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No.11（2016）7.Three-dimensional cell culture using fabrication of high-density collagen fibril matrix gels renaturation of triple-helix collagen from gelatin
Yoshimi Ohyabu，Hirosuke Hatayama，Masataka Ida，Shunji Yunoki，Yosuke Hiraoka
Fibril formation of collagen is frequently used for encapsulating cells and 3D cell culture, but the weak mechanical properties of collagen fibrillar gel limits its use in regenerative medicine. To improve the mechanical properties by increasing the density of the collagen gel, we have developed a novel method for creating high-density collagen gels by renaturing denatured collagen. Pepsin- digestion collagen was denatured gently to avoid cleavage of the molecules to produce uncleaved gelatin (UCG). UCG having a denatured collagen triple-helix structure exhibited a more rapid and efficient renaturation of the triple-helix structure than cleaved gelatin (CG) over a broad range of temperature settings. The UCG solution formed an opaque gel having high-density reconstituted collagen fibrils at 28–32°C and a transparent gel similar to CG at ˂25°C. The unique gelation properties of UCG enabled the encapsulation of cultured cells by CFM with a high cell viability even at a high concentration (>5%) and high elasticity (1.28 ± 0.15 kPa at 5% and 4.82 ± 0.38 kPa at 8%). The elastic modulus was higher than that of conventional CFM containing 0.5% collagen. High-strength CFM may provide more durable hydrogels for tissue engineering and regenerative medicine.
Keywords
Collagen, Gelatin, Renaturation of triple-helix, Fibril, Three-dimensional cell culture
