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. 2011 Dec 1;225(2):415-25.
doi: 10.1016/j.bbr.2011.07.051. Epub 2011 Aug 5.

Effect of intrastriatal mesenchymal stromal cell injection on progression of a murine model of Krabbe disease

Shawna E Wicks  1 , Heaven Londot, Bo Zhang, Jennifer Dowden, Jessica Klopf-Eiermann, Jeanne M Fisher-Perkins, Cynthia B Trygg, Brittni A Scruggs, Xiujuan Zhang, Jeffrey M Gimble, Bruce A Bunnell, Paul J Pistell
Affiliations

Effect of intrastriatal mesenchymal stromal cell injection on progression of a murine model of Krabbe disease

Shawna E Wicks et al. Behav Brain Res. .

Abstract

One of a family of devastating lysosomal storage disorders, Krabbe disease is characterized by demyelination, psychosine accumulation, and inflammation. Affected infants rarely survive longer than 2 years. Using the twitcher mouse model of the disease, this study evaluated the potential of intrastriatal injection of adipose or bone marrow-derived mesenchymal stromal cells (MSCs) as a treatment option. Neonatal pups were injected with MSCs at 3-4 days of age and subjected to a battery of behavioral tests beginning at 15 days. While MSC injection failed to increase lifespan of twitchers, improvements in rotarod performance and twitching severity were observed at 27-38 days of age using MSCs derived from bone marrow. This study tested several different tasks developed in adult mice for evaluation of disease progression in immature twitchers. Rotarod was both reliable and extremely sensitive. Automated gait analysis using the Treadscan program was also useful for early evaluation of differences prior to overt gait dysfunction. Finally, this study represents the first use of the Stone T-maze in immature mice. Validation of rotarod and automated gait analysis for detection of subtle differences in disease progression is important for early stage efforts to develop treatments for juvenile disorders.

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Figures

Fig. 1
Fig. 1
Injection of BMSC and ASC did not alter disease progression in twitchers as measured by A) survival analysis using a terminal point of 20% weight loss, and B) weight gain. Genotype differences in weight between twitcher and normal mice became significant at days 21–26 (T2). * indicates significant (p<0.001) difference between twitcher and normal mice. Details of the statistical tests employed are provided in the text.
Fig. 2
Fig. 2
BMSC and ASC injection had no effect on A) twitching frequency in twitcher mice. BMSC, but not ASC, injection significantly lessened the severity of twitching (B) between days 27 and 38 (T3 and T4). Neither injection improved performance on the wire hang apparatus (C). ‘a’ denotes significant (p<0.05) difference between BMSC and HBSS twitchers by Bonferroni posthoc analysis and by a priori t-test.
Fig. 3
Fig. 3
Rotarod is a sensitive measure for detecting differences in motor performance between twitcher and normal mice (A), even at the earliest time points. The twitcher groups are presented alone in panel B to improve resolution. BMSC, but not ASC, injection significantly improves rotarod performance of twitchers from days 27–38 (T3 and T4). * indicates significant (p<0.001) difference between twitcher and normal mice. ‘a’ denotes significant (p<0.05) difference between BMSC and HBSS twitchers by a priori t-test.
Fig. 4
Fig. 4
Automated gait analysis by Treadscan software is capable of discriminating between twitcher and normal genotypes by several measures. A) Percent Stance is significantly different between twitchers and normals at all ages tested. B) Homolateral coupling detects differences between twitchers and normals at all ages tested, but may be a somewhat variable measure to use prior to T2. C) Minimum longitudinal distance from body axis is less sensitive, discriminating between twitchers and normals only at T3 (days 30–35). D) Maximum longitudinal distance from body axis clearly detects genotype differences at all ages. * indicates significant (p<0.05) difference between twitcher and normal mice. ^ indicates significant (p<0.05) injection effect. ‘a’ denotes significant (p<0.05) difference between BMSC and HBSS twitchers by Bonferroni posthoc analysis and by a priori t-test. ‘b’ denotes significant (p<0.05) difference between BMSC and ASC twitchers by Bonferroni posthoc analysis.
Fig. 5
Fig. 5
Treadscan analysis of footprint parameters following manual footprint selection is a sensitive means to detect genotype differences. In A) toe spread is significantly different between genotypes at all ages tested, but may be less reliable at T1 (days 18–23) than internal toe spread (B), which appears more consistent at T1. * indicates significant (p<0.001) difference between twitcher and normal mice. ^ indicates significant (p<0.05) injection effect. ‘a’ denotes significant (p<0.05) difference between BMSC and HBSS twitchers by a priori t-test. ‘b’ denotes significant (p<0.05) difference between BMSC and ASC twitchers by Bonferroni posthoc analysis. ‘c’ denotes significant (p<0.05) difference between ASC and HBSS twitchers by Bonferroni posthoc analysis.
Fig. 6
Fig. 6
The mouse Stone T-maze can be used to measure learning ability in young mice; however, neither BMSC nor ASC injection significantly affects learning (A). Twitcher mice have no learning impairments at either days 23–26 (A), or at days 27–29 (B).
Fig. 7
Fig. 7
Detection of injected ASCs and BMSCs by GFP fluorescence. GFP positive cells are apparent immediately after injection into the striatum at 4X magnification (A). Cells can also be detected at more posterior brain locations at this time at 4X (B). At 8 days post-surgery, some BMSC cells could be detected by GFP signal at 20X (representative images in C & D). GFP positive cells were not detected at 20 days post-surgery.
See this image and copyright information in PMC

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