MiR-146a Promotes Tolerogenic Properties of Dendritic Cells and through Targeting Notch1 Signaling

ABSTRACT

Background

MiR-146a has been shown to negatively regulate innate immune, inflammatory response and antiviral pathway, however, its role in the tolerogenic responses remains largely unknown. This study aimed to investigate the role of miR-146a in the OVA-induced allergic inflammation of dendritic cells (DCs).

Methods

Bone marrow-derived DCs (BMDCs) were treated with OVA (100 µg/ml) for 24 h. MiR-146a expressions were assessed by quantitative RT-PCR. BMDCs were transfected with miR-146a mimics or inhibitor. Cell surface markers were analyzed by flow cytometry. Cytokine levels were determined by ELISA assay. Mixed lymphocyte culture assay was adopted to assess CD4 + T-cell differentiation. The 3ʹ UTR luciferase reporter assay was utilized to determine the miRNA target sequence.

Results

OVA treatment significantly up-regulated miR-146a in BMDCs in a dose- and time-dependent manner. In the OVA-treated DCs, overexpression of miR-146a (mimics transfection) down-regulated the surface markers (CD80, CD86) and increased production of anti-inflammatory cytokines TGF-β1 and IL-10 but decreased pro-inflammatory cytokine IL-12. MiR-146a overexpression promoted immature DC to induce regulatory T cells (Treg) differentiation. By contrast, transfection of miR-146a inhibitor into DC exhibited the opposite trends. Notch1 was a direct target of miR-146a, and Notch1 knock-down induced similar effects as miR-146a mimics transfection in BMDCs. Moreover, the effect of miR-146a inhibitor on OVA-induced DC was attenuated by Notch1 knock-down.

Conclusion

miRNA-146a promoted tolerogenic properties of DCs, at least partially, through targeting Notch1 signaling.

KEYWORDS:

• Dendritic cells
• tolerogenic responses
• miR-146a
• Notch1
• Treg

Compliance with Ethical Standards

All animal experiments were performed in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals and were approved by the Scientific Investigation Board of Sun Yat-Sen University (Guangzhou, China).

Conflict of Interest

The authors declare that they have no conflict of interest.

Additional information

Funding

This work was supported by the National Natural Science Foundation of China [Grant number U0832007].