Abstract 173: Effects of Ischemic Preconditioning on Microglial Phenotype and Transcriptome
Background: Ischemic preconditioning (IPC) is a robust neuroprotective phenomenon in which a brief period of cerebral ischemia confers transient tolerance to subsequent ischemic challenge. Microglia are critical in stroke pathophysiology however their role in IPC is unknown.
Methods: We performed middle cerebral artery occlusion (MCAO) or sham surgery on 12 - 14 week old wild-type male mice following established paradigms for IPC ± stroke. We assessed infarct volume with 2,3,5-triphenyltetrazolium staining and used ex vivo flow cytometry (exFC) to quantify immune cell populations in cortex following IPC. We performed immunofluorescent microscopy (IFM) with stereology (optical dissector), Cavalieri point counting and Ki67 staining to characterize ionized calcium-binding adapter 1 molecule (IBA1)-positive immune cell number, morphology and proliferation, respectively. Finally, we performed mouse ST gene arrays (Affymetrix) on RNA isolated from sorted cortical microglia and carried out bioinformatic (Ingenuity Systems) and promoter analyses to characterize the effects of IPC on the microglial transcriptome.
Results: IPC reduced infarct volume from 53±6 (mean±SEM) to 26±8 mm3 (p<0.01, n = ≥5 mice per group) and induced robust increases in the number of ipsilateral cortical microglia and macrophages relative to both sham and contralateral controls (n = ≥13 and ≥4 mice per group for exFC and IFM, respectively). IPC also induced marked increases in IBA1+ cell volume (processes and somata). Microarray analysis demonstrated marked changes in microglial transcriptome [2,230 gene probesets (6.5% of total) were significantly regulated]. The top three IPC-induced canonical gene expression pathways were: (i) DNA replication, recombination and repair, (ii) Cellular assembly and (iii) Cell cycle.
Conclusions: Our novel in vivo microglia-specific, IPC-targeted, genomic dataset indicates profound activation of cell proliferation-related gene expression. This finding correlates well with the IPC-induced increases we found in microglia/macrophage cell number. These unexpected findings indicate that cell cycle regulation of CNS immune cells may be important in IPC-mediated neuroprotection.
Author Disclosures: J.R. Weinstein: None. R.V. Lee: None. S. Noor: None. R. Dodge: None. J. Colman: None. A. Savos: None. S.P. Murphy: None. T. Möller: None.
- © 2014 by American Heart Association, Inc.