HMPAO-labeled CD133⁺ cells and measurement of the distribution of radioactivity by γ-counter in dystrophic injected mice. (A) Viability of radiolabeled cells. Flow cytometry analysis with PI and annexin V confirmed the 97% viability observed with trypan blue staining. (B) When we blocked the VCAM-1 molecule with the anti-VCAM-1 antibody, we obtained a significant decrease in radioactivity 12 hours after intra-arterial injection in the muscles and in the different organs, such as brain, lungs, kidneys, spleen, and liver, compared with values obtained from untreated scid/mdx mice. A decrement in radioactivity in organs of mice treated with anti-ICAM-1 was also detected, whereas in the muscle we registered a significant improvement in counts per minute per gram. Animals injected with CD133^- cells used as control showed a decrement in the recruitment of intra-arterial injected cells after exercise (C-F). Human dystrophin expression 60 days after intra-arterial injection of human CD133⁺ stem cells is shown. Error bars indicate standard deviation. (C-D) Colocalization of the human dystrophin (red), human lamin A/C nuclei (green), and Hoechst (blue) demonstrated the formation of normal human myofibers in dystrophic muscles after transplantation of the human blood-derived CD133⁺ cells. The number of human dystrophin-positive myofibers in quadriceps of exercised mice (D, F) was higher than muscle of unexercised mice (C, E). Low magnification revealed mature human dystrophin muscle fibers (red) near αSMA-positive muscle arteries (green) after the intra-arterial transplantation of CD133⁺ stem cells into unexercised (E) and exercised (F) dystrophic scid/mdx mice. Scale bars represent 100 μm (C), 75 μm (D), and 50 μm (E-F).