CONTENT REFLECTION: Throughout this process, I was extremely curious about whether genipin would be a viable solution to repairing the biochemical degradation caused by decellularization. I am glad that I did find that there is some evidence of genipin’s ability to do this. However, there is so much more that can be done with this research including looking at stem cell seeding and how it reacts with the tissue soaked in genipin, as well as completing this process with human tissue. I am glad that I can take the knowledge I gained from this year’s research and apply it to biomedical engineering research. Although I'm studying neuroscience next year, I can apply similar processes of tissue engineering to the brain. I will be working at CU Boulder over the summer in a neuroscience lab to gain experience doing this type of research.
PRODUCT: The academic paper and POD was an extensive undertaking, mainly the literature review. However, through the large literature review conducted, I took away a lot of information, as well as the skills to put pieces of information together. To create the methods surrounding genipin and the GAG assay, I had to take little pieces of information present across multiple different research papers, to come up with methods that work for cartilage. This included switching the GAG assay I would be using to something more friendly for cartilage tissue samples. I also took away a lot of information about the cell toxicity of each compound I would be using and how that would affect my experiment. With the academic paper, taking the time to reflect and analyze on the results of my research using other research was extremely valuable as it allowed me to see the links between different methods and subject areas.
PROCESS: This research enabled me to think about the applications of biomedical engineering to the world. There were so many topics I was interested in exploring, which signaled to me the endless possibilities with bioengineering projects. As a researcher, this entire process was extremely valuable as it led me to gain new skills in the lab and as a human. I can now skillfully use a lot of the equipment present in the lab right now and I gained the necessary skills to use a microplate reader. As a student, I was able to learn the ways of navigating through writing a paper and presenting my research. As a human, I strengthened a multitude of skills like communication and time management. These new skills will aid me in the research world and I am forever grateful to the AP Research and biotechnology program for allowing me to do this. In the future, I hope to apply bioengineering to neurological research. In the summer, I will be working in a lab at CU Boulder. In the fall of 2022, I will be majoring in neuroscience to gain the necessary skills to apply both fields of study.
Here is the abstract from my research:
Osteoarthritis, otherwise known as OA, is a common and painful disorder characterized by the degeneration of diarthrodial joints. Over time, the weakened tissue is less able to withstand mechanical loading, to the point that even normal activities produce abnormal wear and damage. One of the approaches to treating OA is the design of an engineered tissue, through decellularization, that reproduces the functional properties of healthy tissue while reducing the chances of tissue rejection. However, these decellularizing methods can be considered harsh, removing essential proteins needed for the subsequent cell seeding processes. One way to reduce this degradation is through crosslinking. This research aimed to develop a functional construct (made out of porcine articular cartilage) that will withstand the biochemical degradation from decellularization by using the crosslinking agent, genipin. Ultimately, I hypothesize that increasing concentrations of genipin will prevent the biochemical degradation of the porcine articular cartilage as evidenced by the GAG assay. A dose-dependent and time-dependent study of genipin was done after the specimens were decellularized for 48 hours in 2% SDS. For the 15 min study and the 30 min study, specimens were incubated in 0, 5, and 10 mM genipin. Afterward, a glycosaminoglycans (GAG) assay was completed to quantify the GAG levels to determine the biochemical integrity of the tissue. In order to identify if there is a significant difference in GAG preservation levels between the different time and dosage intervals of genipin, a Hotelling’s T-squared (T²) test will be performed. A statistically significant difference will occur if the p-value in GAG levels is ≤ 0.05, rejecting the null hypothesis that genipin does not contribute to lessening biochemical degradation.
Key Words: tissue engineering, decellularization, genipin, GAG assay, articular cartilage
The link to the final presentation will be coming soon!
Acknowledgments: I would like to thank Nikki Dobos, Shawndra Fordham, and Susanne Petri for their help in answering questions about the experimental design of this research and for pointing me in new directions I would have never thought of. I would also like to thank Dr. Alonzo Cook, an associate professor in the chemical engineering department at Brigham Young University, as well as Calvin Ewing, an alumnus from Rock Canyon’s Biotechnology program, for their guidance in this project in regards to the complication overseen in decellularization and staining last year. Without their insights from our project last year, the thought process behind this research would not have been possible. I would like to thank Chondrex too, who offered a generous discount for the glycosaminoglycans assay. Lastly, I am grateful to Rock Canyon High School and Douglas County School District for the funding and lab space that was provided for this research project.