Correlations of relative alterations in CBF following CCAO, MCAO, reperfusion, neurological score, weight loss, age and starting weight to ischemic infarct volume.

<p>Correlations of relative alterations in CBF following CCAO, MCAO, reperfusion, neurological score, weight loss, age and starting weight to ischemic infarct volume.</p

Representative laser-Doppler flowmetry during Koizumi or Longa intraluminal filament MCAO

<p>(A) Representative laser-Doppler flowmetry during Koizumi’s method of intraluminal filament middle cerebral artery occlusion (MCAO). (B) Representative laser-Doppler flowmetry during Longa’s method of intraluminal filament MCAO. Perfusion Units (PU) are arbitrary units of cerebral blood flow. a. Baseline. b. Immediately post CCAO. c. Immediately pre-MCAO. d. Immediately post-MCAO. e. Immediately pre-MCA reperfusion. f. 5 min post-MCA reperfusion. g. CCA reperfusion (Longa method only).</p

Relative alterations in CBF following CCAO, MCAO, during occlusion and following reperfusion for Koizumi vs. Longa at 4 h after reperfusion.

<p>Relative alterations in CBF following CCAO, MCAO, during occlusion and following reperfusion for Koizumi vs. Longa at 4 h after reperfusion.</p

Relative alterations in CBF following CCAO, MCAO, during occlusion and following reperfusion for 15 min, 30 min, 45 min and 60 min occlusions measured at 4 h or 24 h after reperfusion.

<p>Relative alterations in CBF following CCAO, MCAO, during occlusion and following reperfusion for 15 min, 30 min, 45 min and 60 min occlusions measured at 4 h or 24 h after reperfusion.</p

Post-reperfusion time-course of ischemic lesion volume detected by TTC following 60 min intraluminal filament MCAO using the Koizumi method

<p>(A) Representative TTC stained brain sections indicating areas of healthy tissue (red) and ischemic injury (white) for each group. (B) Total volume of ischemic lesion in the ipsilateral hemisphere, expressed as a percentage of the total contralateral hemisphere volume, 30 min, 4 h, 12 h and 24 h after reperfusion following sham surgeries or 60 min MCAOs, with thin or thick silicone coated filaments, via the Koizumi method. (C) No alteration in neurological severity scores between animals assessed at 4 h, 12 h and 24 h after reperfusion, following a 60 min occlusion. (D) Body weight loss post-MCAO significantly increased from 24 h as compared to 4 h and 12 h after reperfusion. Each value represents the mean ± the standard error of the mean (SEM). *<i>p < 0</i>.<i>05</i>. N = 3–4 for sham surgeries and n = 5–8 for animals undergoing occlusion.</p

Representative laser-Doppler flowmetry traces of SAH and premature reperfusion

<p>(A) A typical representative laser-Doppler flowmetry during Koizumi’s method of middle cerebral artery occlusion (MCAO), during a 60 min occlusion. Insert: representative resulting ischemic lesion analysed by TTC staining. (B) A typical representative laser-Doppler flowmetry during Longa’s method of MCAO, during a 60 min occlusion. (C) Representative laser-Doppler flowmetry of subarachnoid haemorrhage (SAH) during the Koizumi method of MCAO, with resulting ischemic injury. (D) Representative laser-Doppler flowmetry of false-positive SAH. Inserts: image of the same animal, with a brain bleed resulting from vessel perforation, yet leading to no discernible ischemic injury. (E) Representative laser-Doppler flowmetry showing arbitrary units of CBF falling gradually over the course of a 60 min MCAO occlusion time, conducted via the Koizumi method. (F) Representative laser-Doppler flowmetry indicating premature reperfusion, due filament movement out of place, followed by immediate recovery of MCAO by repositioning the filament. Perfusion Units (PU) are arbitrary units of cerebral blood flow.</p

Relative alterations in CBF following CCAO, MCAO, during occlusion and following reperfusion, for thin and thick filaments at 30 min, 4 h, 12 h, and 24 h after reperfusion.

<p>Relative alterations in CBF following CCAO, MCAO, during occlusion and following reperfusion, for thin and thick filaments at 30 min, 4 h, 12 h, and 24 h after reperfusion.</p

Ischemic lesion volume detected by TTC following 15 min, 30 min, 45 min or 60 min intraluminal filament MCAO using the Koizumi method

<p>(A) Representative TTC stained brain sections indicating areas of healthy tissue (red) and ischemic injury (white) for each group. (B) Total volume of ischemic lesions in the ipsilateral hemisphere, expressed as a percentage of the total contralateral hemisphere volume, measured at 4 and 24 h after reperfusion, following 0 min, 15 min, 30 min, 45 min or 60 min of MCAO with a thick silicone coated filament, via the Koizumi method. (C) Significant increases in neurological severity scores following 45 or 60 min MCAO, compared to 15 min MCAO. (D) Body weight loss significantly increased from 4 h to 24 h after reperfusion in every group. Each value represents the mean ± the standard error of the mean (SEM). *<i>p < 0</i>.<i>05</i>. N = 3 for sham surgeries and n = 4–7 for animals undergoing occlusion.</p

Evaluation of synthetic vascular grafts in a mouse carotid grafting model

<div><p>Current animal models for the evaluation of synthetic grafts are lacking many of the molecular tools and transgenic studies available to other branches of biology. A mouse model of vascular grafting would allow for the study of molecular mechanisms of graft failure, including in the context of clinically relevant disease states. In this study, we comprehensively characterise a sutureless grafting model which facilitates the evaluation of synthetic grafts in the mouse carotid artery. Using conduits electrospun from polycaprolactone (PCL) we show the gradual development of a significant neointima within 28 days, found to be greatest at the anastomoses. Histological analysis showed temporal increases in smooth muscle cell and collagen content within the neointima, demonstrating its maturation. Endothelialisation of the PCL grafts, assessed by scanning electron microscopy (SEM) analysis and CD31 staining, was near complete within 28 days, together replicating two critical aspects of graft performance. To further demonstrate the potential of this mouse model, we used longitudinal non-invasive tracking of bone-marrow mononuclear cells from a transgenic mouse strain with a dual reporter construct encoding both luciferase and green fluorescent protein (GFP). This enabled characterisation of mononuclear cell homing and engraftment to PCL using bioluminescence imaging and histological staining over time (7, 14 and 28 days). We observed peak luminescence at 7 days post-graft implantation that persisted until sacrifice at 28 days. Collectively, we have established and characterised a high-throughput model of grafting that allows for the evaluation of key clinical drivers of graft performance.</p></div

Endothelialisation.

<p>A: Scanning electron microscopy images of the luminal side of the graft. Scale bar = 500 μm. Inset scale bar = 50 μm. B: Quantification of CD31 coverage represented as a percentage of total luminal circumference. Data expressed as mean ± SEM and the mid section of the grafts were analysed using one-way ANOVA, n = 7 animals/timepoint. C: Representative images of cross sections with CD31 staining in red, nucleus in blue. White dotted lines indicate the graft wall. Scale bar = 100 μm.</p