In Vitro Ligament–Bone Interface Regeneration Using a Trilineage Coculture System on a Hybrid Silk Scaffold

Abstract

The ligament-bone interface is a complex structure that comprises ligament, fibrocartilage, and bone. We hypothesize that mesenchymal stem cells cocultured in between ligament and bone cells, on a hybrid silk scaffold with sections suitable for each cell type, would differentiate into fibrocartilage. The section of scaffold for osteoblast seeding was coated with hydroxyapatite. A trilineage coculture system (osteoblasts-BMSCs-fibroblasts) on a hybrid silk scaffold was established. RT-PCR results and immunohistochemistry results demonstrated that BMSCs cocultured between fibroblasts and osteoblasts had differentiated into the fibrocartilaginous lineage. The morphological change was also observed by SEM observation. A gradual transition from the uncalcified to the calcified region was formed in the cocultured BMSCs from the region that directly interacted with fibroblasts to the region that directly interacted with osteoblasts. The role of transforming growth factor β3 (TGF-β3) in this trilineage coculture model was also investigated by supplementing the coculture system with 10 ng/mL TGF-β3. The TGF-treated group showed similar results of fibrocartilaginous differentiation of BMSCs with coculture group without TGF-β3 supplement. However, no calcium deposition was found in the cocultured BMSCs in the TGF-treated group. This may indicate TGF-β3 delayed the mineralization process of chondrocytes.

Strategies to engineer tendon/ligament-to-bone interface: Biomaterials, cells and growth factors

“…Given the importance of heterotypic cell communication for enthesis regeneration, it has been hypothesized that cellular cross-talk may be able to promote MSC differentiation via intercellular signals and paracrine factors [46]. To analyze the effect of heterotypic cell interaction on BMSCs differentiation, He et al Accordingly, calcium deposition was found only when BMSCs were co-cultured with osteoblasts, but not with fibroblasts [111]. These findings suggest that heterotypic cellular communication plays a key role in influencing MSC fate and demonstrates the feasibility of co-culture strategies for the generation of a fibrocartilage-like tissue [4].…”

Nanoengineered biomaterials for repair and regeneration of orthopedic tissue interfaces

“…[117] Superior mechanical properties and biocompatible characteristics have made silk a promising material for interface regeneration strategies. [118,119] The presence of osteoblasts and fibroblasts in the adjacent regions, allowed BMSCs to differentiate into both the lineages, thereby giving rise to a transitional region.…”

“…This hybrid silk scaffold supported a tri-lineage environment and the researchers aimed to construct grafts for ACL treatments. [117] Another nontraditional approach to bone-ligament regeneration involved fabrication of two-and three-dimensional electrospun scaffolds. [120] PCL and PLGA were respectively chosen for the aligned and random portions of the scaffold and the transition region contained both materials (Figure 8a,b).…”

The combined use of kartogenin and platelet-rich plasma promotes fibrocartilage formation in the wounded rat Achilles tendon entheses

“…As a result, the ability to withstand mechanical loads is reduced, thereby leading to re-injuries. 2,8 Therefore, regenerating the enthesis is a common clinical problem, and current approaches for engineering tissue using cells (mesenchymal stem cells (MSCs)) 9,10 and scaffolds (hybrid silk, 9 nanofibre, 11 ) have achieved only limited success in this area. 12 …”