Ligament Tissue Engineering Using a Novel Porous Polycaprolactone Fumarate Scaffold and Adipose Tissue-Derived Mesenchymal Stem Cells Grown in Platelet Lysate

Abstract

Purpose: Surgical reconstruction of intra-articular ligament injuries is hampered by the poor regenerative potential of the tissue. We hypothesized that a novel composite polymer ''neoligament'' seeded with progenitor cells and growth factors would be effective in regenerating native ligamentous tissue. Methods: We synthesized a fumarate-derivative of polycaprolactone fumarate (PCLF) to create macro-porous scaffolds to allow cell-cell communication and nutrient flow. Clinical grade human adipose tissue-derived human mesenchymal stem cells (AMSCs) were cultured in 5% human platelet lysate (PL) and seeded on scaffolds using a dynamic bioreactor. Cell growth, viability, and differentiation were examined using metabolic assays and immunostaining for ligament-related markers (e.g., glycosaminoglycans [GAGs], alkaline phosphatase [ALP], collagens, and tenascin-C). Results: AMSCs seeded on three-dimensional (3D) PCLF scaffolds remain viable for at least 2 weeks with proliferating cells filling the pores. AMSC proliferation rates increased in PL compared to fetal bovine serum (FBS) ( p < 0.05). Cells had a low baseline expression of ALP and GAG, but increased expression of total collagen when induced by the ligament and tenogenic growth factor fibroblast growth factor 2 (FGF-2), especially when cultured in the presence of PL ( p < 0.01) instead of FBS ( p < 0.05). FGF-2 and PL also significantly increased immunostaining of tenascin-C and collagen at 2 and 4 weeks compared with human fibroblasts. Summary: Our results demonstrate that AMSCs proliferate and eventually produce a collagen-rich extracellular matrix on porous PCLF scaffolds. This novel scaffold has potential in stem cell engineering and ligament regeneration.

Osteogenic Stimulation of Human Adipose-Derived Mesenchymal Stem Cells Using a Fungal Metabolite That Suppresses the Polycomb Group Protein EZH2

“…The stromal fraction of adipose tissue contains between 1% and 10% MSCs which possess the ability for multi‐lineage differentiation and immunomodulation. Hence, these cells are useful for cell transplantations to treat skeletal degenerative disorders such as osteoarthritis and for interactions with biopolymer scaffolds to support skeletal tissue engineering applications . Our group has previously validated in vitro differentiation as well as cell surface marker expression and the transcriptome of clinical grade adipose‐derived MSCs .…”

Biodegradable polymer nanocomposites for ligament/tendon tissue engineering

“…Wagner et al [86] produced 3D porous polycaprolactone fumarate (PCLF) scaffolds to mimic the anterior cruciate ligament. Porous scaffold molds were designed using SolidWorks CAD software and 3D-printed.…”

Extracellular matrix protein production in human adipose-derived mesenchymal stem cells on three-dimensional polycaprolactone (PCL) scaffolds responds to GDF5 or FGF2

“…These include: i) decellularization of autografts, and the use of either ii) natural polymer or iii) synthetic polymer scaffolds. A-cellular scaffolds can be designed to meet the mechanical demands within the joint, but lack cellular conductive and inductive properties [9]. The combination of these scaffold structures with progenitor cells and growth factors, however, represents a promising solution to native ligamentous regeneration[10].…”