Jamie Twiggs and Dr. Robert Seegmiller, Department of Physiology and Developmental Biology
Stickler Syndrome is a severe disease with no cure that occurs in 1/7500 births resulting in extreme ocular, craniofacial, musculoskeletal and cardiovascular problems for those diagnosed. It is caused by mutations in the collagen genes, most often a premature stop codon in the collagen 2A1 gene, although in some cases it is caused by mutations in the collagen 11 A1 and A2 genes. The purpose of this research was to discover an effective method of gene therapy in mice models by reinserting a functional collagen gene into mice embryos using a viral vector and plasmids. I hypothesized that if the gene therapy in the animal model was successful then it would correct the collagen problems that cause the symptoms of Stickler Syndrome.
After receiving this grant I moved forward in this study, working closely with other students in Dr. Seegmiller’s undergraduate research lab. We discussed possible ways to transfect the functional collagen gene into a mouse embryo with Dr. Barrow and were able to come up with an initial plan. However, we soon learned that in order to conduct the experiment in a way that would yield viable results it would cost quite a bit more than we originally expected, and there was not enough money to proceed. Because there was no way to fund the Stickler Syndrome project, I received permission from the PDBio Department Chair and the ORCA office to begin working on a different project. While awaiting approval from these offices I continued to manage the Mouse Colony, which included breeding mice for other projects within the lab, isolating DNA from mice, and running PCR and gel electrophoresis regularly. I also worked with other students in my lab who were conducting an experiment with DDR2 Mice. Unfortunately we were never able to successfully genotype the DDR2 mice, which resulted in this experiment being put on hold as well.
This past semester I have been working under Dr. Seegmiller and Dr. Kooyman with David McDonald, and Kyle Jarman to more fully understand the pathway of Osteoarthritis. We have formed a study that will show how the presence or absence of TFG Beta affects the progression of Osteoarthritis in Spondyloepiphyseal dysplasia congenita mice. The results of this study will provide information necessary in moving forward in the search to better treat and correct the symptoms associated with Osteoarthritis in the human model. I expected that with an increase of osteoarthritis we would see an increase in TGF Beta in the SEDC mice. After staining slides of the knee joints of SEDC mice from ages 3 months, 6 months, and 9 months we did not see a direct correlation between TGF Beta and the expression of osteoarthritis. Because these results were not expected we decided that we needed to increase the number of slides that we were analyzing. Over the past few weeks we have been collecting a larger sample size and staining these slides for other components that are known to be key players in the osteoarthritis pathway such as HtrA1, S100, and DDR2. We will be staining the remaining slides in January. The larger sample size will allow us to better see if there is a correlation between TGF Beta and the expression of osteoarthritis and more fully understand what that correlation means.
I have aided in this experiment by reading many articles concerning TGF Beta and SEDC mice as well as taking photographs of the slides that we have stained and then Mankin scoring them. This means that I have catalogued the slides of different mice into categories of varying degrees of osteoarthritis expression. I will continue to do this with the slides that will be stained in January. I also spent many hours in the lab searching for the right slides to stain and then organized them in a manner that will allow us to stain them more efficiently. Other tasks have included meeting weekly with fellow lab partners and discussing what needs to be done and what conclusions can be made from the data we have already acquired. We have learned that contrary to our original hypothesis, TGF Beta does not seem to be highly expressed in mice that are known to have osteoarthritis. If this conclusion continues to be consistent throughout our study it will change some ideas that are currently held regarding osteoarthritis and it’s pathway. I have also been speaking frequently with my mentor, Dr. Seegmiller. He has directed my research and helped in formulating the hypothesis of the experiment. When this project is completed I am hopeful that we will be able to publish a paper and possibly present the results at a conference. My timeline is to finish by the end of
Winter semester 2013.
Over the past year in my research experience I have learned many valuable things and have had an overall positive and successful experience. Although I was not able to carry out my original plan of gene therapy in a mouse model, I was able to work closely with faculty members and learn from their examples and teaching. I also learned that there is always more than one solution to a problem in science, which is a lesson that will continue to be beneficial in my life as I further my education and move on in the health care field. I have become proficient in many lab procedures and I have learned the importance of working hard both as a team and as an individual.