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. Author manuscript; available in PMC: 2016 May 18.
Published in final edited form as: Gene Ther. 2015 Jul 24;23(1):1–9. doi: 10.1038/gt.2015.81

Figure 1.

Figure 1

Panel a. Protocol for longitudinal monitoring of brain repair in vivo using MRI in mice with BCAO or sham operation. We made weekly measurements (up to 10 weeks, w10): for cerebral atrophy (ventriculomegaly) without MR-CA at w4 or w5, expression of scAAV2-CMV-hG-CSF, baseline MRI using SPION-Ran studies. The protocol can be extended to 24 weeks in some studies for leakage of the BBB using Gd-DATA. Panel b. Abnormal DWI is obvious at high b values; DWIs before and after BCAO are shown (frame 1). High-lighted areas showing significant below-threshold rADC after BCAO-60 represent regions of interest (ROI) for calculating the volume of metabolic disturbance (VMD, summation of all ROI of five MR slices of 1 mm each) as the lesion size (mm3). The results of BCAO-60, including abnormal water diffusion in the cerebrospinal fluid (CSF) in normal tissue as well as tissue with edema are illustrated, as detected by MRI (frame 2). Panel c-1. Representative T2-weighted images (caudal view). Enlarged lateral ventricles show cerebral atrophy without visible abnormal T2 MRI from mice at weeks 4 and 5 following sham operation and BCAO (one of five MR images from each mouse). Panel c-2. Normalized lateral ventricular size (LVS). We measured ventriculomegaly (n = 16) as normalized lateral ventricular size (LVS, or by size ratio of lateral ventricle to corresponding brain volumes in percent, %). Panel c-3. Scatter plot of LSV and VDM from mice before (n = 4) and after (n = 12) BCAO-60. The scatter plot shows the correlation between LVS (%) at w4–w5 post BCAO and lesion volume (VMD) in the same mice. Panel d. Corner test results of BCAO mice (normothermia, n = 8 survivors of 32; hypothermia, n = 6; sham-operation, n = 4) when facing a 30° corner. BCAO mice without treatment showed significant asymmetric turning (~90%) beginning one week after the procedure and persisting for at least four weeks. Mice with treatment (BCAO + hypothermia) exhibited behavior not significant differently from that seen in the sham-operated mice. Panels e–h. Validations of noninvasive measurements. Hypothermia treatment reversed DWI and rADC (e) as well as VMD (f). The expression of MMP-9 antigen one day post-BCAO in postmortem samples (g, arrows point to vascular MMP-9, green) validates brain lesion reduction with hypothermia treatment, shown here as normothermia (g1) vs. hypothermia (g2). (Endothelia = red; Nuclei = Hoechst stain).