Novel insight into stem cell trafficking in dystrophic muscles

Andrea Farini, Chiara Villa, Adrian Manescu, Fabrizio Fiori, Alessandra Giuliani, Paola Razini, Clementina Sitzia, Giulia Del Fraro, Marzia Belicchi, Mirella Meregalli, Franco Rustichelli, Yvan Torrente, Andrea Farini, Chiara Villa, Adrian Manescu, Fabrizio Fiori, Alessandra Giuliani, Paola Razini, Clementina Sitzia, Giulia Del Fraro, Marzia Belicchi, Mirella Meregalli, Franco Rustichelli, Yvan Torrente

Abstract

Recently published reports have described possible cellular therapy approaches to regenerate muscle tissues using arterial route delivery. However, the kinetic of distribution of these migratory stem cells within injected animal muscular dystrophy models is unknown. Using living X-ray computed microtomography, we established that intra-arterially injected stem cells traffic to multiple muscle tissues for several hours until their migration within dystrophic muscles. Injected stem cells express multiple traffic molecules, including VLA-4, LFA-1, CD44, and the chemokine receptor CXCR4, which are likely to direct these cells into dystrophic muscles. In fact, the majority of intra-arterially injected stem cells access the muscle tissues not immediately after the injection, but after several rounds of recirculation. We set up a new, living, 3D-imaging approach, which appears to be an important way to investigate the kinetic of distribution of systemically injected stem cells within dystrophic muscle tissues, thereby providing supportive data for future clinical applications.

Keywords: CD133+ stem cells; dystrophic muscles; iron nanoparticles; micro-CT.

Figures

Figure 1
Figure 1
Micro-CT procedure. (A) Tomograms of the tight region at 0, 2, 13, and 24 hours after injection; the femur zone has been subtracted to calculate the volume fraction of the Endorem-marked cells. Some marked cells are necessarily cut out in this way, but their amount is negligible with respect to the total amount present in the region of interest. (B) Histogram and detail for the investigated mouse, injected with 500,000 cells. Abbreviation: CT, computed tomography.
Figure 2
Figure 2
Endorem+ CD133+ stem cells. Notes: CD133+ stem cells labeled with iron oxide nanoparticles (Endorem). Cells were stained with Prussian blue and taken at 40× magnification.
Figure 3
Figure 3
Micro-CT images of injected muscle. (A) Reconstruction of the muscle injected with human cells and its muscle compartments. (B) Images of micro-CT showing the distribution of the cells after each time-point. Abbreviation: CT, computed tomography.
Figure 4
Figure 4
Quantification of different volumes in the investigated ROI. Notes: Overlapping of the fat + muscle signal for the four measurements in mice injected with 500,000 cells: from the left, the first peak corresponds to air; the second one is an overlapping of the signal coming from fat and muscle. In the magnification, the third one comes from labeled cells and bone. Abbreviation: ROI, region of interest.
Figure 5
Figure 5
Endorem volume fraction in injected muscles. Note: The apparent Endorem volume fraction increased up to two hours, then in reached a plateau and remained constant until 24 hours. Abbreviation: ROI, region of interest.
Figure 6
Figure 6
Q-PCR analysis of injected muscles and organs. Q-PCR of injected scid/mdx mice euthanized 0, 30 minutes, 60 minutes, 90 minutes, and 2 hours after cellular transplantation: the graphs are related to (A) muscles of injected leg; (B) muscles of contralateral leg; (C) DIA; and (D) organs. Abbreviations: DIA, diaphragm; Q-PCR, real-time polymerase chain reaction.

