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Updated: May 29, 2026
Isolation of Cortical Microglia with Preserved Immunophenotype and Functionality From Murine Neonates
Published on: January 30, 2014
Microglial cells contribute to endogenous brain defenses after acute neonatal focal stroke
Joel V Faustino1, Xia Wang, Cali E Johnson
1Department of Neurology, University of California, San Francisco, San Francisco, California 94143-0663, USA.
Abstract:
Macrophages are viewed as amplifiers of ischemic brain injury, but the origin of injury-producing macrophages is poorly defined. The role of resident brain macrophages-microglial cells-in stroke remains controversial. To determine whether microglial cells exert injurious effects after neonatal focal stroke, we selectively depleted these cells with intracerebral injection of liposome-encapsulated clodronate before transient middle cerebral artery occlusion in postnatal day 7 rats. Phagocytosis of apoptotic neurons by activated microglia was poor in animals with unmanipulated microglia, and depletion of these cells did not increase the number of apoptotic neurons. Lack of microglia increased the brain levels of several cytokines and chemokines already elevated by ischemia-reperfusion, and also increased the severity and volume of injury, suggesting that microglial cells contribute to endogenous protection during the subacute injury phase. Then, to determine whether accumulation of reactive oxygen species in microglia adversely affects phagocytosis of dying neurons and contributes to injury, we delivered reduced glutathione (GSH) into microglia, again using liposomes. Remarkably, pharmacologically increased intracellular GSH concentrations in microglia induced superoxide accumulation in lipid rafts in these cells, further increased the brain levels of macrophage chemoattractants, and exacerbated injury. Together, these data show that microglia are part of the endogenous defense mechanisms and that, while antioxidants can protect the injured neonatal brain, high levels of reducing equivalents in activated microglia, GSH, trigger superoxide production, favor the reorganization of lipids, amplify local inflammation and exacerbate injury.
Insights
Microglia protect the neonatal brain after stroke, contrary to previous beliefs. However, high levels of glutathione in microglia can paradoxically increase injury by promoting inflammation and oxidative stress.
Area of Science:
- Neuroscience
- Immunology
- Stroke Research
Background:
- Macrophages are implicated in amplifying ischemic brain injury, but their origin and role, particularly resident microglial cells, remain debated.
- The specific contribution of microglial cells to injury or protection following neonatal stroke requires clarification.
Purpose of the Study:
- To investigate the role of microglial cells in neonatal focal stroke injury.
- To determine if microglial cells exert injurious effects or provide endogenous protection.
- To examine the impact of altered intracellular glutathione levels in microglia on stroke outcomes.
Main Methods:
- Selective depletion of microglial cells using intracerebral injection of liposome-encapsulated clodronate before inducing transient middle cerebral artery occlusion in P7 rats.
- Assessment of phagocytosis of apoptotic neurons by microglia.
- Measurement of cytokine and chemokine levels.
- Delivery of reduced glutathione (GSH) into microglia via liposomes to investigate the effects of elevated intracellular GSH.
Main Results:
- Microglial depletion did not increase the number of apoptotic neurons, indicating poor phagocytosis in unmanipulated microglia.
- Lack of microglia exacerbated stroke severity and injury volume, suggesting a protective role during the subacute phase.
- Increased intracellular GSH in microglia led to superoxide accumulation, enhanced chemoattractant levels, and worsened brain injury.
Conclusions:
- Microglial cells function as endogenous defense mechanisms in the subacute phase of neonatal stroke.
- While antioxidants may protect the injured brain, excessive levels of reduced glutathione (GSH) in activated microglia can paradoxically amplify inflammation and exacerbate injury.
- Targeting microglial function requires careful consideration of intracellular redox balance to avoid adverse effects.
