RNA结合蛋白LIN28对糖尿病肾病的保护机制研究

王艳丽; 李雁; 叶芳; 孙燕; 侯俊霞; 程洁萍; 艾比班; 李金峰; 张新菊

doi:10.11659/jjssx.11E022103

您当前的位置：

首页 >

文章列表页 >

RNA结合蛋白LIN28对糖尿病肾病的保护机制研究

基础研究 | 更新时间：2023-12-13

- RNA结合蛋白LIN28对糖尿病肾病的保护机制研究
- Protective mechanism of RNA binding protein LIN28 on diabetic nephropathy
- 局解手术学杂志 2023年32卷第12期页码：1048-1053
- 作者机构：
  
  新疆生产建设兵团医院内分泌科，新疆乌鲁木齐 830002
- 作者简介：
  
  张新菊，E-mail：x152729@163.com
- 基金信息：
  
  新疆生产建设兵团科技局社会发展领域(2018AB024)
- DOI：10.11659/jjssx.11E022103
  中图分类号： R692.9
- 纸质出版日期：2023-12-25，
  
  收稿日期：2022-11-18，
扫描看全文
王艳丽,李雁,叶芳等.RNA结合蛋白LIN28对糖尿病肾病的保护机制研究[J].局解手术学杂志,2023,32(12):1048-1053.

WANG Yan-li,LI Yan,YE Fang,et al.Protective mechanism of RNA binding protein LIN28 on diabetic nephropathy[J].JOURNAL OF REGIONAL ANATOMY AND OPERATIVE SURGERY,2023,32(12):1048-1053.
王艳丽,李雁,叶芳等.RNA结合蛋白LIN28对糖尿病肾病的保护机制研究[J].局解手术学杂志,2023,32(12):1048-1053. DOI： 10.11659/jjssx.11E022103.

WANG Yan-li,LI Yan,YE Fang,et al.Protective mechanism of RNA binding protein LIN28 on diabetic nephropathy[J].JOURNAL OF REGIONAL ANATOMY AND OPERATIVE SURGERY,2023,32(12):1048-1053. DOI： 10.11659/jjssx.11E022103.

摘要

目的

探索RNA结合蛋白LIN28对糖尿病肾病（DN）的保护机制。

方法

在HEK 293T细胞中通过腺病毒介导过表达或敲低LIN28。通过RNA-Seq转录组测序和生物信息学分析获得过表达LIN28后显著改变的基因和功能信号通路。体外过表达LIN28实验分组为Control组、Treatment组、Adv-LIN28-OE+D-核糖组和Adv-LIN28-NC+D-核糖组；敲低LIN28实验分组为Control组、Treatment组、Adv-shRNA LIN28+D-核糖组和Adv-shRNA NT+D-核糖组。通过ELISA法测定上述各组细胞上清液人晚期糖基化终产物（AGEs）的水平。通过Western blot法测定上述各组细胞内RAGE、NF-κB和MMP-2的表达水平。

结果

RNA-Seq转录组测序和GO分析及KEGG分析结果显示，过表达LIN28导致HEK 293T细胞内AGE-RAGE信号通路显著下调（

0.05）。ELISA结果显示，与Control组相比，Treatment组细胞产生大量AGEs（

0.05）；与Treatment组相比，Adv-LIN28-OE+D-核糖组细胞AGEs水平明显降低（

0.05），Adv-shRNA LIN28+D-核糖组细胞AGEs水平显著升高（

0.05）。Western blot结果显示，与Control组相比，Treatment组细胞内RAGE、NF-κB和MMP-2的表达水平显著升高（

0.05）；与Treatment组相比，Adv-LIN28-OE+D-核糖组细胞内RAGE、NF-κB和MMP-2的表达水平显著降低（

0.05），Adv-shRNA LIN28+D-核糖组细胞内RAGE、NF-κB和MMP-2的表达水平则显著升高（

0.05）。

结论

在体外细胞水平上通过腺病毒过表达LIN28，能够导致数千种基因发生显著性差异表达，特别是能够抑制在DN疾病进展中十分关键的糖尿病-并发症相关AGE-RAGE信号通路，发挥保护DN的作用。

Abstract

Objective

To explore the protective mechanism of RNA binding protein LIN28 on diabetic nephropathy （DN）.

