Background & Aims

Postamputation pain is currently managed unsatisfactorily with neuron-targeted pharmacological and interventional therapies. Non-neuronal pain mechanisms have emerged as crucial factors in the development and persistence of chronic pain. Consequently, these mechanisms offer exciting prospects as innovative therapeutic targets.
We examined the hypothesis that engaging mesenchymal stem cells (MSCs), either by their direct use, or after in vitro exposure, or in vivo treatment with the oligodeoxynucleotide IMT504, would foster local neuroimmune interactions, leading to a potential reduction in postamputation pain. We included in the analysis, three different sites in the pain neuroaxis, two in the periphery (the injured nerve and corresponding dorsal root ganglia (DRG)), and one centrally located, the dorsal horn of the spinal cord.

Methods

MSC samples from 4 different donors undergoing hip replacement, and 1 neuroma sample from a patient suffering postamputation pain of over a year duration, were utilized for in vitro or ex vivo analyses of pro-migratory and anti-inflammatory phenotype-inducing actions of IMT504 over MSCs. Migration assays, plus ELISA and qRT-PCR analysis of cells, neuroma and conditioned media resulting from culturing experiments were carried out. In vivo studies included use of male, 8-week-old Sprague Dawley rats undergoing full axotomy of the right sciatic nerve and receiving two consecutive subcutaneous doses of IMT504 (6 mg/kg) on days 0 and 1 after injury. Rats were scored during 28 days, observing for signs of autotomy (self-mutilation behavior). Flow cytometry of injured nerves and DRGs was employed to address MSC tissue homing. Immunofluorescence and ELISA analysis of the spinal cord, injured nerves or DRGs was employed to address changes in glial reactivity and the immune microenvironment.

Results

We show, on an ex vivo neuroma model from a phantom limb pain patient, that IMT504 engages human primary MSCs, enhancing their migratory capacity and promoting their acquisition of an anti-inflammatory phenotype, ultimately and positively affecting the inflammatory microenvironment of the injured nerve. Reverse translation experiments recapitulated these effects. Thus, in an in vivo rat model, IMT504 exhibited strong efficacy in preventing autotomy (self-mutilation) behaviors. Accordingly, the exogenous systemic administration of MSCs phenocopied the behavioral effects of IMT504. Rats treated with IMT504 exhibited substantial accumulation of MSCs in the neuroma and associated DRG, as well as the establishment of an anti-inflammatory phenotype in these compartments. These peripheral effects were paralleled by reduced glial reactivity and presence of pro-inflammatory cytokines in the dorsal horn of the spinal cord, suggestive of diminished central pro-nociceptive influences.

Conclusions

Our findings underscore the mechanistic relevance of MSCs and the translational therapeutic potential of IMT504 to engage non-neuronal cells for prevention of postamputation pain. The present study suggests that IMT504-dependent recruitment of endogenous MSCs within severely injured nerves and DRGs may prevent post-amputation pain by modifying the inflammatory scenario at relevant sites in the pain pathway. Reinforcing data in rat and human tissues supports the potential therapeutic value of IMT504 in patients suffering postamputation pain.

References

•List EB, Krijgh DD, Martin E, Coert JH: Prevalence of residual limb pain and symptomatic neuromas after lower extremity amputation: a systematic review and meta-analysis. Pain 162:1906-1913, 2021.
•Brumovsky PR, Casadei M, Coronel MF, Leiguarda C, Montaner A, Rubione J, Villar M: Modulation of mesenchymal stem cells, glial cells and the immune system by oligodeoxynucelotides as a novel multi-target therapeutic approach against chronic pain. In: Atta-ur-Rahman, Amtul Z (eds): Frontiers in Clinical Drug Research – CNS and Neurological Disorders. Singapore, Bentham Books, 2022. pp 226-268.
•Grace PM, Tawfik VL, Svensson CI, Burton MD, Loggia ML, Hutchinson MR: The Neuroimmunology of Chronic Pain: From Rodents to Humans. J Neurosci 41:855-865, 2021.

Presenting Author

Bernardo Miguel

Poster Authors

Bernardo Miguel

Dr. med.

Instituto de investigaciones en medicina traslacional

Lead Author

Mailin Casadei PhD

Instituto de Investigaciones en Medicina Traslacional, CONICET-Universidad Austral, Argentina

Lead Author

Julia Rubione PhD

Instituto de Investigaciones en Medicina Traslacional, CONICET-Universidad Austral, Argentina

Lead Author

Esteban Fiore PhD

Instituto de Investigaciones en Medicina Traslacional, CONICET-Universidad Austral, Argentina

Lead Author

Diego Mengelle MD.

Hospital Universitario Austral, Universidad Austral, Argentina

Lead Author

Roberto A. Guerri-Guttenberg MD

PhD

Hospital Universitario Austral, Universidad Austral, Argentina

Lead Author

Alejandro Montaner PhD

Instituto de Ciencia y Tecnología “César Milstein”, CONICET-Fundación Pablo Cassará

Lead Author

Marcelo Villar

Universidad Austral, Faculty of Biomedical Sciences

Lead Author

Luis Constandil-Córdova PhD

Universidad de Santiago de Chile, Santiago de Chile, Chile

Lead Author

Alfonso Romero-Sandoval

Wake Forest University School of Medicine, Winston Salem, North Carolina, United States

Lead Author

Pablo Brumovsky

Instituto de Investigaciones en Medicina Traslacional, CONICET-Universidad Austral, Argentina

Lead Author

Topics

  • Specific Pain Conditions/Pain in Specific Populations: Pain in Amputees