Recombinant Extracellular Matrix Protein Fragments Support Human Embryonic Stem Cell Chondrogenesis

Abstract

We previously developed a 14-day culture protocol under potentially GMP, chemically defined conditions, to generate chondroprogenitors from human embryonic stem cells (hESCs). In vivo work has confirmed the cartilage repair capacity of these cells in a nude rat osteochondral defect model. Aiming to enhance hESC-chondrogenesis, we screened a range of extracellular matrix (ECM) molecules for their ability to support differentiation of hESCs toward chondrocytes. We identified two novel ECM protein fragments that supported hESC-chondrogenesis: Fibronectin III (fibronectin 7-14 protein fragments, including the RGD domain, syndecan-binding domain, and heparin-binding domain) and fibrillin-1 (FBN1) fragment PF8 (encoded by exons 30–38, residues 1238–1605, which contains the RGD motif but not heparin-binding site). These two protein fragments support hESC-chondrogenesis compared with the substrates routinely used previously (a mixture of fibronectin and gelatin) in our directed chondrogenic protocol. We have identified recombinant fibronectin fragment (FN III) and FBNI fragment (PF8) as alternative coating substrates to promote expression of genes known to regulate chondrocytes and code for chondrocyte ECM components. These recombinant protein fragments are likely to have better batch to batch stability than full-length molecules, especially where extracted from tissue/serum.

3D Culture of Bone Marrow-Derived Mesenchymal Stem Cells (BMSCs) Could Improve Bone Regeneration in 3D-Printed Porous Ti6Al4V Scaffolds

“…This is because of the ECM properties of Matrigel, which can promote the osteogenic differentiation of BMSCs directly. The ECM has already been reported to provide structural support for cells and physical and biochemical cues to regulate cell phenotype [ 23 , 24 ]. BMSCs cultured in Matrigel can interact with multiple ECM components within the Matrigel, such as laminin and TGF- β .…”

Gelatin Endorses the Pluripotent Stem Cells from the Reprogrammed Fibroblasts of Adult Mice Differentiation into Subtypes of Neurons in the Feeder-Free Condition for a Long-Term

“…The recombinant human (rh) laminin-511 [35], rh vitronectin [36], and rh E-cadherin [37] were considered as the noteworthy landmarks when at the first time they were applied in the culture of hPSCs as the xenogeneic-free substrates. In addition, the recombinant protein fragments recombinant fibronectin fragment (FN III) and FBNI fragment (PF8) supported the human embryonic stem cell chondrogenesis [38]. Furthermore, the mixtures of fibronectin, Figure 2: Gelatin endorses the iPSCs to differentiate into consequent several phased neural cells.…”

A Preliminary Evaluation of the Pro-Chondrogenic Potential of 3D-Bioprinted Poly(ester Urea) Scaffolds

“…Previous studies have demonstrated that hydrophilic surfaces enhance chondrogenic phenotypes by limiting protein adsorption and cell spreading, promoting a more spherical morphology [ 51 , 55 , 56 ]. Such a spherical phenotype is correlated with synthesis of a chondrogenic matrix [ 57 , 58 , 59 ]. Contact angle measurements revealed PEU to be significantly more hydrophilic than PCL and confocal microscopy images suggest that seeded cells exhibit reduced spreading on PEU scaffolds.…”