Abstract：
We investigated the role of miR-150-5p in osteoarthritic (OA) chondrocytes, as well as the possible regulatory role of long non-coding RNAs (lncRNAs) in miR-150-5p expression. TargetScan, StarBase, DIANA-LncBase, and Open Targets databases were used to predict miR-150-5p target genes, lncRNAs/miR-150-5p interactions, and OA-related genes. Protein-protein interaction (PPI) network was constructed using the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING). Gene ontology (GO) and pathway analysis were performed using Enrichr database. A publicly available RNA-seq dataset was retrieved to identify differentially expressed lncRNAs in damaged vs intact cartilage. We re-analyzed the retrieved RNA-seq data and revealed 177 differentially expressed lncRNAs in damage vs intact cartilage, including Nuclear Paraspeckle Assembly Transcript 1(NEAT1). MiR-150-5p, NEAT1, b-catenin, matrix metallopeptidase 13 (MMP-13), and ADAM metallopeptidase with thrombospondin type 1 motif 5 (ADAMTS-5) expressions were assessed by reverse transcription-quantitative PCR (RT-qPCR) and western blot assay. Knockout and transfection experiments were conducted to investigate the role of NEAT1/miR-150-5p/b-catenin in cartilage degradation. Bioinformatics analysis revealed that b-catenin was an OA-related miR-150-5p target. MiR-150-5p overexpression in OA chondrocytes resulted in decreased expression of b-catenin, as well as MMP-13 and ADAMTS-5, both being Wnt/b-catenin downstream target genes. NEAT1/miR-150-5p interaction was predicted by bioinformatics analysis, while NEAT1 knockout led to increased expression of miR-150-5p in OA chondrocytes. Moreover, inhibition of miR-150-5p reversed the repressive effects of NEAT1 silencing in b-catenin expression in OA chondrocytes. Our results support a possible catabolic role of NEAT1/miR-150-5p interaction in OA progression by regulating b-catenin expression.
摘要：
我们研究了miR-150-5p在骨关节炎（OA）软骨细胞中的作用，以及长链非编码RNA（lncRNA）在miR-150-5p表达中的可能调节作用。TargetScan,StarBase,DIANA-LncBase,和开放目标数据库用于预测miR-150-5p靶基因，lncRNAs/miR-150-5p相互作用，和OA相关基因。使用用于检索相互作用基因/蛋白质(STRING)的搜索工具构建蛋白质-蛋白质相互作用(PPI)网络。使用Enrichr数据库进行基因本体论(GO)和通路分析。检索了公开可用的RNA-seq数据集以鉴定受损与完整软骨中差异表达的lncRNA。我们重新分析了检索到的RNA-seq数据，揭示了177个差异表达的lncRNAs在损伤和完整的软骨中，包括核旁斑装配文字记录1(NEAT1)。MiR-150-5p,NEAT1,b-catenin,基质金属肽酶13（MMP-13），通过逆转录定量PCR（RT-qPCR）和蛋白质印迹法评估具有血小板反应蛋白1型基序5（ADAMTS-5）表达的ADAM金属肽酶。进行基因敲除和转染实验以研究NEAT1/miR-150-5p/b-catenin在软骨降解中的作用。生物信息学分析显示，b-catenin是OA相关的miR-150-5p靶标。MiR-150-5p在OA软骨细胞中的过表达导致b-catenin的表达降低,以及MMP-13和ADAMTS-5，两者都是Wnt/b-catenin下游靶基因。通过生物信息学分析预测NEAT1/miR-150-5p相互作用，而NEAT1基因敲除导致miR-150-5p在OA软骨细胞中的表达增加。此外，miR-150-5p的抑制逆转了NEAT1沉默对OA软骨细胞b-catenin表达的抑制作用.我们的结果支持NEAT1/miR-150-5p相互作用通过调节b-catenin表达在OA进展中的可能分解代谢作用。
