Tag Archives: undergraduate summer research

Sex Differences in Asthma

Research Project

This summer, I was involved in research with Dr. Silveyra at Penn State College of Medicine. We looked at microRNAs, which are small pieces of RNA involved in various processes in the cell, including regulation of how much of a certain protein is made. MicroRNA-106a is a microRNA that influences asthma in mouse lung immune cells, which cause inflammation. miRNA-106a does so by preventing an anti-inflammatory protein, IL-10, from being made. Because we knew that more adult females have asthma than adult males, we hypothesized that miRNA-106a would also be expressed differently between males and females. To investigate this hypothesis, we allowed several mice to develop asthma by repeatedly exposing them to house dust mites for 5 weeks. We then exposed the mice, along with additional mice that did not develop asthma as a control, to either ozone or filtered air for 3 hours. After that, we harvested the lungs and extracted the RNA from the tissue. We separated out miRNA-106a from the total RNA and determined its relative amount compared to the amount of another RNA that is always present in cells. We discovered several differences in how much miRNA-106a was present between males and females within treatment groups. Along with other factors, these differences in miRNA-106a levels in mice may play a role in the differences seen in human asthma, which affects about 8% of all people in the US. It could even lead to a new treatment for asthma that is specific to men or women.

Realities of Research

Doing research full time in a lab was more enjoyable than I expected. Each day looked different for me in terms of the tasks I needed to complete. I learned several new techniques, including RNA extraction and real time PCR, and I used these techniques very frequently throughout the summer. During the experiments, we had to repeat our real time PCR plates several times due to a lot of random error, and we had to repeat the RNA extraction for several samples because we didn’t get the amount of RNA we wanted. It was a long process, with many setbacks, but we finally got results from our experiments. We did see differences in the amount of miRNA-106a between males and females, but we did not see the differences that we expected. Because of this, we understood that miRNA-106a may not be causing all the differences seen between men and women with asthma, but other factors may cause the differences as well.

Life of a Scientist

As said earlier, the part I most appreciated about my summer as a scientist was that each day brought new tasks. However, experiments can take a long time to complete, with several setbacks and problems that need to be addressed along the way. There is no instant gratification; I had to work for every bit of data I had. But it is still rewarding when one can publish or present research at a conference, and other scientists listen and ask questions. To be a scientist, one must know how to work as a team and communicate clearly so everyone understands what their role is in the lab. I felt that my team was very good at doing that, and our lab environment was better for it. My lab team was a huge help to me in teaching me techniques, assisting me in carrying them out, and helping me to make sense of my results. All in all, the life of a scientist, though with many obstacles, is rewarding, and I would encourage anyone to check it out.

Rachel Steckbeck is a junior at Messiah College in Mechanicsburg, PA. She is a 2018 Undergraduate Summer Research Fellow (UGSRF), and worked in Dr. Silveyra’s lab at Penn State College of Medicine in the summer of 2018 for 10 weeks. UGSRF is funded by the American Physiological Society. In the future, Rachel hopes to attend medical school and work at a local hospital or practice.
Ways to reduce Myocardial Reperfusion Injury

Research Project

This summer I am working in the Department of Anesthesiology at Stanford University. The main aim of my assigned project is to study the role of TRPV-1 (Transient Receptor Potential Vanilloid-1) in myocardial reperfusion injury. TRPV-1 is considered as a main channel that mediates cellular responses to different stimuli such as pain and heat. Regulation of TRPV-1 is important because it controls mitochondrial membrane potential and reperfusion injury. Modulation of TRPV-1 interaction site with calcineurin is essential to reduce the injury. My main objective for the summer is to study the effect of previously synthesized peptide drugs on the interaction of TRPV-1 with calcineurin. We hope that this project will help to reduce damage from cardiac reperfusion injury since it is responsible for up to 50% of myocardial infarct size.

Realities of Research

Stanford was my dream and I spent this summer in this dream. As I had an inadequate knowledge in the real physiological lab before, I was lucky to have an opportunity to experience many new techniques and skills. The most captivating experience was rats. Unfortunately, my home university does not have animal resources and students are deprived of working with animals. Hence I am planning to pursue PhD next year, I was very anxious about my incompetence and inadequacy. Luckily, my assigned project for the summer required me to isolate fresh cardiomyocytes from adult rats and perform different cell death assays on them. I was very excited about upcoming surgeries but forgot about one significant nuance- rats itself. Rats were afraid of me, so do I. Ironically, my fear made them even more stressful, so eventually I ended up with zero cells. I repeated surgeries dozens of time forcing myself to be braver and bolder. After continuous failure, I got used to them and I am obtaining enough cells these days to continue further. Now, I understand that any skill can be acquired with determination and perseverance unless it is highly desired.

Life of a Scientist

People do the science for many reasons. For some, it gives a prestige and high social status, but for some, it gives a hope for the better future. When I first told my family a decision of becoming a scientist and pursuing an academia, I was warmly welcomed with benevolent joy about my safe future. For sure, being a scientist seems to be the most auspicious occupation for women hence it requires neither intense physical nor intellectual activity. That was when my first fairytale illusions about the science career have started. Years later, after experiencing both flukes and failures I understood that life of a scientist indeed requires something very special- struggle with Yourself.

