Yun Chen, Kun Huang, Wenjun Ji, Miao Huang, Jincheng Sima, Jin Li, Hao Song, Wei Xiong, Chao-Qun Ma
{"title":"LncRNA NR2F1-AS1通过miR-423-5p参与骨折愈合中的成骨分化。","authors":"Yun Chen, Kun Huang, Wenjun Ji, Miao Huang, Jincheng Sima, Jin Li, Hao Song, Wei Xiong, Chao-Qun Ma","doi":"10.1007/s10616-025-00786-8","DOIUrl":null,"url":null,"abstract":"<p><p>To investigate the function and mechanism of action of LncRNA NR2F1-AS1 involved in osteogenic differentiation process. An in vitro model was constructed by osteogenic differentiation-induced stimulation (OS) on the hFOB1.19 cell line. Real-time fluorescence quantitative PCR (RT-qPCR) was performed to detect the expression of NR2F1-AS1, miR-423-5p and osteogenic differentiation markers (RUNX2, OCN, OPN). Cell Counting Kit-8 (CCK8) method and flow cytometry were observed cell proliferation and apoptosis, respectively. Enzyme linked immunosorbent assay (ELISA) tested alkaline phosphatase (ALP) activity. Dual-Luciferase Report (DLR) assay and RNA immunoprecipitation (RIP) verified gene interactions. Bioinformatics methods predicted downstream target genes and their pathways of action. OS increased osteogenic differentiation markers and NR2F1-AS1 expression and decreased miR-423-5p levels. Transfection of si- NR2F1-AS1 promoted OS osteoblast apoptosis, but inhibited cell proliferation, ALP activity and osteogenic differentiation marker expression. NR2F1-AS1 is mostly present in the cytoplasm and is involved in the osteogenic differentiation process by down-regulating miR-423-5p. The use of miR-423-5p inhibitor can resist apoptosis induced by silencing NR2F1-AS1, promote osteoblast proliferation, activate ALP activity, and induce osteogenic differentiation process in osteoblasts. Bioinformatics prediction identified 82 target genes that might be involved in osteogenic differentiation, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis suggested that they were mainly associated with inter-synaptic formation and signaling. NR2F1-AS1 may promote osteoblast proliferation stimulate ALP activity, and induce osteogenic differentiation by down-regulating miR-423-5p.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s10616-025-00786-8.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"135"},"PeriodicalIF":2.0000,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12222589/pdf/","citationCount":"0","resultStr":"{\"title\":\"LncRNA NR2F1-AS1 is involved in osteogenic differentiation in fracture healing via miR-423-5p.\",\"authors\":\"Yun Chen, Kun Huang, Wenjun Ji, Miao Huang, Jincheng Sima, Jin Li, Hao Song, Wei Xiong, Chao-Qun Ma\",\"doi\":\"10.1007/s10616-025-00786-8\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>To investigate the function and mechanism of action of LncRNA NR2F1-AS1 involved in osteogenic differentiation process. An in vitro model was constructed by osteogenic differentiation-induced stimulation (OS) on the hFOB1.19 cell line. Real-time fluorescence quantitative PCR (RT-qPCR) was performed to detect the expression of NR2F1-AS1, miR-423-5p and osteogenic differentiation markers (RUNX2, OCN, OPN). Cell Counting Kit-8 (CCK8) method and flow cytometry were observed cell proliferation and apoptosis, respectively. Enzyme linked immunosorbent assay (ELISA) tested alkaline phosphatase (ALP) activity. Dual-Luciferase Report (DLR) assay and RNA immunoprecipitation (RIP) verified gene interactions. Bioinformatics methods predicted downstream target genes and their pathways of action. OS increased osteogenic differentiation markers and NR2F1-AS1 expression and decreased miR-423-5p levels. Transfection of si- NR2F1-AS1 promoted OS osteoblast apoptosis, but inhibited cell proliferation, ALP activity and osteogenic differentiation marker expression. NR2F1-AS1 is mostly present in the cytoplasm and is involved in the osteogenic differentiation process by down-regulating miR-423-5p. The use of miR-423-5p inhibitor can resist apoptosis induced by silencing NR2F1-AS1, promote osteoblast proliferation, activate ALP activity, and induce osteogenic differentiation process in osteoblasts. Bioinformatics prediction identified 82 target genes that might be involved in osteogenic differentiation, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis suggested that they were mainly associated with inter-synaptic formation and signaling. NR2F1-AS1 may promote osteoblast proliferation stimulate ALP activity, and induce osteogenic differentiation by down-regulating miR-423-5p.