Assessment of Chitosan/ Gelatin Complexes as Tissue Engineering Scaffolds for Cartilage Regeneration

• Main Authors: Shu-Wen Whu, 胡淑文
• Other Authors: 徐善慧
• Format: Others
• Language: en_US
• Online Access: http://ndltd.ncl.edu.tw/handle/19285418611675536817

博士 === 國立中興大學 === 化學工程學系所 === 96 === Constructs based on chondrocytes and biomaterial scaffolds were developed for cartilage tissue engineering. One of the keys for success is to select suitable materials for fabrication of the scaffolds. In this study, chitosan-gelatin polyelectrolyte complexes were evaluated as tissue engineering scaffolds for cartilage regeneration in vitro and in vivo. The crosslinker for gelatin was selected among glutaradehyde, bisepoxy and water-soluble carbodiimide (WSC), based upon the growth of chondrocytes on the crosslinked gelatin. WSC was found to be the most ideal crosslinker for the system. The complex scaffolds with chitosan/gelatin ratio equal to one possessed the proper degradation profile and mechanical stability. Chondrocytes proliferated well and secreted extracellular matrix in chitosan-gelatin complex scaffolds crosslinked by WSC. Rabbit implantation confirmed the suitability of using chitosan-gelatin complex scaffolds for cartilage tissue engineering. For better culture condition, it was hypothesized that good mass transfer and the physiological shear provided by the rotating-type bioreactor were important for the neocartilage formed in the scaffolds to exhibit satisfactory mechanical strength and compression modulus; However, the dynamic culture condition was not prerequisite for the constructs to develop a histological resemblance to the real tissues. Then gelatin was observed to promote the human chondocytes proliferation; while chitosan was observed to maintain the human chondocytes morphology. For more cell source, human bone marrow mesenchymal stem cells (hBMSC) were seeded into two scaffolds, including blended polymers of PLGA50/50 and PLLA modified by type II collagen (BCII) and chitosan-gelatin complexes (CG). Cell numbers in CG scaffolds were higher than those in BCII scaffolds. The materials of the scaffolds had no effect on TGF-β3 induced hBMSC transformation into differentiated cells. The dynamic culture system promoted cell proliferation, but not cell differentiation. Finally, there is no current method for non-destructive quality control of tissue-engineered cartilage. This study explored a way to utilize a dynamic mechanical analyzer and rheological analysis to assess the cartilage tissues from different anatomic locations (e.g. arthrosis, costa and ear). Cartilage from different locations showed different storage modulus. Higher storage modulus was observed in positions that offered a greater loading force. Hyaline cartilage, either from arthrosis or costa, had similar values in loss tan. On the other hand, elastic cartilage (from ear) showed a distinct value of loss tan from that of arthrosis or costa. In spite of the much higher matrix content and cell number for ear cartilage vs. arthrosis cartilage, the mechanical properties of ear cartilage were much lower than those of arthrosis cartilage. Tissue-engineered constructs were cultured for 1 and 28 days, where the cell number, matrix content and storage modulus all increased with the culture time, but were still much lower than the values in the real cartilage. The values of loss tan of all constructs, however, approached those of real cartilage. It thus appeared that values of loss tan may serve as one of the major performance indice for tissue-engineered cartilage.