IA LepA injection via ICA limits ipsilateral CA1 cell death following ischemia and reperfusion injury in mice.

(A) Schematic description of a mouse model to test the effects of targeted LepA treatment on ischemia and reperfusion injury: (1) Bilateral ligation of the common carotid artery (CCA) and external carotid artery (ECA), (2) removal of the ligature from right CCA after 12 minutes of ischemia, (3) concurrent injection at reperfusion using either saline or LepA into the right ICA through the right proximal ECA stump. The left CCA remained ligated throughout the experiment. This model was used to generate right hemispheric IR injury, alongside with simultaneous sustained ischemia in left hemisphere. (B) Representative laser doppler flowmetry measurement of the IR model used in the current study. Note the instant drop in flux following bilateral carotid ligation (1), and the gradual increase in flux flowing reperfusion of the right CCA, until reaching baseline levels. (C) Representative H&E staining of the hippocampal CA1 region of mice treated with saline or LepA. The right hemisphere was subjected to ischemia and reperfusion, and the left hemisphere was exposed to ischemia only (presented images are from same slide for each treatment group). (D) Pyramidal cells in right hippocampal CA1 region, subjected to (i) IR/Saline (ii) IR/LepA (iii) control (i.e., non-operated). (E) Viable cell count in ipsilateral CA1 region proved to be significant (n = 14 in the saline treatment group, n = 15 in the LepA treatment group and n = 3 in the control group, Kruskal-Wallis H = 18.77, p < 0.0001), with less viable cells in IR/saline compared to IR/LepA (p = 0.0023) and to control (p = 0.0009). No differences were found between the ischemia only (i.e., contralateral hemisphere) of saline-treated versus LepA-treated mice (Mann-Whitney U = 96.50, p = 0.7172). Also, in IR/LepA there were more viable cells in the ipsilateral hemisphere than in the contralateral hemisphere (Wilcoxon test, W = -69.00, p = 0.0293), while this was the opposite in the IR/saline group (Wilcoxon test, W = 61.00, p = 0.0563). Data is represented as mean for all data points. Scale bars are 500 μm and 100 μm for low and high magnification, respectively.