Combined use of bone marrow-derived mesenchymal stromal cells (BM-MSCs) and platelet rich plasma (PRP) stimulates proliferation and differentiation of myoblasts in vitro: new therapeutic perspectives for skeletal muscle repair/regeneration

Abstract

Satellite cell-mediated skeletal muscle repair/regeneration is compromised in cases of extended damage. Bone marrow mesenchymal stromal cells (BM-MSCs) hold promise for muscle healing but some criticisms hamper their clinical application, including the need to avoid animal serum contamination for expansion and the scarce survival after transplant. In this context, platelet-rich plasma (PRP) could offer advantages. Here, we compare the effects of PRP or standard culture media on C2C12 myoblast, satellite cell and BM-MSC viability, survival, proliferation and myogenic differentiation and evaluate PRP/BM-MSC combination effects in promoting myogenic differentiation. PRP induced an increase of mitochondrial activity and Ki67 expression comparable or even greater than that elicited by standard media and promoted AKT signaling activation in myoblasts and BM-MSCs and Notch-1 pathway activation in BM-MSCs. It stimulated MyoD, myogenin, α-sarcomeric actin and MMP-2 expression in myoblasts and satellite cell activation. Notably, PRP/BM-MSC combination was more effective than PRP alone. We found that BM-MSCs influenced myoblast responses through a paracrine activation of AKT signaling, contributing to shed light on BM-MSC action mechanisms. Our results suggest that PRP represents a good serum substitute for BM-MSC manipulation in vitro and could be beneficial towards transplanted cells in vivo. Moreover, it might influence muscle resident progenitors' fate, thus favoring the endogenous repair/regeneration mechanisms. Finally, within the limitations of an in vitro experimentation, this study provides an experimental background for considering the PRP/BM-MSC combination as a potential therapeutic tool for skeletal muscle damage, combining the beneficial effects of BM-MSCs and PRP on muscle tissue, while potentiating BM-MSC functionality.

Platelet biology in regenerative medicine of skeletal muscle

“…A schematic diagram showing a skeletal muscle stem cell's (ie, satellite cell) possible response to growth factors, based on current published evidence . Located between the basal lamina and the sarcolemma of the muscle fibre, the satellite cell may come into contact with hundreds of growth factors and cytokines in response to platelet‐rich plasma ( PRP ) administration.…”

“…It has been shown, however, that PDGF‐BB upregulates the proliferation of satellite cells (ie, skeletal muscle stem cells) and may affect differentiation negatively . Conversely, PRP application on satellite cells was shown to improve differentiation, indicating that the role of PRP on muscle progenitor cell differentiation has yet to be elucidated . Additionally, the VEGF and PDGF pathways both interact through the phosphoinositide 3‐kinase/protein kinase B/mammalian target of rapamycin pathway to induce proliferation of satellite cells (Figure ) .…”

“…Some of the cell types in Table include myogenic progenitor cells, bone‐derived periosteal cells, osteosarcoma, endothelial, trabecular bone cells, human adipose‐derived mesenchymal stem cells, fibroblasts, tenocytes, myo‐endothelial cells, pericytes, C2C12 cells, adipose‐derived stem cells and muscle satellite cells. Variable levels of differentiation have been reported for different cell types, for example increased differentiation for rat bone marrow cells, human skeletal muscle myoblasts, rat muscle satellite cells, rabbit bone marrow mesenchymal stem cells and C2C12 myoblasts or maintained effect with hMDPCs, myo‐endothelial cells and pericytes compared to control conditions . Conversely, some studies reported that platelets inhibited differentiation of C2C12 myoblasts .…”

“…This discrepancy in the current literature may provide the basis for a thorough consideration of the technical aspects in platelet applications that may affect the final outcome in a study. For example, despite the majority of studies applying 10% PRP, a recent study used higher concentrations of platelets in smaller volumes (ie, 1%‐2%) to induce differentiation, assuming that the concentration of growth factors was not altered . Finally, an interesting study carried out by Miroshnychenko et al has led to a new insight in PRP and PPP effects in vitro on human skeletal muscle myoblast cells.…”

Thioredoxin mitigates H2O2‐induced inhibition of myogenic differentiation of rat bone marrow mesenchymal stem cells by enhancing AKT activation

“…Bone marrow mesenchymal stem cells (BMSCs) have multiple differentiation potentials, which are dependent on the cell environment. Previous studies have shown that BMSCs can differentiate into muscle cells in a given condition, which is positively regulated by muscular transcription factors myogenic differentiation 1 (MyoD1) and myogenin [1,2]. During the process of aging, aberrant oxidative stress can result in the peroxidation of muscular tissues, leading to their atrophy [3].…”