UC Irvine

Exploring Microglial Homeostasis and Repopulation Dynamics Using Colony-Stimulating Factor 1 Receptor Inhibition

By

Allison Rachel Najafi

Doctor of Philosophy in Biological Sciences

University of California, Irvine, 2017

Associate Professor Kim Green, Chair

Microglia are the primary immune effector cells in the CNS, responsible for the majority of inflammatory responses in the brain. Microglial homeostatic signals and replicative properties are largely unknown - however, early studies indicate that microglial turnover is low and proliferation is rare (Lawson et al., 1992), which could result in the acquisition of age-related deficiencies and senescence. As microglia age they become dysregulated, exhibiting an altered inflammatory profile, reduced phagocytic efficiency, and impaired migratory abilities (Godbout and Johnson, 2004; Frank et al., 2006; Damani et al., 2011; Njie et al., 2012). Given the dysregulation of aged microglia, activation of these cells could incite a chronic self-perpetuating cycle of inflammation and toxicity, contributing to neuronal death. We have recently discovered that microglia are fully dependent upon colony stimulating factor-1 receptor (CSF1R) signaling for their survival in the adult brain, and through the administration of small molecule inhibitors of the CSF1R, we have previously demonstrated we can eliminate >99% of all microglia brainwide (Elmore et al. 2014). Furthermore, my data show that the microglia-depleted brain has a profound ability to repopulate with new cells once CSF1R inhibitors are withdrawn, indicating that microglial elimination is fully reversible.

The goal of my dissertation is to determine the source and properties of repopulated microglia in order to establish whether these cells can be used for potential therapeutic purposes – either to resolve chronic neuroinflammatory processes or, more ambitiously, to reverse microglial senescence in the aged brain. Indeed, I have discovered that microglial repopulation arises both from nestin+ progenitor cells and surviving microglia in the adult brain. To assess the regenerative capacity of these progenitor cells, I treated mice with multiple cycles of CSF1R inhibition followed by withdrawal of inhibitors - each cycle consists of 7 days of inhibition followed by a 7- or 28-day withdrawal period. These studies determined that there is a limited capacity for microglial repopulation. However, this capacity can be expanded by increasing recovery time between cycles.

In addition, by increasing the exposure to CSF1R inhibitors, I have found that I can eliminate 100% of microglia from the adult brain, and that this results in a drastically altered repopulation pattern. Not only is the number of returning Iba1+ cells greatly diminished following complete microglial ablation, but cells initially only appear in white matter tracts along the rostral migratory stream (RMS) and projecting axons, and at early timepoints do not express the microglia-specific marker P2ry12. Eventually, these Iba1+ cells spread throughout the brain parenchyma, although cortical microglia numbers, even with extended time, do not reach those of control and continue to exhibit an altered RNA profile relative to control. These results not only highlight the presence of distinct microglial subtypes within the CNS and suggest multiple routes of repopulation, but also reveal that we have an unprecedented level of control over CNS myeloid cells.