Coming to the lab every day and continuously repeating the same routine experiments over and over again can frustrate even the nerds. But for me, this is a real beauty of the science. When the experiments don’t work for weeks and the moment when you start quitting, surprisingly everything begins to work by lucky chance. You get happy and renew the cycle from the very beginning without even a tiny memory of those hapless days. Ironically, the cycle goes on further and combo of successful/unsuccessful experiments continue to form a chain reaction in your life. I agree with Mare Curie that “A scientist in the laboratory is not only a technician: he is also a child placed before natural phenomena which impress him like a fairy tale”. If you still believe in miracles and get amazed by the natural laws, follow your passion and just chase the science!

 

Reference

Hurt, C.M., Lu, Y., Stary, C.M., Piplani, H., Small, B.A., Urban, T.J., Qvit, N., Gross, G.J., Mochly-Rosen,D.,  Gross, E.R. Transient Receptor Potential Vanilloid 1 Regulates Mitochondrial Membrane Potential and Myocardial Reperfusion Injury. J Am Heart Assoc 5(9), 2016.

Zhuldyz Zhanzak is a senior majoring in Molecular Biology at Nazarbayev University, Kazakhstan. She is a 2018 Undergraduate Research Excellence Fellowship (UGREF) Fellow working in Dr. Eric Gross lab at Stanford University School of Medicine, CA. Zhuldyz’s fellowship is funded by the APS. Following the graduation, Zhuldyz plans to pursue PhD in immunology and follow a career as a scientist in the academia.
Targeting NHE3 in the Hfe mouse model of hemochromatosis

Research Project

Over the summer I have been working in Dr. Bryan Mackenzie’s lab at the University of Cincinnati College of Medicine in the Pharmacology and Systems Physiology department. My focus has been on finding novel therapeutic targets in the treatment and/or prevention of iron overload disorders. Hemochromatosis is just one of several common hereditary disorders that result in iron overload and subsequent liver disease, heart damage, and a slew of other health disorders across several body systems. Since there is no regulated mechanism for iron excretion, iron homeostasis of the body is regulated by iron absorption. The main mechanism by which non-heme iron (the majority of iron humans consume from their diets) is absorbed from the small intestine is the divalent metal ion transporter-1 (DMT1) (3). DMT1 is already a validated therapeutic target in hereditary hemochromatosis. More recently, the Mackenzie lab identified a role for the Na+/H+ exchanger-3 (NHE3) in intestinal absorption of non-heme iron by DMT1.

 

Intestinal iron uptake (redrawn from reference 1)

The Mackenzie lab demonstrated that NHE3 generates the driving force for iron absorption via DMT1 by transporting protons into the small intestine (2). My project this summer has built upon this prior knowledge using an Hfe knockout mouse model. Rendering this single gene inoperative in mice results in a very good model of iron overload. We are then able to test the hypothesis that iron overload in the Hfe KO mouse can be corrected by ablation of NHE3. The idea is that without NHE3, the driving force for DMT1-mediated iron uptake is reduced, thereby reducing iron absorption. If the data supports this hypothesis, we can conclude that ablating NHE3 prevents iron loading in the Hfe mouse model of hereditary hemochromatosis. Even further, we can reveal intestinal NHE3 as a potential therapeutic target in the treatment of iron overload.

Realities of Research

Throughout the summer I have learned countless basic research techniques and have mastered various skills that are good to possess in the Mackenzie Lab. I have learned how to ween and care for our colonies of mice. I now know how to genotype them using tail clippings and polymerase chain reaction (PCR). This step is especially important because it determines which mice are our experimentals, or the mice we collect blood and tissue samples from on experiment days. I was placed in charge of the mouse experiments, learning how to effectively set-up and delegate responsibilities. Being my first full-time experience in a research lab, I appreciated this opportunity to take charge and have ownership over such a large task. I also had the task of running serum iron assays and non-heme iron assays on freezers full of samples that have been collected. This task proved to be challenging and slightly stressful at times, as I found errors in our established assay protocols and analytical procedures. For example, some mice had very little blood collected on experiment days, meaning I had very little serum to run the assays. When the protocols had detrimental errors, I felt I had wasted irreplaceable serum at times. Luckily, I have corrected our protocols and have finally organized/assayed most of our samples. I have also not only learned how to operate a software program called SigmaPlota to analyze my data, but have learned how to edit our labs established transformations within this software. This summer full of learning techniques, troubleshooting, and analyzing my data has been an invaluable experience. I have transformed into a proficient, independent, and productive lab member in just a few months’ time.

