P070/O09 miR-221-3p is upregulated in rheumatoid arthritis and affects JAK3/STAT3 signaling in M2-macrophages. (March 2019) - Quero, L
Abstract : Career situation of first and presenting author: Post-doctoral fellow. Introduction: Recurrent activation of synovial macrophages is considered as intrinsic factor that precedes rheumatoid arthritis (RA)-related autonomous cytokine cascades. microRNAs (miRs) and Toll-like receptors (TLRs) play a pivotal role in regulating the inflammatory response in innate immune cells and are at dysregulated levels in RA patients. Objectives: Here we explored TLR-regulated miRs that tune the inflammatory response of M2-type macrophages. Methods: Based on a miR-screen in TLR-stimulated human M2-macrophages we selected miR-221-3p to analyze its expression profile in clinical samples from RA, other inflammatory arthritis (OIA), osteoarthritis (OA) and healthy donors (HD) by qPCR. We challenged M1- and M2-macrophages with elevated miR-221-3p and/or miR-155-5p expression thereby mimicking RA-related conditions. Cytokine/chemokine secretion of IL-6, IL-10, IL-12, IL-8 and CXCL13 was measured using ELISA. Changes of inflammatory pathways including JAK/STAT were evaluated by Western Blot. Results: TLR4-stimulation of M2-macrophages results in suppression of miR-221-3p, while in vivo expression in synovial tissue and fluids is significantly increased in RA versus OA or OIA. M2-macrophages with high levels of miR-221-3p lose their anti-inflammatory response and support a M1-like profile. miR-221-3p in combination with miR-155-5p leads to the secretion of M1-specific IL-12 and lowered IL-10. miR-221-3p is affecting the JAK/STAT pathway in M2-macrophages by downregulating JAK3 protein leading to decreased STAT3 activation. Treatment of M2-macrophages using JAK3 inhibitors display the same shift in cytokine/chemokine pattern as increased miR-221-expression. Conclusions: TLR4-activated M2-macrophages exposed to elevated miR-221-3p expression diminished anti-inflammatory response partly by suppressing JAK3/STAT3 pathway. Thus, miR-221-3p is a homeostatic regulator of M2-M1 inflammatory function, which is dysregulated in RA condition. Acknowledgements: Tissue RNA from RA and OA patients was kindly provided by Caroline Ospelt, University of Zurich, Center of Experimental Rheumatology, Switzerland. Disclosure of Interest: None declared.