MiR-146a Regulates Cytokine Responses in Activated B Cells Through IRAK1

Objectives: MicroRNAs (miRs) play an important role in immune system balance and have been implied in the regulation of various T cell functions. However, it is unclear whether miRs regulate B cell immune responses in the context of periodontitis. This study is to explore the role of miR-146a on the inflammatory cytokine production of P. gingivalis LPS-challenged B cells.
Methods: Primary B cells were harvested from the spleen of mice. Expression of pro-inflammatory cytokines, miR-146a, IL-1 receptor associated kinase-1 (IRAK1), and TNF receptor associated factor (TRAF) 6 in B cells cultured in the absence or presence of P. gingivalis LPS were detected by real-time quantitative polymerase chain reaction (RT-qPCR) or enzyme-linked immunosorbent assay (ELISA). miR-146a mimic and inhibitor were used to detect the changes of pro-inflammatory cytokines via RT-qPCR and ELISA. Bioinformatics, RT-qPCR, luciferase reporter gene assay and gain-of-function assay were applied to explore the binding target of miR-146a.

Results: P. gingivalis LPS increased the expression of miR-146a, interleukin (IL)-1, -6 and -10, IRAK1, but not TRAF6 in B cells. Addition of miR-146a mimic could decrease not only the levels of these cytokines but also IRAK1 expression, meanwhile, miR-146a inhibitor increased the levels of these cytokines. Overexpression of IRAK1 reversed the inhibitory effect of miR-146a on IL-1, -6 and -10. Furthermore, miR-146a was able to bind with IRAK1 3’ untranslated region (UTR) as detected by reporter gene assay.
Conclusions: miR-146a could inhibit pro-inflammatory cytokine secretion in P. gingivalis LPS-challenged B cells through down-regulating IRAK1 signaling, suggesting that miR-146a functions as a negative regulator in periodontal inflammation.