Life of a Scientist

I have enjoyed my day-to-day life in the lab this summer. I was pleased to learn that a 9-to-5 workday is certainly possible, with proper planning. I also understood that some late nights/early mornings are unavoidable, especially when things didn’t go as planned and required a little extra time. I discovered that the environment in the lab was much more relaxed than I anticipated. I was able to control my day to day activities with little intervention, unless I needed assistance or extra guidance. This assistance from other lab members is something I really appreciated. Although everyone has their own projects and own experiments to run, a helping hand was always there when I needed it. I liked this collaborative, team-based environment and believe it greatly increased all I was able to learn in the lab this summer. The only con in the life of a scientist that I can think of is the down time that sometimes occurs, as I’m the type of person that likes to stay busy. Luckily, I was able to fill my time by reviewing relevant literature in the field or discussing my progress with other lab members. Overall, I really enjoyed getting to experience the life of a scientist for a summer and look forward to finishing up my project/exploring new directions in the lab.

 

References

  1. Mackenzie B, Garrick MD. Iron Imports. II. Iron uptake at the apical membrane in the intestine. American Journal of Physiology-Gastrointestinal and Liver Physiology289, 2005.
  1. Shawki A, Anthony SR, Nose Y, Engevik MA, Niespodzany EJ, Barrientos T, Öhrvik H, Worrell RT, Thiele DJ, Mackenzie B. Intestinal DMT1 is critical for iron absorption in the mouse but is not required for the absorption of copper or manganese. American Journal of Physiology-Gastrointestinal and Liver Physiology309, 2015.
  1. Shawki A, Engevik MA, Kim RS, Knight PB, Baik RA, Anthony SR, Worrell RT, Shull GE, Mackenzie B. Intestinal brush-border Na /H exchanger-3 drives H -coupled iron absorption in the mouse. American Journal of Physiology-Gastrointestinal and Liver Physiology311, 2016.
Sydney Stone is a junior majoring in Medical Sciences at the University of Cincinnati in Cincinnati, OH. She is a 2018 Short-Term Research Education Program to Increase Diversity in Health-Related Research Fellow (STRIDE) working in Dr. Bryan Mackenzie’s lab at the University of Cincinnati College of Medicine. Her summer of research was supported by APS and a grant from the National Heart, Lung, and Blood Institute (Grant #1 R25 HL115473-01). Upon graduation, Sydney plans to pursue an MD/PhD or MD/MS in order to become a physician scientist. She hopes to combine proficiency in physiology with invaluable access to patients in order to follow her passion for preventing and curing human disease.
Life Behind the Lab Coat: Part 2

Are you curious about the world of science? Do you crave to find new and exciting things about physiology? Well, I do!  Last summer, I had the opportunity to work in a research lab at the University of South Florida Morsani College of Medicine. Furthermore, I was able to renew my APS STRIDE fellowship and continue to work in Drs. Jessica Dominguez Rieg and Timo Rieg’s lab.

Research Project

During my time at the lab, my research project expanded on my work from last summer involving a transporter called the sodium/hydrogen exchanger (NHE). NHE is used to exchange sodium for hydrogen across a cell’s membrane.  There are several different isoforms of NHE; however, my project focused on the isoform NHE3. When NHE3 is genetically deleted from a mouse, called a whole animal knockout, they absorb or reabsorb sodium from the intestine or kidney. Consequently, the NHE3 knockout mice have severe diarrhea, low blood volume, and low blood pressure1. Furthermore, since NHE3 transports hydrogen, it also plays an important role in acid-base regulation. Moreover, NHE3 is also significant for balancing fluid and the regulation of blood pressure.  Last year, my studies concluded that the inducible deletion of NHE3 only in the intestine causes fluid accumulation in the intestine and metabolic acidosis.

This year, my research project remained focused on the role of intestinal versus renal NHE3 and its effects on phosphate homeostasis and blood pressure regulation. Moreover, my research is significant as nothing like this has been done before!  Additionally, there are currently drugs on the market that block NHE3, one example is Tenapanor, which helps to lower blood pressure or high blood phosphate levels; however, the mechanisms of how they work are unclear. As of last year, I saw significant changes in acid-base regulation. Hopefully, this year I will be able to determine how NHE3 affects phosphate homeostasis. Furthermore, I hope I am able to produce more data in order to help answer these ongoing questions about NHE3 and its effects on phosphate homeostasis and blood pressure regulation. Thus far, I have been conducting physiological experiments to determine changes in fluid and food intake, collecting blood and urine for further analysis.  My project is still being conducted but I am excited to see the results!

Life of a Scientist

I love being part of a research lab! Working with people who share the same passion and love discovering new things is amazing! Although I love doing research, the hours can be long. Having to work 8 plus hours a day during the week can be tiring and then having to come in again on the weekends, but I’ll tell you it’s all worth it! One thing I learned is experiments are a process and take time. Last year I thought working in a lab would be similar to being in the labs at the university that were required for your major, but they’re not. Even though they are very different, I still loved every moment I came into work! I am so thankful for this opportunity that APS and my mentors allowed me to expand my knowledge and skills. I am forever grateful for this experience.