Methods

LIN28 was overexpressed or knocked down by adenovirus in HEK 293T cells. The gene and functional signaling pathway significantly changed after overexpression of LIN28 were obtained through RNA Seq transcriptome sequencing and bioinformatics analysis. HEK 293T cells were divided into the Control group， the Treatment group， the Adv-LIN28-OE+D-ribose group and the Adv-LIN28-NC+D-ribose group in in vitro LIN28 overexpression studies. And HEK 293T cells were divided into the Control group， the Treatment group， the Adv-shRNA LIN28+D-ribose group and the Adv-shRNA NT+D-ribose group in in vitro LIN28 knockdown studies. ELISA was used to detect the levels of human advanced glycation end products （AGEs） of cells supernatants in the above groups. Western blot was used to detect the expression levels of RAGE， NF-κB and MMP-2 of cells in the above groups.

Results

The results of RNA-Seq transcriptome sequencing， GO analysis and KEGG analysis showed that overexpression of LIN28 resulted in a significant down-regulation of AGE-RAGE signaling pathway in HEK 293T cells （

0.05）. The results of ELISA showed that compared with the Control group， the cell in the Treatment group produced a large amount of AGEs （

0.05）； compared with the Treatment group， the AGEs level of cells in the Adv-LIN28-OE+D-ribose group was significantly decreased （

0.05）， while the AGEs level of cells in the Adv-shRNA LIN28+D-ribose group was increased（

0.05）. The results of Western blot showed that compared with the Control group， the expression levels of RAGE， NF-κB and MMP-2 of cells in the Treatment group were significantly increased （

0.05）； compared with the Treatment group， the expression levels of RAGE， NF-κB and MMP-2 of cells in the Adv-LIN28-OE+D-ribose group were significantly decreased （

0.05）， while the expression levels of RAGE， NF-κB and MMP-2 of cells in the Adv-shRNA LIN28+D-ribose group were significantly increased（

0.05）.

Conclusion

Overexpression of LIN28 by adenovirus at the cellular level in vitro can lead to significant differential expression of thousands of genes. In particular， it can inhibit the diabetes and complications-related AGE-RAGE signaling pathway， which is critical in the progression of DN disease， and play a role in protecting DN.

关键词

糖尿病肾病LIN28腺病毒AGE-RAGE信号通路NF-κBMMP-2

Keywords

diabetic nephropathyLIN28adenovirusAGE-RAGE signaling pathwayNF-κBMMP-2

references

American Diabetes Association.Classification and diagnosis of diabetes：standards of medical care in diabetes-2018 ［J］.Diabetes Care，2018，41（Suppl 1）：S13-S27.doi：10.2337/dc18-S002http://dx.doi.org/10.2337/dc18-S002.

Chatterjee S，Khunti K，Davies MJ.Type 2 diabetes ［J］.Lancet，2017，389（10085）：2239-2251.doi：10.1016/S0140-6736（17）30058-2http://dx.doi.org/10.1016/S0140-6736（17）30058-2.

Ganasegeran K，Hor CP，Jamil MFA，et al.A systematic review of the economic burden of type 2 diabetes in malaysia ［J］.Int J Environ Res Public Health，2020，17（16）：5723.doi：10.3390/ijerph17165723http://dx.doi.org/10.3390/ijerph17165723.

Zhang XX，Kong J，Yun K.Prevalence of diabetic nephropathy among patients with type 2 diabetes mellitus in China：a meta-analysis of observational studies ［J］.J Diabetes Res，2020，2020：2315607.doi：10.1155/2020/2315607http://dx.doi.org/10.1155/2020/2315607.

Koye DN，Magliano DJ，Nelson RG，et al.The global epidemiology of diabetes and kidney disease ［J］.Adv Chronic Kidney Dis，2018，25（2）：121-132.doi：10.1053/j.ackd.2017.10.011http://dx.doi.org/10.1053/j.ackd.2017.10.011.

Bommer C，Sagalova V，Heesemann E，et al.Global economic burden of diabetes in adults：projections from 2015 to 2030 ［J］.Diabetes Care，2018，41（5）：963-970.doi：10.2337/dc17-1962http://dx.doi.org/10.2337/dc17-1962.

