Abstract TP115: Cellular and Molecular Characterization of Microglia in Secondary Thalamic Injury After Ischemic Stroke

Abstract

Introduction: Secondary thalamic injury develops progressively after cortical ischemic stroke. This secondary injury associates with somatosensory dysfunctions and hinders recovery. However, the underlying pathological mechanisms are unclear. Previously our time course histological study characterized neuronal degeneration and inflammatory responses in the secondary thalamic injury after cortical ischemic stroke. In this study, we further investigated the cellular and molecular changes of microglia during the development of secondary thalamic injury. Methods: Left middle cerebral artery was permanently occluded to generate cortical ischemic stroke on male C57BL6J mice. Brain sections were collected on post-stroke days (PD) 1, 3, 7, 14 and 28. Immunostaining was used to detect microglia/macrophage activation (Iba-1) in the ipsilesional thalamus (iTH). Flow cytometry was used to discriminate between microglia and peripheral macrophages at PD28. iTH at PD7 and PD28 were processed for NanoString neuropathology transcriptome analysis (780 genes) to investigate the molecular changes. Microglial activation related genes were validated by qPCR. Results: In iTH, very few activated microglia/macrophages were detected at PD1. Clusters of microglia/macrophages appeared at PD7, became denser at PD14 and peaked at PD28. Flow cytometry analysis confirmed that at PD28, most of the microglial/macrophage population in iTH was CD11b + CD45 low , indicating they were mainly resident microglia. Furthermore, a subtype of CD11c+ microglia population were identified in iTH. NanoString analysis revealed significant changes in iTH transcriptome, with 50 differentially expressed genes at PD7 and 191 at PD28 when compared to naive mice. Several microglial activation related genes (CX3CR1, CSF1R and TLR2) were significantly increased at PD28 iTH compared to naïve and PD7. Conclusion: We have characterized the cellular and molecular changes of microglia in secondary thalamic injury. We demonstrated that the activated microglia/macrophages in iTH were mainly resident microglia exhibiting increased microglial specific genes. Targeting microglial activation may alleviate secondary thalamic injury and promote long-term stroke recovery.