Monthly Archives: January 2018

My Summer of Science

In the summer of 2017 I spent my time working an unconventional full-time job in the lab growing as a scientist. I was given an opportunity to be mentored by an established research professor who I look up to greatly. She helped me step by step go through the process of coming up with a project, understanding the current research in the field, and guiding me through the behind the scenes documentation to get fully approved by local, state, and federal agencies.

My project that I am undertaking is geared towards ultimately understanding the roll of dietary choices and its implications on oxidative stress, leading to potential for cardiovascular disease. My lab and I are utilizing birds as a model to demonstrate the effects of a high-carbohydrate diet and its ability to cause a hyperglycemic condition. For the study, Mourning Doves will be used due to their high density in the Tempe, AZ area and much of the studies leading up to this used the same birds. They have a unique ability to combat the oxidative stress that is commonly seen due to high blood sugar. We hope to better understand how their body combats the negative effects of hyperglycemia in efforts to process the information regarding the protective mechanisms of the vascular system.

Realities of Research

The idea of moving research forward can be very attractive. The actual process of progressing it may not be as attractive. Research is a very meticulous, detail-oriented task that needs to be well planned, documented, and thought-out for it to properly work. There is no book you can read that gives you the exact answer to what you are doing. This is the world of the unknown and if properly done could potentially help shift the understanding for those to come after you. However, a major hiccup that I believe is frequently over looked as a young scientist going into a research lab is that it’s quite common to get negative results, or even though the data seems to support what you thought, it is not statistically valid based on the mathematical analysis. This is a huge part of research and can be very testing of resiliency as the process of getting this far could take months or years.

Life of a Scientist

Although, it is common to see science hitting the news headlines frequently on your phone and social media feed, it is a much longer process of meticulous work than what is publicized. In the efforts of using an animal in the aide of progressing the scientific community there is a very stringent process of approvals needed to work with them. For example, Mourning Doves are migratory birds, so my project requires an approval by the Arizona State Game and Fish Department, the U.S. Fish and Wildlife services, and the Institutional Animal Care and Use Committee (IACUC) before I can begin the research itself. I was surprised as I began to work in the lab by all the intricacies of administrative background work that went into being able to conduct research. While repeating experiments take a lot of time and effort, I now feel as if there is equal if not more effort done in paperwork and formalities than physical experimentation. I, myself am still waiting to begin the physical hands on portion of the research. I have worked through the course of the summer gaining valuable experience in the creation of the protocol and understanding the approval process. I have also been given the opportunity this summer to work very closely with another research project that is much larger than my own.

This summer has been an invaluable experience in research that I would not have gained otherwise without the gracious efforts of the APS.

References

  1. Braun, and Sweazea. “Glucose Regulation in Birds.” Comparative Biochemistry and Physiology, Part B1 (2008): 1-9. Web.
  2. Smith, Toomey, Walker, Braun, Wolf, Mcgraw, and Sweazea. “Naturally High Plasma Glucose Levels in Mourning Doves ( Zenaida Macroura) Do Not Lead to High Levels of Reactive Oxygen Species in the Vasculature.” Zoology3 (2011): 171-76. Web.
  3. Sweazea, Braun, and Sparr. “Novel Role of Insulin in the Regulation of Glucose Excretion by Mourning Doves (Zenaida Macroura).” Zoology122 (2017): 58-62. Web.
Will Clark is a senior biochemistry student at Arizona State University in Tempe, AZ. He is a 2017 Integrative Organismal Systems Physiology (IOSP) fellow conducting research in the lab of Dr. Karen Sweazea at Arizona State University. His long-term goal is to become a Physicians Assistant, and join the health care workforce to help improve the health and well-being of others.
Stemming Beyond the Brain

This summer, I am very thankful to be a recipient of the Undergraduate Summer Research Fellowship (UGSRF). This fellowship has given me the opportunity to conduct research under the guidance of Dr. Justin Grobe at the University of Iowa. My research project studies the role of AT1a receptors on the vasopressin-producing cells found in the hypothalamus region of the brain.

