MicroRNA-181a-5p Curbs Osteogenic Differentiation and Bone Formation Partially Through Impairing Runx1-Dependent Inhibition of AIF-1 Transcription

Endocrinol Metab (Seoul). 2023 Feb;38(1):156-173. doi: 10.3803/EnM.2022.1516. Epub 2023 Jan 6.

Abstract

Backgruound: Evidence has revealed the involvement of microRNAs (miRNAs) in modulating osteogenic differentiation, implying the promise of miRNA-based therapies for treating osteoporosis. This study investigated whether miR-181a-5p influences osteogenic differentiation and bone formation and aimed to establish the mechanisms in depth.

Methods: Clinical serum samples were obtained from osteoporosis patients, and MC3T3-E1 cells were treated with osteogenic induction medium (OIM) to induce osteogenic differentiation. miR-181a-5p-, Runt-related transcription factor 1 (Runx1)-, and/or allograft inflammatory factor-1 (AIF-1)-associated oligonucleotides or vectors were transfected into MC3T3-E1 cells to explore their function in relation to the number of calcified nodules, alkaline phosphatase (ALP) staining and activity, expression levels of osteogenesis-related proteins, and apoptosis. Luciferase activity, RNA immunoprecipitation, and chromatin immunoprecipitation assays were employed to validate the binding relationship between miR-181a-5p and Runx1, and the transcriptional regulatory relationship between Runx1 and AIF-1. Ovariectomy (OVX)-induced mice were injected with a miR-181a-5p antagonist for in vivo verification.

Results: miR-181a-5p was highly expressed in the serum of osteoporosis patients. OIM treatment decreased miR-181a-5p and AIF-1 expression, but promoted Runx1 expression in MC3T-E1 cells. Meanwhile, upregulated miR-181a-5p suppressed OIM-induced increases in calcified nodules, ALP content, and osteogenesis-related protein expression. Mechanically, miR-181a-5p targeted Runx1, which acted as a transcription factor to negatively modulate AIF-1 expression. Downregulated Runx1 suppressed the miR-181a-5p inhibitor-mediated promotion of osteogenic differentiation, and downregulated AIF-1 reversed the miR-181a-5p mimic-induced inhibition of osteogenic differentiation. Tail vein injection of a miR-181a-5p antagonist induced bone formation in OVX-induced osteoporotic mice.

Conclusion: In conclusion, miR-181a-5p affects osteogenic differentiation and bone formation partially via the modulation of the Runx1/AIF-1 axis.

Keywords: MicroRNAs; Osteogenesis; Osteoporosis; Ovariectomy; Transcription factors.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Differentiation / genetics
  • Core Binding Factor Alpha 2 Subunit / genetics
  • Female
  • Mice
  • MicroRNAs* / genetics
  • MicroRNAs* / metabolism
  • Osteogenesis / genetics
  • Osteoporosis* / genetics

Substances

  • Core Binding Factor Alpha 2 Subunit
  • MicroRNAs
  • Runx1 protein, mouse
  • Aif1 protein, mouse