My practicum site was at the Cheema Lab in Georgetown University Medical School at the Lombardi Comprehensive Cancer Center (LCCC). At the Cheema Lab, I worked on a project titled "Influence of Head and Neck Cancer-derived Extracellular Vesicles on Macrophage Polarization".
As an overview about my project, Head and Neck Cancer is a type of cancer that originates from mucosal epithelial cells that line the oral cavity, pharynx, larynx, and sinonasal tract. It is the 7th most common cancer. Some symptoms include a sore or lump in the mouth or throat, constant sore throat, difficulty swallowing, and trouble breathing. To study this cancer, I grew the FaDu cell line of Head and Neck Cancer, which is derived from a tumor in the Hypopharynx.
I mentioned that I was interested in the extracellular vesicles released by this cancer. Extracellular vesicles, or EVs, are small membrane bound sacs released by cells into the external environment. They contain cellular components and are involved in cell-to-cell communication, transport of materials, and the immune response. One of the cells involved in the immune response are macrophages.
Macrophages are a type of immune cell that have various roles. These roles include phagocytosis, which is where the macrophage will detect foreign and damaged material and engulf the material (as shown in this diagram), activating other immune cells towards an immune response, tissue repair, healing, and maintaining homeostasis.
Macrophages are derived from bone marrow monocytes. This means that macrophages will take different roles depending on the signals in the environment. This process is called macrophage polarization, a process where macrophages adopt distinct functional phenotypes in response to signals in the microenvironment. For example, when you get an injury, those damaged cells will release cytokines, which are signal molecules. Monocytes are recruited to the site, when encountering these cytokines, they differentiate into macrophages and adopt a phenotype specific for a function.
There are two extremes of phenotype that macrophages can have, M1 and M2. The M1 phenotype performs phagocytosis and acts as the killer cells. It is often involved in killing pathogens and cancerous cells. The M2 phenotype acts as the repair cells and often is pro-tumor if a tumor is present. With Head and Neck Cancer, and many other cancers, the M2 phenotype of macrophages is involved with tumor survival.
Now, with all this background information, my project is about looking at Cancer-derived EVs and their influence in Macrophage polarization. Research has shown that cancer-derived EVs result in macrophage polarization such that they obtain a phenotype that will support the tumor microenvironment. These macrophages are called Tumor-Associated Macrophages (TAMs), which support tumor growth instead of attacking the tumor.
I did this by growing Head and Neck Cancer cells and Normal Oral Epithelial Cells (as a control group). After growth, I isolated the extracellular vesicles (EVs) and characterized them by NTA analysis, which looked at the size of EVs in my samples to confirm they are EVs, and immunoblot, which looked for markers typically found on extracellular vesicles. After confirming that my samples were EVs, I treated them to monocytes to push the monocytes into macrophages. I then ran a western blot of the treated monocytes and looked for expression of GAPDH, a protein expressed by all macrophages, and Arginase-1, a protein expressed by tumor associated macrophages. The results showed that the macrophages resulting from the Head and Neck Cancer derived EVs had higher Arginase-1 expression compared to the macrophages resulting from the Normal Oral epithelial EVs. This shows that the Head and Neck cancer-derived EVs resulted in TAMs. This research's significance is that it can lead to a new area of research of potentially finding a way to target these extracellular vesicles from cancer cells so that the immune system can be protected and function properly against the cancer rather than for it.
I found this opportunity by searching for undergraduate research opportunities. I came across an application website at Georgetown University LCCC for a program that relates to undergraduate research and decided to apply. The program matches you to a professor based on your indicated interest and I was matched with Prof. Cheema. In my case, my site supervisor was given based on interests, however I did my own research beforehand to look at what research is done at the LCCC. I think this helped me get into the program. My advice would be to make sure that you are interested in the work they do and mention your interests. Take the time to read their backgrounds on their website and a couple of papers. Even if you don’t fully understand it, the fact that you took the time to read past papers will show your genuine interest.
At this practicum site, I learned many wet-lab techniques (listed below). The techniques that I most enjoyed and found fun and interesting were cell-culture, EV isolation, RNA isolation, Western Blot, and NTA analysis. I also realized that one of the things that I like most about this experience was the fact that alot of this work was hands on. Although it may be tiring, since you need to be there at the right time ready to move to the next thing, since the cells won't wait for you, it was also a lot of fun to do in the moment. I was also exposed to many new things such as research in head and neck cancer, macrophages and their role in cancer proliferation, clinical application of cancer research, as well as participating in scientific discussions and meetings.
Techniques I learned:
Beyond science, this experience confirmed my interest in research and wet-lab research. Although it was challenging, it was rewarding at the end to see it all come together and how all the pieces led to a conclusion. In the future, I hope to do research and I am interested in going to graduate school for Biochemistry. Overall, I am very grateful for this opportunity and I really enjoyed the entirety of it.