Vasopressin (AVP) is a hormone, produced in the magnocellular neurons within the hypothalamus, playing a key role in maintaining homeostasis. This hormone maintains homeostasis by increasing blood pressure through upregulation of water reabsorption and vasoconstriction. The AT1a receptor responds to high levels of angiotensin II, ANGII, by constricting the cells with AT1a as part of the renin-angiotensin-aldosterone system, RAAS. A previous study conducted in our lab has shown AT1a receptor localization on vasopressin-producing cells in the supraoptic nucleus of the hypothalamus1. In order to observe the role of AT1a receptors, the experiment compared the response of AT1aAVP-KO mice with control mice under the ICV infusion of ANGII or aCSF at 50ng/hr. Specifically, I am looking for responses in water-intake, urine output, blood pressure, and vasopressin secretion. This experiment will provide further insights to understand AVP secretion in response to brain RAAS and could potentially help prevent hypertension, exhibited patients with obesity, diabetes, and/or preeclampsia.

Figure 1: qPCR machine for gene expression analysis

I have learned many valuable skills from conducting research over the course of this summer thanks to my awesome mentors and principal investigator. They have devoted their time to teach me how to use Excel in analyzing the results of specific qPCR gene expressions and creating protocols for many experiments utilized in the lab. Nevertheless, there are many other essential skills that I have yet to learn such as interpreting results, reading scientific journals, and other lab protocols. They are important in all aspects of research but honing in these skills will require time and practice. For instance, several experiments that I performed yielded poor results and required additional extra time devoted towards troubleshooting the problem. Although frustrating at times, I have learned troubleshooting is one of the most important skills in research because not all the experiments will yield the desired results. While learning and refining my skills, the research project has progressed into the final stages towards completion and I am very excited to find out what results from the samples reveal regarding the role of AT1A receptors.

Figure 2: Tail-cuff system setup for blood pressure measurement

As a member of the lab during this summer, I have gained an insight into becoming a scientist. Every morning is devoted towards measuring the blood pressure in the mice cohorts, which need the measurements to be taken daily for three weeks prior to the surgery and two weeks after. The afternoon is spent performing qPCRs on specific genes from tissues collected earlier, analyzing data, learning new protocols, and/or reading publications pertaining to my research area. There are many basic aspects that scientists value greatly such as labeling tubes, checking the timer, and pipetting the exact amount of solutions. The focus on minute details is a trait developed over the years dedicated towards conducting research, and I have definitely noticed improvements in my attention to detail since I first began conducting research. In the next couple of weeks, I will have the opportunity to collect, organize, and analyze the results during the finals days of this experiment. I would like to again thank APS for the opportunity to conduct research at the University of Iowa this summer.

References

  1. Linggonegoro D, Sandgren J, Claflin K, Perschbacher K, Ni J, Pearson N, Pierce G, Santillan M, Grobe J. Physiological Significance of Angiotensin AT1A Receptors in Vasopressin-Producing Cells of the Supraoptic Nucleus. Experimental Biology.
  2. Guyton AC, Hall JE. Textbook of Medical Physiology. 10th ed. Philadelphia, PA: W.B. Saunders Company, 2010.
I am rising junior at the University of Iowa, majoring in biomedical engineering. I will be pursuing a biomechanics and biomaterials track with an interest in implant design and drug delivery. This summer, I had the opportunity to conduct research under my principal investigator, Dr. Justin Grobe, with the assistance of UGSRF fellowship funding by the American Physiological Society. My project investigated the role of AT1a receptors on the vasopressin-producing cell in cardiometabolic control.
The Life of a Summer Student Researcher

Childhood obesity has become a major health issue in the United States recently. Research studies show that childhood obesity is associated with an increased risk of declined renal function, which is defined as renal injury. Since renal hyperfiltration, which is when the kidneys are working more to than needed, this leads to excessive amounts of protein to be produced in the urine (proteinuria). This condition is typically associated with obesity. The current study will determine if we prevent the renal hyperfiltration, can we decrease proteinuria and/or renal injury?

Working on ground breaking research and being in an environment that is focused and effective was eye-opening for me. I enjoyed being around scientists that had similar mindsets and were able to help me develop a scientific mind of my own. It surprised me that everyone was so open and welcoming to a new unexperienced college student. But it made me more comfortable, especially when some of my experiment’s failed, but they were very helpful and encouraging throughout the entire process. During my time in the lab the initial study I worked on was not successful, so I was give the study I have now and the results that were collected were what my research host expected and it was a successful study that will be continued.

