My Internship

Abstract

According to the National Library of Medicine, a staggering 100,000 to 200,000 ACL sprains and tears are reported annually in the United States. Our research is led by principal Investigator Stephen Schlecht, Ph.D. to study the anterior cruciate ligament (ACL), including its development, function, and response to loading. The overarching goals of research in the Schlecht Lab are to develop new clinical diagnostics for ACL injury prevention in adolescent and young adult recreational and competitive athletes and to improve primary ACL reconstruction outcomes. We accomplish this with the use of animal models and their corresponding response to induced fatigue. First, we looked at molecular changes in collagen after fatiguing mice and rats while varying the duration of recovery time. An increase in unraveled collagen in the knee was observed. Moreover, our publication on this study demonstrated an increase in collagen unraveling negatively correlates with ACL strength. Currently, we are in the process of investing in genomic expression of damaged knee tissue after induced fatigue. We hope to see the different expressions of genes responsible for strength and recovery that could serve as a marker for potential injury.

Overview

  The Schlecth Lab is looking for ways to better prevent and recover from ACL damage. In doing so, we work alongside the University of Michigan where they implement our findings in mice on human cadavers. We are currently looking to see if there is a biological marker for predispositions for ACL tears. We hope to see a change in the natural healing process after ACL damage collagen levels in the knee on different days subsequent to damage. We accomplish this by loading (damaging) mice and analyzing their knees at 3,6,9, and 12 days after loading. In order to quantify our results the mice's knees are segmented and stained to illustrate the presence of collagen in the knee. It is known that when a knee is damaged, collagen floods the knee inducing inflammation. After the segmented knees are stained, a fluorescent imaging process is used to visualize increased levels of collagen. Once the fluorescent images of the knees are captured, computer software is used to manually crop out the ACL. It is common that the PCL incurs some damage which may also be susceptible to staining as well as other tissue and fibrocartilage. From here, we run the cropped ACL images through software that measures the presence of stained tissue. We can then compare the damaged/stained tissue area to the total area of the initial cropped ACL. The ratio of damaged tissue: to total ACL tissue is compared on different days after load. We are currently cropping the ACLs and should be able to produce some results of this study in the coming weeks. 

Learning and skills

During my time in Dr. Schlecht’s lab, I hope to learn more about the ACL complex, and general lab techniques. I am readily learning and applying new information about the knee and will hopefully continue to grow my knowledge of the ACL complex and the knee as a whole. I think this will help me later down the road as my foundation for the education of the ACL is rooted in research that is not readily available in standard classroom settings. I also would like to become more familiar with various lab techniques. Being familiar with different types of lab equipment and procedures will allow me to possibly branch off into different, more specialized aspects of clinical research and further my education. Additionally, I am looking forward to the translational aspect of our research. I find it really interesting how the research we do in the lab is translated over to humans. To do this, my internship site works with the University of Michigan for the translation research aspect, and I am looking forward to getting involved in their weekly discussions.

Working in Dr. Shclect's lab has improved many topics covered in my past and present course material. More specifically, working with the ACL involves understanding its structure and many different biological and chemical applications. The CHP staining process involves chemistry seen in many entry-level chemistry labs such as diluting alcohol and mixing solutions. After staining, the imaging process requires a previous understanding of how to use a microscope in order to capture images of segmented mice knees. Additionally, the differentiation of various types of tissues illustrate in the microscopic images is one of the topics being covered in my current biology k103 course. The experiences have reinforced my knowledge of the topic as well as applied my understanding to an influential cause. Personally, I believe my professional skill set has developed immensely throughout the course of the semester. I have had many work and internship experiences but not at this level of education and professionalism. The way in which individuals in this field conduct themselves on a daily basis is drastically different from what was known to me. This has been a welcome adjustment for me as I am hoping to pursue a prestigious professional position in the medical field. One of my favorite experiences to date is using the Image J software to measure the area of each ACL segment. While tedious, I find it extremely rewarding knowing that the data I collect is one of the final stages as well as one of the most influential steps in our current study.

Recently, we have continued our analysis of mice knee fatigue. We are assessing the impacts of 30% and 60% forces on the presence of unraveled collagen using the CHP staining process outlined previously. Again, I was tasked with staining sections of tissue and compiling the data for a broader comparison.

The Workplace

The individuals I work alongside have a variety of educational backgrounds. Primarily, I work with Taeyong (Ted) Ahn, Ph.D., Postdoctoral Fellow, and Ben Loflin, MS, Research Analyst I. Our research is led by principal Investigator Stephen Schlecht, Ph.D. I truly enjoy coming to the lab and working with these individuals. Since joining, I have noticed that they are a very tight-knit group of colleagues, and it has been a pleasure getting to know them. Up until this point, I have been pleasantly surprised with my responsibilities. I feel as if I am an influential team member and relied upon heavily. Most recently I was tasked with cropping and measuring the ACL areas of mice knees. This is a very tedious and time-consuming process that is vital to the results of the study. I am gracious in the responsibility delegated to me and I hope to continue to make an impact.

After a full semester, I can confidently say I have improved my professional skill set. Unlike my previous experiences, many of the people I am surrounded by are highly educated and very busy. I came to realize that everybody has external stuff going on that may affect their availability. Rather than sitting on my hands waiting for a response I have learned to take more initiative. As my personal confidence grows, my supervisor has entrusted me to handle tasks independently without direct supervision. Communicating professionally and efficiently has quickly been stressed as being of the utmost importance. It is also one of my first positions where technical terms/jargon are frequently used to adequately describe an observation. This has helped me grow my communication skills immensely in the workplace.

As my time in the lab progresses I feel more comfortable with the tasks assigned to me. Often, similar procedures and protocols are followed to analyze ACL fatigue. I can tell that my lab is more confident in my skills as I am left to conduct my work largely independently. As stated previously, many of my tasks are things that I have done before so my confidence has grown throughout the semester. In our genomic study, we assessed the quality of samples extracted by four members of the lab including myself. My sample was found to have the greatest content of protein and therefore made me the primary lab tech on this protocol for extracting RNA from an ACL tissue segment.  The subtle nod from the principal investigator was very much appreciated and made me feel like a valuable asset to the team. I also feel as if my basic laboratory skills have improved largely in part to regularly practicing these techniques and refining them along the way.

Successes and Challenges

Many successes and challenges have been encountered throughout the semester. As this is my first clinical lab position, I had to be ready to learn on the fly and be comfortable making mistakes. I laugh now, but I accidentally knocked over some samples when placing slides into the refrigerator. Luckily, these samples in the refrigerator in were all sealed, and nothing was spilled or damaged. In the grand scheme of things, this was a pretty minor mishap but something I was able to learn from and hopefully not do again. While I am a firm believer that mistakes are the greatest learning opportunities, it is good to recognize and appreciate your success at times. As stated previously, measuring the stained ACL segments was a very time-consuming process. Upon successful completion of this was a minor win in my book. From there, I was able to see tangible results about natural ACL recovery over a 3, 6, 9, and 12-day period.

As the semester marches on, I continue to complete various tasks assigned to me. For example, our next component of the analysis is observing the genomic expression from RNA of segmented ACL tissue recovered from fatigued mice at varying time points. This may be a challenge for me as my background in RNA analysis is not very extensive. The past couple of weeks at my internship have largely been educational in preparation for this new study. This involved reading up on current literature as well as the protocol for isolating and harvesting the RNA from tissue to be tested. This past week I successfully collected and purified RNA samples from two practice tissue segments that were leftover from a previous study. Currently, I am continuing to read up on information that will allow me to better understand the data when we get it back for analysis.