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Updated: Apr 11, 2026
Isolation and Flow Cytometric Assessment of Neuroimmune Interactions in a Mini-Stroke Murine Model
Published on: June 20, 2025
Functional differences between microglia and monocytes after ischemic stroke
Rodney M Ritzel1, Anita R Patel2, Jeremy M Grenier3
1Department of Neurology, University of Connecticut Health Center, 263 Farmington Avenue, Farmington, CT, 06030, USA. rritzel@uchc.edu.
Background:
The brain's initial innate response to stroke is primarily mediated by microglia, the resident macrophage of the CNS. However, as early as 4 h after stroke, the blood-brain barrier is compromised and monocyte infiltration occurs. The lack of discriminating markers between these two myeloid populations has led many studies to generate conclusions based on the grouping of these two populations. A growing body of evidence now supports the distinct roles played by microglia and monocytes in many disease models.
Methods:
Using a flow cytometry approach, combined with ex-vivo functional assays, we were able to distinguish microglia from monocytes using the relative expression of CD45 and assess the function of each cell type following stroke over the course of 7 days.
Results:
We found that at 72 h after a 90-min middle cerebral artery occlusion (MCAO), microglia populations decrease whereas monocytes significantly increase in the stroke brain compared to sham. After stroke, BRDU incorporation into monocytes in the bone marrow increased. After recruitment to the ischemic brain, these monocytes accounted for nearly all BRDU-positive macrophages. Inflammatory activity peaked at 72 h. Microglia produced relatively higher reactive oxygen species and TNF, whereas monocytes were the predominant IL-1β producer. Although microglia showed enhanced phagocytic activity after stroke, monocytes had significantly higher phagocytic capacity at 72 h. Interestingly, we found a positive correlation between TNF expression levels and phagocytic activity of microglia after stroke.
Conclusions:
In summary, the resident microglia population is vulnerable to the effects of severe ischemia, show compromised cell cycle progression, and adopt a largely pro-inflammatory phenotype after stroke. Infiltrating monocytes are primarily involved with early debris clearance of dying cells. These findings suggest that the early wave of infiltrating monocytes may be beneficial to stroke repair and future therapies aimed at mitigating microglia cell death may prove more effective than attempting to elicit targeted anti-inflammatory responses from damaged cells.
Insights
Stroke triggers distinct responses from brain microglia and infiltrating monocytes. Monocytes aid early debris clearance, while microglia adopt a pro-inflammatory role, suggesting therapeutic targets for stroke recovery.
Area of Science:
- Neuroscience
- Immunology
- Stroke Research
Background:
- Stroke initiates a rapid inflammatory response involving resident microglia and infiltrating monocytes.
- Distinguishing these myeloid populations is crucial as they play distinct roles in stroke pathology.
- Previous studies often grouped microglia and monocytes, potentially obscuring their individual contributions.
Purpose of the Study:
- To differentiate and functionally characterize microglia and monocytes post-stroke.
- To investigate the dynamic changes and roles of these cell types in the ischemic brain.
Main Methods:
- Utilized flow cytometry with CD45 expression to distinguish microglia from monocytes.
- Performed ex-vivo functional assays to assess cellular activities.
- Monitored cell populations and functions over 7 days following middle cerebral artery occlusion (MCAO).
Main Results:
- Microglia decreased while monocytes increased significantly in the stroke-affected brain by 72 hours post-MCAO.
- Infiltrating monocytes were the primary source of proliferating macrophages (BRDU-positive) in the ischemic area.
- Microglia produced more reactive oxygen species and TNF, whereas monocytes were the main IL-1β producers; monocytes exhibited higher phagocytic capacity.
Conclusions:
- Resident microglia are vulnerable to ischemia, exhibiting impaired cell cycle and a pro-inflammatory phenotype.
- Infiltrating monocytes are key players in early debris clearance, potentially aiding stroke repair.
- Therapeutic strategies could focus on preserving microglia or leveraging monocyte functions for stroke recovery.
