A transcriptional toolbox for exploring peripheral neuroimmune interactions

Abstract

Correct communication between immune cells and peripheral neurons is crucial for the protection of our bodies. Its breakdown is observed in many common, often painful conditions, including arthritis, neuropathies, and inflammatory bowel or bladder disease. Here, we have characterised the immune response in a mouse model of neuropathic pain using flow cytometry and cell-type-specific RNA sequencing (RNA-seq). We found few striking sex differences, but a very persistent inflammatory response, with increased numbers of monocytes and macrophages up to 3 1/2 months after the initial injury. This raises the question of whether the commonly used categorisation of pain into “inflammatory” and “neuropathic” is one that is mechanistically appropriate. Finally, we collated our data with other published RNA-seq data sets on neurons, macrophages, and Schwann cells in naive and nerve injury states. The result is a practical web-based tool for the transcriptional data mining of peripheral neuroimmune interactions. http://rna-seq-browser.herokuapp.com/

Stem Cell Therapy for Modulating Neuroinflammation in Neuropathic Pain

“…The endoneurial accumulation of neutrophils at sites of peripheral nerve injury is important in the early pathogenesis of hyperalgesia. Further, neuroimmune interactions occur as a result of peripheral nerve injury and are important in the subsequent development of NP [ 10 ].…”

Probing the peripheral immune response in mouse models of oxaliplatin-induced peripheral neuropathy highlights their limited translatability

“…In potentially the only peripherally focused, flow cytometry-heavy investigation of OIPN, oxaliplatin-induced increases were noted specifically in circulating T cell populations, though such differences were not detected in either the lumbar DRG or sciatic nerves (Makker et al, 2017). However, their flow cytometric investigations did not extend to myeloid cell types in the periphery, such as macrophages, which have been implicated in the development and even maintenance of neuropathic pain in a variety of models (Kiguchi et al, 2019;Ristoiu, 2013), including nerve injury (Liang et al, 2020;Yu et al, 2020).…”

“…Lastly, the right and left inguinal lymph nodes were exposed and dissected out into F12, trying to separate them as much as possible from the surrounding fat. Tissues were kept in F12 on ice until all animals in a given batch were processed, they were then dissociated largely in accordance with previously described methods (Liang et al, 2020). Briefly, tissues were transferred into 50µl of digestion mix and incubated at 37°C, shaking at 220RPM for 45 minutes (see Table 1 and Table 2 for digestion mixes used for each tissue type).…”

“…Staining of samples for flow cytometry was conducted as described previously (Liang et al, 2020). Briefly, to distinguish live cells, samples were incubated in a fixable yellow viability dye (Invitrogen, # L34959) for 30 minutes, followed by 30 minutes incubation in a mix of directly conjugated antibodies and Fc block (see Table 4 for antibody panel and concentrations).…”

“…Harvesting and culture of bone marrow derived macrophages (BMDMs) Harvesting and culture of BMDMs was conducted as described previously (Liang et al, 2020). Briefly, femur and tibia bones from both hind limbs were collected and the bone marrow was flushed out with cold PBS.…”