Lauren Gray – Independent Project

Lauren Gray – Independent Project

Class of 2017

Introduction to Topic

The threat of sulfur mustard, commonly called mustard gas, is rising in the use of the United States’ adversaries. In order to create new screenings and medical innovations to test the severe blisters and edema associated with mustard gas, an In-Vitro model of the human skin was developed eliciting a response to create potential remedies.  The invention of an In-Vitro human skin model will aid in the on-site treatment of soldiers affected by mustard gas and those in hospitals, while limiting the need to inflict harm on laboratory animals. The final product is a working In-Vitro model that were used test new medical innovations related to mustard gas.

Project Description

While interning at USAMRICD, a hypodermal component of an In-Vitro human skin model was created from Human Mesenchymal stem cells. The HMSCs were proven valid by differentiating them into osteocytes, adipocytes, and neurocytes. This was done because true HMSCs have the ability to turn into almost any cell found in the body. These stem cells were differentiated into adipocytes by feeding the stem cells adipogenesis media, then were layered with a vascular system printed from a bioprinter.

The bioprinter was formatted to print cells by reducing the air pressure and material that is printed. Most of this work was done in a sterile environment as to avoid contamination of the cells. Once the hypodermal layer is complete, the dermal layer and epidermal layer were placed on top, effectively creating a working human skin model to aid in mustard gas testing and to reduce the use of animals in laboratory testing. This project took  place over the course of the summer in 2016 from June of 2016 to August of 2016.

Experience Description

While shadowing at USAMRICD, I was exposed to a real working government regulated laboratory, the first I had seen ever. It was very interesting to see how the military impacted the protocols of lab, meaning that the purpose of the lab’s various studies was to aid in any militarists biological threat. The interns were placed  into groups, so luckily I was placed in a group with two other biomeders. Together we created the In-Vitro human skin model, with each intern specializing in one layer of the skin. I selected the hypodermal layer, not knowing how challenging it would be. At first, the problem presented to us seemed overwhelming, but once we learned more about the skin and its many layers, it seemed less intimidating.

A typical day was as follows: we would begin our day in a cubicle researching a published article on each of our layers, taking notes and educating ourselves. Some papers were up to 30 pages long. Then our mentor would have a teaching session with us; discussing our articles and teaching us laboratory techniques that we would us in the lab later. After our session, we would venture into the lab, getting comfortable using micropipettes under the hood- a sterile environment. There I learned to culture the stem cells and turn them into different tissue. After the lab time, we would take notes on what we did that day, and work on our journals.

I learned the hard way to pay attention to protocol. When I was passing, or moving my cells into a bigger flask, I accidentally left the cells in their original container, only moving the media to the bigger one, and threw the original away. Essentially I killed sixty million HMSCs and set the hypodermal layer portion of the project back a week. I was devastated and felt like I had let the group down. My mentor was understanding and showed me where they housed the HMSCs, proving that it was okay to mess up because it was a learning process and that they had more than enough to replace what I killed.

Because of the rigorous demands of the military in our mentor, he had to leave the interns alone a lot in the lab; I got very acquainted with it. I was a little frustrated at how many times we were left alone or forgotten. It felt like the military duties were always first and the science always came last. Before completing this internship I had wanted to do ROTC for the Air Force, and becoming a lab technician, but this experience taught me that I want to focus more on science than on the military, ultimately changing my mind. I now want to be in the research field, but with a doctorate so I can publish my findings.

Overall I learned the thorough and detail oriented lab techniques that were necessary in completing the skin model. Upon completion of the project, I was very proud that I had created something that would reduce animal suffering, while benefiting the soldiers that are over seas fighting for our country.

Innovation Description

The process of creating the hypodermal layer was very difficult. It required weeks of raising the HMSCs to maturity. To make make sure they are properly fed, they have to be checked up on every day to make sure the cells are not dying and have adequate moisture. This process requires that an AmScope is turned on, adjusted to take microscopic pictures, and set to capture the correct colors and type of cells. Doing this everyday was a time consuming process. In addition to taking pictures everyday, the cells had to be kept from becoming too confluent, as having too many cells can cause them to form spiral bundles and turn into bone. To keep them from turning into bone, the cells were passed into another, bigger container at least once a week. Because of this tedious process, hours were spent in the lab monitoring, feeding, and taking pictures of the cells. When not waiting for the cells to grow, further research was spent on stem cells and related material to the innovation. This proves that the innovation was an original concept, as every detail was researched before entering the lab.

