The increase of microRNA-21 during lung fibrosis and its contribution to epithelial-mesenchymal transition in pulmonary epithelial cells

Respiratory Research

Masakazu Ichinose²  Takashi Suzuki³  Kiyoshi Takagi³  Tomonori Makiguchi²  Naoya Fujino¹  Takaya Suzuki⁴  Yukiko Tando¹  Toru Takahashi¹  Chiharu Ota¹  Hiroshi Kubo¹  Mitsuhiro Yamada² 

[1] Department of Advanced Preventive Medicine for Infectious Disease, Tohoku University Graduate School of Medicine, 2-1 Seiryoumachi, Aobaku 980-8575, Sendai, Japan;Department of Respiratory Medicine, Tohoku University Graduate School of Medicine, 1-1 Seiryoumachi, Aobaku 980-8574, Sendai, Japan;Department of Pathology and Histotechnology, Tohoku University Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai 980-8575, Miyagi-ken, Japan;Department of Thoracic Surgery, Institute of Development, Aging and Cancer, Tohoku University, 4-1 Seiryoumachi, Aobaku 980-8575, Sendai, Japan

关键词: Lung single cell separation;    Alveolar type II cells;    Pulmonary fibrosis;    Epithelial-mesenchymal transition;    microRNAs;   

Others  :  792756
DOI  :  10.1186/1465-9921-14-95

 received in 2013-03-25, accepted in 2013-09-23,  发布年份 2013

【 摘 要 】

Background

The excess and persistent accumulation of fibroblasts due to aberrant tissue repair results in fibrotic diseases such as idiopathic pulmonary fibrosis. Recent reports have revealed significant changes in microRNAs during idiopathic pulmonary fibrosis and evidence in support of a role for microRNAs in myofibroblast differentiation and the epithelial-mesenchymal transition in the context of fibrosis. It has been reported that microRNA-21 is up-regulated in myofibroblasts during fibrosis and promotes transforming growth factor-beta signaling by inhibiting Smad7. However, expression changes in microRNA-21 and the role of microRNA-21 in epithelial-mesenchymal transition during lung fibrosis have not yet been defined.

Methods

Lungs from saline- or bleomycin-treated C57BL/6 J mice and lung specimens from patients with idiopathic pulmonary fibrosis were analyzed. Enzymatic digestions were performed to isolate single lung cells. Lung epithelial cells were isolated by flow cytometric cell sorting. The expression of microRNA-21 was analyzed using both quantitative PCR and in situ hybridization. To induce epithelial-mesenchymal transition in culture, isolated mouse lung alveolar type II cells were cultured on fibronectin-coated chamber slides in the presence of transforming growth factor-β, thus generating conditions that enhance epithelial-mesenchymal transition. To investigate the role of microRNA-21 in epithelial-mesenchymal transition, we transfected cells with a microRNA-21 inhibitor. Total RNA was isolated from the freshly isolated and cultured cells. MicroRNA-21, as well as mRNAs of genes that are markers of alveolar epithelial or mesenchymal cell differentiation, were quantified using quantitative PCR.

Results

The lung epithelial cells isolated from the bleomycin-induced lung fibrosis model system had decreased expression of epithelial marker genes, whereas the expression of mesenchymal marker genes was increased. MicroRNA-21 was significantly upregulated in isolated lung epithelial cells during bleomycin-induced lung fibrosis and human idiopathic pulmonary fibrosis. MicroRNA-21 was also upregulated in the cultured alveolar epithelial cells under the conditions that enhance epithelial-mesenchymal transition. Exogenous administration of a microRNA-21 inhibitor prevented the increased expression of vimentin and alpha-smooth muscle actin in cultured primary mouse alveolar type II cells under culture conditions that induce epithelial-mesenchymal transition.

Conclusions

Our experiments demonstrate that microRNA-21 is increased in lung epithelial cells during lung fibrosis and that it promotes epithelial-mesenchymal transition.

【 授权许可】

   
2013 Yamada et al.; licensee BioMed Central Ltd.

【 图 表 】

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