Being a scientists is like riding a roller coaster every day that you enter into the lab. Some days you are so busy with experiments that you forget to eat lunch. But then other days are very slow, because you are waiting on data or results. It is a great environment to learn not only about science but about yourself, how you manage time, how you interact with people, and how well you work alone. I think the best part about being a scientist is that every day is a new challenge because every day you are working on something that could possibly change the world. I think that worst part is that even if you do everything correctly the data can still not turn out how you want it. But overall I really enjoyed the team aspect of working in a lab. Everyone in the lab helps everyone on their projects and vice versa, because everyone wants to see the lab/experiments succeed because it could have a positive impact on the world.

Alyssa Pennington is a senior majoring in Chemistry at Jackson State University. Alyssa is working in Dr. Jan Michael Williams lab at the University of Mississippi Medical Center in Jackson, MS. Alyssa is a second year Short-Term Education Program for Underrepresented Persons (STRIDE) Fellow which is funded by the APS and a grant from the National Heart, Lung and Blood Institute (Grant # 1 R25 Hl115473-01). After graduation Alyssa plans to pursue a career in medicine and research.
Investigating the Role of Piezo2 in Serotonin Release

At the mention of serotonin, people commonly think of its role as regulating anxiety, mood, and happiness in the brain. However, 95% of the body’s serotonin is produced by enterochromaffin (EC) cells present in the tissue lining of the gut (1) before being taken up by platelets that circulate it throughout the body (2). The EC cell releases serotonin in response to mechanical and chemical stimulation, and this release differs depending on whether the body has been fed or is fasted (3). Gut-derived serotonin is important for regulating:

  • Stretch and contraction of the gut
  • Gut sensation
  • The ability for platelets to clot blood
  • Bone development
  • Blood pressure
  • Metabolism (1)

There are still many questions regarding how the EC cell releases serotonin. Recent studies from Dr. Beyder’s laboratory of Mayo Clinic discovered that a mechanosensitive ion channel called Piezo2 is present in human and mouse EC cells, and that Piezo2 is important for serotonin release in EC cell models after a mechanical force like pressure is applied (4, 5). In the broad scheme of things, understanding the role of Piezo2 in EC cell serotonin release and in regulation of blood serotonin levels could lead to future drug developments for gastrointestinal diseases.

A visual representation of serotonin release from EC cells and platelet uptake. Reference: Modified from Mawe & Hoffman. Nature Rev Gastro & Hep. 2013. (1)

Dr. Beyder’s laboratory created a mouse model with the Piezo2 gene knocked out of the gastrointestinal tissue lining the gut. These knockout mice can be compared to the control mice that do not have the Piezo2 gene removed to see how Piezo2 affects the physiology of the gut. My aims for my ten week research fellowship were:

  • To determine the role of Piezo2 in mechanosensitive EC cell serotonin release from primary EC cells from fed and fasted epithelial Piezo2 knockout and control mice
  • To determine the role of Piezo2 in systemic serotonin levels in fed and fasted epithelial Piezo2 knockout and control mice

As is the case with most research, everything did not turn out as planned!

In the first few weeks of my research project, I had already faced unexpected complications. I was planning to collect cells from mouse colon and place them on a special plate with the proper nutrients and environment to help them grow into what is called a primary culture. However, we were not able to take pictures to show that the primary cultures were present since we did not have a microscope lens that could fit the wells in the plate. With the limited time I had for the project, my team decided to focus on measuring whole gut serotonin instead. This highlights how quickly a research project can change and the many approaches a scientist can take to solve a problem. There were also times I would try a new technique to yield better results, but would ultimately go back to the original technique. Overall, the general research process was to look back to see how an experiment was done in the past, and then complete a series of trial and error cycles from there to optimize the protocol for my specific project.

After collecting and preparing samples, I ran a test called an enzyme-linked immunosorbent assay (ELISA) that quantifies the amount of serotonin in a sample.

Amid the journal clubs and experiments, this summer I learned about the collaborative efforts required to be successful as a scientist. I shared equipment and bench space with other researchers in the lab, so we had to coordinate a schedule so that everyone could complete their experiments. I enjoyed getting to know the other members of the lab, many with diverse backgrounds and specialties. Every week the department had a lab meeting where someone from each lab would take turns presenting research updates. It was a great opportunity to learn about other projects that related to the enteric nervous system and new adjustments made to fix problems. That being said, life as a researcher has its challenges. There were days I would spend hours preparing an experiment, only to made a mistake that could have been prevented if I would have asked the right person. If I had to give any advice from my summer experience, I would say don’t be afraid to ask for help – especially if you are feeling discouraged! Everyone in a lab develops his or her own expertise in something, which makes it impossible to know everything. But one of the beauties of science is that there is still a lot for us to learn, and sometimes the most frustrating failures can bring unexpected discoveries.

