Involvement of miR‐140‐3p in Wnt3a and TGFβ3 signaling pathways during osteoblast differentiation in MC3T3‐E1 cells

Abstract

The Wnt/β-catenin signaling and TGFβ signaling pathways play a key role in osteoblast differentiation. The miRNAs play important roles in regulating gene expression at the post-transcriptional level through fine-tuning of protein-encoding gene expression. However, involvement of miRNAs is not established for Wnt3a and TGFβ signaling pathways in osteoblast differentiation. Here, we examined the role of miRNAs expressed differentially after Wnt3a expression during osteoblast differentiation. Over-expression of the Wnt3a gene increased ALP transcription, but decreased Col1, Runx2, and OCN transcription in osteoblastic MC3T3-E1 cells. Expression profiling and quantitative PCR for miRNAs showed that miR-140-3p decreased in Wnt3a-over-expressing osteoblastic cells. Wnt3a over-expression increased TGFβ3 expression, whereas transfection of the miR-140-3p mimic into MC3T3-E1 cells significantly inhibited TGFβ3 expression. Luciferase assay for the TGFβ3 transcript showed that TGFβ3 was a direct target of miR-140-3p. miR-140-3p mimic transfection resulted in significantly increased OCN transcription, but did not affect ALP, Col1, and Runx2 transcription in MC3T3-E1 cells. rTGFβ3 treatment decreased OCN transcription in MC3T3-E1 cells. These results suggest that the miR-140-3p is involved in osteoblast differentiation as a critical regulatory factor between Wnt3a and TGFβ3 signaling pathways.

Potential Novel Prediction of TMJ-OA: MiR-140-5p Regulates Inflammation Through Smad/TGF-β Signaling

“…It suggested that TGF-β3 may regulated by miR-140-5p in multiple ways, including Smad3 pathway. Novel evidences demonstrating that miR-140-3p negatively controlled TGF-β3, which proved to be a direct target of miR-140-3p (Fushimi et al, 2018). That’s may explained why co-transfection of miR-140-5p mimics and Si-Smad3 showed a lower expression of TGF-β3.…”

The Role of MicroRNAs in Bone Metabolism and Disease

“…However, when overexpressed, miR-140-3p stimulated the transcription of OCN in MC3T3-E1 cells. Thus, miR-140-3p regulated osteoblast differentiation by acting as a regulatory factor between Wnt3a and TGFβ3 signaling pathways [92]. According to the study, miR-133a-5p overexpression was shown to significantly lower the expression of osteoblast differentiation markers as well as ECM mineralization in MC3T3-E1 cells by targeting 3 UTR of Runx2 and reduce Runx2 expression at both mRNA and protein levels [93].…”

The Therapeutic Potential of MicroRNAs as Orthobiologics for Skeletal Fractures

“…It is worth mentioning that four of the miRNAs (miR‐140‐3p, miR‐148a, mir‐218, and miR‐214), previously discussed in the context of fracture repair, are also listed as involved in bone remodeling. For example, miR‐140‐3p was found in osteoblast‐derived exosomes and in osteoblastic MC3T3 cells and has been shown to promote osteoblast differentiation . Additionally, miR‐148a and miR‐218 were found in osteoblast precursor exosomes and in human BMSCs where the former promotes osteoclastogenesis and the latter enhances osteoblast differentiation .…”

“…For example, miR-140-3p was found in osteoblast-derived exosomes and in osteoblastic MC3T3 cells and has been shown to promote osteoblast differentiation. (67) Additionally, miR-148a and miR-218 were found in osteoblast precursor exosomes and in human BMSCs where the former promotes osteoclastogenesis (68) and the latter enhances osteoblast differentiation. (69) Lastly, miR-214-3p is found in osteoclast-derived exosomes and inhibits osteoblastic bone formation ( Figure 4).…”

“…Serum from osteoporotic fracture patientsupregulated (33) Osteoblast-derivedpromotes osteoblast differentiation (67) Callus from rat nonunion fracturesdownregulated (35) Callus from mouse transverse fracturesupregulated (36)…”