PURM Research Summary
This past summer I had the opportunity to work under the leadership of Dr. Hydar Ali at the dental school. His lab studies the role of mast cells in host defense and chronic inflammatory disease like anaphylaxis, asthma, atopic dermatitis, etc. Mast cells are granulated immune cells primarily found in tissue exposed to the external environment such as skin, mucosal surfaces of the gut and lungs, and the tissue surrounding blood vessels. The Ali Lab specifically focuses on the newly discovered Mas-related G protein coupled receptor X2 (MRGPRX2), and how its expression in mast cells contributes to allergic and inflammatory disease.
The overarching goal of my project was to identify the most promising humanized mouse model for in vivo study of the MRGPRX2 receptor. My project compared two different humanized mouse models: BLT and the NBSGW mice. NBSGW mice are engrafted with human hematopoietic stem cells in their bone marrow, and they carry a mutation in the c-kit gene that promotes the growth of human hematopoietic stem cells without irradiation. BLT mice are also engrafted with human hematopoietic stem cells, but they are more advanced in that the mice are irradiated and then transplanted with human fetal liver and thymus fragments. Knowing which of these two models develops the most human mast cells with MRGPRX2 expression can lead to more effective studies of the MRGPRX2 receptor.
Through this project, I gained a lot of experience with dual immunofluorescence staining and microscopy. While the staining procedures became pretty straightforward to me over time, I was surprised to discover how difficult of a process fluorescent microscopy and imaging was. Identifying and counting cells may sound like a simple process, but one has to consider the several issues that can arise. For example, immunofluorescent staining often has a lot of non-specific staining or autofluorescence from the tissue itself. This creates a lot of background in the images, making it difficult to distinguish between positive and false signals. Therefore, you cannot just mindlessly count any dots you see — you have to verify the presence of a nucleus, make sure the cell shape matches that of a mast cell, look at the tissue at a higher magnification, and make any changes to the procedure to decrease any non-specific staining. This process taught me the importance of adaptation and perseverance in science, since science requires you to constantly improve your methods and techniques to achieve better results. I am grateful that PURM gave me this opportunity to learn not only laboratory procedures, but also the critical thinking and scientific process that goes behind these procedures. I am thankful towards all of the members of the Ali Lab who provided me with a valuable introduction to immunology research at Penn.