1 Department of Pediatrics.
2 Department of Medicine.
Am J Respir Crit Care Med. 2018 Mar 1;197(5):632-643. doi: 10.1164/rccm.201704-0732OC.
MicroRNAs (miRNAs) destabilize mRNA transcripts and inhibit protein translation. miR-145 is of particular interest in cystic fibrosis (CF) as it has a direct binding site in the 3'-untranslated region of CFTR (cystic fibrosis transmembrane conductance regulator) and is upregulated by the CF genetic modifier TGF (transforming growth factor)-β.
To demonstrate that miR-145 mediates TGF-β inhibition of CFTR synthesis and function in airway epithelia.
Primary human CF (F508del homozygous) and non-CF airway epithelial cells were grown to terminal differentiation at the air-liquid interface on permeable supports. TGF-β (5 ng/ml), a miR-145 mimic (20 nM), and a miR-145 antagonist (20 nM) were used to manipulate CFTR function. In CF cells, lumacaftor (3 μM) and ivacaftor (10 μM) corrected mutant F508del CFTR. Quantification of CFTR mRNA, protein, and function was done by standard techniques.
miR-145 is increased fourfold in CF BAL fluid compared with non-CF (P < 0.01) and increased 10-fold in CF primary airway epithelial cells (P < 0.01). Exogenous TGF-β doubles miR-145 expression (P < 0.05), halves wild-type CFTR mRNA and protein levels (P < 0.01), and nullifies lumacaftor/ivacaftor F508del CFTR correction. miR-145 overexpression similarly decreases wild-type CFTR protein synthesis (P < 0.01) and function (P < 0.05), and eliminates F508del corrector benefit. miR-145 antagonism blocks TGF-β suppression of CFTR and enhances lumacaftor correction of F508del CFTR.
miR-145 mediates TGF-β inhibition of CFTR synthesis and function in airway epithelia. Specific antagonists to miR-145 interrupt TGF-β signaling to restore F508del CFTR modulation. miR-145 antagonism may offer a novel therapeutic opportunity to enhance therapeutic benefit of F508del CFTR correction in CF epithelia.
MicroRNAs（miRNAs）使 mRNA 转录本不稳定，并抑制蛋白质翻译。miR-145 在囊性纤维化（CF）中特别引人注目，因为它在 CFTR（囊性纤维化跨膜电导调节剂）的 3'非翻译区有一个直接结合位点，并且由 CF 遗传修饰因子 TGF（转化生长因子）-β上调。
证明 miR-145 介导 TGF-β 抑制气道上皮细胞中 CFTR 的合成和功能。
将原代 CF（F508del 纯合子）和非 CF 气道上皮细胞在可渗透支持物上的气液界面生长至终末分化。使用 TGF-β（5ng/ml）、miR-145 模拟物（20nM）和 miR-145 拮抗剂（20nM）来操纵 CFTR 功能。在 CF 细胞中，lumacaftor（3μM）和 ivacaftor（10μM）纠正了突变型 F508del CFTR。通过标准技术定量 CFTR mRNA、蛋白和功能。
与非 CF 相比，CF BAL 液中的 miR-145 增加了四倍（P<0.01），CF 原代气道上皮细胞中的 miR-145 增加了十倍（P<0.01）。外源性 TGF-β使 miR-145 表达增加两倍（P<0.05），使野生型 CFTR mRNA 和蛋白水平减半（P<0.01），并使 lumacaftor/ivacaftor F508del CFTR 校正无效。miR-145 过表达同样降低了野生型 CFTR 蛋白合成（P<0.01）和功能（P<0.05），并消除了 F508del 校正因子的益处。miR-145 拮抗剂阻断 TGF-β 对 CFTR 的抑制作用，并增强 lumacaftor 对 F508del CFTR 的校正作用。
miR-145 介导 TGF-β 抑制气道上皮细胞中 CFTR 的合成和功能。miR-145 的特异性拮抗剂阻断 TGF-β 信号转导，恢复 F508del CFTR 的调节。miR-145 拮抗剂可能为增强 CF 上皮中 F508del CFTR 校正的治疗益处提供一种新的治疗机会。