 

Citations:

  1. McDonough AA. Mechanisms of proximal tubule sodium transport regulation that link extracellular fluid volume and blood pressure. Am J Physiol Regul Integr Comp Physiol 298: R851–R861, 2010. doi:10.1152/ajpregu.00002.2010.
Alexandria Valdez is a Senior majoring in Biomedical Science and minoring in Psychology at the University of South Florida in Tampa, FL. Alexandria is a 2018 second year Short-Term Research Education Program to Increase Diversity in Health-Related Research (STRIDE) Fellow working in Dr. Jessica Dominguez Rieg and Dr. Timo Rieg’s lab at the University of South Florida Morsani College of Medicine in Tampa, FL. Alexandria’s fellowship is funded by the APS and a grant from the National Heart, Lung and Blood Institute (Grant #1 R25 HL115473-01). Upon graduation, Alexandria wishes to pursue a career in biomedical science as a research scientist.
The Fascinating Relationship Between Inflammation and Respiratory Plasticity

Research Project

I am grateful to say that this summer I had the opportunity to work with Dr. Irene C. Solomon at Stony Brook University, in the Department of Physiology and Biophysics, as an awarded fellow from the American Physiological Society’s Short-Term Research Education Program to Increase Diversity in Health-Related Research (STRIDE) Fellowship. Under the mentorship and guidance of Dr. Solomon, I was able to explore the scientific field of neuroinflammation and the underlying neural mechanisms controlling our central respiratory system.

Figure 1. Intermittent Hypoxia (IH) can induce respiratory motor plasticity, by increasing phrenic motor neuron and diaphragm activity

Acute Intermittent Hypoxia (AIH), also known as the delivery of low levels of oxygen in repetitive bouts, has been demonstrated to alter neural ventilatory control, and therefore, elicit different physiological changes within the respiratory system that can persist into normoxia (also otherwise known as normal O2/CO2/N2 levels).[1] These alterations are more formally known as neural plasticity, which is the nervous system’s response to varying, natural stimuli and/or the initiation and progression of diseases, injuries, and pathology.[2] What that means is that the nervous system attempts to compensate for the lower levels of oxygen by increasing the expression of neurotransmitters like serotonin and adenosine, which then affects the upregulation of specific growth/trophic factors within the Gq and Gs pathways involved in eliciting neural plasticity. More specifically, the G protein coupled receptors (GPCR), serotonin 5HT2 receptors and adenosine A2A receptors, are being targeted within the Gq and Gs, respectively.[3] Although there seems to be some interplay between these two pathways, the Gq pathway and the activation of the serotonin 5HT2 receptors appear to play a larger role in stimulating the physiological changes found during and after AIH.[4] The physiological changes that follow (i.e.increased phrenic motor neuron and diaphragm activity), are utilized within the central respiratory system to essentially breathe “better,” or in other words, consume more oxygen, in such hypoxic conditions.

Additionally, recent evidence from rodent studies, which have implemented different protocols/models for the acute administration of bacterial endotoxin lipopolysaccharide (LPS) — a toxic substance bound to bacterial cell wall, usually causing inflammation in different regions of the brain when

Possible mechanism, including the growth/trophic factor, BDNF, for motor plasticity following AIH in rats and humans

released via bacterium rupture or disintegration[1] — indicates that inflammatory processes can also alter ventilatory control, and subsequently, affect the level of respiratory plasticity that can be achieved. The interactions between LPS-induced inflammation, and accompanying neuroinflammation, and various aspects of ventilatory control, however, are not well understood. It is important to understand how LPS affects the respiratory system since LPS-induced neuroinflammation remains to be a compounding factor of many diseases, such as sepsis, acute lung injury (ALI), and/or acute respiratory distress syndrome (ARDS)[2], and serves as a cause of mortality across a wide variety of respiratory malfunctions (e.g. previously mentioned, OSA). Therefore, the fundamental hypothesis guiding this study is that inflammatory processes can impair multiple aspects of central respiratory neural control, resulting in a reduced ability to adequately and appropriately compensate for disease/pathology. The overall goals of my study are to (1) characterize and quantify the effects (basal ventilatory and respiratory-related neural activities) of moderate (11%O2) AIH in spontaneously breathing male rodents to the 2-hit LPS-induced neuroinflammation model, in which the rats will endure 5 cycles of 3 minutes of 11% O2, 2% CO2, and 87% N2 on and off, and (2) to characterize and quantify the chemical control of breathing via immunohistochemical analyses of various elements from collected tissue samples (e.g. respiratory brainstem cytokines expression, along with neural markers from the Gq pathway). 

