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.