Fibroblast Growth Factor-2–Overexpressing Myoblasts Encapsulated in Alginate Spheres Increase Proliferation, Reduce Apoptosis, Induce Adipogenesis, and Enhance Regeneration Following Skeletal Muscle Injury in Rats

Abstract

The fibroblast growth factor 2 (FGF-2) is known as pleiotropic cytokine with myoblast proliferative properties. In the present study, we tested the hypothesis that gene transfer of human FGF-2 via transplantation of genetically modified L8-myoblast encapsulated in alginate modulates the skeletal muscle recovery after crush injury in Wistar rats. Therefore, we performed a crush injury to the soleus muscle and transplanted alginate spheres containing myoblasts genetically modified to overexpress human FGF-2 (FGF-2) or a luciferase (LUC) cDNA at the site of injury. Animals that underwent muscle injury without transplantation of alginate spheres served as control (control). At day 4 after trauma the FGF-2 group showed significant higher mean values of cell proliferation (bromodeoxyuridine immunohistochemistry) and significant lower values of cell apoptosis (terminal deoxynucleotidyl transferase nick end labeling histology) compared to animals receiving luciferase-overexpressing myoblasts. At the same time point adiponectin expression (ACRP30 immunohistochemistry) was increased in the FGF-2 group exclusively. The p75(NTR) expression (p75(NTR) immunohistochemistry) significantly improved in both the FGF-2 and LUC group compared to the control group. Functional analysis of the injured muscle did not reveal a significant increase of the muscle force in the FGF-2 group compared to the control and LUC group 14 days after injury. In vitro analysis for 14 days of the FGF-2-modified spheres demonstrated at day 7 and day 14 a significant increase of the relative cell count as well as of the relative viable cell count in the FGF-2 myoblast spheres compared to luciferase myoblast spheres. Additionally, the expression of FGF-2 (enzyme-linked immunosorbent assay analysis) and luciferase (chemiluminescence analysis) persisted in vitro for 4 and 14 days, respectively. These results demonstrate that FGF-2-overexpressing myoblasts cannot considerably improve muscle strength but are able to modulate the proliferation as well as the apoptosis of injured muscle tissue mainly by conducting adipogenesis.