Once the cells were mature, they had to prove their validity. To do this, they had to be differentiated into three different types of tissue: bone, neuron, and adipose. This consuming process involved passing the cells into different flasks, feeding them the appropriate media to turn them into their respective cells, and then monitoring them daily to make sure they were growing. Once all of the cells had been grown, more pictures were taken, and then the samples were thrown away because they had proven the validity of the cells. New HMSCs were then grown to maturity, restarting the process mentioned above.

The the cells were mature a second time, and then fed adipogenesis media. The Adipogenesis media was made by hand every time it needed to be used, so everyday. It involved mixing up different chemicals in exact ratios, warming them as to not shock the cells, then removing the original media and replacing it with the hand made media. After a few days of this, it was realized that the cells were not developing vesicles, so the type of media used was changed into one derived from pig and rabbit proteins. This also had to be made by hand. Once the cells were finally turned, the bioprinter printed out a vascular system made of rigid proteins. The bioprinter was made by a college intern, and the SEAP interns helped assemble.

Then the adipocytes were layered at the bottom of welled plates, so that they were under the wells. This required the use of one of the smallest micropipettes housed, as the adipocytes had to be feed under the well without disturbing the dermal and epidermal layer above. The risk of forcing media in too fast also threatened to shear the cells, of which damage was minimal. The expertise and skill required to delicately feed the cells in this manner was acquired by the hours spent under the fumigation hood practicing sterile techniques, and if a mistake was made, the process had to be started over.

Once the adipocytes were adhered to the bottom of the well plate, the vascular system was then layered. While the vascular system did not layer correctly, it did work for the short amount of time that it was placed in the hypodermal layer.

The innovation itself is pink in color due to the fact that the hypodermal cells do not have  a color, but the media is red in color. The hypodermal layer is under the dermal and dermal layer, requiring a special feeding time and strict scheduling of media change. It was in a well, but will be expanded to fit in bigger containers to have mustard gas tested on it. The response to the mustard gas will be more human than the skin of guinea pigs, which are the primary animals used in mustard gas testing.

The closest competitor with this innovation is USAMRICD itself. No other facility has attempted to make this model with the hypodermal component included (Eaton). There are rudimentary designs with the dermal and epidermal component, but none completed like the innovation described. If taken further, there is a possibility of the innovation being patented, with  possibility of SEAP interns name on it. If completely successful, lab procedures and ways of achieving innovation will be classified, as they will become Army Laboratory practice.  

Project Topic

Introduction to Topic

The threat of sulfur mustard, commonly called mustard gas, is rising in the use of the United States’ adversaries. In order to create new screenings and medical innovations to test the severe blisters and edema associated with mustard gas, an In-Vitro model of the human skin was developed eliciting a response to create potential remedies.  The invention of an In-Vitro human skin model will aid in the on-site treatment of soldiers affected by mustard gas and those in hospitals, while limiting the need to inflict harm on laboratory animals. The final product is a working In-Vitro model that were used test new medical innovations related to mustard gas.

Project Overview

Project Description

While interning at USAMRICD, a hypodermal component of an In-Vitro human skin model was created from Human Mesenchymal stem cells. The HMSCs were proven valid by differentiating them into osteocytes, adipocytes, and neurocytes. This was done because true HMSCs have the ability to turn into almost any cell found in the body. These stem cells were differentiated into adipocytes by feeding the stem cells adipogenesis media, then were layered with a vascular system printed from a bioprinter.

The bioprinter was formatted to print cells by reducing the air pressure and material that is printed. Most of this work was done in a sterile environment as to avoid contamination of the cells. Once the hypodermal layer is complete, the dermal layer and epidermal layer were placed on top, effectively creating a working human skin model to aid in mustard gas testing and to reduce the use of animals in laboratory testing. This project took  place over the course of the summer in 2016 from June of 2016 to August of 2016.

Experience

Experience Description

While shadowing at USAMRICD, I was exposed to a real working government regulated laboratory, the first I had seen ever. It was very interesting to see how the military impacted the protocols of lab, meaning that the purpose of the lab’s various studies was to aid in any militarists biological threat. The interns were placed  into groups, so luckily I was placed in a group with two other biomeders. Together we created the In-Vitro human skin model, with each intern specializing in one layer of the skin. I selected the hypodermal layer, not knowing how challenging it would be. At first, the problem presented to us seemed overwhelming, but once we learned more about the skin and its many layers, it seemed less intimidating.

A typical day was as follows: we would begin our day in a cubicle researching a published article on each of our layers, taking notes and educating ourselves. Some papers were up to 30 pages long. Then our mentor would have a teaching session with us; discussing our articles and teaching us laboratory techniques that we would us in the lab later. After our session, we would venture into the lab, getting comfortable using micropipettes under the hood- a sterile environment. There I learned to culture the stem cells and turn them into different tissue. After the lab time, we would take notes on what we did that day, and work on our journals.

