Single-cell RNA-seq reveals distinct injury responses in different types of DRG sensory neurons.

Ganlu Hu,Zhigang Xue,Xianmin Zhu,Guoping Fan,Youjin Hu,Kevin Huang,Guizhen Du

doi:10.1038/srep31851

Abstract

Peripheral nerve injury leads to various injury-induced responses in sensory neurons including physiological pain, neuronal cell death, and nerve regeneration. In this study, we performed single-cell RNA-sequencing (scRNA-seq) analysis of mouse nonpeptidergic nociceptors (NP), peptidergic nociceptors (PEP), and large myelinated sensory neurons (LM) under both control and injury conditions at 3 days after sciatic nerve transection (SNT). After performing principle component and weighted gene co-expression network analysis, we categorized dorsal root ganglion (DRG) neurons into different subtypes and discovered co-regulated injury-response genes including novel regeneration associated genes (RAGs) in association with neuronal development, protein translation and cytoplasm transportation. In addition, we found significant up-regulation of the genes associated with cell death such as Pdcd2 in a subset of NP neurons after axotomy, implicating their actions in neuronal cell death upon nerve injury. Our study revealed the distinctive and sustained heterogeneity of transcriptomic responses to injury at single neuron level, implicating the involvement of different gene regulatory networks in nerve regeneration, neuronal cell death and neuropathy in different population of DRG neurons.

Highlights

Peripheral nerve injury leads to various injury-induced responses in sensory neurons including physiological pain, neuronal cell death, and nerve regeneration
sciatic nerve transection (SNT) was performed on one side of hind limbs so that dorsal root ganglion (DRG) neurons on the contralateral side at the same level of the lumbar vertebrate bone served as an intact control
Using weighted gene network analysis, we identified injury-responsive genes related to abnormal pain and cell death in small-size nociceptors as well as shared gene modules related to regeneration that are co-regulated among all three subtypes of sensory neurons

Summary

Introduction

Peripheral nerve injury leads to various injury-induced responses in sensory neurons including physiological pain, neuronal cell death, and nerve regeneration. We performed singlecell RNA-sequencing (scRNA-seq) analysis of mouse nonpeptidergic nociceptors (NP), peptidergic nociceptors (PEP), and large myelinated sensory neurons (LM) under both control and injury conditions at 3 days after sciatic nerve transection (SNT). Our study revealed the distinctive and sustained heterogeneity of transcriptomic responses to injury at single neuron level, implicating the involvement of different gene regulatory networks in nerve regeneration, neuronal cell death and neuropathy in different population of DRG neurons. Analysis of bulk samples is not capable of distinguishing different cell types which undergo either cell death and neuropathic pain or alternatively nerve regeneration and functional recovery due to injury-evoked transcriptional changes. We found that differentially regulated genes are functionally correlated with the fates of neuron subtypes after injury including shared up-regulation of regeneration associated genes (RAGs) in all subtypes of neurons (i.e., LM, PEP and NP) as well as preferential induction of cell death related genes in small NP neurons

Methods

Results

Conclusion