Andrew Miner and Dr. Allan Judd, Physiology and Developmental Biology
Steroidogenic Acute Regulatory Protein (StAR) is a protein involved in the genesis of cortisol, testosterone, estrogen and other steroids. In fact, it is the rate-limiting step in the process of steroidogenesis. StAR is responsible for transporting cholesterol from the outside of the mitochondrial matrix to the inside, where it undergoes chemical processes that turn it into the steroids listed above. The body is thought to increase the amount of StAR in mitochondrial membranes when it receives certain signals in order to create more steroids. My topic involves the mechanism for this upregulation.
Interleukin-6 (IL-6), a cytokine involved in inflammation, fevers, and other immune responses, is hypothesized by us to play a role in this steroidogenesis. We received a plasmid (a circular piece of DNA) from some generous individuals at Texas Tech University that contained the StAR promoter region linked to the luciferase gene, the same gene that makes the protein that helps fireflies glow in the dark. Using a machine called a luminometer, we should be able to record how our cell culture of adrenal cortex cells responds to IL-6.
There are a number of reasons for our belief that IL-6 is involved in the long-term regulation of Steroidogenesis. Firstly, under chronic stress, typical hormones that increase stress steroids do not remain high, but the stress steroids themselves do. This leads us to believe that some other mechanisms besides those traditional methods must be involved in the process. In the past, Allan Judd has shown that IL-6 in expressed in the adrenal gland along with its receptors. This, along with some other factors too technical to mention in a report like this, led us to believe that IL-6 could cause a long-term increase in stress steroid hormones like cortisol.
We used a lipofectamine procedure to introduce the StAR/Luciferase plasmid into our adrenal cortex cells. We exposed some cells to IL-6 over time and other cells to a control medium. The cells that were exposed to IL-6 showed a significant increase in StAR activity, as measured by the amount of light they emitted under the correct light-generating conditions. At some doses and with some time courses, we saw as much as a tripling of StAR activity. These results were recorded several times, but then we ran into some problems in the lab.
The first problem came when we started to see massive amounts of luciferase activity even in the control systems. We thought the problem may lie in the luminometer or that I may have mixed up the tubes themselves. But after careful replication of the experiment, it became apparent that something else had happened. We found through much literature research that there have been cases of the plasmid we were using “kicking out” the regulatory promoter and replicating the luciferase at a mad pace. To solve this, we got a new plasmid with no history of such uncontrollable activity and tried some new experiments. About this time, our cells became infected and would not grow in the fashion we needed them to. Instead of lying in one nice flat layer, they were clumping together. We even tried a new cell line, which took several weeks to set up, but the project never recovered from those set backs, despite the early success. Currently, the lab is trying to reintroduce plasmid to the cells to continue on with the findings we have already established. We hope to publish these findings, along with some other findings from the lab, in the near future.
These events gave me much experience in the lab and helped me in all my upper-division courses on cell biology, molecular biology and Neuroscience. In addition, my ability to discuss research techniques was an invaluable asset when I interviewed for medical schools and could discuss all the things I had done in the lab. At my favorite medical school, my interviewer was a StAR researcher, and in large part due to this, I will be attending UT Southwestern medical school this fall, one of the greatest research institutions in the world. In addition to this, this research has helped me find a job as a research assistant in a dermatologic lab this summer, where I will prepare for a career in dermatology.
I wish to thank BYU’s office of research and development, their sponsors, and my mentor, Dr. Allan Judd for allowing me the opportunity to learn and develop more in this burgeoning field of Endocrinology. The support allowed me to perform better in my classes, approach great researchers and research opportunities at some of the world’s finest medical institutions with confidence, and understand more about the wonders of the human body. This program is a great asset to the University and its students. By fostering creative thinking and genuine research, Brigham Young University is sending brighter and better professionals into the world.