References

  1. Mawe GM, Hoffman JM. Serotonin signalling in the gut–functions, dysfunctions and therapeutic targets. Nature Reviews: Gastroenterology and Hepatology 10: 473-86, 2013; 10.1038/nrgastro.2013.105.
  2. Berger M, Gray JA, Roth BL. The expanded biology of serotonin. Annu Rev Med. 60: 355-66, 2009; 10.1146/annurev.med.60.042307.110802.
  3. Sumara G, Sumara O, Kim JK, Karsenty G. Gut-derived serotonin is a multifunctional determinant to fasting adaptation. Cell Metab. 16: 588-600, 2012; 10.1016/j.cmet.2012.09.014.
  4. Wang F, Knutson K, Alcaino C, Linden DR, Gibbons SJ, Kashyap PK, Grover M, Oeckler R, Gottlieb PA, Li HJ, Leiter AB, Farrugia G, Beyder A. Mechanosensitive ion channel Piezo2 is important for enterochromaffin cell response to mechanical forces. J Physiol. 595: 79-91, 2016; 10.1113/JP272718 PMCID: PMC27392819.
  5. Alcaino C, Knutson K, Gottlieb PA, Farrugia G, Beyder A. Mechanosensitive ion channel Piezo2 is inhibited by D-GsMTx4. Channels. 11: 245-253, 2017; 10.1080/19336950.2017.1279370.
Alecia Alto attends the University of Minnesota Rochester in Rochester, MN. She is a 2017 Undergraduate Summer Research Fellow (UGSRF) doing research in Dr. Arthur Beyder’s lab at Mayo Clinic in Rochester, MN.  After graduating with a bachelor or science in health sciences, she is considering a career as a physician scientist.

 

Frozen In Time: The Story of a Tree Frog and its Potential to Help Humans Freeze Their Organs, and Possibly Themselves

I know what you’re thinking. What in the world does a tree frog have to do with humans freezing their organs? And yes, before beginning my undergraduate research project, I too would likely be thinking this exact same thought. But nature allows animals to do a very funny thing: adapt according to their environment. And the gray tree frog that you are itching to read about in the following paragraphs has adapted in one of the most extreme ways found in the animal kingdom: it has the ability to freeze itself and survive.

Credit: John White, University of California, Berkeley

My 2017 summer research project, funded through the American Physiological Society, is focused on Cope’s gray tree frog, Hyla chrysoscelis. H. chrysoscelis has developed a dramatic strategy to combat the cold during the winter months, known as freeze tolerance. Freeze tolerance permits this tree frog to withstand the cold by allowing it to convert 50-65% of its total fluid body water into ice. During its time spent frozen, Cope’s gray tree frog experiences fundamental changes in its physiology and biochemistry, including the abandonment of:

  • Blood circulation
  • Breathing
  • Nerve conduction
  • Brain activity
  • A heartbeat

Yes, you read that right. It has no heartbeat while it is frozen.

However, as wintertime comes to a close and external temperatures rise, the thawing process begins, vital signs reactivate, and normal life resumes for this frog. My research project for the summer is aimed at understanding a specific cellular process involved in the whole body freezing of H. chrysoscelis, which may provide future scientists with the capacity to not only freeze human organs in the future, but also freeze our entire bodies.

H. chrysoscelis experiences a variety of stresses during freezing and thawing, most of them being osmotic stresses. This is because the majority of the fluid surrounding the frog’s cells freezes, leaving a higher concentration of solutes outside the cell than prior to freezing. To combat this disruption of fluid homeostasis, H. chrysoscelis has evolved a transmembrane protein known as aquaglyceroporin HC-3 that acts as a channel for water and solutes to flow through, which helps to re-establish a fluid homeostatic equilibrium. In non-science terms, this protein, HC-3, helps to keep the cells of this frog from shriveling up like raisins or bursting open like a balloon with too much air.

