Topographical and morphological patterns of post reperfusion molecular penumbra

Question: Provide a topographical and morphological characterization of post-reperfusion molecular penumbra in the rat stroke model of transient middle cerebral artery (MCA) occlusion Methods: Young adult male Wistar rats (n=25) underwent 90 minutes of transient intraluminal MCA occlusion, followed by 24 hours of reperfusion. Immunohistochemistry for Heat Shock Protein 70 (HSP70) and Microtubule-Associated Protein 2 (MAP2), TUNEL staining and Cresyl violet staining were performed in the same or adjacent brain coronal sections within the entire vascular territory supplied by MCA. Results: Peri-lesional regions showed multiple areas of HSP70 immunostaining of variable extent and localization in the neocortex, mostly in superficial cortical layers. HSP70-positive cells were qualified as neurons (both morphologically and by MAP2 staining) and apparently maintained a tissue organization. We interpreted such regions as ‘mini-penumbras’, likely saved by reperfusion, hypothesis supported by lack of TUNEL staining. ‘Mini-penumbras’ surrounded spots or boundaries of low density marked cells interpreted as ‘mini-cores’, penumbra which has progressed to infarction in spite of reperfusion. Within the ischemic lesion, infarcted cortical and subcortical areas presented a large number of TUNEL-positive cells and a low density of regularly distributed HSP70 marked cells, mainly identified as endothelial cells and rarely as neurons. Conclusions: Qualitative assessment of residual molecular penumbra after 24 hours of reperfusion demonstrated a high variability in staining topography, pattern and extension, along with the emerging concept of heterogeneous penumbra. The preferential localization of “mini-penumbras” in the superficial cortical areas might reflect the variable contribution of leptomeningeal collateral circulation during the ischemic phase.