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Updated: Jun 22, 2026
Isolation and Flow Cytometric Analysis of Immune Cells from the Ischemic Mouse Brain
Published on: February 12, 2016
Mast cells are early responders after hypoxia-ischemia in immature rat brain
Yuxuan Jin1, Ann J Silverman, Susan J Vannucci
1Department of Pathology and Cell Biology, Columbia University Medical Center, New York, NY, USA.
Background And Purpose:
Perinatal hypoxia-ischemia (HI) produces acute and prolonged inflammation of the brain. Mast cells (MCs), numerous in the pia and CNS of neonatal rats, can initiate inflammation attributable to preformed mediators. MCs contribute to HI brain damage in the neonatal rat; MC stabilization protects through 48 hours of reperfusion. Here we hypothesize that HI induces early MC migration, activation, and release of proinflammatory molecules.
Methods:
HI was induced by right CCA ligation and 75 minutes 8% oxygen. Histochemistry and immunocytochemistry described the time course of early cellular changes in the CNS. For neuroprotection by MC stabilization, pups were treated with Cromolyn (CR) during the initial 24 hours post-HI; brains were examined through 4 weeks.
Results:
Brain MC number and activation were elevated in ipsilateral hemisphere immediately after HI (P<0.05), before detection of cleaved caspase-3 in neurons (NeuN+; 2 hours post-HI), astroglial activation (GFAP+ with swollen cell body, 4 hours post-HI), or microglial activation (OX42+, 4 hours post-HI). TNF-alpha-positive MCs were present in a subpopulation of MCs in control animals and the percent of TNF-alpha MCs increased dramatically ipsilaterally immediately after HI (P<0.01). Microglial TNF-alpha was evident at 4 hours; endothelial cells had no detectable TNF-alpha until 48 hours post-HI. Cromolyn prevented MC migration, reduced brain damage/neuronal loss, glial activation, and brain atrophy through 4 weeks of recovery (P<0.05).
Conclusions:
MCs are early responders to HI in neonatal brain. MC stabilization provides lasting protection and suggests a new target for therapeutic interventions.
Insights
Hypoxia-ischemia (HI) in neonatal brains triggers early mast cell (MC) activation and inflammation. Stabilizing mast cells with Cromolyn offers lasting neuroprotection against brain damage.
Area of Science:
- Neuroscience
- Immunology
- Neonatal Research
Background:
- Perinatal hypoxia-ischemia (HI) causes significant acute and chronic brain inflammation in neonates.
- Mast cells (MCs), prevalent in the neonatal rat brain, possess pre-formed mediators that can initiate inflammation.
- Previous studies indicate MCs contribute to HI-induced brain damage, with MC stabilization showing protective effects up to 48 hours post-reperfusion.
Purpose of the Study:
- To investigate the hypothesis that HI induces early mast cell migration, activation, and the release of pro-inflammatory molecules in the neonatal brain.
- To evaluate the long-term neuroprotective effects of mast cell stabilization following perinatal HI.
Main Methods:
- Hypoxia-ischemia (HI) was induced in neonatal rats via carotid artery ligation and systemic hypoxia.
- Histochemistry and immunocytochemistry were employed to track early cellular changes in the central nervous system (CNS).
- Mast cell stabilization was achieved using Cromolyn (CR) treatment during the initial 24 hours post-HI, with subsequent brain analysis up to 4 weeks.
Main Results:
- Mast cell number and activation significantly increased in the ipsilateral hemisphere immediately following HI, preceding neuronal and glial activation markers.
- A notable increase in TNF-alpha-positive mast cells was observed immediately post-HI, with microglial TNF-alpha appearing at 4 hours.
- Cromolyn treatment effectively prevented mast cell migration, reduced neuronal loss, glial activation, brain damage, and atrophy over a 4-week recovery period.
Conclusions:
- Mast cells are identified as early responders to hypoxia-ischemia in the developing neonatal brain.
- Mast cell stabilization demonstrates potential for long-lasting neuroprotection, suggesting a novel therapeutic target for HI-induced brain injury.