Realities of Research

In the beginning of my fellowship, I spent a great deal of my time acquiring new methodological techniques and learning surgical procedures often used in conducting the physiological experiments that are run in lab. Some of the essential skills included establishing EMG signals using tongue and diaphragm electrodes, which is short for electromyography and is an electrodiagnostic medicine technique for evaluating and recording the electrical activity produced by skeletal muscles and the nerve cells that control them (motor neurons)[1], giving intraperitoneal injections (IP), intratracheal injections (IT)., and subcutaneous (sc) injections, conducting femoral and jugular cannulations, administering i.v. drug/saline supplements, and collecting tissue samples that were stored in liquid nitrogen. After acquiring the necessary skills, I was able to spend some time developing a thorough protocol that would help outline the experiments I would be running, and determine what my main hypothesis would be based off of previous findings from the lab and publications I read. I was also given the opportunity to develop my presentation skills as I would prepare and give a presentation on a paper during our bi-weekly lab meetings with the Collins Lab, who we extensively work with, and engage in meaningful conversations on new insights my fellow peers would bring up. I am currently going the data analysis of the rats’ BPM frequency and cyclic height (amplitude) for signs of either an increase or decrease in respiratory motor plasticity post-hypoxia. Immunohistochemical staining of the tissue samples collected will be used to further analyze respiratory brainstem cytokines expression for signs of neuroinflammation and potential decreases in neuroplasticity.

Life of a Scientist

Throughout my experience at the Solomon Lab, I have come to realize that the strange and mysterious world of scientific research is not one of only cut-throat competition and solitude, but a juxtaposition of individualized responsibilities and teamwork. There is something about research that, oddly enough, reminds me of my favorite childhood cartoon show, Scooby Doo. Just like how the Mystery Gang goes around in their fruity van looking for clues to solve the case and figure out the culprit of the crime, a research lab — surprisingly — works in a similar manner. See, being a part of the journey in trying to solve the big mystery is a team effort, unlike the stereotypical image most people conjure up in their minds of an old, crazy gray-haired scientist mixing chemicals together in an underground lab. Just as every member in the Mystery Gang was important to cracking the case, all members in a lab must be willing to cooperate and take on the smaller assignments in order to contribute to the larger picture and find the missing clues to the puzzle. And that’s what I love so much about scientific research: the camaraderie that forms among peers in a lab — sometimes even extending beyond the doors of your very own lab and with the people of the lab across yours. Overall, I am not only grateful for receiving the opportunity to delve deep into the world of physiology and neurobiology research, but also for the opportunity to meet such fantastic people who have helped me grow as a scientist and as an individual.

References

  1. Moya EA and Powell FL. (2018) Serotonin and Adenosine G-protein Coupled Receptor Signaling for Ventilatory Acclimatization to Sustained Hypoxia. Front. Physiol. 9:860. 06 July 2018. DOI: 10.3389/fphys.2018.00860
  2. Fuller, D. D., & Mitchell, G. S. (2017, January). Respiratory neuroplasticity – Overview, significance and future directions. DOI: 10.1016/j.expneurol.2016.05.022. Retrieved from https://www.ncbi.nlm.nih.gov/pubmed/27208699
  3. Figure 1. Dale, E. A., Mabrouk, F. B., & Mitchell, G. S. (2014, January 29). Unexpected Benefits of Intermittent Hypoxia: Enhanced Respiratory and Nonrespiratory Motor Function. Retrieved from https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4073945/figure/F3/
  4. Figure 2. Dale, E. A., Mabrouk, F. B., & Mitchell, G. S. (2014, January 29). Unexpected Benefits of Intermittent Hypoxia: Enhanced Respiratory and Nonrespiratory Motor Function. Retrieved from https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4073945/figure/F5/
  5. Moya EA and Powell FL. (2018) Serotonin and Adenosine G-protein Coupled Receptor Signaling for Ventilatory Acclimatization to Sustained Hypoxia. Front. Physiol. 9:860. 06 July 2018. DOI: 10.3389/fphys.2018.00860
  6.  Moya EA and Powell FL. (2018) Serotonin and Adenosine G-protein Coupled Receptor Signaling for Ventilatory Acclimatization to Sustained Hypoxia. Front. Physiol. 9:860. 06 July 2018. DOI: 10.3389/fphys.2018.00860
  7. Britannica, T. E. (2014, November 18). Endotoxin. Retrieved from https://www.britannica.com/science/endotoxin
  8. Hong Chen, Chunxue Bai & Xiangdong Wang (2010). The value of the lipopolysaccharide-induced acute lung injury model in respiratory medicine. Expert Review of Respiratory Medicine, 4:6, 773-783, DOI: 10.1586/ers.10.71
  9.  Electromyography (EMG). (2018, May 05). Retrieved from https://www.mayoclinic.org/tests-procedures/emg/about/pac-20393913

Rashi Bhatt is a sophomore majoring in Computational Neuroscience and Public Health, with a minor in Computational Medicine at Johns Hopkins University in Baltimore, MD. She is a Short-Term Research Education Program to Increase Diversity in Health-Related Research (STRIDE) Fellowship working in Dr. Irene C. Solomon’s lab at Stony Brook University in Stony Brook, NY. Rashi’s fellowship was funded by the APS and a grant from the National Heart, Lung and Blood Institute (Grant #1 R25 HL115473-01). After graduation, she plans on attending Johns Hopkins Bloomberg School of Public Health to receive her MPH and hopefully go on to attend medical school, and specialize in neurosurgery and neuro-imaging.

Is Sodium Nitrite Supplementation the Solution to Aging Blood Vessels?

