Thesis (S.M.)--Massachusetts Institute of Technology, Dept. of Biological Engineering, 2012. / Cataloged from PDF version of thesis. Page 104 blank. / Includes bibliographical references. / Tissue engineering can be applied to develop therapeutic techniques for osteoarthritis, a degenerative disease caused by the progressive deterioration of cartilage in joints. An inherent goal in developing cartilage-replacement treatments is ensuring that tissue-engineered constructs possess the same properties as native cartilage tissue. Biochemical assays and imaging techniques can be used to study some of the main components of cartilage and assess the value of potential therapies. Agarose and self-assembling peptides have been used to make hydrogels for in vitro culture of bovine bone marrow stromal cells (BMSCs) which can differentiate into chondrocytes, undergo chondrogenesis, and produce cartilage tissue. So far, differences in cell morphology that characterize chondrogenesis had been observed in peptide hydrogels like KLD and RAD but not in the 2.0% agarose hydrogels typically used for culture. A tissue engineering study was conducted to determine if a suitable environment for cell proliferation and differentiation could be obtained using different agarose compositions. BMSCs were cultured in 0.5%, 1.0%, and 2.0% agarose hydrogels for 21 days following TGF-p1 supplementation. Results indicate that the 0.5% agarose hydrogels are clearly inferior scaffolds when compared to the 1.0% and 2.0% agarose hydrogels, which are generally comparable. Since agarose gels appear to be suboptimal in promoting chondrogenesis, self-assembling peptides should be used in future studies. In addition to the biochemical assays traditionally used in cartilage tissue engineering studies, atomic force microscopy (AFM) can be used to image aggrecan, one of the main components of cartilage. Imaging studies were carried out using fetal bovine epiphyseal aggrecan to optimize previous extraction and sample preparation procedures, as well as an AFM imaging protocol, for samples containing aggrecan. Experiments were conducted with 10, 25, and 50 ptg/mL aggrecan solutions to find the minimum concentration needed to create aggrecan monolayers on APTES-mica that would yield acceptable AFM images (25 [mu]g/mL). AFM instrument and software parameters were optimized to find the working range of the integral and proportional gains (0.2 - 0.4 and 0.6 - 0.8, respectively) and to increase the resolution, showing fields at the 800 nm level. Finally, an image processing protocol relevant to these molecules was established. / by Nicole Casasnovas Ortega. / S.M.
| Identifer | oai:union.ndltd.org:MIT/oai:dspace.mit.edu:1721.1/76172 |
| Date | January 2012 |
| Creators | Casasnovas Ortega, Nicole |
| Contributors | Alan J. Grodzinsky., Massachusetts Institute of Technology. Dept. of Biological Engineering., Massachusetts Institute of Technology. Dept. of Biological Engineering. |
| Publisher | Massachusetts Institute of Technology |
| Source Sets | M.I.T. Theses and Dissertation |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | 104 p., application/pdf |
| Rights | MIT theses are protected by copyright. They may be viewed, downloaded, or printed from this source but further reproduction or distribution in any format is prohibited without written permission., http://dspace.mit.edu/handle/1721.1/7582 |
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