STEP Reflection

STEP Reflection post for Steven Elzein – Undergraduate Research

What? – A detailed description of what you did during your STEP experience.

I participated in undergraduate research as part of my STEP experience. I spent 400+ hours (40+ hours/week for 10 weeks) in the Dr. Ginny Bumgardner laboratory, a transplant immunology lab in the Department of Surgery. I had been a student researcher in this lab for about a year before spending my summer there. Previously, I had worked on multiple lab projects in different capacities, but I had never completed an individual project. For my STEP experience, I undertook a personal project related to transplantation and transplant rejection. Specifically, my project examined the effects of two widely used immunosuppressive drugs (Rapamycin and Tacrolimus) on alloantibody-mediated rejection post-transplant. The title of my project was “Differential Effects of Rapamycin and Tacrolimus on CD8+ T cell-Mediated B Cell Clearance and Post-Transplant Alloantibody Production”. In general, the goal of my project was to uncover exactly what these immunosuppressive drugs were doing to key cellular interactions that regulate antibody-mediated rejection post-transplant.

I was involved in all aspects of the project, from brainstorming, critiquing, and writing up experiments, to actually performing the surgical work and bench work necessary to carry out the experiments, to graphing, analyzing, and reporting results. Planning for this project began in Spring 2014, during which I began writing up experimental trials, applying for funding, and ordering materials. After all of the experiments had been planned and prepared for, I began my STEP experience by performing a survival study during the month of May. This study involved transplanting a cohort of mice with allogeneic (genetically different) hepatocytes in order to incite an immune response, treating the recipients with immunosuppressive drugs, and subsequently monitoring their alloantibody levels and extent of graft survival through blood draws over a period of 30 days. Alloantibody levels were analyzed using the lab’s alloantibody quantification protocol, and graft survival was analyzed by monitoring levels of the ha1at reporter protein (a protein whose level indicates survival of the graft) via ELISA assays. As a result of my year of training in the lab prior to the start of this project, I performed all aspects of this experiment and the ones to follow, including transplant procedures, splenocyte isolations, immunosuppressive treatments, various assays, etc. This initial study included various conditions and served as an initial overview of the important aspects of my project, providing a clearer picture of which experiments to carry out next.

The experiments that followed focused in on the extent of B cell death in the post-transplant immune environment. I conducted two separate in vivo cytotoxicity studies, with each one focusing on one of the two immunosuppressive drugs in question throughout my studies. These studies involved transplanting mice, and then adoptively transferring fluorescently-labeled target cells into recipients and subsequently analyzing the degree of target-cell death one day after adoptive transfer. After the in vivo cytotoxicity studies, I took a little detour from my summer project and helped complete a small study for another project in the lab. This smaller study had the goal of providing more evidence for a manuscript that was recently accepted for publication in the American Journal of Transplantation. Lastly, I conducted an in vitro cytotoxicity study, which served to confirm the previous in vivo data I had obtained and offer more insight on how to best proceed with the next experiments. Unfortunately this study did not pan out, and so work is currently being done to tease out errors in the experiment. Over the course of the summer, I also transplanted mice to serve as organ donors for histological studies to be conducted in the future.

Throughout my entire STEP undergraduate research experience, I kept track of my daily activities and graphed and reported results from each experiment. I also reported my results during weekly laboratory meetings. My studies uncovered that Rapamycin (one of the immunosuppressive drugs in question in my studies) is a more potent inhibitor of alloantibody-producing B cells than Tacrolimus (the other immunosuppressive drug utilized in my studies), as displayed by the fact that Rapamycin completely ablates post-transplant alloantibody, while Tacrolimus does not. In addition, it was found that neither drug is directly cytotoxic to B cells, even at high doses.

So What? – A personal response to your STEP experience, including feelings, thoughts, judgments, and what you have learned about yourself and your assumptions from what you did and how you reacted. 

