摘要
目的 探索咖啡因对斑马鱼幼鱼骨代谢的影响以及具体机制。方法 以100、200、400μmol/L的咖啡因溶液处理受精6 h后的AB系斑马鱼幼鱼,用E3溶液处理同胞幼鱼作为对照组,进行形态组织学观察,阿利新蓝染色评估软骨发育情况,茜素红染色观察骨矿化,单细胞RNA测序检测相关基因表达,并富集差异代谢物相关的基因本体(gene ontology,GO)和京都基因与基因组百科全书(kyoto encyclopedia of genes and genomes,KEGG)代谢通路。数据比较采用单因素方差分析。结果 高浓度咖啡因处理组幼鱼的头尾长度、软骨间距、关节间距、骨矿化面积、钙含量[(14.53±1.91)μm、(11.08±0.97)μm、(19.79±2.22)μm、(10 742.80±3 244.60)、(1 759.40±88.50)μg/g]均低于对照组[(19.4±1.61)μm、(14.99±1.19)μm、(24.46±2.59)μm、(16 831.20±3 055.80)、(2 272.10±100.10)μg/g],差异有统计学意义(P<0.05)。单细胞RNA测序显示咖啡因处理会显著影响不同细胞类型的成骨细胞的基因表达。GO和KEGG富集分析提示咖啡因会通过氧化磷酸化、MAPK信号通路和钙离子信号通路来影响骨代谢。结论 高浓度咖啡因可通过多种机制干扰斑马鱼幼鱼骨发育和代谢,对骨代谢有显著的负面影响。
Objective To investigate the bone metabolism effects of caffeine.Methods At 6 h after fertilization,zebrafish embryos were treated with 100,200,and 400μmol/L of caffeine,and controls were cultured with E3,followed by histological observation,alcian blue staining,alizarin red staining,single-cell RNA sequencing,GO(Gene Ontology)and KEGG(Kyoto Encyclopedia of Genes and Genomes)enrichment analyses.The statistical method was one-way analysis of variance.Results The cranio-caudal length,inter-chondral distance,inter-joint interval,bone mineralization,calcium levels in high caffeine treatment groups[(14.53±1.91)μm,(11.08±0.97)μm,(19.79±2.22)μm,(10742.80±3244.60),(1759.40±88.50)ug/g]significantly reduced as compared with those in the control group[(19.4±1.61)μm,(14.99±1.19)μm,(24.46±2.59)μm,(16831.20±3055.80),(2272.10±100.10)ug/g,P<0.05].Single-cell RNA sequencing revealed that caffeine would significantly influence osteoblast differentiation and function by regulating specific gene expression.GO and KEGG enrichment analyses showed that high-concentration caffeine treatment would affect zebrafish osteoblast function by regulating multiple key biological processes and signaling pathways,such as oxidative phosphorylation,MAPK signaling pathway and calcium signaling pathway.Conclusion It is inferred that high caffeine has a significant negative impact on bone metabolism,potentially interfering with normal bone development and metabolism through multiple mechanisms.
作者
冯诗缘
吴雯
姜宇
FENG Shiyuan;WU Wen;JIANG Yu(Department of General Practice,Wuxi No.2 People’s Hospital,Wuxi 214002,China)
出处
《中国骨质疏松杂志》
北大核心
2025年第12期1741-1745,共5页
Chinese Journal of Osteoporosis
基金
江苏省卫生健康委员会项目(Z2022027)。
