Background & Aims
Neuropathic pain (NP) is a maladaptive condition developing after a spinal cord injury (SCI). Nearly half of people living with SCI develop NP, experiencing symptoms such as allodynia and hyperalgesia. While etiologically complex, dysregulation of the inflammatory response is a key contributor to NP development, promoting central sensitization along pain pathways. Small extracellular vesicles (sEVs) are secreted across cell types and participate in a wide range of homeostatic and pathological processes. Work from our lab suggests that sEVs from polarized M?s attenuate allodynia in a rat SCI model of neuropathic pain. However, the mechanisms by which sEVs produce this effect are unexplored. We hypothesize that administration of sEVs from polarized M?s attenuate NP through modulating the immune response after SCI, altering nociceptor excitability in the dorsal root ganglion (DRG), as well as the anatomical distribution of nociceptive afferents in the dorsal horn.
Methods
We performed von Frey assessment of mechanical pain sensitivity in adult, female Sprague Dawley rats before and after a C5 unilateral contusion SCI, noting decreases in paw withdrawal threshold as an indicator of neuropathic pain. Fourteen days post injury (dpi), SCI rats received 10ug of sEVs derived from either LPS-stimulated or unstimulated RAW 264.7 cells, or a saline vehicle via lumbar puncture. Behavior data was analyzed using a mixed effects approach to determine the effect of each treatment on pain behavior over time. At 18 or 35 dpi, cervical spinal cord and C7-C8 DRG were dissected for immunohistochemical assessments of injury severity, microglial/macrophage activation, and sprouting of CGRP/IB4 positive primary afferent fibers. We are currently exploring the effects of sEV administration on nociceptor excitability after SCI from a subset of rats 4 days after sEV treatment via whole cell patch clamp of isolated IB4+ nociceptors.
Results
Paw withdrawal thresholds to tactile stimuli are decreased after SCI. 4 days after administration of sEVs from LPS-stimulated RAW 264.7 cells, paw withdrawal thresholds were restored compared to rats treated with vehicle or unstimulated RAW 264.7 cell sEVs. This effect was maintained for nearly two weeks before returning to pre-treatment levels and was associated with changes in CGRP/IB4 primary afferent distribution and density in the ipsilateral dorsal horn, as well as alterations to microglial/macrophage activation. Analysis of membrane properties of nociceptors in each treatment group is underway.
Conclusions
Our data suggests that sEVs from LPS-stimulated macrophages could promote analgesia in SCI rats by attenuating DRG nociceptor hyperexcitability and the representation of nociceptive afferents in the spinal dorsal horn. These findings open the possibility of driving endogenous macrophages to an LPS-activation state to promote the amelioration of neuropathic pain after SCI.
References
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2.Viswanath O, Urits I, Burns J, et al. Central Neuropathic Mechanisms in Pain Signaling Pathways: Current Evidence and Recommendations. Adv Ther. 2020;37(5):1946-1959. doi:10.1007/s12325-020-01334-w
3.McDonald MK, Tian Y, Qureshi RA, et al. Functional significance of macrophage-derived exosomes in inflammation and pain [published correction appears in Pain. 2022 Feb 1;163(2):e383-e384]. Pain. 2014;155(8):1527-1539. doi:10.1016/j.pain.2014.04.029
4.Jean-Toussaint R, Lin Z, Tian Y, et al. Therapeutic and prophylactic effects of macrophage-derived small extracellular vesicles in the attenuation of inflammatory pain. Brain Behav Immun. 2021;94:210-224. doi:10.1016/j.bbi.2021.02.005
Presenting Author
Jason Wheeler
Poster Authors
Topics
- Specific Pain Conditions/Pain in Specific Populations: Neuropathic Pain - Central