P13 – Increased Murine Myoblast Proliferation in vitro with Urinary Bladder Matrix: A Mechanism of Tissue Regeneration and Wound Healing

Author(s):
Melody Saeman, Juquan Song, Kevin Despain, Ming-Mei Liu, Joseph Minei, Steven Wolf, UT Southwestern

Background: Porcine-derived urinary bladder matrix (UBM) is a biological scaffold used clinically for wound healing and tissue regeneration after injury. A case report showed the application of UBM to an injured muscle site increased muscle mass and function in a trauma patient.

Hypothesis: The aim of this study was to investigate mechanisms of UBM in regulating muscle regeneration of progenitor cells.

Methods: C2C12 cells from American Tissue and Cell Culture (AATCC) biological resource center were cultured in medium containing Dulbecco’s Modified Eagle Medium (DMEM), 10% fetal bovine serum and 1% glutamine. The cultures were maintained in a humidified atmosphere of 5% CO2 at 37 degrees Celsius. Cells were treated with 0, 5, 50, or 250 ug/ml of UBM (Matricel, Acell Inc.) in culture medium for 18 hours. Sample proteins were extracted from cell lysate and analyzed with SDS-PAGE and western blot. Samples were normalized to controls and then analyzed by Mann-Whitney rank sum test.

Results: Proliferating cell nuclear antigen (PCNA) increases leading strand synthesis in DNA replication, and is a marker of cell proliferation. After normalized to β-actin, we found an increase in PCNA band intensity at higher concentrations of UBM with a median of 1.7 (1.5-3.2, IQR) at 250 ug/ml concentration compared to control, median of 0.9 (0.8-1.2, IQR) (p<0.05). We found no differences at lower concentrations of UBM.

Conclusions: We showed that urinary bladder matrix stimulates proliferation in C2C12 murine myoblast progenitor cells in vitro in a dose response manner demonstrated by an increase in PCNA after 18 hours of treatment. Urinary bladder matrix increases activation of progenitor cells, potentially useful in tissue regeneration after injury.