References

    1. Teo AK, Vallier L. Emerging use of stem cells in regenerative medicine. Biochem J. 2010;428(1):11–23.
    1. Farini A, Razini P, Erratico S, Torrente Y, Meregalli M. Cell based therapy for Duchenne muscular dystrophy. J Cell Physiol. 2009;221(3):526–534.
    1. Jin K, Sun Y, Xie L, et al. Comparison of ischemia-directed migration of neural precursor cells after intrastriatal, intraventricular, or intravenous transplantation in the rat. Neurobiol Dis. 2005;18(2):366–374.
    1. Chen J, Sanberg PR, Li Y, et al. Intravenous administration of human umbilical cord blood reduces behavioral deficits after stroke in rats. Stroke. 2001;32(11):2682–2688.
    1. Benchaouir R, Meregalli M, Farini A, et al. Restoration of human dystrophin following transplantation of exon-skipping-engineered DMD patient stem cells into dystrophic mice. Cell Stem Cell. 2007;1(6):646–657.
    1. Farini A, Meregalli M, Belicchi M, et al. T and B lymphocyte depletion has a marked effect on the fibrosis of dystrophic skeletal muscles in the scid/mdx mouse. J Pathol. 2007;213(2):229–238.
    1. Villa C, Erratico S, Razini P, et al. In vivo tracking of stem cell by nanotechnologies: future prospects for mouse to human translation. Tissue Eng Part B Rev. 2011;17(1):1–11.
    1. Villa C, Erratico S, Razini P, et al. Stem cell tracking by nano-technologies. Int J Mol Sci. 2010;11(3):1070–1081.
    1. Boone JM, Velazquez O, Cherry SR. Small-animal X-ray dose from micro-CT. Mol Imaging. 2004;3(3):149–158.
    1. Badea CT, Drangova M, Holdsworth DW, Johnson GA. In vivo small-animal imaging using micro-CT and digital subtraction angiography. Phys Med Biol. 2008;53(19):R319–350.
    1. Schuster DM, Halkar RK. Molecular imaging in breast cancer. Radiol Clin North Am. 2004;42(5):885–908. vi–vii.
    1. Holdsworth DW, Thornton MM, Drost D, Watson PH, Fraher LJ, Hodsman AB. Rapid small-animal dual-energy X-ray absorptiometry using digital radiography. J Bone Miner Res. 2000;15(12):2451–2457.
    1. Fang S, Wang J, Jiang H, et al. Experimental study on the therapeutic effect of positron emission tomography agent [18F]-labeled 2-deoxy-2-fluoro-d-glucose in a colon cancer mouse model. Cancer Biother Radiopharm. 2010;25(6):733–740.
    1. Peyrin F, Salome M, Cloetens P, Laval-Jeantet AM, Ritman E, Ruegsegger P. Micro-CT examinations of trabecular bone samples at different resolutions: 14, 7 and 2 micron level. Technol Health Care. 1998;6(5–6):391–401.
    1. Salome M, Peyrin F, Cloetens P, et al. A synchrotron radiation microtomography system for the analysis of trabecular bone samples. Med Phys. 1999;26(10):2194–2204.
    1. Torrente Y, Gavina M, Belicchi M, et al. High-resolution X-ray microtomography for three-dimensional visualization of human stem cell muscle homing. FEBS Lett. 2006;580(24):5759–5764.
    1. Boyd SK, Davison P, Muller R, Gasser JA. Monitoring individual morphological changes over time in ovariectomized rats by in vivo micro-computed tomography. Bone. 2006;39(4):854–862.
    1. Louie AY, Huber MM, Ahrens ET, et al. In vivo visualization of gene expression using magnetic resonance imaging. Nat Biotechnol. 2000;18(3):321–325.
    1. Mayerhofer R, Araki K, Szalay AA. Monitoring of spatial expression of firefly luciferase in transformed zebrafish. J Biolumin Chemilumin. 1995;10(5):271–275.
    1. Brown EB, Campbell RB, Tsuzuki Y, et al. In vivo measurement of gene expression, angiogenesis and physiological function in tumors using multiphoton laser scanning microscopy. Nat Med. 2001;7(7):864–868.
    1. Padera TP, Stoll BR, So PT, Jain RK. Conventional and high-speed intravital multiphoton laser scanning microscopy of microvasculature, lymphatics, and leukocyte-endothelial interactions. Mol Imaging. 2002;1(1):9–15.
    1. Kim D, Hong KS, Song J. The present status of cell tracking methods in animal models using magnetic resonance imaging technology. Mol Cells. 2007;23(2):132–137.
    1. Ritman EL. Small-animal CT – its difference from, and impact on, clinical CT. Nucl Instrum Methods Phys Res A. 2007;580(2):968–970.

Source: PubMed

Sponsorer och medarbetare

Medicinska tillstånd

Läkemedelsinterventioner

3
Prenumerera