Enamel matrix derivative enhances tissue formation around scaffolds used for tissue engineering of ligaments

Abstract

The following in vitro translational study investigated whether enamel matrix derivative (EMD), an approved biomimetic treatment for periodontal disease (Emdogain) and hard-to-heal wounds (Xelma), enhanced synovial cell colonization and protein synthesis around a scaffold used clinically for in situ tissue engineering of the torn anterior cruciate ligament (ACL). Synovial cells were enzymatically extracted from bovine synovium and dynamically seeded onto polyethylene terephthalate (PET) scaffolds. The cells were cultured in low-serum medium (0.5% FBS) for 4 weeks with either a single administration of EMD at the start of the 4 week period or multiple administrations of EMD at regular intervals throughout the 4 weeks. Samples were harvested and evaluated using the Hoechst DNA assay, BCA protein assay, cresolphthalein complexone calcium assay, SDS-PAGE, ELISA and electron microscopy. A significant increase in cell number (DNA) (p < 0.01), protein content (p < 0.01) and TGFbeta1 synthesis (p < 0.01) was observed with multiple administrations of EMD. Additionally, SDS-PAGE showed an increase in high molecular weight proteins, characteristic of the fibril-forming collagens. Electron microscopy supported these findings, showing that scaffolds treated with multiple administrations of EMD were heavily coated with cells and extracellular matrix (ECM) that enveloped the fibres. Multiple administrations of EMD to synovial cell-seeded scaffolds enhanced the formation of tissue in vitro. Additionally, it was shown that EMD enhanced TGFbeta1 synthesis of synovial cells, suggesting a potential mode of action for EMD's capacity to stimulate tissue regeneration.

Engineering tendon and ligament tissues: present developments towards successful clinical products

“…The synovial membrane that surrounds the posterior cruciate ligament would be a likely source of cells, especially for ACL. Synovial cells not only have a multi‐lineage potential but also, in an untreated damage, these cells are likely to reach and colonize the injury site (Messenger et al ., ).…”

“…Mechanical loading of human tendon does result in a marked interstitial increase in growth factors that are known to stimulate synthesis of collagen and other ECM proteins (Kjaer et al ., ). Therefore, bioreactors are an important tool in TE strategies, providing dynamic environments and mechanical stimulation to guide tissue remodelling and improve the performance of tendon/ligament tissue substitutes (Chen et al ., ; Doroski et al ., ; Messenger et al ., ).…”

“…() showed that dynamic mechanical stress directed ESCs, seeded onto a collagen‐silk scaffold, into a tenocyte‐like morphology, demonstrated by the positive expression of tendon‐related markers such as collagen I and scleraxis, as well as mechano‐sensory organelles and molecules, namely cilia, myosin and integrins (Chen et al ., ). Several studies have reported that mechanical conditioning positively affected cell proliferation and differentiation and increases tendon/ligament ECM synthesis (Doroski et al ., ; Messenger et al ., ). Cyclic strain significantly upregulated the collagen I, collagen III and tenascin‐C genes of BMSCs after 21 days in culture, whereas genes for other pathways (osteogenic, chondrogenic and adipogenic) did not increase (Doroski et al ., ).…”

Enamel matrix derivative inhibits proteoglycan production and articular cartilage repair, delays the restoration of the subchondral bone and induces changes of the synovial membrane in a lapine osteochondral defect modelin vivo

“…Intriguingly, EMD led to changes in the synovial membrane, including increased villus thickening and changes in the architecture of the synovial villi. In vitro , EMD stimulated the proliferation (Messenger et al ., ) and TGF β 1 synthesis of primary synovial cells (Messenger et al ., ) and fibroblasts (Grayson et al ., ). Clinically, the application of EMD for regenerative periodontal surgery often results in expedited wound healing and minimal post‐operative symptoms, such as pain or swelling (Hoang et al ., ; Rincon et al ., ; Yuan et al ., ).…”

Current Approaches and Future Trends to Promote Tendon Repair

“…2,27,38 Messenger et al concluded that the cyclic tensile strain interferes in the collagen fibres alignment, which aligned along the strain direction, while in the control group the fibres were randomly oriented. 32 JuncosaMelvin et al observed an increase in the gene expression level of type I collagen when the developed stem cell-collagen sponge constructs were subjected to mechanical stimulation in a bioreactor. Moreover, the stimulated constructs also had 2.5 times the linear stiffness and 4 times the linear modulus of nonstimulated constructs.…”