基于蛋白组学探讨哈蟆油对骨质疏松大鼠骨代谢调节作用机制  

Regulatory Mechanism of Oviductus Ranae on Bone Metabolism in Osteoporosis Rats Based on Proteomics

作　　者：李晓华[1] 仇广杰 曹占鸿 姜艳清 李晶[1] 李宜平[1] 杨擎[1] 石晓征[1] 李娜[1] 曲晓波[1] 李银清[1] Li Xiaohua;Qiu Guangjie;Cao Zhanhong;Jiang Yanqing;Li Jing;Li Yiping;Yang Qing;Shi Xiaozheng;Li Na;Qu Xiaobo;Li Yinqing(Changchun University of Chinese Medicine,Changchun 130117;Jilin University of Agricultural,Changchun 130118)

机构地区：[1]长春中医药大学,长春130117 [2]吉林农业大学,长春130118

出　　处：《中药药理与临床》2021年第3期103-108,共6页Pharmacology and Clinics of Chinese Materia Medica

基　　金：长春中医药大学“杏林学者工程”青年科学家项目培养计划项目;吉林省卫生计生委卫生与健康青年科技骨干培养计划项目(编号:2018Q043);吉林省大学生创新创业训练计划项目;吉林省教育厅项目(编号:JJKH20181266KJ);吉林省科技发展计划项目(编号:20170309005YY)。

摘　　要：目的:基于多肽体外标记定量技术(TMT)蛋白定量检测,研究哈蟆油调节大鼠骨代谢,防治绝经后骨质疏松作用。方法:采用蛋白裂解法提取大鼠股骨总蛋白,依次经过胰蛋白酶酶解、TMT试剂标记、HPLC分级、HPLC-MS联用技术鉴定蛋白,利用生物信息学分析方法进行差异表达蛋白注释、GO富集分析、通路富集分析和蛋白结构域富集分析,利用平行反应监测(PRM)技术验证典型特异表达蛋白。结果:蛋白质定量分析共鉴定到5654个蛋白,其中注释到4889个蛋白。与空白对照组比较,模型对照组参与富集度较高的信号通路中差异表达蛋白共22个(fold change>2.0,fold change<1/2.0,P<0.05),其中有6个蛋白发生表达上调,有16个蛋白发生表达下调。与模型对照组比较,哈蟆油0.075 g/kg组参与富集度较高的信号通路中差异表达蛋白共11个(fold change>2.0,fold change<1/2.0,P<0.05),其中有6个蛋白发生表达上调,有5个蛋白发生表达下调。这些差异蛋白重点参与了大鼠体内糖酵解/糖异生、细胞外基质(ECM)-受体相互作用、核糖体代谢、蛋白质消化吸收、脂肪酸降解、补体和凝血级联及心肌病、坏死症、百日咳、病毒病、红斑狼疮等疾病代谢途径。PRM实验验证肌钙蛋白I(Tnni1)、心肌肌钙蛋白C(Tnnc1)、丝氨酸蛋白酶的抑制剂(Serpina3k)、半乳糖凝集素(Galentin-1)及肌球蛋白轻链III(My13)蛋白表达趋势与TMT检测结果一致。结论:哈蟆油对骨质疏松大鼠骨代谢调节作用可能与Tnni1等11个差异表达蛋白相关,其中与Tnni1、Tnnc1、Serpina3k、Galentin-1及My13等5个蛋白密切相关。Objective:Based on the protein quantitative detection technology of TMT,to study the regulatory effect of Oviductus Ranae on the bone metabolism,and the preventive and therapeutic effect on postmenopausal osteoporosis in rats.Methods:The total femur protein was extracted by cell lysis method,followed by trypsin digestion,TMT reagent labeling,HPLC classification and HPLC-MS combination technology on Protein identification.Differentially expressed protein annotation,GO enrichment analysis,pathway enrichment analysis and protein domain enrichment analysis were performed by bioinformatics analysis.The typical and specific expressed proteins were verified by PRM technology.Results:A total of 5654 proteins were identified by protein quantitative analysis,and 4889 proteins were annotated.Compared with the control group,22 proteins with high different expression(fold change>2.0,fold change<1/2.0,P<0.05)were identified in the model group,in which 6 proteins were up-regulated and 16 proteins were down-regulated.Compared with the model group,11 proteins with high different expression(fold change>2.0,fold change<1/2.0,P<0.05)were identified in 0.075 g/kg Oviductus Ranae group,in which 6 proteins were up-regulated and 5 proteins were down-regulated.These differential proteins were mainly involved in the metabolic pathways of glycolysis/gluconeogenesis,ECM-receptor interaction,ribosome metabolism,protein digestion and absorption,fatty acid degradation,complement and coagulation cascades and cardiomyopathy,necrosis,pertussis,viral disease and red envelopes,etc.The expressions of Tnni1,Tnnc1,Serpina3 k,Galentin-1 and My13 detected by PRM experiment showed the similar trends with those by TMT detection.Conclusion:The regulatory effect of Oviductus Ranae on the bone metabolism in osteoporosis rats may be related to 11 differentially expressed proteins,which is closely related to Tnni1,Tnnc1,Serpina3 k,Galentin-1 and My13 proteins.

关 键 词：哈蟆油 蛋白质组学 骨质疏松 多肽体外标记定量技术 作用机制 

分 类 号：R285.5[医药卫生—中药学]