Proliferation of Chondrocytes on a 3-D Modelled Macroporous Poly(Hydroxyethyl Methacrylate)–Gelatin Cryogel

Abstract

Tissue-engineering constructs should be designed to mimic the native tissue environment for cells, the scaffold matching to stiffness and strength of the tissues while maintaining an interconnected porous network and a reasonable porosity. This study presents a new single-step protocol for synthesis of a poly(hydroxyethyl methacrylate)–poly(ethylene glycol) diacrylate–gelatin (HPG) macroporous polymeric scaffold with well-controlled porous structure and good mechanical strength. The pore size of these matrices lies in the range of 30 to 100 μm with an average pore diameter of 80 μm and with an interconnected pore structure as analyzed by scanning electron microscopy. Further, interconnectivity was also confirmed by high solvent uptake capacity, as the cryogel reached its equilibrium within 2 min. The gels also showed substantial mechanical integrity, i.e., the average compressive modulus was 32.73 ± 2.36 kPa at 15% compression of their original length. The degree of weight loss of these cryogels was found to be approx. 88% within 8 weeks of incubation in PBS (pH 7.4) at 37°C. Physio-chemically optimized cryogel was further evaluated for in vitro growth and proliferation of isolated primary goat chondrocytes up to 3 weeks. The cell adherence on cryogel was examined by SEM analysis, while cell–matrix interaction was examined by 4-6-diamidino-2-phenylindole and propidium iodide staining. Furthermore, the cell compatibility and proliferation was evaluated using the MTT assay. Increase in total cellular metabolic activity was observed as shown by continuous increase in glycosaminoglycan and collagen contents with time. Collagen type-I and type-II gene expression analysed for over 3 weeks by RT-PCR showed the prominent expression of collagen type-II. These results suggest the use of synthesised cryogel scaffold as a matrix for chondrocyte attachment and proliferation in 3-D environment and as a delivery system in cartilage-tissue engineering.

Keywords:

• CRYOGEL
• POLY(HYDROXYETHYL METHACRYLATE)
• GELATIN
• CELL–MATERIAL INTERACTION
• BIOCOMPATIBILITY
• CELL PROLIFERATION
• CHONDROCYTE
• COLLAGEN TYPE-I
• COLLAGEN TYPE-II