A Semi-automated Programme for Tracking Myoblast Migration Following Mechanical Damage: Manipulation by Chemical Inhibitors

Abstract

Background: Potential roles for undifferentiated skeletal muscle stem cells or satellite cells in muscle hypertrophy and repair have been reported, however, the capacity, the mode and the mechanisms underpinning migration have not been investigated. We hypothesised that damaged skeletal myoblasts would elicit a mesenchymal-like migratory response, which could be precisely tracked and subsequently manipulated. Methods: We therefore established a model of mechanical damage and developed a MATLAB^TM tool to measure the migratory capacity of myoblasts in a non-subjective manner. Results: Basal migration following damage was highly directional, with total migration distances of 948µm ± 239µm being recorded (average 0-24 hour distances: 491µm ± 113µm and 24-48 hour distances: 460µm ± 218µm). Pharmacological inhibition of MEK or PI3-K using PD98059 (20µM) or LY294002 (5µm), resulted in significant reduction of overall cell migration distances of 38% (p<0.001) and 39.5% (p<0.0004), respectively. Using the semi-automated cell tracking using MATLAB^TM program we validated that not only was migration distance reduced as a consequence of reduced cell velocity, but critically also as a result of altered directionality of migration. Conclusion: These studies demonstrate that murine myoblasts in culture migrate and provide a good model for studying responsiveness to damage in vitro. They illustrate for the first time the powerful tool that MATLAB^TM provides in determining that both velocity and directional capacity influence the migratory potential of cellular movement with obvious implications for homing and for metastases.