Assessing the effects of mRNA silencing of PIEZO1 gene

Author: Addison McLaughlin | Major: Biomedical Engineering | Semester: Spring 2023

Background

My name is Addison McLaughlin and I am going into my junior year of biomedical engineering. This past semester was my first full semester of research. This semester, I studied the PIEZO 1 ion channel. The objective of the study was to examine the effects of silencing a mechanosensitive calcium ion channel, PIEZO 1. Included in this was studying the effects of YODA 1 on the osteogenic differentiation of mesenchymal stem cells (MSCs). YODA 1 is a chemical compound that mimics PIEZO 1 in mice and humans by reducing the mechanical threshold for PIEZO 1 activation. This increases the rate of osteogenic differentiation in MSCs. siRNA transfection was performed on MSCs to silence the PIEZO 1 gene, real time polymerase chain reaction (qPCR) was performed to confirm the silencing, and next the cells will be assessed for differentiation outcomes.

Project Execution

Throughout the semester, I have worked to optimize the protocol for YODA 1. This entailed varying the amount of time that the cells were exposed to YODA 1 and the concentration at which YODA 1 was present. I was also able to learn PCR from a graduate student in our lab before he graduated. This has allowed me to proceed with the portion of my research relating to PIEZO 1.

Trouble Shooting

I ran into many problems throughout my research experience. Though the cells were meant to be adhered to the wells, hyperdepostition of collagen caused the cells to peel off and so I had to start over. I also had issues with the Alizarin Red staining of two different donors that had been treated with YODA 1, where there were few differences between it and the control group, but there were other signs that differentiation had occurred.

Mentorship

My mentor is Dr. Rebekah Margaret Samsonraj. I came to learn about her lab when she was my professor for Introduction to Biomedical Engineering and I joined her lab part way through the fall semester of 2022. She had several projects in the works when I joined her lab. She and I worked together to decide on which one would be best for me, that is how we landed on my chosen topic. My mentor guided me through obstacles and failures in my research by helping me troubleshoot problems as they occurred. She also always encouraged me to read more about my chosen topic and further my understanding of what specifically was happening with the YODA 1 and what each step of the PCR protocol was meant to do

Conclusion and Acknowledgements

I was also helped by my fellow lab mates, Vitali Moldanado and Kadek Ardana. Vitali taught me almost everything I know about cell culture, and Kadek taught me everything there is to know about qPCR. Both Vitali and Kadek were very patient with me and took the time to help make sure I was confident in my abilities.  I am truly grateful for everyone who helped me this past semester.

This past semester has been amazing. Through all that I have learned, I was able to vastly improve my problem solving and perseverance skills, as well as my time management skills. I took each of these problems step by step to try and figure it all out. It helped to cement my future plans as it made me realize that I did want to go to graduate school and that I wanted a career in research and development after completing my education. I plan to continue working in Dr. Samsonraj’s lab throughout the rest of my time at the University of Arkansas, and then I plan to attend graduate school with a focus in tissue engineering.