miR-486参与低氧调控的肺间质纤维化发病过程

MiR-486 regulates the pathogenesis of pulmonary fibrosis under hypoxia

目的 miR-486为首次从胎肝鉴定的miRNA,既往研究发现miR-486在红系白血病细胞等细胞中为一种低氧相关miRNA,本研究旨在于探索低氧条件下小鼠胚肺成纤维细胞3T3细胞miR-486的表达变化,并明确miR-486在低氧诱导肺间质纤维化发病中的调控作用。方法 miR-486的过表达或沉默后用慢病毒技术,转染效率用流式细胞仪及qRT-PCR检测,转染效率均在85%以上。低氧条件及过表达或沉默miR-486 3T3细胞miR-486、FN、α-SMA、CollagenⅠ及CollagenⅢ的表达用实时定量PCR(quantitative real-time PCR,qRT-PCR)方法进行检测。结果与常氧培养3T3细胞相比,低氧6、12、24、48 h 3T3细胞miR-486表达降低(P<0.05),而FN、α-SMA的表达升高(P<0.05)。过表达miR-486可以促进3T3细胞miR-486的表达,并可以下调FN、α-SMA、CollagenⅠ及CollagenⅢ的表达,表达量分别为1.000±0.094 vs. 0.125±0.153,P=0.005;1.000±0.193 vs. 0.044±0.001,P=0.010;1.000±0.057 vs. 0.561±0.009,P=0.048;1.000±0.105 vs.0.275±0.045,P=0.013;而沉默miR-486则抑制3T3细胞miR-486的表达,促进FN、α-SMA、CollagenⅠ及CollagenⅢ的表达,表达量分别为1.000±0.012 vs.48.909±0.584,P=0.005;1.000±0.015 vs. 4.416±0.311,P=0.039;1.000±0.270 vs. 8.892±0.484,P=0.003;1.000±0.005 vs.7.671±1.014,P=0.008。结论 MiR-486参与了调控低氧相关肺间质纤维化疾病进展过程。

Objective MiR-486 was proved to be a hypoxia miRNA in some cells, such as bone marrow derived stem cells and hematopoietic cell. The aim of this study is to observe the effect of miR-486 on the pathogenesis of pulmonary fibrosis under hypoxia. Methods Lentivirus was used to mediate the overexpression or inhibition of miR-486 on 3 T3 cells, Flow cytometry and qRT-PCR were used to check the transfection efficiency, and all the lentivirus transfection efficiency were above 85%. The expression of miR-486, FN,α-SMA, Collagen Ⅰ and Collagen Ⅲ on 3 T3 cells under hypoxia, or transfected by overexpression and inhibition lentivirus were detected by qRT-PCR. Results Compared with hypoxia oh, hypoxia 6, 12, 24, 48 h miR-486 expression decreased(P<0.05), while the FN and α-SMA expression increased(P<0.05). MiR-486 expression was increased in miR-486 overexpression 3 T3 cells(P<0.05). And FN,α-SMA, Collagen Ⅰ and Collagen Ⅱ expression decreased(P<0.05), they were separately(1.000±0.094 vs. 0.125±0.153, P=0.005;1.000±0.193 vs. 0.044±0.001,P=0.010;1.000±0.057 vs. 0.561±0.009, P=0.048;1.000±0.105 vs. 0.275±0.045, P=0.013). FN,α-SMA, Collagen Ⅰ and Collagen Ⅱ expression in miR-486 silence 3 T3 cells were 1.000±0.079 vs. 0.027±0.003,P=0.002;1.000±0.012 vs. 48.909±0.584, P=0.005;1.000±0.015 vs. 4.416±0.311, P=0.039;1.000±0.270 vs. 8.892±0.484, P=0.003;1.000±0.005 vs. 7.671±1.014, P=0.008, mean that miR-486 silence can inhibit miR-486 expression, and promote FN,α-SMA, Collagen Ⅰ and Collagen Ⅱ expression. Conclusion MiR-486 could regulate the pathogenesis of pulmonary fibrosis under hypoxia.