RNA-Seq Analysis of Microglia Reveals Time-Dependent Activation of Specific Genetic Programs following Spinal Cord Injury

Abstract

Neurons have inherent competence to regrow following injury, although not spontaneously. Spinal cord injury (SCI) induces a pronounced neuroinflammation driven by resident microglia and infiltrating peripheral macrophages. Microglia are the first reactive glial population after SCI and participate in recruitment of monocyte-derived macrophages to the lesion site. Both positive and negative influence of microglia and macrophages on axonal regeneration had been reported after SCI, raising the issue whether their response depends on time post-lesion or different lesion severity. We analyzed molecular alterations in microglia at several time-points after different SCI severities using RNA-sequencing. We demonstrate that activation of microglia is time-dependent post-injury but is independent of lesion severity. Early transcriptomic response of microglia after SCI involves proliferation and neuroprotection, which is then switched to neuroinflammation at later stages. Moreover, SCI induces an autologous microglial expression of astrocytic markers with over 6% of microglia expressing glial fibrillary acidic protein and vimentin from as early as 72 h post-lesion and up to 6 weeks after injury. We also identified the potential involvement of DNA damage and in particular tumor suppressor gene breast cancer susceptibility gene 1 (Brca1) in microglia after SCI. Finally, we established that BRCA1 protein is specifically expressed in non-human primate spinal microglia and is upregulated after SCI. Our data provide the first transcriptomic analysis of microglia at multiple stages after different SCI severities. Injury-induced microglia expression of astrocytic markers at RNA and protein levels demonstrates novel insights into microglia plasticity. Finally, increased microglia expression of BRCA1 in rodents and non-human primate model of SCI, suggests the involvement of oncogenic proteins after CNS lesion.

Phenotypic heterogeneity of astrocytes in motor neuron disease

“…Table shows some features of aberrant glial cells, including astrocytes or microglia, abnormally‐expressing markers from different cell lineages have been also reported in Alzheimer's disease, Huntington's disease, central nervous system acute lesions and aging, as well as glioma, brain ischemia and trauma, further suggesting the phenotypic switch is strongly associated with inflammation and tissue remodeling after damage.…”

“…expressing markers from different cell lineages have been also reported in Alzheimer's disease, 75 Huntington's disease, 76 central nervous system acute lesions [77][78][79] and aging, 74,80 as well as glioma, 56,57,81 brain ischemia and trauma, 78,79,82 further suggesting the phenotypic switch is strongly associated with inflammation and tissue remodeling after damage. Compared with microglia/macrophages, astrocytes have been regarded to retain fewer phagocytic abilities.…”

Protective Microglial Subset in Development, Aging, and Disease: Lessons From Transcriptomic Studies

“…In a study of facial nucleus axotomy, the authors also related the observed microglial phenotype (134) to the DAM phenotype, as well as to a phenotype found in the Ck-p25 model (109): 72 genes were regulated in common between all three studies representing almost 75% of all genes upregulated in the facial nucleus axotomy model. Interestingly, in an SCI transcriptomic study, a profile of microglia reminiscent of the CD11c+ phenotype was identified (with upregulation of Gpnmb, Spp1, Lpl, Apoe, Igf1, Lgals3, and Itgax among others) and persisted in a full transection model, whereas it contracted concomitantly to recovery in a hemisection model (135), indicative of the transitory nature of this subset. Conversely, in TBI, the microglial signature was further from the CD11c+ microglia signature, although Itgax was among the upregulated genes 14 and 60 days post-injury, possibly indicating once again a dilution of the signature in all microglia (136).…”

Towards Immunotherapy for Pediatric Brain Tumors

“…As the brain develops, immature microglial cells differentiate as they continue a complex lifelong interplay with neuronal cells, downregulating signal pathways such as C-X-X motif chemokine ligand 12 (CXCL12) and C-X-X motif chemokine receptor 4 (CXCR4), and upregulating others [48], to become mature microglia. RNA sequence analysis has provided novel insights into the plasticity of microglia in models of neurodegeneration (e.g., inducing facial nerve axotomy in mice) or inflammation (e.g., inducing spinal cord injury in non-human primate models) [49][50][51]. For instance, an established unique set of genes that are preferentially expressed by aging microglia were found to be enriched in pathologies such as Alzheimer's disease or multiple sclerosis [49].…”