MIR-185-5P AFFECTS THE GROWTH AND METASTASIS OF COLORECTAL CANCER BY NEGATIVELY REGULATING TNNT1

Tao Wu^#, Chao Jin^#, Peng Gao, Guo-Zhan Jia, Xiong-Chao Fang,  Ying Yang^*, Nan Wang^* 

Department of General Surgery, Tangdu Hospital, the Air Force Medical University, Xi’an 710038, China

jective: The purpose of this study is to elucidate the molecular mechanism of the occurrence and development of colorectal cancer (CRC), and to provide a theoretical basis for possible treatment options for CRC patients. 

Methods: The expression level of TNNT1 in CRC patients was analysed using the UALCAN database, and the survival curve of CRC patients separated by TNNT1 expression level was analysed using the GEPIA database. MiR-185-5p- and TNNT1-overexpressing CRC cell models were constructed by transfecting cells with miR-185-5p mimics and TNNT1 plasmid. The transcription levels of TNNT1 and miR-185-5p in each group of cells were detected by RT-qPCR. The expression of TNNT1 in the cells was detected using Western blot. The possible binding sequence of miR-185-5p to TNNT1 was predicted using the Targetscan database, and the regulatory relationship between miR-185-5p and TNNT1 was further verified using a luciferase reporter gene assay. Cell Count Kit-8 (CCK-8) and BrdU immunofluorescence staining were used to detect cell proliferation in each group, and the migration and invasion levels of each group were detected by Transwell assay. 

Results: The expression of TNNT1 was significantly upregulated in CRC patients, and the survival curve results showed that high expression of TNNT1 was significantly correlated with decrease in the survival time of patients. At the same time, RT-qPCR results showed that the transcription level of TNNT1 was significantly increased in CRC cell lines, while the expression of miR-185-5p was significantly reduced. The overexpression of miR-185-5p significantly inhibited the expression of TNNT1 in SW480 cells and functionally inhibited the proliferation and invasion ability of SW480 cells. Double luciferase reporter gene tests and Western blotting results showed that miR-185-5p could directly act on and negatively regulate the expression of TNNT1. In addition, forced overexpression of TNNT1 completely reversed the inhibitory effect of miR-185-5p on SW480 cells. 

Conclusion: MiR-185-5p inhibits the proliferation and invasion of CRC cells by targeting TNNT1 in CRC, which provides a new therapeutic target for CRC.