未经证实:MicroRNAs(miRNAs)在植物和动物中转录后调节基因表达的能力是公认的。最近,miRNA-信使RNA(mRNA)的调控关系已经在生物过程中得到证实,包括成骨分化。本研究旨在寻找更多参与MC3T3-E1细胞成骨分化的候选miRNA-mRNA对。
UNASSIGNED:从基因表达Omnibus下载的基于MC3T3-E1的微阵列数据集(作为GSE46400)包括具有或不具有14天成骨细胞分化诱导的MC3T3-E1细胞。通过差异表达基因(DEG)搜索多个miRNA-mRNA预测数据库以获得miRNA-DEG调控网络对。将MC3T3-E1细胞培养并在成骨分化培养基中孵育14天。通过实时定量PCR(RT-qPCR)检测候选miRNA和mRNA在MC3T3-E1细胞中的表达。通过双荧光素酶报告基因测定和使用模拟miRNA或其抑制剂的实验来验证miRNA-mRNA相互作用。
UNASSIGNED:通过分析GSE46400数据集的原始数据,我们在有和没有成骨细胞诱导的MC3T3-E1细胞之间鉴定了715个上调的DEGs和603个下调的DEGs。MC3T3-E1细胞成骨分化过程中有7个重叠的miRNA-mRNA对,包括mmu-miR-204-5p-Arhgap11a,mmu-miR-211-5p-Arhgap11a,mmu-miR-24-3p-H2afx,mmu-miR-3470b-Chek2、mmu-miR-3470b-Dlgap5、mmu-miR-466b-3p-Chek1和mmu-miR-466c-3p-Chek1。Arhgap11a,H2afx,Chek2,Dlgap5和Chek1是成骨分化后MC3T3-E1细胞中hub基因下调,通过RT-qPCR结果验证。RT-qPCR还确定miR-204-5p和miR-24-3p的表达下降,同时miR-211-5p的表达升高。miR-3470b,miR-466b-3p,和miR-466c-3p在MC3T3-E1细胞,与未分化的MC3T3-E1细胞相比,具有成骨细胞诱导作用。双荧光素酶报告基因测定证明Arhgap11a是miR-211-5p的靶标。MiR-211-5p通过其模拟物的上调增加了MC3T3-E1细胞中的Arhgap11a表达。
UNASSIGNED:我们的研究表征了miR-211-5p靶向Arhgap11a促进MC3T3-E1细胞的成骨分化,这为骨修复过程中促进成骨过程提供了新的靶点。
UNASSIGNED: MicroRNAs (miRNAs) are well-recognized for their abilities to regulate gene expression post-transcriptionally in plants and animals. Recently, miRNA-messenger RNA (mRNA) regulatory relationships have been confirmed during biological processes, including osteogenic differentiation. This study aimed to find out more candidate miRNA-mRNA pairs involved in the osteogenic differentiation of MC3T3-E1 cells.
UNASSIGNED: An MC3T3-E1-based microarray dataset (accessioned as GSE46400) downloaded from the Gene Expression Omnibus included MC3T3-E1 cells with or without 14-day osteoblast differentiation osteoblast induction. Multiple miRNA-mRNA prediction databases were searched by differentially expressed genes (DEGs) to obtain pairs of a miRNA-DEG regulatory network. The MC3T3-E1 cells were cultured and incubated in the osteogenic differentiation medium for 14 days. The expressions of candidate miRNAs and mRNAs were determined by real-time quantitative PCR(RT-qPCR) in MC3T3-E1 cells. The miRNA-mRNA interactions were verified by dual-luciferase reporter gene assays and experiments using mimics miRNA or their inhibitors.
UNASSIGNED: We identified 715 upregulated DEGs and 603 downregulated DEGs between MC3T3-E1 cells with and without osteoblast induction by analyzing the raw data of the GSE46400 dataset. There were 7 overlapped miRNA-mRNA pairs identified during osteogenic differentiation of MC3T3-E1 cells, including mmu-miR-204-5p-Arhgap11a, mmu-miR-211-5p-Arhgap11a, mmu-miR-24-3p-H2afx, mmu-miR-3470b-Chek2, mmu-miR-3470b-Dlgap5, mmu-miR-466b-3p-Chek1, and mmu-miR-466c-3p-Chek1. The Arhgap11a, H2afx, Chek2, Dlgap5, and Chek1 were hub genes downregulated in MC3T3-E1 cells after osteogenic differentiation, verified by RT-qPCR results. The RT-qPCR also determined declined expressions of miR-204-5p and miR-24-3p concomitant with elevated expressions of miR-211-5p, miR-3470b, miR-466b-3p, and miR-466c-3p in the MC3T3-E1 cells, with osteoblast induction compared with undifferentiated MC3T3-E1 cells. Dual-luciferase reporter gene assays demonstrated Arhgap11a as the target of miR-211-5p. MiR-211-5p upregulation by its mimic increased Arhgap11a expression in MC3T3-E1 cells.
UNASSIGNED: Our study characterizes miR-211-5p targeting Arhgap11a promotes the osteogenic differentiation of MC3T3-E1 cells, which provides novel targets to promote the osteogenesis process during bone repair.