Silencing miR-21-5p in sensory neurons reverses neuropathic allodynia via activation of TGF-β–related pathway in macrophages
Lynda Zeboudj, George Sideris-Lampretsas, Rita Silva, Sabeha Al-Mudaris, Francesca Picco, Sarah Fox, David Chambers, Marzia Malcangio
Lynda Zeboudj, George Sideris-Lampretsas, Rita Silva, Sabeha Al-Mudaris, Francesca Picco, Sarah Fox, David Chambers, Marzia Malcangio
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Research Article Immunology Neuroscience

Silencing miR-21-5p in sensory neurons reverses neuropathic allodynia via activation of TGF-β–related pathway in macrophages

Abstract

Neuropathic pain remains poorly managed by current therapies, highlighting the need to improve our knowledge of chronic pain mechanisms. In neuropathic pain models, dorsal root ganglia (DRG) nociceptive neurons transfer miR-21 packaged in extracellular vesicles to macrophages that promote a proinflammatory phenotype and contribute to allodynia. Here we show that miR-21 conditional deletion in DRG neurons was coupled with lack of upregulation of chemokine CCL2 after nerve injury and reduced accumulation of CCR2-expressing macrophages, which showed TGF-β–related pathway activation and acquired an M2-like antinociceptive phenotype. Indeed, neuropathic allodynia was attenuated after conditional knockout of miR-21 and restored by TGF-βR inhibitor (SB431542) administration. Since TGF-βR2 and TGF-β1 are known miR-21 targets, we suggest that miR-21 transfer from injured neurons to macrophages maintains a proinflammatory phenotype via suppression of such an antiinflammatory pathway. These data support miR-21 inhibition as a possible approach to maintain polarization of DRG macrophages at an M2-like state and attenuate neuropathic pain.

Authors

Lynda Zeboudj, George Sideris-Lampretsas, Rita Silva, Sabeha Al-Mudaris, Francesca Picco, Sarah Fox, David Chambers, Marzia Malcangio

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Figure 8

Intrathecal delivery of antagomir-21–treated BMDMs reverses neuropathic hypersensitivity via upregulation of TGF-βR2 at early stages.

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Intrathecal delivery of antagomir-21–treated BMDMs reverses neuropathic ...
(A) Effect of scramble-treated BMDMs and antagomir-21–treated BMDMs on the development of mechanical hypersensitivity after SNI (n = 10). Data are presented as 50% paw withdrawal thresholds (PWT); mean ± SEM. +P < 0.05, ++P < 0.01, +++P < 0.001 compared with antagomir-21–treated BMDM contralateral thresholds; ***P < 0.001 compared with scramble-treated BMDM thresholds; #P < 0.05, ##P < 0.01 compared with scramble ipsilateral thresholds; 2-way ANOVA followed by Tukey’s multiple-comparison test. (B) Bar graphs represent PWT at 2 hours and (C) 24 hours after intrathecal (i.t.) injection of antagomir-21–treated BMDMs or scramble-treated BMDMs (n = 10). (D) Representative scatterplots of F4/80+CD11b+ macrophages in ipsilateral DRGs 2 hours after i.t. injection of scramble-treated BMDMs or antagomir-21–treated BMDMs (gated on live cells); the bar graphs represent the F4/80+CD11b+ absolute cell number (n = 4–5). (E) Bar graphs of GFP+ BMDM absolute cell number in L3-L4-L5 DRGs 2 hours after i.t. injection (n = 4–5). (F) Representative scatterplots of CD206 and MHCII in CD11b+ F4/80+ macrophages of ipsilateral DRG 2 hours after i.t. injection. (G) Bar graphs represent CD206+MHCII– and (H) MHCII+CD206– absolute cell numbers in CD11b+F4/80+ macrophages 2 hours after i.t. injection (n = 4–5). (I) Representative scatterplots of TGF-βR2 expression in CD11b+F4/80+ macrophages of ipsilateral DRG 2 hours after i.t. injection. (J) TGFBR2+F4/80+ absolute cell numbers (n = 4–5). (K) TGFBR2+CD206+ and (L) TGFBR2+MHCII+ absolute cell numbers in the DRG 2 hours after i.t. injection (n = 4–5). Data are presented as mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001 by 1-way ANOVA followed by Tukey’s multiple-comparison test (B–E, G, H, and J–L).