Research Project

With a quickly aging US population, research to help us live a longer life is on the forefront. Cardiovascular disease is the leading cause of mortality in the US, so finding ways to decrease cardiovascular disease risk could be the answer. At the Integrative Physiology of Aging Laboratory (IPA Lab) at the University of Colorado Boulder, we focus our research on finding ways to decrease this risk. A recent 5 year clinical research trial tested the effects of a sodium nitrite supplement on cardiovascular disease risk in middle aged and older adults. Sodium nitrite provides the body with a very important signaling molecule called nitric oxide, which specifically targets the blood vessels allowing them to dilate and increase blood flow. Nitric oxide production can decrease as a person ages. Past studies in the IPA Lab indicate that increases in nitric oxide are associated with improvements in the function of the blood vessels, thereby decreasing cardiovascular disease risk (1).  Now that the 5-year study is complete, myself and a team of researchers are analyzing endothelial cells collected from subjects. Endothelial cells are found, on the innermost layer of blood vessels and control dilation and constriction of the blood vessels. We are hoping to find mechanisms that sodium nitrate may be acting through to decrease cardiovascular disease risk in middle age and older adults.

Realities of Research

In a perfect world, research would flow without any issues. There would be no problems, no set backs, and no recollecting data. The reality is that this is impossible. There are always mishaps along the way, but the key is overcoming and problem solving these mishaps. Working in the IPA Lab over the past summer, I’ve learned that imaging endothelial cells is an imperfect art. Even if you complete the procedure correctly there are still some slides that don’t stain as well as others or have no cells on them to analyze. Our research team is constantly trying to improve our technique with endothelial cell staining and we have recently made changes to the collection procedure hoping to provide better staining images for the future.

Life of a Scientist

Over the past summer I have also gained insight on what the real life of a physiology researcher is. At times it can be frustrating if your procedure doesn’t work or you don’t receive the results you were expecting, but that’s part of the job. As a researcher it is essential to develop problem-solving skills to work past these frustrating times and to work as a team of researchers helping each other solve problems in the lab. At the same time though it can be one of the most rewarding careers. When you finally finish a paper and publish in a journal, you feel an overwhelming sense of accomplishment and feel as though you and your team of researchers have made a true impact on the future of science.

After staining the endothelial cell slides, I use a microscope to visualize the immunofluorescence of the cells.

References

  1. Seals DR, Jablonski KL, Donato AJ. Aging and vascular endothelial function in humans. Clin Sci 120(9), 357-75 (2011)
Kayla Woodward is a senior majoring in Integrative Physiology at the University of Colorado in Boulder, CO. She is a 2018 Undergraduate Research Excellence Fellow (UGREF) working in Dr. Doug Seals’ Integrative Physiology of Aging Lab at the University of Colorado in Boulder, CO. Kayla’s fellowship is funded by the APS. After graduation this spring, Kayla plans to apply to medical school pursuing a career as physician and researcher.
My Summer Researching and Learning About Worms and Alternative Medicine

My research project explores the possible future uses of the alternative medicine Astragalus, which is a Chinese herb known for its immune stimulating effects, such as decreasing melanoma tumor growth. I am specifically looking for immune system responses in transparent roundworms called C. elegans. I am treating the worms with different concentrations of the Astragalus medicine, then observing how their immune system pathways have reacted. My research expands the information available on alternative medicines and their validity. This project could lead to further, more specific studies as well as studies on other animals and potentially humans. Astragalus could be proven to be a viable medication for disease or replace current cancer treatments and medications.

Working in a research lab has proved to be very interesting and exciting. I started out the summer unsure of myself and the techniques involved with my project. There were many standard procedures and instruments I needed to familiarize myself with to successfully and effectively perform tests. Over time I became more comfortable and could do most tests and procedures on my own, only after being instructed by my research host, of course. My research was unsuccessful at first as I couldn’t get the desired result from polymerase chain reaction (PCR) with the house-keeping gene actin. We reran the end-point PCR and ran another gel but still had no results. After taking another step back and rerunning the cDNA synthesis with my RNA sample, we finally had a successful PCR result. I know there are many techniques I have yet to master in my research journey.

I was surprised by how long certain experiments took and how much everyday maintenance was required to continue working with the worms. Much of my time was spent counting the worms for survival assays or synchronizing the worms to prepare for a test. Also, there was constant upkeep of Nematode Growth Media plates and E. coli stock. I also had to adjust to working independently on my research as opposed to working in a class. I had to prepare all the materials I needed and go through the steps on my own instead of having everything laid out for me. The best part of research was getting successful results and making progress, the worst part was becoming frustrated when results were undesirable or no results were produced at all.

