Transcriptomic analysis of native versus cultured human and mouse dorsal root ganglia focused on pharmacological targets

Abstract

Dorsal root ganglion (DRG) neurons detect sensory inputs and are crucial for pain processing. They are often studied in vitro as dissociated cell cultures with the assumption that this reasonably represents in vivo conditions. However, to our knowledge, no study has ever directly compared genome-wide transcriptomes of DRG tissue in vivo versus in vitro, or between different labs and culturing protocols. We extracted bilateral lumbar DRG from C57BL6/J mice and human organ donors, and acutely froze one side and processed the other side as a dissociated cell culture, which was then maintained in vitro for 4 days. RNA was extracted and sequenced using the NextSeq Illumina platform. Comparing native to cultured human or mouse DRG, we found that the overall expression level of many ion channels and GPCRs specifically expressed in neurons is markedly lower in culture, but still expressed. This suggests that most pharmacological targets expressed in vivo are present in culture conditions. However, there are changes in expression levels for these genes. The reduced relative expression for neuronal genes in human DRG cultures is likely accounted for by increased expression of genes in fibroblast-like and other proliferating cells, consistent with the mitotic status of many cells in these cultures. We did find a subset of genes that are typically neuronally expressed, increased in human and mouse DRG cultures, including genes associated with nerve injury and/or inflammation in preclinical models such as BDNF, MMP9, GAL, and ATF3. We also found a striking upregulation of a number of inflammation-associated genes in DRG cultures, although many were different between mouse and human. Our findings suggest an injury-like phenotype in DRG cultures that has important implications for the use of this model system for pain drug discovery.