(683f) Effects of Igf-1 Modified Ehd Networks on Myoblastic Cell Proliferation

Ten percent of all patients undergoing abdominal incisions subsequently develop abdominal hernias as a surgical complication. Current repair techniques, which utilize nondegradable polymer meshes or xenografts, typically repair the defect with fibrous tissue and as a result provide little mechanical support to the abdomen. Therefore, we propose the use of a biodegradable polymer that facilitates skeletal muscle regeneration as a strategy for abdominal hernia repair. In particular, we have recently developed a class of biodegradable materials based upon 5-ethyl-5-(hydroxymethyl)- β,β-dimethyl-1,3-dioxane-2-ethanol diacrylate (EHD) and investigated their use in abdominal hernia repair. Preliminary studies investigated myoblastic cell attachment to EHD networks as well as insulin like growth factor-1 (IGF-1) induction of myoblastic cell proliferation signaling pathways. To this end, skeletal muscle was harvested and myoblastic cells were isolated from the hind legs of rats. The muscle was digested in collagenase P for 2 hours. The cell suspension was passed in succession through three cell strainers with a mesh size of 100 mm, 70 mm and 40 mm respectively. The cells were spun down and the pellet was resuspended in F-10 Ham media containing 10% Fetal Bovine Serum (FBS) and 1% Penicillin/Streptomycin (growth media). The resulting suspension was plated on a T-25 culture flask for 8 days. After this time, the myoblastic cell population was lifted off the T-25 culture flask and seeded onto the EHD disks weighted down by stainless steel inserts. The EHD networks were fabricated by combining a radical initiator, benzoyl peroxide (BP), dissolved in acetone with EHD. The accelerant N,N-Dimethyl-p,-toluidine (DMT) was added to decrease the gelation time. The solution was poured between two glass plates while gelation occurred and a sheet, approximately 0.5 mm thick, was formed. This sheet was then cut into disks with a diameter of approximately 2 cm. The disks were subsequently washed and sterilized in four stages. First they were rinsed with phosphate buffer saline solution (PBS), then acetone, and again in PBS for 15 minutes each and then placed under UV light for sterilization overnight. In order to investigate growth factor induced myoblastic proliferation, IGF-1 was physically adsorbed onto EHD networks in concentrations of 50 ng/mm² and 75 ng/mm². A myoblastic cell population was then seeded onto these loaded networks the next day and cultured in growth media. The myoblastic cells were lifted off the network at 1, 3 and 5 d; cell proliferation was measured using a picogreen assay for double-stranded deoxyribonucleic acid. Our initial results demonstrated that myoblasts attach to the surface of EHD networks in a manner similar to tissue culture polystyrene surfaces. The results of this work demonstrated that EHD networks support myoblastic cell adhesion and the physical absorption of IGF-1 can be utilized to augment the proliferation of this attached cell population.