Towards the Use of CHO Produced Recombinant Extracellular Matrix Proteins as Bioactive Elements in a 3-D Scaffold for Tissue Engineering

Abstract

The goal of tissue engineering is to regenerate functional tissue to replace injured or diseased tissue using a combination of patient's cells and a physical structure that supports cell attachment and proliferation (scaffold). The goal of this project is to create a new bioactive scaffold which mimics the natural extracellular matrix (ECM). The new scaffold will consist of a synthetic polymer coated with a layer of recombinant ECM proteins produced by CHO cells thus conferring cell recognition signals to the polymer. In a first step, we have established a powerful system to screen a variety of polymers for their ability to allow cell attachment and to promote cell growth using a stable GFP-expressing CHO cell line. This system allows rapid comparisons of polymer chemistries and physical parameters (porosity, weaving structure, etc.) and allows choosing the best performing polymers by simple fluorescence measurement. Subsequently, we cloned genes encoding two main ECM proteins (collagen I and tropoelastin) into mammalian expression vectors. We showed that recombinant elastin and collagen are both expressed and secreted by CHO DG44 cells. In a next step, CHO cells genetically modified to overproduce recombinant ECM proteins will be seeded and grown on synthetic polymers. It is expected that the secreted recombinant ECM proteins will adhere to the scaffold. After removal of the genetically modified CHO cells, the new bioactive scaffold can be seeded with patient's cells to regenerate tissue.