Efficient Isolation of Lymphocytes and Myogenic Cells from the Tissue of Muscle Regeneration

Abstract

Isolation of both lymphocytes and myogenic cells from muscle tissue is required for elucidating the cellular and molecular mechanisms of muscle regeneration. Here, we aimed to establish an optimal method obtaining a high yield of lymphocytes during muscle regeneration. After the muscle injury, we observed higher infiltration of lymphocytic cells in the muscle on day 3 after injury. Then, we compared two different white blood cell isolation methods, the Percoll gradient and CD45-magnetic bead methods, to assess the percentage and number of T and B cells. Flow cytometry analysis showed that the CD45-magnetic bead method has a better efficiency in isolating CD4+, CD8+ T cells, and B cells from injured muscle tissues of wild-type and mdx mice than that by the Percoll gradient method. Moreover, we found that the CD45-negative fraction from wild-type and mdx mice includes myogenic cells. In conclusion, we report that the CD45-magnetic bead method is suitable to isolate T and B cells during muscle regeneration with higher purity and yield and can also isolate myogenic cells within the same sample. This method provides a technical basis for further studies on muscle regeneration, involving lymphocytes and muscle cells, with a wide range of clinical applications.

Keywords

• muscle regeneration
• lymphocytes
• myogenic cells
• Percoll
• CD45 beads