Institute of Otolaryngology Head and Neck Surgery, Affiliated Hospital of Nantong University, Nantong, Jiangsu, China.
Department of Otolaryngology Head and Neck Surgery, Affiliated Hospital of Nantong University, Nantong, Jiangsu, China.
Am J Rhinol Allergy. 2021 Mar;35(2):152-163. doi: 10.1177/1945892420939814. Epub 2020 Jul 5.
Epithelial-to-Mesenchymal Transition (EMT) is considered as a crucial event in disease development and dysregulation of microRNAs (miRNAs) is involved in the regulation of EMT in various human diseases. Emerging evidences congregated over the years have demonstrated that miR-30a-5p was decreased in diseases and its overexpression inhibited the process of diseases via attenuating EMT. Although aberrant expression of miRNAs and occurrence of EMT were previously reported in Nasal Polyps (NPs), the role of miR-30a-5p in EMT of NPs is still remains unclear.
The purpose of our present study was to explore the expression and potential function of miR-30a-5p in EMT of NPs.
The expression of miR-30a-5p and mRNA expression level were detected by quantitative real-time PCR (qRT-PCR) in transforming growth factor β1 (TGF-β1) - induced EMT model and NPs patients. Western Blot (WB) and immunohistochemistry (IHC) were performed to evaluate the protein expression level of EMT markers. The cells mobility was assessed by Wound-Healing assay. Luciferase reporter assay was utilized to verify the relationship between Cyclin-dependent kinase 6 (CDK6) and miR-30a-5p.
Firstly, we observed that miR-30a-5p was down-regulated notably, accompanying with the alteration of EMT markers expression in NPs tissues and EMT model induced by TGF-β1 in primary Human Nasal Epithelial Cells (pHNECs) and A549 cells in vitro. Moreover, the functional assays demonstrated that overexpression of miR-30a-5p significantly inhibited EMT and cells mobility. Subsequently, CDK6 was validated as a direct target of miR-30a-5p. Finally, we performed the rescue experiments indicating that overexpression of CDK6 eliminated the suppressive effects of miR-30a-5p in TGF-β1-induced EMT in pHNECs and A549 cells.
Taken together, our results suggested that EMT was involved in NPs, and overexpression of miR-30a-5p could attenuate EMT via repressing the expression of the CDK6 in pHNECs and A549 cells.
上皮间质转化（EMT）被认为是疾病发展过程中的一个关键事件，而 microRNAs（miRNAs）的失调参与了各种人类疾病中 EMT 的调控。多年来汇集的新证据表明，miR-30a-5p 在疾病中表达降低，其过表达通过抑制 EMT 来抑制疾病的发展。尽管先前在鼻息肉（NPs）中已经报道了 miRNA 的异常表达和 EMT 的发生，但 miR-30a-5p 在 NPs 中的 EMT 中的作用仍不清楚。
本研究旨在探讨 miR-30a-5p 在 NPs 中的 EMT 中的表达和潜在功能。
采用实时定量 PCR（qRT-PCR）检测转化生长因子 β1（TGF-β1）诱导的 EMT 模型和 NPs 患者中 miR-30a-5p 的表达和 mRNA 表达水平。采用 Western Blot（WB）和免疫组织化学（IHC）检测 EMT 标志物的蛋白表达水平。通过划痕愈合试验评估细胞迁移能力。利用荧光素酶报告基因实验验证 Cyclin-dependent kinase 6（CDK6）与 miR-30a-5p 的关系。
首先，我们观察到 miR-30a-5p 在 NPs 组织和体外 TGF-β1 诱导的原代人鼻上皮细胞（pHNECs）和 A549 细胞 EMT 模型中显著下调，同时 EMT 标志物的表达也发生改变。此外，功能实验表明，miR-30a-5p 的过表达显著抑制 EMT 和细胞迁移。随后，验证了 CDK6 是 miR-30a-5p 的直接靶标。最后，我们进行了挽救实验，表明在 pHNECs 和 A549 细胞中过表达 CDK6 消除了 miR-30a-5p 在 TGF-β1 诱导的 EMT 中的抑制作用。
综上所述，我们的研究结果表明 EMT 参与了 NPs 的发生，miR-30a-5p 的过表达通过抑制 CDK6 的表达来抑制 pHNECs 和 A549 细胞中的 EMT。
