Representative endpoint MRI, fluorescence microscopy, and immunohistochemistry acquired at 10x magnification from both implant models.

<p>(A) By day 16, 5/7 immune competent mice had no ¹⁹F signal remaining as shown by the representative MRI. The reference tube is marked by “R”. (B) Fluorescence microscopy of the muscle tissue revealed little red fluorescence remaining in the immune competent mice. No GFP+ mMSC were detectable by fluorescence microscopy, suggesting the original mMSC are no longer present. (C) H&E staining reveals the presence of cells at the implant site which correlate well with the remaining ¹⁹F red fluorescence. (D) Immunohistochemistry staining of adjacent tissue sections with the anti-F4/80 antibody reveals the presence of a few macrophages at this location in the immune competent model. (E) At endpoint, all immune compromised mice had detectable ¹⁹F-MRI signal remaining. (F) We observed more red fluorescence from the ¹⁹F-label at the transplant site in the immune compromised mice. (G) Once again, H&E staining in the immune compromised model correlates well with the regions of red fluorescence. (H) Macrophage staining of the immune compromised model reveals many more F4/80 positive cells at the site of implantation. Furthermore, the fluorescence microscopy of neighboring tissue sections reveals that the red fluorescence from the ¹⁹F agent is in the same location as the macrophages. Once again, scale bars represent 250μm.</p

Representative Day 0 MRI, fluorescence microscopy, and histology acquired as 10x magnification from both implant models.

<p>(A, E) Representative MRI from mice receiving either 2x10⁶ mMSC or 1.5x10⁶ hMSC respectively. The day 0 <i>in vivo</i>¹⁹F-MRI quantification correlates very well with the number of implanted cells. The reference tube is marked by “R”. (B) The red fluorescent fluorine agent is clearly visible in the tissue of the immune competent model, (F) as well as in the immune-compromised model. (C) Furthermore, the GFP+ mMSC are observable within the tissue section. (D) Overlaying the two fluorescent images, reveals the ¹⁹F agent colocalized with the GFP+ mMSC, as expected. (G, H) H&E stained tissue sections corresponding to the fluorescence microscopy clearly show the implant site of the mMSC and hMSC respectively. Scale bars in all images represent 250μm.</p

<i>In vitro</i> validation of ¹⁹F-MRI quantification accuracy.

<p>Quantification was validated in a phantom study using cell pellets ranging from 2x10⁵ to 2x10⁶ MSC. Pellets were imaged three times, with the error bars representing the standard deviation between scans. The ¹⁹F-MRI quantification is in very strong agreement with the true number of cells, and has a Pearson correlation coefficient of 0.99. The red line represents the ideal result of a 1:1 correlation.</p

Comparison of ¹⁹F-labeled cell detection in two transplantation models over time.

<p>(A) Following implantation of 2x10⁶ mMSC, ¹⁹F-MRI was used to quantify the number of cells remaining over 16 days. By day 16, only 2/7 mice had any detectable signal remaining. A significant difference from day 0 is denoted by <b>+</b>, from day 3 by ◆, and from day 9 by ■. (B) The number of detectable cells over a similar time period following a transplant of 1.5x10⁶ hMSC. ¹⁹F signal was found to decrease at a slower rate, with observable signal in all mice at endpoint. Statistical significance is denoted in the same way as A.</p

Cellular viability and loading with the ¹⁹F-agent.

<p>(A) Cellular viability was investigated before and after labeling with the ¹⁹F-agent, Cell Sense. Although a statistically significant difference was observed in hMSC after labeling, the viability remained high (>80%) in all experiments. There was no significant difference in mMSC viability. (B) Cellular loading was determined by performing NMR on a known number of cells alongside a reference peak with a known number of ¹⁹F atoms. We observed variation in cellular loading of both hMSC and mMSC between experiments. However, this variation does not affect in vivo ¹⁹F quantification since each transplant was only compared to its specific cellular loading.</p

Region of interest (ROI) values for ODI, NDI and IsoVF in both the scan and rescan conditions for several representative brain regions.

Each box represents the range from 25th to 75th percentile (Interquartile Range) with the median depicted by the line within the box. The whiskers shown extend to the maximum and minimum value of each measurement.</p

Representative in-plane cross sections from a single subject showing unprocessed raw diffusion image data (4 shot, centric ordered, 2 averages, 25 coronal slices with slice thickness = 500 μm, 250 × 250 μm in plane resolution, FOV 40 x 40 mm, matrix size = 160 x 160, TE = 25 ms, TR = 5.0 s), and corresponding scalar image maps of the following NODDI values: Orientation Dispersion Index (ODI), Neurite Density Index (NDI), and Isotropic Volume Fraction (IsoVF).

Representative in-plane cross sections from a single subject showing unprocessed raw diffusion image data (4 shot, centric ordered, 2 averages, 25 coronal slices with slice thickness = 500 μm, 250 × 250 μm in plane resolution, FOV 40 x 40 mm, matrix size = 160 x 160, TE = 25 ms, TR = 5.0 s), and corresponding scalar image maps of the following NODDI values: Orientation Dispersion Index (ODI), Neurite Density Index (NDI), and Isotropic Volume Fraction (IsoVF).</p

Whole brain average between subject CV maps and histogram.

Values for the between subject condition represent the mean CV within each voxel for the scan and rescan conditions averaged over the two scans. The resulting histogram has been extracted from the averaged scans. Heat maps from a representative slice show the regional variation for each metric.</p

Whole brain voxel-wise histograms representing the i) number of subjects necessary to detect a statistically significant effect with a change in the given metric of 5%, 10%, 15% and 20% and ii) the minimum detectable effect with each metric under a scan-rescan study design given group sample sizes of 6, 8 and 10.

Note the varied scales for the IsoVF metric in each category as opposed to the ODI and NDI metrics.</p

Voxel-wise between and within subject CV histograms within each representative ROI.

Voxel-wise values for the between subject condition represents the mean CV within each ROI for both the scan and rescan conditions averaged over the two scans. Voxel-wise values for the within subject condition represent the mean CV within each ROI averaged over the eight subjects.</p