Overview

Our project focuses on the development of a dual cell factor release system for bone repair using a sequential release mechanism of Vascular Endothelial Growth Factor (VEGF) and Bone Morphogenetic Protein-4 (BMP-4). The design centers on overcoming the limitations of existing bone repair materials by employing a biocompatible collagen hydrogel that mimics the natural processes of angiogenesis and osteogenesis.

Fig 1 Schematic diagram of this project

We conducted three iterative Design-Build-Test-Learn (DBTL) cycles:

1 DBTL Cycle 1

Identification and screening of suitable collagen-binding domains (CBDs) for the sequential release of VEGF and BMP-4. CBD MMPs (for VEGF) and FTD (for BMP-4) were selected based on their binding affinities, ensuring controlled, timely release of each growth factor.

Fig 2 plasmid design

2 DBTL Cycle 2

Development of the dual cell factor release system where CBD MMPs-VEGF facilitates rapid release for angiogenesis, followed by FTD-BMP-4, which supports sustained bone formation. This cycle involved protein expression optimization to ensure the production of bioactive fusion proteins.

Fig 3 Fusion Cell factor expression plasmid

3 DBTL Cycle 3

Creation of a collagen hydrogel matrix capable of delivering the two cell factors in a controlled manner. Type I collagen was expressed in E. coli, and its biocompatibility and structural properties were optimized to enhance cell adhesion and support tissue regeneration.

Through this design, we achieved a functional system that mirrors the natural healing process by promoting early vascularization followed by bone formation, with applications in regenerative medicine.