Investigating the role of microglial senescence in central nervous system injury and regeneration

Abstract

Microglia are the resident macrophages of the central nervous system (CNS). Here, they have many roles, a key one of these being the regulation of myelin health. Myelin is a lipid-rich, insulating layer that wraps around the axons of neurons, allowing for rapid signal transmission.

Although microglia promote remyelination following demyelination, they can become dysfunctional with ageing and neurodegenerative disease in association with impaired remyelination. The mechanisms underpinning this dysfunction are unclear.

One of the hallmarks of ageing is cellular senescence, a state of permanent growth arrest featuring phenotypic alterations and a specific secretory profile. Senescent cells accumulate in tissue with age and are implicated in neurodegenerative diseases such as Alzheimer’s disease (AD) and multiple sclerosis (MS). I asked whether cellular senescence may play a role in the ability of microglia to regulate myelin health as it can promote repair yet also impede it when dysregulated.

While microglia have been shown to exhibit features of senescence with ageing, microglial senescence has not been comprehensively characterised, and how it may differ in distinct disease contexts is also unknown. To address this, I optimised a combination staining protocol using immunofluorescence with various antibody combinations to identify senescent microglia in postmortem human and mouse brain tissue.

I examined post-mortem human AD brain tissue where demyelination occurs, and found increased numbers of senescent microglia compared to controls only in regions of white matter, which is enriched in myelin. Interestingly, this finding was not recapitulated in the APP/PS1 and AppNL-G-F amyloid mouse models, where extremely low numbers of senescent microglia were observed.

To determine whether demyelination may contribute to the microglial senescence observed in the human AD tissue, I assessed microglial senescence in the lysophosphatidylcholine (LPC) mouse model of demyelination where mice are injected with the myelin toxin, LPC, resulting in a focal demyelination lesion in the corpus callosum that remyelinates over the subsequent weeks. In young mice, this myelin damage was sufficient to induce microglial senescence. However, aged LPC model mice, which fail to remyelinate, exhibited relatively increased senescent microglia.

These results indicate that microglial senescence occurs in response to demyelination and during efficient remyelination, yet is increased when remyelination fails, pointing to microglial senescence as a potential regulator of CNS regeneration.

I next sought to investigate the specific role of microglial senescence in CNS injury and regeneration. To this end, I performed two functional experiments in vivo; the first involved the generation of a transgenic mouse model with the aim of inducing senescence in microglia specifically using a Tmem119 tamoxifen-inducible Cre line crossed to a model with floxed Mdm2, a negative regulator of senescence. Despite numerous rounds of troubleshooting, microglial senescence was not induced in this model. Therefore, I performed an alternative functional experiment whereby I used a senolytic cocktail to kill senescent cells in the LPC model and examined how this impacted remyelination efficiency. Preliminary results indicate that the senescent cells may not be important for efficient remyelination to take place.

Further work is required to refine a model of microglial senescence specifically to improve our understanding of the functional implications and mechanisms of microglial senescence in CNS injury and regeneration. However, my findings suggest a potential role for microglial senescence in CNS injury and regeneration, a greater understanding of which could allow for the development of therapeutic strategies for CNS injury and disease.