Anders HJ，Huber TB，Isermann B，et al.CKD in diabetes：diabetic kidney disease versus nondiabetic kidney disease ［J］.Nat Rev Nephrol，2018，14（6）：361-377.doi：10.1038/s41581-018-0001-yhttp://dx.doi.org/10.1038/s41581-018-0001-y.

Lin YC，Chang YH，Yang SY，et al.Update of pathophysiology and management of diabetic kidney disease ［J］.J Formos Med Assoc，2018，117（8）：662-675.doi：10.1016/j.jfma.2018.02.007http://dx.doi.org/10.1016/j.jfma.2018.02.007.

Gerstberger S，Hafner M，Tuschl T.A census of human RNA-binding proteins ［J］.Nat Rev Genet，2014，15（12）：829-845.doi：10.1038/nrg3813http://dx.doi.org/10.1038/nrg3813.

Wilbert ML，Huelga SC，Kapeli K，et al.LIN28 binds messenger RNAs at GGAGA motifs and regulates splicing factor abundance ［J］.Mol Cell，2012，48（2）：195-206.doi：10.1016/j.molcel.2012.08.004http://dx.doi.org/10.1016/j.molcel.2012.08.004.

Zhu H，Shyh-Chang N，Segrè AV，et al.The Lin28/let-7 axis regulates glucose metabolism ［J］.Cell，2019，47（1）：81-94.doi：10.1016/j.cell.2011.08.033http://dx.doi.org/10.1016/j.cell.2011.08.033.

Sung Y，Jeong J，Kang RJ，et al.Lin28a expression protects against streptozotocin-induced β-cell destruction and prevents diabetes in mice ［J］.Cell Biochem Funct，2019，37（3）：139-147.doi：10.1002/cbf.3376http://dx.doi.org/10.1002/cbf.3376.

Tabrez S，Al-Shali KZ，Ahmad S，et al.Lycopene powers the inhibition of glycation-induced diabetic nephropathy：a novel approach to halt the AGE-RAGE axis menace ［J］.Biofactors，2020，41（5）：372-381.doi：10.1002/biof.1238http://dx.doi.org/10.1002/biof.1238.

Kim D，Langmead B，Salzberg SL.HISAT：a fast spliced aligner with low memory requirements ［J］.Nat Methods，2015，12（4）：357-360.doi：10.1038/nmeth.3317http://dx.doi.org/10.1038/nmeth.3317.

Trapnell C，Williams BA，Pertea G，et al.Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation ［J］.Nat Biotechnol，2010，28（5）：511-515.doi：10.1038/nbt.1621http://dx.doi.org/10.1038/nbt.1621.

Love MI，Huber W，Anders S.Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2 ［J］.Genome Biol，2014，15（12）：550.doi：10.1186/s13059-014-0550-8http://dx.doi.org/10.1186/s13059-014-0550-8.

Ilbay O，Nelson C，Ambros V.C.elegans LIN-28 controls temporal cell fate progression byregulating LIN-46 expression via the 5' UTR of lin-46 mRNA ［J］.Cell Rep，2021，36（10）：109670.doi：10.1016/j.celrep.2021.109670http://dx.doi.org/10.1016/j.celrep.2021.109670.

Lawson H，Vuong E，Miller RM，et al.The Makorin lep-2 and the lncRNA lep-5 regulate lin-28 to schedule sexual maturation of the C.elegans nervous system ［J］.Elife，2019，8：e43660.doi：10.7554/eLife.43660http://dx.doi.org/10.7554/eLife.43660.

Wang Y，Zhang B，Gao G，et al.Long non-coding RNA LINC00355 promotes the development and progression of colorectal cancer by elevating guanine nucleotide exchange factor T expression via RNA binding protein lin-28 homolog A ［J］.Front Oncol，2020，10：582669.doi：10.3389/fonc.2020.582669http://dx.doi.org/10.3389/fonc.2020.582669.

Tao L，Fan X，Sun J，et al.Metformin prevented high glucose-induced endothelial reactive oxygen species via OGG1 in an AMPKα-Lin-28 dependent pathway ［J］.Life Sci，2021，268：119015.doi：10.1016/j.lfs.2020.119015http://dx.doi.org/10.1016/j.lfs.2020.119015.

