基于转录组学研究岩藻黄素在扰动剪切应力对EPCs炎性及增殖影响  

Transcriptomic-based analysis of the effects of fucoxanthin on the inflammatory and proliferation of EPCs on oscillatory shear stress

作　　者：傅洁 于洁 张晓芸[1] 李鑫[2] 金成文[2] 刘顺梅 成敏[1] 崔晓栋[1,2] FU Jie;YU Jie;ZHANG Xiao-yun;LI Xin;JIN Cheng-wen;LIU Shun-mei;CHENG Min;CUI Xiao-dong(Medical Physiology Laboratory,School of Basic Medicine Sciences,Weifang Medical University,Weifang 261053,China;Functional Laboratory,School of Basic Medicine Sciences,Weifang Medical University,Weifang 261053,China;Experimental Center for Medical Research,Weifang Medical University,Weifang 261053,China)

机构地区：[1]潍坊医学院基础医学院医学生理学实验室,山东潍坊261053 [2]潍坊医学院基础医学院机能学实验室,山东潍坊261053 [3]潍坊医学院医学研究实验中心,山东潍坊261053

出　　处：《中国海洋药物》2022年第3期34-42,共9页Chinese Journal of Marine Drugs

基　　金：国家自然科学基金项目(81870237,81700406);山东省自然科学基金项目(ZR2020MH20,ZR2020MD0940)资助。

摘　　要：目的 在扰动剪切应力(Oscillatory shear stress, OSS,±4 dynes/cm2, 1 Hz)作用下内皮祖细胞(Endothelial progenitor cells, EPCs)转录组学数据生物信息学分析的基础上,探讨岩藻黄素(Fucoxanthin, FUCO)对OSS作用下的EPCs炎性、增殖以及细胞周期等生物活性改变的影响作用。方法 采用Flexcell flow STR-4000流体系统模拟体内扰动流剪切应力环境;利用Illumina高通量测序平台对相应处理的EPCs进行转录组测序分析,并对筛选到的差异蛋白编码基因进行功能以及通路分析;利用实时荧光定量PCR技术(RT-qPCR)验证相应生物信息学数据;Western blot(WB)检测筛选基因-前列腺素内过氧化物合酶2(PTGS2)蛋白表达;采用EdU和PI染色分别进行EPCs增殖和周期检测。结果 与静止对照组相比,共计差异表达基因1 066个,涉及细胞增殖和死亡、感染性疾病等信号转导通路;GO功能分析显示,该类基因功能主要影响的过程有细胞及细胞组分、细胞进程、细胞器及生物进程等生物活动;根据P<0.05且差异倍数(FC)>2的条件,筛选显著差异基因数17个,并经RT-qPCR以及WB验证,显示PTGS2 mRNA差异表达最为显著(P<0.001),PTGS2蛋白表达差异明显(P<0.05);进一步的结果显示,与OSS组EPCs相比,FUCO处理的OSS组EPCs中PTGS2 mRNA表达显著降低(P<0.001),其蛋白表达被抑制(P<0.05),并且FUCO处理过的OSS组EPCs增殖活性以及DNA合成能力增强。结论 结合转录组数据基础,提示FUCO能够显著降低OSS引发的EPCs炎症反应,尤其明显抑制炎性因子PTGS2表达;促进EPCs细胞增殖和DNA合成,这为FUCO临床干预EPCs进而完善心血管疾病干细胞治疗提供理论依据。Objective On the basis of bioinformatics analysis from the transcriptomics data which imposed endothelial progenitor cells(EPCs) on the action of oscillatory shear stress(OSS, ± 4 dynes/cm2, 1 Hz), we investigated the role of fucoxanthin(FUCO) on the inflammatory, proliferation and cell cycle activities of endothelial progenitor cells(EPCs) on OSS. Methods The Flexcell Flow STR-4000 fluid system was used to simulate the disturbed blood flow shear stress in vivo. Illumina high-throughput sequencing platform was applied to analyze the transcriptome of the EPCs on OSS, and then analyze the functions and pathways of the differentially encoded genes. The corresponding bioinformatics data were verified by real-time fluorescence quantitative PCR(RT-qPCR). Western blot(WB) was used to detect the expression of PTGS2 protein. The cell proliferation and cycle of EPCs were detected by EdU and PI staining, respectively. Results Compared with the static control group, 1 066 differentially expressed genes were involved in signal transductive pathways associated with cell proliferation, death, and infectious diseases. GO enrichment analysis showed that the functions of these genes mainly affected biological activities such as cells and cell components, cell processes, organelles, and biological processes. Based on the condition of P<0.05 and difference multiple(FC) > 2, the number of significantly different genes was screened 17 and verified by RT-qPCR as well as WB, which showed that PTGS2 mRNA was most significantly differentially expressed(P<0.001) and PTGS2 protein was significantly differentially expressed(P<0.05). Further results showed that the mRNA expression of PTGS2 in the OSS group treated by FUCO was decreased(P<0.001) compared with the EPCs of the OSS group, contemporary the protein expression was inhibited(P<0.05). However, the proliferation activity and DNA synthesis capacity in the OSS group EPCs combinedly treated with FUCO were increased.Conclusion Combined with transcriptome data, FUCO could significantly red

关 键 词：岩藻黄素 动脉粥样硬化 内皮祖细胞 细胞周期 细胞增殖 

分 类 号：R915[医药卫生—微生物与生化药学]