Department of Orthopaedics, Central Hospital of Zibo Mining Refco Group Ltd., Zibo, Shandong 255120, P.R. China.
Department of Geriatrics, Central Hospital of Zibo Mining Refco Group Ltd., Zibo, Shandong 255120, P.R. China.
Mol Med Rep. 2019 Oct;20(4):3901-3909. doi: 10.3892/mmr.2019.10630. Epub 2019 Aug 30.
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.
为了研究微小 RNA(miR)-519d-3p 在术后硬膜外瘢痕形成中的作用及其对骨形态发生蛋白和激活素膜结合抑制剂(BAMBI)的调节作用,采用逆转录-定量聚合酶链反应和 Western blot 检测行椎板切除术患者腰椎间盘 miR-519d-3p 和 BAMBI 表达水平。结果表明,患有硬膜外瘢痕的患者腰椎间盘 miR-519d-3p 表达明显增加,而 BAMBI 表达明显降低。随后,将 miR-519d-3p 模拟物转染至硬膜外瘢痕组织中分离的原代成纤维细胞。流式细胞术和细胞计数试剂盒-8 分析表明,miR-519d-3p 的过表达促进了成纤维细胞的增殖、肿瘤坏死因子-α和白细胞介素(IL)-1α的产生,以及 I 型胶原(col I)、α-平滑肌肌动蛋白(α-SMA)和纤维连接蛋白(FN)的表达。miR-519d-3p 反义寡核苷酸转染下调 miR-519d-3p 则产生相反的效果。生物信息学和荧光素酶报告基因分析表明,BAMBI 是 miR-519d-3p 的靶基因:miR-519d-3p 可直接结合 BAMBI mRNA 的 3'UTR 并抑制 BAMBI 蛋白表达。最后,pcDNA-BAMBI 载体和 BAMBI 小干扰 RNA 分别转染原代成纤维细胞以过表达和敲低 BAMBI 基因。结果表明,BAMBI 过表达抑制了成纤维细胞增殖、TNF-α和 IL-1α 的产生以及 col I、α-SMA 和 FN 蛋白的表达,而 BAMBI 敲低则产生相反的效果。综上所述,BAMBI 是 miR-519d-3p 的靶基因,miR-519d-3p 通过抑制 BAMBI 促进转化生长因子-β/母亲抗 decapentaplegic 同源物 9 介导的术后硬膜外瘢痕形成。