Investigating the mechanisms of elevated RNA oxidation impacting pancreatic beta-cells utilizing whole transcriptome sequencing
10.3760/cma.j.issn.0254-9026.2024.07.021
- VernacularTitle:基于全转录组测序分析核糖核酸氧化影响胰岛β细胞的分子机制
- Author:
Faqiang ZHOU
1
;
Anqi WANG
;
Wenze ZHANG
;
Ying LIU
;
Yizhi ZHANG
;
Yujuan LIANG
;
Jianping CAI
;
Wanxia WANG
Author Information
1. 兰州大学第一临床医学院，兰州 730000
- Keywords:
RNA;
Pancreatic beta cell;
Diabetes;
Sequencing
- From:
Chinese Journal of Geriatrics
2024;43(7):889-898
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the impact of elevated glucose-induced RNA oxidation on pancreatic β-cell function, activity, and underlying molecular mechanisms.Methods:Rat pancreatic islet β-cell tumour INS-1 cells were cultured in vitro and subjected to nucleic acid oxidation assessment using isotope dilution ultra-high performance liquid tandem mass spectrometry(ID LC MS/MS)following high glucose exposure.In vitro simulation of increased RNA oxidation in INS-1 cells was achieved using 8-oxoguanosine-5'-triphosphate(8-oxoGTP).Cell proliferation was evaluated through CCK-8 assay, apoptosis was measured via flow cytometry, and gene expression of insulin(INS), pancreatic-duodenal homologous cassette 1(PDX1), cysteine-aspartate proteinase 3(Casp3), and cysteine aspartate protease 6(Casp6)was analyzed at the mRNA level.Additionally, whole transcriptome sequencing was performed to elucidate the molecular mechanisms underlying the impact of RNA oxidation on INS-1 cells.Results:Elevated glucose levels induced an increase in RNA oxidation within INS-1 cells.This heightened RNA oxidation led to the inhibition of INS-1 cell proliferation, a reduction in mRNA levels of INS and PDX1 genes, and the promotion of apoptosis-related casp3 and casp6 gene mRNA synthesis.Transcriptome sequencing analysis unveiled that the elevated RNA oxidation caused differential expression of mRNA, lncRNA, miRNA, and circRNA in INS-1 cells.This included a significant down-regulation of transcription factors such as Mafa, Pdx1, Pax6, and Mnx1, alongside an up-regulation of various miRNAs like rno-miR-124-3p, rno-miR-133a-3p, rno-miR-3120, rno-miR-212-3p, and rno-miR-7a-2-3p.These molecular changes contributed to the altered expression of associated lncRNAs, ultimately hindering insulin synthesis and secretion, as well as β-cell proliferation.Conclusions:Increased RNA oxidation down-regulates the levels of key β-cell transcription factor mRNAs, contributes to the differential expression of related non-coding RNAs(ncRNAs), particularly lncRNAs, impacts β-cell insulin synthesis and secretion, hinders cell proliferation, and serves as a significant factor in β-cell dysfunction and decreased activity in type 2 diabetes mellitus(T2DM).
