Skin-resident dendritic cells mediate postoperative pain via CCR4 on sensory neurons.

Inflammatory pain, such as hypersensitivity resulting from surgical tissue injury, occurs as a result of interactions between the immune and nervous systems with the orchestrated recruitment and activation of tissue-resident and circulating immune cells to the site of injury. Our previous studies identified a central role for Ly6Clow myeloid cells in the pathogenesis of postoperative pain. We now show that the chemokines CCL17 and CCL22, with their cognate receptor CCR4, are key mediators of this response. Both chemokines are up-regulated early after tissue injury by skin-resident dendritic and Langerhans cells to act on peripheral sensory neurons that express CCR4. CCL22, and to a lesser extent CCL17, elicit acute mechanical and thermal hypersensitivity when administered subcutaneously; this response abrogated by pharmacological blockade or genetic silencing of CCR4. Electrophysiological assessment of dissociated sensory neurons from naïve and postoperative mice showed that CCL22 was able to directly activate neurons and enhance their excitability after injury. These responses were blocked using C 021 and small interfering RNA (siRNA)-targeting CCR4. Finally, our data show that acute postoperative pain is significantly reduced in mice lacking CCR4, wild-type animals treated with CCR4 antagonist/siRNA, as well as transgenic mice depleted of dendritic cells. Together, these results suggest an essential role for the peripheral CCL17/22:CCR4 axis in the genesis of inflammatory pain via direct communication between skin-resident dendritic cells and sensory neurons, opening therapeutic avenues for its control.

Chronic mechanical hypersensitivity in experimental autoimmune encephalomyelitis is regulated by disease severity and neuroinflammation.

Chronic pain severely affects quality of life in more than half of people living with multiple sclerosis (MS). A commonly-used model of MS, experimental autoimmune encephalomyelitis (EAE), typically presents with hindlimb paralysis, neuroinflammation and neurodegeneration. However, this paralysis may hinder the use of pain behavior tests, with no apparent hypersensitivity observed post-peak disease. We sought to adapt the classic actively-induced EAE model to optimize its pain phenotype. EAE was induced with MOG35-55/CFA and 100-600 ng pertussis toxin (PTX), and mice were assessed for mechanical, cold and thermal sensitivity over a 28-day period. Spinal cord tissue was collected at 14 and 28 days post-injection to assess demyelination and neuroinflammation. Only mice treated with 100 ng PTX exhibited mechanical hypersensitivity. Hallmarks of disease pathology, including demyelination, immune cell recruitment, cytokine expression, glial activation, and neuronal damage were higher in EAE mice induced with moderate (200 ng) doses of pertussis toxin, compared to those treated with low (100 ng) levels. Immunostaining demonstrated activated astrocytes and myeloid/microglial cells in both EAE groups. These results indicate that a lower severity of EAE disease may allow for the study of pain behaviors while still presenting with disease pathology. By using this modified model, researchers may better study the mechanisms underlying pain.

Circadian control of pain and neuroinflammation.

The importance of a neuroinflammatory response to the development and maintenance of inflammatory and neuropathic pain have been highlighted in recent years. Inflammatory cells contributing to this response include circulating immune cells such as monocytes, T and B lymphocytes, and neutrophils, as well as microglia in the central nervous system. Pain signals are transmitted via sensory neurons in the peripheral nervous system, which express various receptors and channels that respond to mediators secreted from these inflammatory cells. Chronobiological rhythms, which include the 24-hr circadian cycle, have recently been shown to regulate both nervous and immune cell activity and function. This review examines the current literature on chronobiological control of neuroinflammatory processes, with a focus on inflammatory and neuropathic pain states. While the majority of this work has stemmed from observational studies in humans, recent advances in using animal models have highlighted distinct mechanisms underlying these interactions. Better understanding interactions between the circadian and neuroimmune systems can help guide the development of new treatments and provide improved care for patients suffering from acute and chronic pain.

Advanced Dynamic Weight Bearing as an Observer-independent Measure of Hyperacute Hypersensitivity in Mice.

Background: Standard methods assessing pain in rodents are often observer dependent, potentially resulting in biased outcomes. Advanced dynamic weight bearing (ADWB) offers an observer-independent approach that can provide objective, reliable data in preclinical pain research. Aims: The aim of this study was to characterize the use of ADWB in assessing murine responses to allyl isothiocyanate (AITC)-induced hyperacute hypersensitivity and identify best practices for use of the device. Methods: Male C57BL/6J mice received intraplantar injections of saline or 0.1% AITC solution and were assessed using the ADWB system; simultaneous observer-dependent durations of paw licking and biting were measured. ADWB data were analyzed using the proprietary software from Bioseb and correlated to observer-dependent results, with parameters assessed to optimize data collected. Results: ADWB detected pain-directed changes in weight and surface area distribution in AITC-treated mice, with paw weight and surface area placement correlating to paw licking and biting. Optimization of adjustable threshold parameters allowed for reduced coefficients of variability and increased duration of validated data. Conclusions: The ADWB assay provides an efficient and unbiased measure of chemical-induced hyperacute hypersensitivity in mice. ADWB detection parameters influence amount of validated data and variability, a consideration for data analysis in future studies.

