Tag Archives: muscle

PoWeRful mice and the effect of satellite cell depletion
Alec Dupont
Junior, biomedical science major
Auburn University

My Research Project

My project involved examining the adaptation of skeletal muscle to resistance exercise in mice that had been depleted of muscle stem cells (satellite cells). Generally, muscle growth is accompanied by an increase in protein synthesis and the differentiation of satellite cells into muscle nuclei. During this project, we examined if growth happens without the addition of satellite cells into muscle. As certain clinical populations have reduced satellite cell content and muscle mass, our project aimed to provide insights into how muscles respond to a growth stimulus with the loss of this cell population.

We used Progressive Weighted Wheel running (PoWeR) as a model for resistance exercise. PoWeR involves voluntary running activity of the mice in weighted running wheels. The weight placed on the running wheel is gradually increased over the course of four to eight weeks, overloading the musculature and causing a growth response called muscle hypertrophy. Using a genetic mouse model that allowed for the selective depletion of satellite cells, we compared sedentary- and resistance-exercised mice in groups of satellite cell-replete (vehicle treated) and -depleted (tamoxifen treated) mice. We compared muscle hypertrophy and other physiological adaptations between groups to determine the effects of satellite cell depletion. At the completion of this project, we hoped to gain a further understanding of the role satellite cells play in muscle growth.

Realities of Research

My main focus for the summer was using muscle tissue from the PoWeR mice, and making it possible to obtain data and useful information. I accomplished this through immunohistochemistry, a laboratory technique where we cut cross sections of the muscle and stain them for proteins of interest. This staining allowed us to visualize the sections under the microscope, image them and quantify the images using different forms of software. This technique presented certain challenges because the tissue must be carefully prepared and stored to prevent degradation. Poor quality tissue introduced variability outside of what is normal to the mice models. For example, having to overcome challenges and work to optimize a stain meant visualizing newly formed RNA in muscle nuclei. The stain can appear too dull and the quality would not be high enough to draw conclusions unless the procedure was optimized. Overcoming these challenges provided stunning images and reliable data. We found that although satellite cells were not absolutely required for muscle growth in response to weighted wheel running, there was a decrease in growth in the satellite cell depleted mice.

Life as a Scientist

The day-to-day life of a research scientist presented me with a constantly changing experience that was more engaging than the traditional classroom setting. There was always a new aspect of the project to investigate. It was incredibly satisfying to see your work come together in data that tell a cohesive story. The process of getting there was occasionally tedious though. For example, we’d normalize our data to the number of fibers in the muscle cross section and when the software couldn’t count for us, we were forced to count by hand. When the sections were between 600 and 800 fibers in a study with 48 mice, that part of research tended to drag. But that was only a minor inconvenience to a necessary bump in the road towards a satisfying research project.

Alec Dupont is a junior at Auburn University in Auburn, Alabama, studying biomedical science. He is a 2019 Undergraduate Summer Research Fellow (UGSRF) working under Dr. Charlotte Peterson at the Center for Muscle Biology at the University of Kentucky in Lexington. Alec’s work is funded by the American Physiological Society’s UGSRF program and a grant from the National Institute of Health to Dr. Charlotte Peterson and Dr. John McCarthy (AR060701).

Assessing a Therapeutic Target for the Treatment of Duchenne Muscular Dystrophy

Duchenne muscular dystrophy (DMD) is a severe, progressive muscle wasting disease that affects about 1 in every 5,000 boys, and it is the most common fatal X-linked disease in the world. Individuals affected by this disease will start developing muscle weakness by the age of 5, usually lose the ability to walk by their early teens, and ultimately, they will succumb to respiratory or cardiac failure by the age of 30. In terms of treatment, however, there are very few FDA approved drugs and therapies that can help alleviate, delay, or lessen the symptoms associated with this disease. What my project aims to do, is identify a new potential therapeutic target in dystrophic muscle. Previous studies have shown that dystrophic muscle, as compared to normal muscle, is unable to properly dispose of the damaged organelles within the cell1. In order to help the cells properly reduce damaged organelles, I am increasing the activity of the protein PGC-1α, which in the past has been shown to be successful in improving neurological function by helping to clear and get rid of damaged cellular components in brain cells affected by Huntington’s disease2. The ultimate goal of this project is to identify if this is a potential treatment target to help individuals suffering from DMD.

What are the pros and cons of working in a lab?

Working in a lab can be both an incredibly rewarding and frustrating process. This summer I have learned how to do immunohistochemistry and how to western blot. These two techniques have laid the ground work for any new scientific endeavors that I may go on to pursue later in my career. However, what this summer experience has taught me more than anything, is that not everything, and in fact, most things usually do not go as planned when working in science. For example, I have run many western blots over the course of the summer, and before I develop, I usually have an idea of what I think the blot is going to look like. Sometimes the blot comes out exactly the way I thought it was going to, and sometimes it comes out completely different. It’s during those frustrating times when I feel I have grown as a scientist. I have to go and talk to my other lab members about why the data looks the way it does. We usually sit down, look at previous literature, and talk about what may be potentially causing the data to look the way it does. As this happens, my research team and I go back to our research question and shape it to fit our new and unexpected results.

My day-to-day activities change every day. There really is no set schedule that I follow because my schedule is really dictated by the type of experiments that I am running that day. Some days are experiment heavy and some days are reading and writing heavy; it all just depends on the day. I think this is what surprised me most. When I began researching this summer, I figured I would follow a very set schedule. What I did not realize was that I would need to set aside a substantial amount of time to analyze data and compare it to similar experiments in other literature. I also get to talk with my other lab members and PI about the results and brainstorm with them about my data. I think that this is the best part about working in a lab. It is very rewarding when we work together as a team and brainstorm about what we think may be happening. The downside to working as a scientist is the frustration associated with experiments, which you have spent a substantial amount of time on, not working. Patience is important in life, but this experience has taught me that it is especially important in science.

References

  1. Palma CD, Morisi F, Cheli S, Pambianco S, Cappello V, Vezzoli M, Rovere-Querini P, Moggio M, Ripolone M, Francolini M, Sandri M, Clementi E. Autophagy as a new therapeutic target in Duchenne muscular dystrophy. Cell Death and Disease 3, 2012.
  2. Spada ARL. PPARGC1A/PGC-1α, TFEB and enhanced proteostasis in Huntington disease. Autophagy 8: 1845–1847, 2012.
Amanda Ludwig is a junior at Purdue University in West Lafayette, Indiana where she is majoring in Biology. She is a 2017 Undergraduate Research Summer Fellow, and she spent her summer working for Dr. Joshua Selsby at Iowa State University in Ames, Iowa. Amanda’s summer fellowship was funded through the American Physiological Society. Amanda’s post-graduation plans are still unknown, but she is interested in pursuing a career in medicine or biomedical research.