My hypothesis demands a week long experiment, involving a variety of techniques that enable me to begin with the blood of this frog, fresh from its brachial artery, and shrink all the way down to the molecular level to analyze HC-3, the protein that I suspect to be a vital player in the freezing process. The first part of my experiment requires culturing red blood cells from Hyla chrysoscelis over a period of 48 hours, which in non-science terms translates to: keep 90% of these cells alive over a period of two days. Sounds simple, right? Well, after the first two weeks of failed attempts in maintaining proper viability of the cells in culture, I was left devastated, demoralized, and with numerous flasks of cells contaminated with who-knows-what. Science, without mercy, and in just a period of two weeks, showed me just how difficult it would be to successfully complete my week long experiment by halting me after just the first two days.

As the summer has progressed, I have finally achieved viable cell cultures for the full 48 hours through troubleshooting and controlling for one variable at a time, leading me to the conclusion that a certain reagent I was adding to the media my cells were kept in had been contaminated earlier in the summer. Currently, I am now focusing on more demanding techniques, such as western blotting and immunocytochemistry, while still always encountering new obstacles to overcome and problems to solve. New to research, and even newer to researching full time, I have come to enjoy the independence and autonomy that I’ve found in the laboratory. Every day is a new opportunity, and a new challenge, demanding my full attention and effort to successfully complete my experiments for the day. Through the few successes and many failures of my summer of research thus far, I have gained a deeper appreciation for science and continue to aspire to engage in scientific research throughout the rest of my professional career.

References

  1. White, John. “Hyla chrysoscelis, Cope’s Gray tree frog.” University of California, Berkley Regents (2006).
Dante Pezzutti is a rising senior majoring in Pre-Medicine at the University of Dayton in Dayton, Ohio. He is a 2017 Undergraduate Summer Research Fellow (UGSRF) working in Dr. Carissa Krane’s laboratory at U.D. over the summer. Dante’s fellowship is funded through the American Physiological Society. After graduation, Dante plans to attend medical school to become a physician, and also aspires to engage in bench-to-bedside translational research as a medical doctor.
Research Project Narrated for My Parents

My summer research project, which will span much of the next year, revolves around Classical Ferroportin Disease. Firstly, before explaining the minutia of Classical Ferroportin Disease it is important to know what ferroportin is; ferroportin is an iron transport protein that is responsible for exporting iron out of cells. Classical Ferroportin Disease is a loss of function disease, meaning that a mutation has occurred in the protein that makes ferroportin less efficient [1]. Seeing as though ferroportin is exporting iron out of cells, patients who have Classical Ferroportin disease can get iron into their cells, but their cells struggle to get iron back out of the cells. This ferroportin malfunction results in a net accumulation of iron in cells above what is physiologically healthy; macrophages of the liver and spleen become particularly iron overloaded because they metabolize a lot of iron. The disease can present clinically as anemia because the body is unable to acquire the adequate iron it needs for red blood cell production since the iron is being sequestered in the liver and spleen macrophages with no way to be properly exported [2]. The disease can also cause diseases of the liver such as fibrosis and cirrhosis because iron overload of liver cells is detrimental to liver health and functioning.