I learned the hard way to pay attention to protocol. When I was passing, or moving my cells into a bigger flask, I accidentally left the cells in their original container, only moving the media to the bigger one, and threw the original away. Essentially I killed sixty million HMSCs and set the hypodermal layer portion of the project back a week. I was devastated and felt like I had let the group down. My mentor was understanding and showed me where they housed the HMSCs, proving that it was okay to mess up because it was a learning process and that they had more than enough to replace what I killed.

Because of the rigorous demands of the military in our mentor, he had to leave the interns alone a lot in the lab; I got very acquainted with it. I was a little frustrated at how many times we were left alone or forgotten. It felt like the military duties were always first and the science always came last. Before completing this internship I had wanted to do ROTC for the Air Force, and becoming a lab technician, but this experience taught me that I want to focus more on science than on the military, ultimately changing my mind. I now want to be in the research field, but with a doctorate so I can publish my findings.

Overall I learned the thorough and detail oriented lab techniques that were necessary in completing the skin model. Upon completion of the project, I was very proud that I had created something that would reduce animal suffering, while benefiting the soldiers that are over seas fighting for our country.

Innovation

Innovation Description

The process of creating the hypodermal layer was very difficult. It required weeks of raising the HMSCs to maturity. To make make sure they are properly fed, they have to be checked up on every day to make sure the cells are not dying and have adequate moisture. This process requires that an AmScope is turned on, adjusted to take microscopic pictures, and set to capture the correct colors and type of cells. Doing this everyday was a time consuming process. In addition to taking pictures everyday, the cells had to be kept from becoming too confluent, as having too many cells can cause them to form spiral bundles and turn into bone. To keep them from turning into bone, the cells were passed into another, bigger container at least once a week. Because of this tedious process, hours were spent in the lab monitoring, feeding, and taking pictures of the cells. When not waiting for the cells to grow, further research was spent on stem cells and related material to the innovation. This proves that the innovation was an original concept, as every detail was researched before entering the lab.

Once the cells were mature, they had to prove their validity. To do this, they had to be differentiated into three different types of tissue: bone, neuron, and adipose. This consuming process involved passing the cells into different flasks, feeding them the appropriate media to turn them into their respective cells, and then monitoring them daily to make sure they were growing. Once all of the cells had been grown, more pictures were taken, and then the samples were thrown away because they had proven the validity of the cells. New HMSCs were then grown to maturity, restarting the process mentioned above.

The the cells were mature a second time, and then fed adipogenesis media. The Adipogenesis media was made by hand every time it needed to be used, so everyday. It involved mixing up different chemicals in exact ratios, warming them as to not shock the cells, then removing the original media and replacing it with the hand made media. After a few days of this, it was realized that the cells were not developing vesicles, so the type of media used was changed into one derived from pig and rabbit proteins. This also had to be made by hand. Once the cells were finally turned, the bioprinter printed out a vascular system made of rigid proteins. The bioprinter was made by a college intern, and the SEAP interns helped assemble.

Then the adipocytes were layered at the bottom of welled plates, so that they were under the wells. This required the use of one of the smallest micropipettes housed, as the adipocytes had to be feed under the well without disturbing the dermal and epidermal layer above. The risk of forcing media in too fast also threatened to shear the cells, of which damage was minimal. The expertise and skill required to delicately feed the cells in this manner was acquired by the hours spent under the fumigation hood practicing sterile techniques, and if a mistake was made, the process had to be started over.

Once the adipocytes were adhered to the bottom of the well plate, the vascular system was then layered. While the vascular system did not layer correctly, it did work for the short amount of time that it was placed in the hypodermal layer.

The innovation itself is pink in color due to the fact that the hypodermal cells do not have  a color, but the media is red in color. The hypodermal layer is under the dermal and dermal layer, requiring a special feeding time and strict scheduling of media change. It was in a well, but will be expanded to fit in bigger containers to have mustard gas tested on it. The response to the mustard gas will be more human than the skin of guinea pigs, which are the primary animals used in mustard gas testing.

The closest competitor with this innovation is USAMRICD itself. No other facility has attempted to make this model with the hypodermal component included (Eaton). There are rudimentary designs with the dermal and epidermal component, but none completed like the innovation described. If taken further, there is a possibility of the innovation being patented, with  possibility of SEAP interns name on it. If completely successful, lab procedures and ways of achieving innovation will be classified, as they will become Army Laboratory practice.  

By | 2017-05-15T14:59:24+00:00 May 15th, 2017|Biomed Capstone Project 2017|0 Comments

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