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s10616-025-00786-8.</p>\",\"PeriodicalId\":10890,\"journal\":{\"name\":\"Cytotechnology\",\"volume\":\"77 4\",\"pages\":\"135\"},\"PeriodicalIF\":2.0000,\"publicationDate\":\"2025-08-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12222589/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Cytotechnology\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.1007/s10616-025-00786-8\",\"RegionNum\":4,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2025/7/2 0:00:00\",\"PubModel\":\"Epub\",\"JCR\":\"Q3\",\"JCRName\":\"BIOTECHNOLOGY & APPLIED MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Cytotechnology","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1007/s10616-025-00786-8","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2025/7/2 0:00:00","PubModel":"Epub","JCR":"Q3","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
LncRNA NR2F1-AS1 is involved in osteogenic differentiation in fracture healing via miR-423-5p.
To investigate the function and mechanism of action of LncRNA NR2F1-AS1 involved in osteogenic differentiation process. An in vitro model was constructed by osteogenic differentiation-induced stimulation (OS) on the hFOB1.19 cell line. Real-time fluorescence quantitative PCR (RT-qPCR) was performed to detect the expression of NR2F1-AS1, miR-423-5p and osteogenic differentiation markers (RUNX2, OCN, OPN). Cell Counting Kit-8 (CCK8) method and flow cytometry were observed cell proliferation and apoptosis, respectively. Enzyme linked immunosorbent assay (ELISA) tested alkaline phosphatase (ALP) activity. Dual-Luciferase Report (DLR) assay and RNA immunoprecipitation (RIP) verified gene interactions. Bioinformatics methods predicted downstream target genes and their pathways of action. OS increased osteogenic differentiation markers and NR2F1-AS1 expression and decreased miR-423-5p levels. Transfection of si- NR2F1-AS1 promoted OS osteoblast apoptosis, but inhibited cell proliferation, ALP activity and osteogenic differentiation marker expression. NR2F1-AS1 is mostly present in the cytoplasm and is involved in the osteogenic differentiation process by down-regulating miR-423-5p. The use of miR-423-5p inhibitor can resist apoptosis induced by silencing NR2F1-AS1, promote osteoblast proliferation, activate ALP activity, and induce osteogenic differentiation process in osteoblasts. Bioinformatics prediction identified 82 target genes that might be involved in osteogenic differentiation, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis suggested that they were mainly associated with inter-synaptic formation and signaling. NR2F1-AS1 may promote osteoblast proliferation stimulate ALP activity, and induce osteogenic differentiation by down-regulating miR-423-5p.
Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00786-8.
期刊介绍：
The scope of the Journal includes:
1. The derivation, genetic modification and characterization of cell lines, genetic and phenotypic regulation, control of cellular metabolism, cell physiology and biochemistry related to cell function, performance and expression of cell products.
2. Cell culture techniques, substrates, environmental requirements and optimization, cloning, hybridization and molecular biology, including genomic and proteomic tools.
3. Cell culture systems, processes, reactors, scale-up, and industrial production. Descriptions of the design or construction of equipment, media or quality control procedures, that are ancillary to cellular research.
4. The application of animal/human cells in research in the field of stem cell research including maintenance of stemness, differentiation, genetics, and senescence, cancer research, research in immunology, as well as applications in tissue engineering and gene therapy.
5. The use of cell cultures as a substrate for bioassays, biomedical applications and in particular as a replacement for animal models.
求助内容：
应助结果提醒方式：
京公网安备 11010802042870号