Contexte: Les méthodes standard d'évaluation de la douleur chez les rongeurs dépendent souvent de l'observateur, ce qui peut fausser les résultats. La mise en charge dynamique avancée offre une approche indépendante de l'observateur qui peut fournir des données objectives et fiables dans la recherche préclinique sur la douleur.Objectifs: L'objectif de cette étude était de caractériser l'utilisation de la mise en charge dynamique avancée dans l'évaluation des réponses murines à l'hypersensibilité hyperaiguë induite par l'isothiocyanate d'allyle et de répertorier les meilleures pratiques d'utilisation de l'appareil.Méthodes: Des souris C57BL/6J mâles ont reçu des injections intraplantaires de solution saline ou de solution d'isothiocyanate d'allyle à 0,1 % et ont été évaluées à l'aide du système de mise en charge dynamique avancée; les durées simultanées de léchage et de morsure des pattes, dépendantes de l'observateur, ont été mesurées. Les données obtenues par la mise en charge dynamique avancée ont été analysées à l'aide du logiciel propriétaire de Bioseb et corrélées aux résultats dépendants de l'observateur, avec des paramètres évalués pour optimiser les données collectées.Résultats: L'essai réalisé à l'aide de la mise en charge dynamique avancée a détecté des changements de poids et de distribution de surface liés à la douleur chez les souris traitées à l'isothiocyanate d'allyle, le poids des pattes et le placement de la surface étant corrélés au léchage et à la morsure des pattes. L'optimisation des paramètres de seuil ajustables a permis de réduire les coefficients de variabilité et d'augmenter la durée des données validées.Conclusion: L'essai réalisé à l'aide de la mise en charge dynamique avancée fournit une mesure efficace et impartiale de l'hypersensibilité hyperaiguë induite par les produits chimiques chez la souris. Les paramètres de détection du système de mise en charge dynamique avancée influencent la quantité de données validées et la variabilité, ce qui doit être pris en compte pour l'analyse des données dans les études futures.

Spinal cord injury in mice affects central and peripheral pathology in a severity-dependent manner.

ABSTRACT: Chronic pain is a common medical complication experienced by those living with spinal cord injury (SCI) and leads to worsened quality of life. The pathophysiology of SCI pain is poorly understood, hampering the development of safe and efficacious therapeutics. We therefore sought to develop a clinically relevant model of SCI with a strong pain phenotype and characterize the central and peripheral pathology after injury. A contusion (50 kdyn) injury, with and without sustained compression (60 seconds) of the spinal cord, was performed on female C57BL/6J mice. Mice with compression of the spinal cord exhibited significantly greater heat and mechanical hypersensitivity starting at 7 days postinjury, concomitant with reduced locomotor function, compared with those without compression. Immunohistochemical analysis of spinal cord tissue revealed significantly less myelin sparing and increased macrophage activation in mice with compression compared with those without. As measured by flow cytometry, immune cell infiltration and activation were significantly greater in the spinal cord (phagocytic myeloid cells and microglia) and dorsal root ganglia (Ly6C+ monocytes) after compression injury. We also decided to investigate the gastrointestinal microbiome, as it has been shown to be altered in patients with SCI and has recently been shown to play a role in immune system maturation and pain. We found increased dysbiosis of the gastrointestinal microbiome in an injury severity-dependent manner. The use of this contusion-compression model of SCI may help advance the preclinical assessment of acute and chronic SCI pain and lead to a better understanding of mechanisms contributing to this pain.

Weight bearing as a measure of disease progression in experimental autoimmune encephalomyelitis.

Motor disability in multiple sclerosis is often modeled using experimental autoimmune encephalomyelitis (EAE) and assessed using the clinical score (CS), an observer-dependent tool that can lead to potential bias. The Advanced Dynamic Weight Bearing (ADWB) system was evaluated as an observer-independent measurement of EAE symptoms. ADWB detected weight shifts onto the front paws as mice develop hindlimb motor disability. CS and ADWB were strongly correlated, indicated that these measures are comparable and suggesting that ADWB may be an appropriate observer-independent tool for the assessment of EAE progression.

γδ T Cells Modulate Myeloid Cell Recruitment but Not Pain During Peripheral Inflammation.

Circulating immune cells, which are recruited to the site of injury/disease, secrete various inflammatory mediators that are critical to nociception and pain. The role of tissue-resident immune cells, however, remains poorly characterized. One of the first cells to be activated in peripheral tissues following injury are γδT cells, which serve important roles in infection, disease, and wound healing. Using a mouse line lacking these cells, we sought to identify their contribution to inflammatory pain. Three distinct models of peripheral inflammatory pain were used: intraplantar injection of formalin (spontaneous inflammatory pain), incisional wound (acute inflammatory pain), and intraplantar injection of complete Freund's adjuvant (chronic inflammatory pain). Our results show that absence of γδT cells does not alter baseline sensitivity, nor does it result in changes to mechanical or thermal hypersensitivity after tissue injury. Myeloid cell recruitment did show differential changes between models of acute and chronic inflammatory pain. These results were consistent in both male and female mice, suggesting that there are no sex differences in these outcomes. This comprehensive characterization suggests that γδT cells do not contribute to basal sensitivity or the development and maintenance of inflammatory pain.

Chronobiological regulation of psychosocial and physiological outcomes in multiple sclerosis.

There is mounting scientific evidence showing the importance of innate biological rhythms on disease onset and progression. Perhaps the most important of these is the circadian rhythm, a cycle of oscillations lasting approximately 24 h. Recent work has shown that circadian rhythms are intrinsically linked to the immune system in a bidirectional fashion, and that disruption of these cycles can contribute to changes in pathology and quality of life (including fatigue, mood, and disability). This is particularly true in diseases of the nervous and immune systems. We review here the current preclinical and clinical literature to highlight interactions between circadian rhythms and multiple sclerosis, as well as its animal model, experimental autoimmune encephalomyelitis. We highlight potential benefits of chronotherapy (the temporal administration of immunomodulatory drugs) in an effort to increase treatment efficacy and reduce the negative side-effects of the drugs that often burden those suffering from the disease.