Customized Platelet-Rich Plasma for Skeletal Muscle Injuries
Authors: Hicks Justin James , Li Hongshuai , Philippon Marc J. , Hurwitz Shepard R. , Huard Johnny , Hogan MaCalus Vinson 
Published: July 2016

Objectives: Skeletal muscle injuries are among the most common sports-related trauma. Current treatment strategies result in formation of fibrous tissue that hinders the healing process before complete recovery. Incomplete recovery impairs muscle function and predisposes to re-injury. Platelet-Rich-Plasma (PRP) contains a multitude of growth factors and is an autologous source of growth factors for various tissue repairs. It is well established that PRP contains beneficial growth factors for muscle repair; however, it also contains high concentrations of deleterious growth factors for optimal muscle healing, such as transforming growth factor-beta 1 (TGF-β1). TGF-β1 leads to increased fibrosis impeding muscle healing. We therefore hypothesized that neutralization of TGF-β1’s action within PRP could improve PRP’s beneficial effect on skeletal muscle repair. Methods: Sixteen week old in-bred Fisher rats were used. Three rats were used for PRP isolation. 10 ml of blood were extracted from abdominal aorta and mixed with citrate phosphate dextrose solution. PRP were isolated by twice centrifugation. 24 rats were randomly assigned to four groups. A small incision was made along the tibialis anterior (TA) muscle; 50 µl cardiotoxin (CTX) (0.15ug/ul) was injected intramuscularly to the TA. One day after CTX injection, the animals were treated with PBS (control), plain PRP (PRP group), customized PRP+Ab-1x, and PRP+Ab-5x. Animals were sacrificed, and TA muscles were dissected on week 1 and 2 for assessment of muscle regeneration, fibrosis, macrophage infiltration, and satellite cell activation. Results: We observed significantly more regenerative myofibers in the PRP and customized PRP groups compared to control (Fig 1A-C). Collagen deposition (fibrosis) was detected in all groups at week 1 and week 2 after injury; while customized PRP group showed significantly decreased collagen deposition at week 1 and week 2 when compare to control and PRP groups (Fig. 1D-F). PRP was also shown to activate satellite cells at day 7 while customized PRP extended satellite cell activation through day 14 (Fig. 1G, H). Macrophage infiltration was significantly decreased in PRP group when compared to control at week 1 after injury; however the macrophage infiltration was increased in customized PRP group when compare with PRP group, specifically, we observed an increased in the percentage of the M2 macrophage subtype in the customized PRP group at week 1 but not week 2(Fig. 1I, J). Conclusion: This study demonstrates that substantial fibrosis occurs in skeletal muscle following CTX injury. PRP treatment significantly increases muscle regeneration by promoting angiogenesis, satellite cell activation, and reducing the infiltration of inflammatory macrophages. However, augmented fibrosis was also observed. Neutralizing TGF-β1 within PRP also significantly promotes muscle regeneration while significantly reducing fibrosis. It has been suggested that M2 macrophages are involved in the suppression, resolution and reversal of fibrosis. Hence, the increase in M2 macrophages in the customized PRP group may have contributed to the observed decrease in fibrosis. Our results demonstrate that neutralization of TGF-β1 within PRP is a promising approach to improve PRP’s beneficial effect on skeletal muscle repair while decreasing deleterious fibrosis.