Exploration of the mechanism of NORAD activation of TGF-β1/Smad3 through miR-136-5p and promotion of tacrolimus-induced renal fibrosis.

BACKGROUND

Tacrolimus is a potent immunosuppressant, but has various side effects, with nephrotoxicity being the most common. Renal fibrosis is an important process of tacrolimus nephrotoxicity. Therefore, it is important to identify the factors that contribute to renal fibrosis after tacrolimus nephrotoxicity, and control its development.

METHODS

The present study aims to determine whether tacrolimus may speed up the course of renal fibrosis by upregulating noncoding RNA activated by DNA damage (NORAD) to compete with miR-136-5p, and activating the TGF-β1/Smad3 pathway. Furthermore, rat models and cell models were established. Then, the expression levels of NORAD and miR-136-5p were determined by RT-qPCR, while the expression of the TGF-β1/Smad3 pathway was determined by western blot and RT-qPCR. In order to investigate the interaction between NORAD and miR-136-5p, as well as miR-136-5p and SYK, two luciferase reporters were employed. The renal fibrosis of mice was observed using Masson and PAS staining. The expression of inflammatory factors IL-1, IL-6, MCP-1 and TNF-α was detected by ELISA.

RESULTS

In the experiments, NORAD was upregulated, while miR-136-5p was downregulated after tacrolimus induction. The expression of the TGF-β1/Smad3 pathway correspondingly changed after the induction by tacrolimus. In the experiments, the expression of NORAD and miR-136-5p, and the trend for renal fibrosis were consistent with the results in the experiments. Furthermore, the inflammatory factors correspondingly changed with the severity of renal fibrosis. Moreover, the expression trend of the TGF-β1/Smad3 pathway in tacrolimus-induced rats was consistent with that in the experiments.

CONCLUSION

Through and experiments, the present study was able to successfully prove that tacrolimus upregulates NORAD to compete with miR-136-5p, resulting in a decrease in miR-136-5p expression, which in turn activates the TGF-β1/smad3 pathway, and finally induces the aggravation of renal fibrosis.