Abstract
This research is designed to investigate the regulatory effect of miR-18a to the target gene connective tissue growth factor (CTGF, or CCN2), by participating in TGF-β1 signaling pathway and explore the pathogenic mechanism of miR-18a in pulmonary injury induced by nano-SiO2 based on our early study. miR-18a and expression of TGF-β1 in Chinese hamster lung (CHL) fibroblasts cells stimulated by supernatants of NR8383 cells exposed to 40 μg/ml nano-SiO2 for 24 h demonstrated 1.58 ± 0.22-fold and 1096.00 ± 2.60 pg/ml increase compared with blank control group analyzed by real-time quantitative PCR (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA), respectively. Expression increase of miR-18a and reduction of CCN2 mRNA expression levels and protein gray value ratio detected by Western blotting in CHL cells transfect miR-18a mimics for 48 h. The reverse of CHL cell transfection miR-18a inhibit is also true. The result of miR-18a and CCN2 binding sites tested by luciferase reporter gene assay shows that the report relative fluorescence value of miR-18a mimics wild type on CCN2 is 0.50 ± 0.02 with the control of mimics NC and mutant relative fluorescence report value 0.86 ± 0.04 (P < 0.05). Expression levels of miR-18a, CCN2 mRNA, and protein gray value ratio decreased in CHL cells treated by TGF-β1, respectively, and vice versa treated by TGF-β1corepressor. The results suggest that CCN2 is the target gene regulated by miR-18a and miR-18a participates in TGF-β1 signaling pathway by regulating the expression of CCN2 negatively through CCN2 3’UTR site, and thus may be involved in the development process of pulmonary injury.
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[Fund Project] Supported by the National Natural Science Foundation of China (Grant No.81273046), the Fundamental Research Funds for the Central Universities, the Preventive Medicine Research Projects of Jiangsu Province (Grant No.Y2012039), and the Major State Basic Research Development Program of China (973Program, Grant No.2011CB933404).
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Yang, H., Li, W., Zhang, Y. et al. Regulatory role of miR-18a to CCN2 by TGF-β1 signaling pathway in pulmonary injury induced by nano-SiO2 . Environ Sci Pollut Res 25, 867–876 (2018). https://doi.org/10.1007/s11356-017-0344-0
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DOI: https://doi.org/10.1007/s11356-017-0344-0