It is known that Classical Ferroportin exhibits autosomal dominant inheritance; my summer research project is interested in shedding light upon how Classical Ferroportin Disease is inherited via an autosomal dominant route. There are two models to explain the autosomal dominance that classical ferroportin disease exhibits. One model is a haploinsufficiency model, the other is a dominant-negative effect model [1]. These terms may sound complex but they are fundamentally simple. The haploinsufficiency model can simply be thought of in terms of dosage. When you make a cake the recipe may call for five eggs, if you were to only put one egg into the cake, said cake would turn out abysmally. Haploinsufficiency is exactly the same way; you inherit one copy of ferroportin from each parent. You need two good copies to be a well baked cake and make enough functional ferroportin protein, but if you only get one good copy and one mutant copy then you will be an abysmal cake with not enough functional ferroportin to properly export iron. All in all, in the haploinsufficiency model you don’t have enough functional ferroportin to keep up with the export rates needed to be healthy. The second model, the dominant-negative effect, is a little more complex but still fundamentally simpler than the name implies. This model is best envisioned as an arts and crafts project. Your task is to make a sphere out of two objects and super glue. THE TWO OBJECTS MUST COME TOGETHER TO CREATE THE PRODUCT. In the working art studio you’d be provided with two hemispheres and super glue; you would with ease construct the sphere and happily go about your day. In the art studio run by people lacking infant geometry knowledge, you are provided with one hemisphere, one cylindrical object, and super glue. No matter how hard you try with the materials provided you cannot construct the desired spherical object. Out of frustration you glue the block to the sphere permanently and leave full of rage to throw the mushroom shaped object at a wall.  The complete sphere, representing a healthy person without ferroportin disease, can be made when you are provided with two good hemispheres, one healthy ferroportin gene from each parent. The second art model, where the person is forced to construct a mushroom shaped object, represents Classical Ferroportin Disease via a dominant-negative effect. The person was forced to glue the hemisphere, a healthy ferroportin gene product, to a cylinder, a mutant ferroportin gene product, creating a non-sphere and thus a ferroportin protein unable to export iron. They key distinction between the haploinsufficiency model and the dominant-negative model is that in the haploinsufficiency diseased state you simply do not have enough of the good product, whereas in the dominant negative model the bad product hinders the good product from working all together. Overall my research project revolves around trying to figure out which model Classical Ferroportin Disease follows and my tools to do so are Xenopus laevis oocytes, human mRNA injections, confocal fluorescent microscopy, and scanning electron microscopy. This project falls into a huge sector of molecular biology and physiology that is cellular transport and it will aid in the treatment and understanding of one type of a family of diseases that fall under the name of Hemochromatosis.

Realities of Research

Experimental design is key; hard work pays handsomely in the form of pretty graphs; don’t start to genotype at 3 PM’ and experiments will be repeated a frustrating amount of times until those variance bars look pretty.

The importance of well thought out experimental design became a frustrating reality about 5 weeks into my project. I had been tirelessly performing experiments for weeks in a way that could only be analyzed via a one-way ANOVA. An ANOVA is a commonly used statistical test used to analyze data; all statistical tests require an individualized experimental design in order to be analyzed via a specific test. One day at lunch my PI and I were discussing my progress and he looks at me worriedly and says “How have you been setting up your experiments and how have you been analyzing the data?” He then went on to inform me that my experimental design had a critical weak point and that a one-way ANOVA would not provide the insight needed to make a strong claim on my hypothesis. So the rest of the lunch was spent redesigning my experimental approach so that I could run a two-way ANOVA, yet just another statistical test. This taught me that experimental design is worth troubling your mind over for a long while because good data is meaningless unless it is masterfully collected with a specific statistical analysis in mind.

A lab reality surprise I encountered was how rewarding good data and graphs can be. I grew up playing baseball and soccer and I excelled at them, therefore I know the feeling of hitting a homerun and scoring a goal. They are full body highs encompassed with an adrenaline surge and confidence boost. I was surprised how close to those feelings I got when I got my first very usable data set and graphs. It’s very comparable to sports because you train in much the same methodical way for so long so that when you finally see results it feels amazing. Hard work does in fact pay handsomely with good data and graphs.

Another lab reality is planning your day saves your dinner date. I learned this the hard way multiple times when I started an experiment, genotyping, or prep way too late in the day.  It has become increasingly apparent that planning is your best friend in the lab. Fifteen minutes of thoughtful planning at the start of the day may save you hours each day. AKA don’t hold off on very pressing genotyping until 3 PM unless you plan to be in the lab until 9 PM.

Lastly, a frustrating lab reality is that some experiments will have to be repeated so many times you that start to worry you’ll be running the same experiment with the same experimental groups forever, or even worse you fear they will never give you interpretable data…. gasp. This is a frustrating reality that can never truly be realized until you have suffered first-hand through running the same experiment for weeks on end. It always gets better though; see baseball homerun feelings above.

Life of a Scientist

The most striking part of the day to day of a scientist is how social it is. I always assumed scientists were reclusive and withdrawn. I was wrong, outright wrong. The lab I work in is bustling with life. Intellectual exchange and idea bouncing occurs incessantly as we pipet about running assays. We come together at lunch and effortless fill the air with debate and jovial disputes. But the chit chat goes beyond academics, which I was most surprised about. Beyond bouncing ideas off each other and discussing relevant academic papers, a plethora of enticing conversations about extracurricular activities also buzz loud. We talk about music, the best coffee places, the trendiest foods, sports, and philosophy. Science is inescapably social in a beautiful way. Another thing I love about being a scientist is that your schedule is decently malleable as long as you get plan well. You are able to catch lunch with a friend or leave a little early one day if you make up for that time and work somewhere else; I am trying to get at the fact that science is not a strict 9-5 because as long as you get your work done no one is going to hound you about when you do it. This is a huge plus because it plays very well into an adequate work-life balance.