My summer research experience was both busy and rewarding. I spent a lot of time learning, reading, and conducting experiments in the laboratory, and I accomplished a great deal as a result. First and foremost, I made it through my first full-time working experience. I had never worked full-time before, so this research experience also doubled as a work experience. While a full-time work schedule took some getting used to at first, it turned out to be beneficial and allowed me to maintain continual focus on my research.

I would categorize my entire research experience this summer as a learning experience. No matter how much I thought I knew or accomplished, there was always something new that I was learning each day. . First, I learned first-hand that research takes time. Before I started this summer experience, I was unaware of how much time was put into planning, executing, and perfecting each experiment. For example, for my studies in particular, not only did I have to spend a couple of days planning logistics and preparing materials for an experiment, but I needed to also wait one week after the transplant for the immune cells to become activated before I conducted any studies. Recipient mice also needed to be monitored and treated with immunosuppressive drugs daily. Furthermore, it takes time to isolate and prepare cells for an experiment, as well as analyze, graph, and interpret the results from that particular experiment. All in all, if everything goes smoothly, it can generally be expected that one experiment will last approximately two weeks. This was something that I did not expect coming into the summer research experience. However, now that I have obtained a sense of how much time each experiment takes, I have a newfound appreciation for all of the published manuscripts that I read, especially in the field of transplant immunology. When one thinks about how much time each experiment takes, coupled with the fact that manuscripts normally comprise many experiments, it is remarkable how much research is taking place in this field and others, and I am certainly glad to have learned this aspect of the research process this past summer.

Second, I learned that a researcher, over the course of any particular study, will more likely than not run into several roadblocks along the way to obtaining their results. No matter how much planning goes into an experiment, it doesn’t always turn out the way you had planned. Through my full-time research this summer, I experienced the process of running into roadblocks with research and having to use critical thinking skills in order to work my way around them. In addition, it is almost always necessary to determine what went wrong in a particular experiment before continuing. For example, I’ve had more cells than usual die during a staining step of an in vivo cytotoxicity assay. It was determined later on that the stain had been applied to the cells for too long, causing many of them die. In addition, the nature of my studies contains inherent room for roadblocks. My studies involve surgical procedures in which cells are transplanted into recipient mice. As with any surgical procedure, there is always the risk of complications resulting from the incision, anesthesia, or the procedure itself. With that being the case, it is not uncommon for a recipient mouse to pass away from time to time, thus hindering the study. Yet, even with all of the potential aspects that can go wrong with an experiment, this summer experience has helped me learn that setbacks should be expected in research, and that I should use the critical thinking skills I gained this summer and elsewhere to overcome any hindrances in my progress.

Now What? – Discuss how the things you experienced and learned during your STEP experience will affect your academic, personal, and life goals moving forward. 

While my STEP experience gave me the opportunity to get started on an undergraduate research project, one summer was insufficient to complete my studies and there is still more work to be done. As such, my STEP experience has directly influenced my academic goals, especially for the upcoming year. I plan to continue doing research on this same project until it is complete, and the work will likely extend into the upcoming academic year. This research will likely culminate in several poster presentations this school year, including the Fall Undergraduate Student Research Poster Forum, the Denman, and the Great Lakes Transplant Immunology Forum to take place at Ohio State in the fall.

My STEP experience has also influenced my personal goals with regards to completing a project. Even though one summer of full-time research wasn’t enough to complete all of the experiments in my study, it started me on the path to completing my study in full. As a result of my STEP experience, I have adopted the personal goal of continuing to conduct experiments in an effort to complete the study in its entirety. The ultimate goal for my work on this project is for it to culminate into a published manuscript in a recognized transplant journal.

Before my STEP experience began, I was sure that I would like to pursue a career in medicine. However, I was unsure whether or not I would also like to pursue research as a component to my future career, perhaps as a part of an MD/PhD pathway. By allowing me to focus all of my energy on research, my STEP experience has given me insight into what it’s like to be a researcher. As a result, I am more confident in my ability to conduct research and have made it a life goal to include research as a component of my future career.

 

 

 

 

 

 

 

Using the hood