摘要
[目的]基于高通量测序技术,探究绝经后骨质疏松症(postmenopausal osteoporosis,PMOP)肾阴虚证的潜在机制。[方法]分离培养PMOP肾阴虚证和骨量正常对照组骨髓间充质干细胞(bone mesenchymal stem cells,BMSCs),从BMSCs外泌体中提取RNA,利用芯片测序技术和生物信息学分析mRNA的表达谱和功能。使用定量逆转录聚合酶链式反应(quantitative reverse transcription polymerase chain reaction,RT-qPCR)分析20例PMOP肾阴虚证患者和20例骨量正常对照组受试者差异表达的mRNA。[结果]与骨量正常对照组比较,PMOP肾阴虚证BMSCs外泌体检测出174个差异表达的mRNA,其中92个上调,82个下调;RT-qPCR鉴定出9个关键m RNA(ATM、COL8A1、GABARAPL1、HLA-DQA1、HSPA1L、MAX、PPP2R2D、PTHLH和WWTR1),与基因芯片数据相一致,两组之间差异有统计学意义(P<0.001)。功能分析表明,PMOP肾阴虚证患者差异表达的mRNA可能靶向参与丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)、自噬调节、磷脂酰肌醇3激酶-蛋白激酶B(phosphatidylinositol 3 kinase-protein kinase B,PI3K-AKT)和叉头框转录因子O(forkhead box transcription factor O,FoxO)亚族等信号通路。[结论] PMOP肾阴虚证患者BMSCs外泌体中差异表达的基因可能作为新型的生物标志物,为PMOP肾阴虚证的靶向治疗提供有价值的信息。
[Objective] To explore the underlying mechanism of kidney Yin deficiency syndrome in postmenopausal osteoporosis(PMOP)based on high throughput sequencing technology. [Methods] Bone mesenchymal stem cells(BMSCs) from PMOP patients with kidney Yin deficiency syndrome and controls were isolated and cultured, and exosomes were extracted. Differential expression mRNA of BMSCs exosomes was identified by high throughput sequencing technology, and functional analysis of differential expression mRNA was performed by bioinformatics technology. Quantitative reverse transcription polymerase chain reaction(RT-qPCR) was further used to verify differentially expressed mRNAs in 20 pairs of clinical samples. [Results] Compared with normal bone mass control group, 174differentially expressed mRNA were detected by BMSCs exosomes with kidney Yin deficiency syndrome of PMOP, of which 92 were upregulated and 82 were down-regulated. The nine key mRNAs(ATM, COL8A1, GABARAPL1, HLA-DQA1, HSPA1L, MAX, PPP2R2D,PTHLH and WWTR1) identified by RT-qPCR were consistent with the microarray data, there was significant difference between the two groups(P <0.001). Functional analysis showed that the differentially expressed mRNA of PMOP patients with kidney Yin deficiency syndrome may target and participate in mitogen-activated protein kinase(MAPK) signaling pathway, autophagy regulation,phosphatidylinositol 3 kinase-protein kinase B(PI3K-Akt) signaling pathway and forkhead box transcription factor O(FoxO) signaling pathway. [Conclusion] The differentially expressed genes in BMSCs exosomes of PMOP patients with kidney Yin deficiency syndrome may be used as novel biomarkers and provide valuable information for the targeted therapy of PMOP patients with kidney Yin deficiency syndrome.
作者
方莉萍
姜丹生
林树梁
吴俊琪
王金华
浮苗
FANG Liping;JIANG Dansheng;LIN Shuliang(Jinhua Municipal Central Hospital,Jinhua 321000,China)
出处
《浙江中医药大学学报》
CAS
2022年第10期1077-1085,共9页
Journal of Zhejiang Chinese Medical University
基金
浙江省中医药科技计划项目(2021ZA142)
浙江省医药卫生科技计划项目(2021KY1180)
金华市中心医院中青年科研启动基金项目(JY2019-2-08)。
关键词
外泌体
绝经后骨质疏松症肾阴虚证
骨髓间充质干细胞
mRNA
基因芯片
生物信息学
exosome
kidney Yin deficiency syndrome of postmenopausal osteoporosis
bone marrow mesenchymal stem cells
mRNA
microarray
bioinformatics