Alyssa Knudson is a junior at Coe College majoring in Biology and Molecular Biology and minoring in Music and Chemistry. She is a part of the Coe College Cross Country and Track and Field teams. She is also involved in Pre-Health Club, Biology Club, and Intergenerational Connections at Coe and a member of the co-ed service fraternity Alpha Phi Omega. She has participated in a 10 week research program working under Dr. Cassy Cozine the summer after her sophomore year. After college she plans to go on to graduate school and get her degree in Genetic Counseling.
Stressed Rats and a Stressed Undergraduate

Chronic stress leads to a greater likelihood of the development of many conditions including post-traumatic stress disorder (PTSD), anxiety, irritable bowel syndrome (IBS), functional dyspepsia and other gastrointestinal (GI) dysfunction. This indicates a likely rearrangement of neural pathways and regulation, although the mechanisms of how this happens are not yet known. As an APS Undergraduate Summer Research Fellow, I worked for ten weeks under my research mentor Dr. R. Alberto Travagli studying the neurochemical oxytocin’s role in stress adaptation. My project focused on the regulation of oxytocin signals between the brain and GI tract under conditions of chronic stress in rats. In other words, I studied whether oxytocin has a different effect on the brain and gut of rats after they have been stressed.

Following a 5-day stress treatment on each rat, oxytocin was microinjected in the dorsal vagal complex (i.e. the brain area that directly signals the GI tract). The response to these injections on gastric tone and motility in two areas of the stomach were then recorded and analyzed. The research is still ongoing, but we hope to answer a few questions: How does the regulation of oxytocin change after stress adaptation? Does oxytocin work through different neural pathways after homotypic stress (i.e. same stress each day) or heterotypic stress (i.e. different stress each day)? Since females have a greater likelihood of developing GI disorders, do sex/estrogen levels affect the regulation of oxytocin under stress? Although we are still collecting data, I am very excited to see the results when completed and honored to participate in this research!

What surprised you most about working in the lab?

Upon starting this project, I was surprised by how much skill is required to complete the tasks at hand. Although the technology we use to inject oxytocin and record the gastric response is quite advanced, it can easily be faulted by a human mistake. For example, if I did not suture the sensors tight enough to the stomach, the responses were difficult to read and interpret. There was a huge learning curve to carrying out the research day-to-day and then it was another challenge to ensure I was as consistent as possible between animals. Additionally, I was surprised by how much my project changed between the beginning and end of the summer. For example, early on we injected a new pathway-blocker out of curiosity, expecting it to have little to no effect on oxytocin injections. Surprisingly, however, in one treatment group it seems to be blocking the effects of oxytocin. After, we used that pathway-blocker for every animal and its effects may be crucial to our final conclusions.

I am very grateful to the American Physiological Society for providing me this opportunity because it has made me realize how challenging a career as a basic research scientist is! This summer has exposed me to how exhausting, long, and physically demanding lab research can be. But, I love the big-picture parts of research; designing the experiments, analyzing the results, and adjusting when results are not as predicted. It is amazing to work on research that could be part of a bigger solution (i.e. understanding of anxiety/IBS/colonic pain), especially when you can collaborate with other researchers and pool data to come to even more conclusions within each study. However, I will admit that lab research is grueling work and, like the rats, I was a little stressed at times! I look forward to next year’s summer project so that I can experience translational or clinical research and gain a more holistic view of the research world.

Julia Zimmerman is a sophomore majoring in Neuroscience at Swarthmore College in Swarthmore, PA. She is a 2017 Undergraduate Summer Research Fellow (UGSRF) working in Dr. R. Alberto Travagli’s lab at Pennsylvania State University College of Medicine in Hershey, PA, funded by the APS. After graduation, Julia intends to pursue work as an MD-MPH, or MD-PhD, bridging basic research into the clinical world.
Understanding Muscle Force During Cyclic Movements: Does Titin Play a Role?

During cyclic every day movements, such as running, jumping, and walking, our muscles go through cycles of shortening and stretching. While there has been extensive research on muscle function for the last 50 years, there is no current muscle model that can accurately predict natural movements. For example, when active muscle is stretched, it produces more force than expected based on current theories of muscle contraction. Likewise, when active muscle shortens, it produces less force than predicted by current theories. For years, scientists have been measuring properties of muscles under highly controlled conditions. The classic force-velocity relationship shows that force generated by a muscle is inversely related to the velocity of the shortening. However, this relationship changes during natural, more life-like movements. Recent work suggests that for a given velocity, muscle force is higher during cyclic contractions than the traditional force-velocity relationship. My research investigates the role of the elastic protein titin in the force-velocity relationship measured under different conditions. Using a mouse model with a mutation in titin, I conducted in vitro muscle experiments to compare the force-velocity relationship in cyclic and controlled (isotonic) conditions. Hopefully, my results will shed light on titin’s role as a spring in active muscle. If titin truly does store energy like a spring, this could account for the extra force and lack of force in the stretch-shortening cycles. This research will allow us to better understand movement on a whole organism scale, which can prove quite useful in prosthetic design and bioengineering, for example.

Much like the active muscle, doing research in a lab goes through cycles, except instead of stretch-shortening cycles, it is periods of challenge and reward. Some days, you go into lab, collect great data, and leave feeling utterly fulfilled. However, other days, you go into lab and it seems as though you spent your entire day trouble-shooting. Mainly though, our experiments worked and we were able to collect useable data. We have yet to fully analyze our results, but preliminary results seem to support our expectations.

