue to treat patients with liver-kidney deficiency type knee osteoarthritis effectively alleviated knee pain,increased the range of motion,and improved walking ability.However,the exact mechanism by which oral administration of Guilu Erxian Glue can produce local therapeutic effects on the knee joint is still unclear.OBJECTIVE:To investigate the effects of Guilu Erxian Glue on gut microbiota in rats with knee osteoarthritis and to evaluate its mechanism using 16S rDNA sequencing and machine learning analysis.METHODS:Totally 18 female SD rats were randomly divided into three groups:blank group,model group,and Guilu Erxian Glue group,with 8 rats in each group.A knee osteoarthritis model was prepared using the destabilization of the medial meniscus surgical method.After successful modeling,the Guilu Erxian Glue group was given a decoction of Guilu Erxian Glue by gavage,while the blank and model groups were given an equal amount of distilled water.After 28 days of continuous intervention,high performance liquid chromatography was used to detect the active ingredients of Guilu Erxian Glue.MRI imaging was used to observe the condition of rat knee articular cartilage.Fecal samples were collected;DNA was extracted using a kit,amplified and purified by PCR,and an Illumina sequencing library was constructed.The Illumina MiSeq platform was used for high-throughput sequencing to generate raw sequence data.After obtaining the raw data,QIIME2 software was used to process the data.Linear Discriminant Analysis Effect Size analysis and random forest algorithm were used to screen for differential species in microbial data.KEGG and MetaCyc functional pathway analyses were used to explore the association between key microbial communities and experimental groups.Linear discriminant analysis effect values and random forest algorithm were used to screen f

关 键 词：龟鹿二仙胶 膝骨关节炎 肠道菌群 16SrDNA测序 机器学习 Α多样性 Β多样性 差异通路 

分 类 号：R452[医药卫生—治疗学] R318[医药卫生—临床医学] R285.5