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Visualization and Quantification of TGFβ/BMP/SMAD Signaling under Different Fluid Shear Stress Conditions using Proximity-Ligation-Assay
Published on: September 14, 2021
miR‑519d‑3p promotes TGFβ/Smad mediated postoperative epidural scar formation via suppression of BAMBI
Longbiao Yang1, Qingyuan Gao1, Feng Lv1
1Department of Orthopaedics, Central Hospital of Zibo Mining Refco Group Ltd., Zibo, Shandong 255120, P.R. China.
Abstract:
To investigate the role of microRNA (miR)‑519d‑3p in postoperative epidural scar formation and its regulation of the bone morphogenetic protein and activin membrane‑bound inhibitor (BAMBI), miR‑519d‑3p and BAMBI expression levels in the lumbar disc of patients who had undergone laminectomy were detected with reverse transcription‑quantitative polymerase chain reaction and western blotting. The results demonstrated that miR‑519d‑3p expression was significantly increased, whereas BAMBI expression was sharply reduced in the lumbar discs of patients suffering from epidural scars. Subsequently, the miR‑519d‑3p mimic was transfected into primary fibroblasts isolated from epidural scar tissues. Flow cytometric and Cell Countin Kit‑8 analyses indicated that overexpression of miR‑519d‑3p promoted the proliferation of fibroblasts, the production of tumor necrosis factor‑α and interleukin (IL)‑1α, and the expression of type I collagen (col I), α‑smooth muscle actin (α‑SMA) and fibronectin (FN). Downregulation of miR‑519d‑3p by the miR‑519d‑3p antagomir transfection had the opposite effect. Bioinformatics and luciferase reporter gene analyses demonstrated that BAMBI is a target gene of miR‑519d‑3p: miR‑519d‑3p directly binds to the 3'‑untranslated region of BAMBI mRNA and suppressed BAMBI protein expression. Finally, the pcDNA‑BAMBI vector and BAMBI small interfering RNA were respectively transfected into primary fibroblasts to overexpress and knockdown the BAMBI gene. It was demonstrated that BAMBI overexpression suppressed fibroblast proliferation, TNF‑α and IL‑1α production, and the expression of col I, α‑SMA and FN proteins, whereas, BAMBI knockdown had the opposite effect. In conclusion, it was noted that BAMBI is a target of miR‑519d‑3p and miR‑519d‑3p promotes transforming growth factor β/mothers against decapentaplegic homolog 9‑mediated postoperative epidural scar formation via suppression of BAMBI.
Insights
MicroRNA-519d-3p promotes epidural scar formation by suppressing BAMBI. This microRNA increases fibroblast proliferation and collagen production, contributing to scar tissue development after surgery.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Postoperative epidural scar formation is a significant clinical challenge following spinal surgery.
- MicroRNAs (miRNAs) play crucial roles in cellular processes, including tissue repair and fibrosis.
- Bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI) is implicated in regulating cell growth and differentiation.
Purpose of the Study:
- To elucidate the role of miR-519d-3p in postoperative epidural scar formation.
- To investigate the regulatory relationship between miR-519d-3p and BAMBI.
- To understand the molecular mechanisms underlying scar development mediated by this miRNA-target interaction.
Main Methods:
- Detection of miR-519d-3p and BAMBI expression in patient samples using RT-qPCR and Western blotting.
- In vitro studies involving transfection of miR-519d-3p mimic/antagomir and BAMBI overexpression/knockdown in primary fibroblasts.
- Bioinformatics analysis and luciferase reporter assays to confirm direct targeting of BAMBI by miR-519d-3p.
Main Results:
- miR-519d-3p expression was upregulated, while BAMBI expression was downregulated in epidural scars.
- Overexpression of miR-519d-3p promoted fibroblast proliferation, inflammatory cytokine production (TNF-α, IL-1α), and extracellular matrix deposition (collagen I, α-SMA, fibronectin).
- BAMBI was identified as a direct target of miR-519d-3p, and its suppression mediated the pro-fibrotic effects.
Conclusions:
- miR-519d-3p promotes postoperative epidural scar formation by directly targeting and suppressing BAMBI.
- The miR-519d-3p/BAMBI axis influences fibroblast behavior, cytokine production, and extracellular matrix synthesis.
- Targeting miR-519d-3p or restoring BAMBI expression may offer therapeutic strategies for preventing epidural scarring.
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