Abstract 20541: MicroRNA-216a Mediates the Activation of Telomerase in Macrophages Differentiation via the Smad3/NF-κB Pathway
Background: The monocyte/macrophage is a central participant in the inflammatory process that is involved in all stages of atherosclerosis. Our previous work showed that telomerase reverse transcriptase (TERT) expression was particularly observed in the macrophage-rich region of atherosclerotic lesions. Interestingly, we have identified that miR-216a was remarkably up-regulated in macrophages differentiation, paralleling with the increased expression of TERT. In the present study, we aimed to examine the role of miR-216a in the activation of telomerase during macrophages differentiation.
Methods and results: To investigate whether TERT is expressed particularly in macrophages of the atherosclerotic lesion, we performed immunofluorescent test on the human carotid atherosclerotic plaques, and the co-localization of TERT with macrophage CD68 immuno-reactivity was verified. We next assessed the regulatory effect of miR-216a on TERT expression, and the results showed that miR-216a overexpression markedly increased TERT mRNA expression and telomerase activity by 4.5-fold using the telomerase repeat amplification protocol (TRAP) assay, while inhibition of miR-216a resulted in a significant decrease of telomerase activity. Furthermore, we identified a predicted binding site for miR-216a in the untranslated region of Smad3, then we found miR-216a overexpression significantly decreased Smad3 protein and downstream IκBα protein expression, and vice versa. Next, we observed that miR-216a promoted NF-κB nuclear translocation by immunofluorescent analysis. NF-κB is one of important transcription factors of TERT. To confirm whether miR-216a mediates the activation of telomerase through the Smad3/NF-κB pathway, Smad3 siRNA was transfected, and the results showed that silencing Smad3 promoted NF-κB nuclear translocation and increased TERT expression. On the contrary, inhibiting miR-216a simultaneously, Smad3 and IκBα protein level were rescued, which indicated miR-216a promotes NF-κB nuclear translocation and telomerase activation through targeting Smad3.
Conclusions: Our data indicated that miR-216a mediates the activation of telomerase in macrophages differentiation via the Smad3/NF-κB pathway.
Author Disclosures: S. Yang: None. J. Li: None. Y. Chen: None. R. Hui: None. W. Zhang: None.
- © 2016 by American Heart Association, Inc.