Repopulation of a microglia-depleted central nervous system : molecular characterization during homeostasis and disease

Abstract: Microglia are predominant tissue resident macrophages within the central nervous system (CNS), and contribute to both CNS development and homeostasis. During disease conditions microglia undergo transcriptional re-programming and their dysfunction is implicated in a multitude of disorders, such as multiple sclerosis (MS). How microglia could be therapeutically targeted is a current research focus. Recent experimental microglial depletion methods using conditional genetic targeting and pharmacological therapies have broadened our perspective of these multi-tasking microglia. Newly repopulated microglia following experimental microglial ablation hold great promise for reducing neuroinflammation and treating a variety of neurological disorders. In Study 1 our results indicated that microglia could be ablated (approximately 95%) by systemic use of tamoxifen in Cx3cr1CreER/+Rosa26DTA/+ mice. Microglial repopulation ensued through both the proliferation of surviving microglia in the CNS, and from the infiltration of Ly6Chi monocytes. Under this condition infiltrating monocytes could be shaped into microglia-like cells by the CNS microenvironment. Furthermore, isolated newly repopulated resident microglia and infiltrating microglia-like cells following experimental depletion exhibited differential functionality in vitro, such as phagocytic capacity and cytokine production. In Study 2 we used the microglial depletion and repopulation model mentioned above and demonstrated that the presence of infiltrating microglia-like cells following ablation could exacerbate experimental autoimmune encephalomyelitis (EAE) symptoms in Cx3cr1CreER/+Rosa26DTA/+ female mice. This was not evident in male mice, indicating a potential sex effect. Under this condition there was a higher expression of major histocompatibility complex class II and a greater secretion of proinflammatory cytokines during the acute period in the female mice. In Study 3 we discovered a novel subpopulation of microglia that escape the genetic modification of Cx3cr1 in Cx3cr1CreER-EYFP/+Rosa26DTA/+ mice. Following microglial depletion using tamoxifen, newly repopulated Cx3cr1highEYFP– microglia had an advantage over Cx3cr1CreER-EYFP/+ and Cx3cr1lowEYFP+ microglia. We also found that microglial repopulation was tightly regulated by the CX3CL1-CX3CR1 signaling. The numbers of repopulated CNS-resident microglia were significantly decreased, while the numbers of infiltrating microglia-like cells were increased during repopulation in mice devoid of Cx3cr1. In Study 4 we demonstrated that experimentally removing microglia using both Cx3cr1CreER/+Rosa26DTA/+ mice and PLX3397 treatment had crucial effects on circulating monocytes and splenic macrophages, a finding that had previously received little attention. We therefore proposed that clinical translation of preclinical studies using microglial depletion should take peripheral effects into consideration.