Systemic interleukin 1 receptor blockade regulates muscle pain development

in a mouse model of ischemia and reperfusion

Jordan E. Lamb1, Jessica L. Ross1, Renita C. Hudgins1, Luis F. Queme1, Zachary K. Ford1, Michael P. Jankowski1,2
Dept. of Anesthesia1, Cincinnati Childrens Hospital Medical Center, and Dept. of Pediatrics2, University of Cincinnati, Cincinnati, OH 45229

Introduction
Musculoskeletal pain is an exceedingly common complaint for
patients who seek medical treatment; however, the specific
mechanisms that lead to muscle pain development are not
fully understood.
Recent studies have suggested that issues of peripheral
circulation are an underlying cause of muscle pain in patients
with disorders such as peripheral vascular disease, sickle cell
anemia, complex regional pain syndrome, and fibromyalgia.
We have previously found in our mouse model of disrupted
peripheral circulation that ischemia and reperfusion injury (I/R)
altered chemosensitivity and mechanical responsiveness in
thinly myelinated (group III) and unmyelinated (group IV)
muscle sensory neurons, which correlated with increased
pain-related behaviors.
I/R also induced upregulation of specific receptors and
channels involved in sensory processing in the dorsal root
ganglia (DRG) as well as growth factors and cytokines within
the affected muscle tissue.
We have previously shown that increased interleukin 1
(IL1), a pro-inflammatory cytokine, within the ischemic
muscle drives upregulation of DRG acid sensing ion channel 3
(ASIC3) via enhanced interleukin 1 receptor (IL1r1)
expression in sensory neurons to induce peripheral
sensitization and generate ischemic muscle pain.

Results

Ex Vivo recording preparation

I/R injury increases IL1 levels in the muscles

*p value < 0.05 vs nave; One-way ANOVA with Tukeys post hoc
Occlusion of Brachial Artery
#p value < 0.05 compared to IL1RA+I/R, *p value < 0.05 vs nave; 2-way RM-ANOVA with Holm-Sidak post hoc

I/R-induced upregulation of DRG IL1r1 is blocked by nervespecific siRNA injection or systemic IL1RA delivery

Systemic treatment with IL1RA reverses I/R induced changes

in muscle afferents similar to nerve specific inhibition of IL1r1

Prolonged exercise blocks increases in muscle IL1

induced by I/R

*p value < 0.05 vs nave; One-way ANOVA with Tukeys post hoc
**p value 0.001 vs all other conditions; Two-way ANOVA with Holm-Sidak
Exercise x Injury Interaction p<0.001

Increased DRG ASIC3 after I/R is also blocked by PenIL1r1

siRNA injection or systemic IL1RA delivery

Conclusions

Hypothesis
Systemic blockade of IL1r1 with the IL1 receptor antagonist
(IL1RA) will prevent the injury-induced upregulation of ASIC3 in
muscle afferents and prevent the development of peripheral
sensitization and pain-related behaviors after I/R.

*p value < 0.05 vs nave; Chi-squared analysis or Kruskal-Wallis One-Way

ANOVA on Ranks with Dunns post hoc

Blockade of IL1r1 reduces I/R induced changes in muscle

pain-related behaviors

Methods
IL1RA intraperitoneal (IP) injections (3 mg/kg based on doseresponse analysis)
Surgical brachial artery occlusion (6hr) & reperfusion (18hr)
Median and ulnar nerve-specific Penetratin 1-conjugated
siRNA injections
Analysis of spontaneous pain (forepaw guarding), mechanical
hypersensitivity (von Frey filament stimulation of forepaws),
muscle function (grip strength), and voluntary activity levels
(monitored wheel running)
Ex vivo forepaw muscles/median and ulnar nerves/DRG/
spinal cord electrophysiological recording
Real-time quantitative RT-PCR of whole DRGs
Western blot of forepaw muscles and affected DRGs
Statistical analyses

IL1r1 inhibition restores voluntary activity levels

reduced by I/R

These data suggest that the IL1r1 pathway regulates I/Rinduced peripheral sensitization and pain-related behaviors.
Inhibition of muscle IL1 signaling may be a potential
therapeutic target for ischemic myalgia.

Future Directions

*p value < 0.05 vs nave; One-way ANOVA with Tukeys post hoc

I/R-induced upregulation of purinergic receptors in DRGs is

partially blocked by PenIL1r1 injection or IL1RA treatment

Investigate effect of IL1RA on other I/R related myalgia

conditions in mice
Test IL1RA effects on I/R in female rodents
Assess the potential value in utilizing both
pharmacological therapies targeting the IL1r1 pathway in
addition to exercise regimens to treat ischemic myalgia
Determine exact origin of IL1 in ischemic muscle with
targeted knockdown of specific immune cell function

Acknowledgements

*p value < 0.05 vs nave; One-way ANOVA with Tukeys post hoc

*p value < 0.05 vs nave, #p value <0.05 vs nave and control I/R; 2-way RM-ANOVA with Holm-Sidak post hoc

This research was supported by grants from the Rita Allen

Foundation and the American Pain Society (MPJ), the
International Association for the Study of Pain Early Career
Grant (MPJ), and the NIH/NIAMS (R01AR045581-01A1).