References

  1. Hal Drakesmith, Elizabeta Nemeth, Tomas Ganz, Ironing out Ferroportin, Cell Metabolism, Volume 22, Issue 5, 2015, Pages 777-787
  2. Mayr, Roman et al. “Ferroportin Disease: A Systematic Meta-Analysis of Clinical and Molecular Findings.” Journal of Hepatology5-3, 2010
John Bonamer is a senior majoring in Medical Sciences at the University of Cincinnati in Cincinnati, Ohio. He is a UGSRF fellow working in Dr. Bryan Mackenzie’s lab at the University of Cincinnati. John’s Fellowship is funded by the APS. After graduation John plans to pursue an MD/PhD with special interest in Urology.
My Summer Experience Observing Changes to Sarcomeres with Titin Regulation

Muscles are comprised of repeating units called sarcomeres, which can be seen as bands in myofibrils. Prior research has shown that the consistency of the lengths of these bands correlates to muscle strength – the more consistent the bands, the stronger the muscle. My project focuses on the relationship between the consistency of these sarcomeres and titin, the longest known protein which is proposed to maintain sarcomere length. Our lab developed an inducible skeletal muscle specific Bmal1 knockout mouse that demonstrates a to a longer titin isoform. By altering titin and comparing the variance of lengths of these sarcomeres to a control group, I hope to demonstrate a correlation between sarcomere length and the expression of titin. My project takes a histological approach to study changes in sarcomere length and A-band centrality in the skeletal muscle by staining for alpha actinin and myosin. These images are overlaid and peak fluorescence is measured. Knowledge from this experiment would contribute to knowledge about titin and could eventually lead to more targeted drugs or therapies. Results of my experiment thus far indicate that there is a trend toward a longer sarcomere length and no significant change in variance, but there are still other stains and trials I would like to do to strengthen my findings. If my hypothesis is supported then my project would support the assertion that titin has a structural role in the sarcomere.

The myosin stain allowed me to see if there were any differences in the location in the A-band in relation to the Z-line.

The alpha actinin stain labels the z-lines, allowing the sarcomere to be visualized and measured.

Realities of Research

Research in Dr. Esser’s lab was very different from what I had anticipated. Generally, people think of scientific research as solitary, but what I have found is there is a lot of collaboration in academic research. When we encountered an error in the RNA sequencing program of the model, and we reached out to other people at the university in order to help understand and correct this error. Additionally, we borrowed equipment such as the cryostat (a machine used to cut muscle at cold temperatures) and microscopes from other labs. Lab meetings were also a useful part of my experience because they allowed for brainstorming when I faced challenges. In addition to the RNA sequencing obstacle, there was also a problem with a control antibody stain because there was fluorescence in an unexpected region. After discussing the problem, we came up with the hypothesis that the secondary stain for one of the regions was attaching to the conjugated antibody of another region, which was why we were seeing the unexpected fluorescence. To correct this, I applied the antibodies in two steps instead of combining them. All of these obstacles were easier to overcome because I had people around to assist me, an invaluable asset that I did not expect.

Day-to-Day of a Scientist

I look forward to coming in to the lab because I like the challenge of overcoming the unforeseen obstacles. Designing ways to solve problems is definitely my favorite part of working in a lab. The production of ideas about how an experiment or procedure should work and testing the methods is exciting. While these obstacles can be a nice challenge, they can also become tedious. The problem with the RNA sequencing was especially frustrating because it was out of our hands. Despite these complications, my research was overall very enjoyable. I am very appreciative of the APS for allowing me to pursue my interests this summer, and would encourage others to do a program because the experience was very rewarding.

Joseph Mijares, also known as Robby, is a junior at the University of Florida in Gainesville, FL. He is grateful to be a 2017 Undergraduate Summer Research Fellow in Dr. Karyn Esser’s Lab at the University of Florida.  Robby plans on applying to several MD and MD-PhD programs after his junior year, with hopes of attending after graduation.