Wu XQ，Zhang DD，Wang YN，et al.AGE/RAGE in diabetic kidney disease and ageing kidney ［J］.Free Radic Biol Med，2021，171：260-271.doi：10.1016/j.freeradbiomed.2021.05.025http://dx.doi.org/10.1016/j.freeradbiomed.2021.05.025.

de Vos WM，Tilg H，Van Hul M，et al.Gut microbiome and health：mechanistic insights ［J］.Gut，2022，71（5）：1020-1032.doi：10.1136/gutjnl-2021-326789http://dx.doi.org/10.1136/gutjnl-2021-326789.

Chen Y，Guo TL.Dietary advanced glycation end-products elicit toxicological effects by disrupting gut microbiome and immune homeostasis ［J］.J Immunotoxicol，2021，18（1）：93-104.doi：10.1080/1547691X.2021.1959677http://dx.doi.org/10.1080/1547691X.2021.1959677.

Qi W，Niu J，Qin Q，et al.Glycated albumin triggers fibrosis and apoptosis via an NADPH oxidase/Nox4-MAPK pathway-dependent mechanism in renal proximal tubular cells ［J］.Mol Cell Endocrinol，2015，405：74-83.doi：10.1016/j.mce.2015.02.007http://dx.doi.org/10.1016/j.mce.2015.02.007.

马壮，修光辉，熊伟，等.血管内皮糖萼的研究进展［J］.广东医学，2019，40（14）：2113-2116.doi：10.13820/j.cnki.gdyx. 20183583http://dx.doi.org/10.13820/j.cnki.gdyx.20183583.

Dou L，Jourde-Chiche N.Endothelial toxicity of high glucose and its by-products in diabetic kidney disease ［J］.Toxins （Basel），2019，11（10）：578.doi：10.3390/toxins11100578http://dx.doi.org/10.3390/toxins11100578.

Guo B，Shan SK，Xu F，et al.Protective role of small extracellular vesicles derived from HUVECs treated with AGEs in diabetic vascular calcification ［J］.J Nanobiotechnology，2022，20（1）：334.doi：10.1186/s12951-022-01529-zhttp://dx.doi.org/10.1186/s12951-022-01529-z.

Wang Y，Zhu X，Yuan S，et al.TLR4/NF-κB signaling induces GSDMD-related pyroptosis in tubular cells in diabetic kidney disease ［J］.Front Endocrinol （Lausanne），2019，10：603.doi：10.3389/fendo.2019.00603http://dx.doi.org/10.3389/fendo.2019.00603.

Zhou Y，Ma XY，Han JY，et al.Metformin regulates inflammation and fibrosis in diabetic kidney disease through TNC/TLR4/NF-κB/miR-155-5p inflammatory loop［J］.World J Diabetes，2021，12（1）：19-46.doi：10.4239/wjd.v12.i1.19http://dx.doi.org/10.4239/wjd.v12.i1.19.

Ramnath RD，Butler MJ，Newman G，et al.Blocking matrix metalloproteinase-mediated syndecan-4 shedding restores the endothelial glycocalyx and glomerular filtration barrier function in early diabetic kidney disease ［J］.Kidney Int，2020，97（5）：951-965.doi：10.1016/j.kint.2019.09.035http://dx.doi.org/10.1016/j.kint.2019.09.035.

Bai S，Xiong X，Tang B，et al.Exosomal circ_DLGAP4 promotes diabetic kidney disease progression by sponging miR-143 and targeting ERBB3/NF-κB/MMP-2 axis ［J］.Cell Death Dis，2020，11（11）：1008.doi：10.1038/s41419-020-03169-3http://dx.doi.org/10.1038/s41419-020-03169-3

浏览量

下载量

CSCD

文章被引用时，请邮件提醒。

提交

工具集

关联资源

富勒醇促进小鼠皮肤创面愈合的疗效及机制

二肽基肽酶抑制剂类似物对LPS诱导的小胶质细胞炎症反应的抑制作用

Ad-shRNA-NgR转染EAE大鼠脑组织安全性评估

糖尿病肾病发病机制及治疗的研究进展