Poster L8 - The Effects of Composite Bioengineered Scaffolds on Myoblast Alignment and Myotube Formation

Saturday, October 26, 2024

10:00 AM – 11:00 AM EST

Location: Exhibit Hall E, F & G

Primary Investigator(s)

Perla Ayala, PhD

Associate Professor
California State University Long Beach, California, United States

Presenting Author(s)

ED

Edwin L. Danahy

Research Assistant
California State University, Long Beach
Santa Clarita, California, United States
Alp Tahincioglu (he/him/his)

Research Assistant
California State University, Long Beach
Edgewater, New Jersey, United States

Co-Author(s)

AB

Ashling Bolt

Research Assistant
California State University, Long Beach, United States
Abby Gregorio (she/her/hers)

Research Assistant
California State University, Long Beach, United States

Introduction: Severe tissue damage can result in Volumetric Muscle loss (VML) which commonly results in significant fibrosis. Skeletal muscle regeneration after significant trauma is often deficient and can lead to loss of function and disability. Current surgical approaches to treat severe muscle loss are associated with donor site morbidity and functional deficiency. There is a significant need for the development of improved bioengineered implants that promote skeletal muscle tissue regeneration. Our goal is to develop a bioengineer scaffold that provides biomimetic components for effective cell delivery and structural support to guide muscle regeneration. The current approach is to develop a mechanically robust collagen-based composite scaffold composed of porcine collagen and decellularized skeletal muscle extracellular matrix (dSMM) that can promote the growth of skeletal muscle tissue. The hypothesis is that through a biomimetic muscle environment enhanced cell proliferation, cell alignment, and myotube formation will be achieved.

Materials and

Methods: The composite tissue consists of porcine derived collagen type 1 and decellularized muscle tissue dSMM formed into sheets on micro-grooved PDMS molds. Collagen type I is extracted and purified from porcine skin using an acid solubilization method. The dSMM is derived from porcine abdominal skeletal muscle tissue using an SDS/pepsin decellularization protocol. The scaffold is cast as a gel onto PDMS molds with a micro-channeled base, and then the gel is processed into a thin sheet. For this work, both mouse C2C12 myoblasts and Human Skeletal Muscle Cell (HSkMC) myoblasts are used to evaluate the bioengineered scaffold. Cells are seeded on collagen-only and bioengineered composite scaffolds for 14 days to study the effect of scaffold composition on the formation of muscle tissue. Samples are stained using Alexa-Phalloidin to identify the F-actin, immune-assayed for α-actinin and myosin, and DAPI is to stain the nuclei. Quantitative Polymerase Chain Reaction (qPCR) is performed to analyze gene expression of ⍺-actinin and myosin.

Results, Conclusions, and Discussions: The incorporation of dSMM into the collogen micro-channeled scaffold resulted in increased cell proliferation, and cell alignment for both C2C12 myoblasts and Human skeletal muscle myoblasts after 14 days. Current results also show an increased α-actinin and Myosin expression in C2C12 cells cultured on composite scaffolds compared to the collagen-only scaffolds. These results demonstrate that the composite scaffold is effective in promoting myotube formation and that incorporating dSMM could improve muscle tissue regeneration outcomes. The use of physiologically relevant biomaterials to fabricate the scaffold and the incorporation of physical features to promote alignment are effective in supporting the proliferation of myoblasts and the formation of myotubes. Current work focuses on further characterizing these bioengineered composite scaffolds using qPCR to analyze gene expression, and on optimizing dSMM incorporation. Through this optimization, we can contribute to the advancement of scalable implants that could provide a solution to VML and related muscle repair challenges.

Acknowledgements (Optional): This research was supported by the national Institute of General Medical Sciences of the National Institutes of Health [grant number: 1 SC2GM139719-01]; and the Building Infrastructure Leading to Diversity (BUILD) Initiative [grant numbers: 5TL4GM118980-10 and 5RL5GM118978-10]. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.