Fate mapping via CCR2-CreER mice reveals monocyte-to-microglia transition in development and neonatal stroke

Abstract

Whether monocytes contribute to the brain microglial pool in development or after brain injury remains contentious. To address this issue, we generated CCR2-CreER mice to track monocyte derivatives in a tamoxifen-inducible manner. This method labeled Ly6Chi and Ly6Clo monocytes after tamoxifen dosing and detected a surge of perivascular macrophages before blood-brain barrier breakdown in adult stroke. When dosed by tamoxifen at embryonic day 17 (E17), this method captured fetal hematopoietic cells at E18, subdural Ki67+ ameboid cells at postnatal day 2 (P2), and perivascular microglia, leptomeningeal macrophages, and Iba1+Tmem119+P2RY12+ parenchymal microglia in selective brain regions at P24. Furthermore, this fate mapping strategy revealed an acute influx of monocytes after neonatal stroke, which gradually transformed into a ramified morphology and expressed microglial marker genes (Sall1, Tmem119, and P2RY12) for at least 62 days after injury. These results suggest an underappreciated level of monocyte-to-microglia transition in development and after neonatal stroke.

Cortical stroke produces secondary injury and long-lasting gliosis in the ipsilateral thalamus

“…Subsequently, the microglial cell population can return through proliferation of remaining MG or migration of MG from nearby regions [49][50][51] . In addition, CCR2 + peripheral monocytes can infiltrate the injury region and then differentiate into macrophages or microglia-like cells 34, 52,53 . The differentiation of the macrophages is affected by the local environment as well as the type and time course of injury 54 .…”

Immovable Object Meets Unstoppable Force? Dialogue Between Resident and Peripheral Myeloid Cells in the Inflamed Brain

“…Microglia are now known to arise from uncommitted KIT + erythromyeloid precursors (EMP) ( 6 ) ( Figure 1 ), which seed the brain from the YS at embryonic day 9.5 (E9.5) in the mouse ( 5 ), well before other glial cells and before the formation of the blood-brain barrier (BBB) ( 6 , 7 ). However, other evidence suggests that microglia are not exclusively YS-derived, and that a small population arise from Hoxb8 + progenitors in the E12.5 fetal liver ( 8 ) or from fetal HSC-derived monocytes ( 9 ). Subsequent to the formation of the brain, microglia are renewed in-situ throughout life, independently of BM-derived HSCs ( 10 – 12 ).…”

“…P2RY12 was upregulated in models of pseudorabies virus encephalitis ( 168 ) and neuropathic pain ( 169 ), whilst P2RY12 ( 170 ) and TMEM119 ( 163 , 171 ) were stably expressed during stroke. However, both markers have been shown to be expressed by peripherally-derived myeloid cells in the CNS ( 9 , 163 ), with TMEM119 also expressed by other non-CNS cell types ( 172 ). TMEM119, originally shown to be expressed in mouse osteoblasts, is additionally expressed in human bone tissue, DCs, osteosarcoma, and lymphoid tissue ( 173 , 174 ).…”

“…Thus, enabling the identification of cells following the downregulation of relevant genetic markers. Fate-mapping approaches have been used in a number of neuroinflammatory models to distinguish resident from infiltrating myeloid cells ( 9 , 163 ). For example, using Cxcr4 CreER/Wt ; R26 CAG-LSL-tdT mice in a stroke model, HSC-derived myeloid cells were traceable by tdTomato (tdT) fluorescence ( 163 ).…”

“…Moreover, the ubiquitously active CAG promoter in R26 CAG-LSL-tdT enabled MDMs to be traced, despite their downregulation of CXCR4 in the CNS during stroke. A similar approach was used in neonatal stroke and development using bi-transgenic CCR2-CreER tg/+ ; R26R-EGFP tg/+ mice, where Ly6C hi and Ly6C lo cells could be mapped despite downregulation of CCR2 ( 9 ). Although fate-mapping is a powerful approach that can be used to study microglial functions in-vivo , these models can be time-consuming and costly to generate, as well as requiring cell-specific markers to target particular cell types.…”