二氧化硅纳米颗粒诱导血管内皮细胞自噬激活的转录组分析及实验验证  

Transcriptome analysis and experimental verification of autophagy activation induced by silica nanoparticles

作　　者：阮晨 吴冉[3] 王来友 杜佳 曹敬博 Ruan Chen;Wu Ran;Wang Laiyou;Du jia;Cao Jingbo(Henan Key Laboratory of Industrial Microbial Resources and Fermentation,Nanyang Institute of Technology,Nanyang 473004,China;School of Biological and Chemical Engineering,Nanyang Institute of Technology,Nanyang 473004,China;School of Information Engineering,Nanyang Institute of Technology,Nanyang 473004,China;College of Mathematics and Statistics,Liaoning University,Shenyang 110036,China)

机构地区：[1]南阳理工学院河南省工业微生物资源与发酵技术重点实验室,河南南阳473004 [2]南阳理工学院生物与化学工程学院,河南南阳473004 [3]南阳理工学院信息工程学院,河南南阳473004 [4]辽宁大学数学与统计学院,沈阳110036

出　　处：《河南师范大学学报(自然科学版)》2025年第3期143-148,I0009,共7页Journal of Henan Normal University(Natural Science Edition)

基　　金：河南省自然科学基金(242300420500);河南省高等学校重点科研项目(23A180023);国家自然科学基金(22201146);2024-2025年南阳市科技计划项目.

摘　　要：为寻找二氧化硅纳米颗粒诱导细胞自噬激活的调控蛋白,用20 nm的二氧化硅纳米颗粒处理大鼠血管内皮细胞株,对细胞样品进行转录组测序分析及试验.根据CCK-8方法测定的细胞死亡率及自噬Marker蛋白的表达量确定诱导时长,由此制备细胞样品并进行转录组测序及分析预测,通过RNA干扰实验筛选具有自噬调控功能的蛋白质.研究结果表明,20 nm二氧化硅纳米颗粒处理细胞的较优诱导时间为24 h,自噬Marker蛋白LC3B-Ⅱ的表达随着纳米颗粒处理时间的增加而增高;在转录组测序结果中共筛选到295个差异表达基因,差异基因的KEGG富集分析结果显示大部分基因参与多个自噬调控信号通路;通过RNA干扰实验和免疫印迹实验验证,结果表明干扰Arrdc4基因的表达可以降低细胞内LC3B-Ⅱ的表达水平.综合考虑所有实验结果,Arrdc4可能通过KEGG显著富集的信号通路影响二氧化硅纳米颗粒诱导的细胞自噬,其调控机制仍有待深入挖掘.In order to find the regulatory proteins of autophagy activation induced by silica nanoparticles,the rat vascular endothelial cell line was treated with 20 nm silica nanoparticles,and the cell samples were subjected to transcriptome sequencing analysis and subsequent verification.The optimal induction time was determined according to the cell death rate by CCK-8 kits and the expression of autophagy Marker protein.Then cell samples which were treated with 20 nm silica nanoparticles were prepared for transcriptome sequencing and analysis.The differentially expressed genes were selected for RNA interference experiments to screen the proteins with autophagy regulatory function.The results showed that the optimal induction time of cells treated with 20 nm silica nanoparticles was 24 h.The expression level of autophagy Marker protein LC3B-II increased with the increase of nanoparticle treatment time.A total of 295 differentially expressed genes were identified in the transcriptome sequencing results.KEGG enrichment analysis of the differentially expressed genes showed that most of the genes were mainly involved in multiple autophagy regulatory pathways.The subsequent experiments were performed on the differentially expressed genes,which were verified by RNA interference experiment and Western blot.The experimental results showed that interfering the expression of Arrdc4 gene could reduce the expression level of LC3B-Ⅱ in cells.In this study,we conducted transcriptome sequencing analysis and subsequent experiments to verify the cell autophagy induced by silica nanoparticles,and found that interference with Arrdc4 gene expression could reduce the protein expression of LC3B-Ⅱ in cells.Considering all the experimental results,Arrdc4 may affect silica nanoparticles-induced autophagy through KEGG significantly enriched signaling pathways,and the regulatory mechanism remains to be further explored.

关 键 词：二氧化硅纳米颗粒 自噬 转录组 Arrdc4基因 

分 类 号：Q291[生物学—细胞生物学]