In general, I have found my lab group experience to be very similar to my experience with playing college soccer. Both activities involve a group of people working toward a common goal. While in soccer, your team is working together to win, in the lab, there are many scientists working together to uncover a truth. Collecting and analyzing data is a collaborative effort and, to me, that was the best part of summer research. Working as part of a lab team allows you the opportunity to constantly learn and build off of others. It teaches you to adapt, be open to new ideas, and to use your time efficiently. The worst part of day-to-day life in the lab, is that sometimes data collection does not go as planned and you need to figure out what went wrong.  However, this aspect doesn’t seem so bad when you have your lab team to help brainstorm.

Overall, my time in the lab has been an incredible experience. It has helped me grow as both an individual and as a scientist and has stimulated my interest in future research opportunities. It is an experience I would highly recommend to other undergraduate students!

Lindsay Piwinski attends Pitzer College in Claremont, CA. She is a 2017 Undergraduate Summer Research Fellow (UGSRF) doing research with Drs. Jenna Monroy (Pitzer College) and Kiisa Nishikawa (Northern Arizona University, Flagstaff, AZ). She hopes to attend graduate school in the future and continue pursuing research.
Impact of Injury on Inflammation

For my research project, I will compare the levels of a known marker of inflammation in and around motor neurons in rats with and without cervical spinal cord injury. We will examine rats with chronic C2 incomplete spinal cord injuries, and compare them to uninjured tissues. We will be examining the frozen and preserved tissue under a microscope to quantify the different levels and locations of inflammatory markers at the different time points. The results of this experiment are important because they will enable us to better treat those who have suffered from the devastating effects of spinal cord injuries. This experiment is necessary to determine if p38 MAP kinase (a specific known marker of inflammation) is activated following spinal cord injury. The results will allow us to determine how to proceed in our search for successful rehabilitative treatments for patients. If p38 MAP kinase is activated following injury, it may call for future studies to investigate treatment of this specific cause of neuroinflammation in order to improve the outcomes of the rehabilitative treatments our lab is studying. Neuroinflammation can decrease the positive effects of rehabilitative efforts, and therefore is something we need to study so we can reduce this inflammation and better treat those who are suffering from spinal cord injury.

Life in the Lab

My experience in the lab was very educational. I did not realize how many different things actually go into research. I became familiar with behind-the-scenes tasks that you do not realize need to be done. For example, we spent a large majority of our time sectioning tissue, which I never realized would be such a large and time-consuming part of the experimental process. In addition, I became independent in many different techniques and procedures used in our lab. I learned proper animal handling and care, the methods used for immunohistochemistry, as well as proper imaging strategies and techniques on the Keyence microscope that we use in our lab. I also learned how to problem solve when problems arose. I found that one of the biggest challenges in research is how time-consuming and detail-oriented everything is. It is necessary to plan very far in advance and plan other aspects of your day around what is needed in lab. I was studying for the MCAT while completing my research project this summer, and I found it very difficult to dedicate time to studying. However, I quickly learned to manage my time wisely and study during down-time in lab and in the evenings. I believe this skill is not only valuable for research, but will also help me throughout my life.

Although what I did day-to-day varied, there were certain tasks that remained constant. For example, animal care and running exposures was something that needed to be completed by someone every day, so I was usually around to help out with those two things. In addition, the weeks were organized in a way that there usually was not more than one big task going on at a time. For example, there were weeks focused on surgeries, as well as others focused on perfusions and harvesting. What I did between rounds of animal care throughout the day varied depending on the week and what needed to be completed. Some days were filled with staining, while others dealt with microscopy. In my opinion, the best part of working in a lab was how often you were able to see your hard work pay off. Although the experiments tend to take at least a few months to complete, there are many milestones where you begin to see the outcomes of all of your hard work. Personally, I thought the hardest part of my time in research was not having set-in-stone days. Your schedule can vary every day depending on the point of the project you are in and what needs to be done, so you need to be able to adjust your plans and schedule around your lab responsibilities.

One of the best parts about working in a lab is being a part of a large team with a common goal. It is much more rewarding to accomplish a goal when everyone is working on it together in my opinion, and it’s nice to always have people around who are willing to help you and your project succeed. Research is usually not a one-person job, and for good reason; there is so much that goes on to ensure that a project is successful, and everyone in lab is needed. Throughout my research experience, I have developed a deep appreciation for how important research is to the functionality of many different aspects of society. Advancements in everything from technology to medicine would not be possible if scientists were not working hard in lab each and every day, and I am glad I will be able to take this appreciation for research into my future career. Research is a tough task, but it is truly life-changing in more ways than one.

Ashley Holland is an undergraduate at the University of Florida in Gainesville, FL. She is working in the lab of Drs. Gordon S. Mitchell and Elisa Gonzalez-Rothi at the University of Florida under the UGREF fellowship funded by APS. After graduation, Ashley hopes to attend medical school and use her skills acquired during her research experience to further the medical field and help her patients in new and innovative ways.