Differential Transcriptional Profiling of Damaged and Intact Adjacent Dorsal Root Ganglia Neurons in Neuropathic Pain

Abstract

Neuropathic pain, caused by a lesion in the somatosensory system, is a severely impairing mostly chronic disease. While its underlying molecular mechanisms are not thoroughly understood, neuroimmune interactions as well as changes in the pain pathway such as sensitization of nociceptors have been implicated. It has been shown that not only are different cell types involved in generation and maintenance of neuropathic pain, like neurons, immune and glial cells, but, also, intact adjacent neurons are relevant to the process. Here, we describe an experimental approach to discriminate damaged from intact adjacent neurons in the same dorsal root ganglion (DRG) using differential fluorescent neuronal labelling and fluorescence-activated cell sorting (FACS). Two fluorescent tracers, Fluoroemerald (FE) and 1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine perchlorate (DiI), were used, whose properties allow us to distinguish between damaged and intact neurons. Subsequent sorting permitted transcriptional analysis of both groups. Results and qPCR validation show a strong regulation in damaged neurons versus contralateral controls as well as a moderate regulation in adjacent neurons. Data for damaged neurons reveal an mRNA expression pattern consistent with established upregulated genes like galanin, which supports our approach. Moreover, novel genes were found strongly regulated such as corticotropin-releasing hormone (CRH), providing novel targets for further research. Differential fluorescent neuronal labelling and sorting allows for a clear distinction between primarily damaged neuropathic neurons and “bystanders,” thereby facilitating a more detailed understanding of their respective roles in neuropathic processes in the DRG.

Therapeutic effects of an anti-ADAMTS-5 antibody on joint damage and mechanical allodynia in a murine model of osteoarthritis

“…Indeed, ADAMTS-5 can be expressed by DRG neurons [24, 25] and chondroitin sulfate proteoglycans have been shown to interact with peripheral nerves [26]. We demonstrated that there was no acute effect of ADAMTS-5 treatment on mechanical allodynia (Fig 3B), but long-term effects cannot be ruled out and future work will seek to understand this in greater detail.…”

An evolutionarily conserved gene family encodes proton-selective ion channels

“…Thus, we expect OTOP channels are restricted to cell types that use changes in intracellular pH for cell signaling or to regulate biochemical or developmental processes. Along with a role in formation of vestibular otoconia (14), OTOP1 has been shown to protect mice from obesity-induced metabolic dysfunction (15), and it is upregulated in dorsal root ganglion cells in response to cell damage (20). The knowledge that this gene family encodes proton channels can be used to understand its contribution to physiology and disease.…”

Histone deacetylase inhibitors prevent persistent hypersensitivity in an orofacial neuropathic pain model

“…39,40 Overexpression of Atf3, as part of the inducible nerve RAG response, along with its “effector” RAGs, Sprr1a, and Gal, has been observed previously following both facial nerve damage 41 and sciatic nerve injury 42 in mice. Atf3 mutant mice have decreased neurite outgrowth on primary DRG neurons.…”