Overview
Human Practice and Integrated Human Practice, emphasizing how our project is good for the world and vice versa how the feedbacks drive the project’s evolvement, act as important links between our iGEM project and our real life. In this year’s SHSBNU-China project, our team members connected with people from all walks of life to impact and be impacted. Through our Human Practices, we learned about the needs, and received valuable feedbacks from different stakeholders, such as doctors, manufacturers，synthetic biology experts，and policy makers. We incorporated these insights into our project design, implementation, and evaluation, making it more relevant and feasible. Our Human Practices not only helped us improve our project, but also inspired us to think beyond the lab. We realized that synthetic biology is not only a scientific endeavor, but also a social one. By engaging with diverse perspectives and voices, we gained a deeper understanding of the context and implications of our project, and developed a sense of responsibility for the future of synthetic biology. We introduce our integrated Human Practices in five stages: “Project Ideation”, “Project Design and Experiment”, “Project Hardware”, “Project Implementation”, and “Public Awareness”.
Human Practices Highlights
Stage I：Project Ideation
In the brainstorming process of the project, we focused on the issue of heart diseases. Among them, diseases related to the heart valve stand out. The current treatments for these diseases are limited, and the cost of valve replacement is high.
Activity 1. Is there a market demand for a new type of heart valve material?
[Stakeholder]: Online interview with Mr. Shang Qingxue from Edward Lifescience [Team’s Reflection]: We confirmed that the heart valve market needs a material with better mechanical strength and biocompatibility. Edwards Lifescience specializes in areas such as heart surgery, emergency care, and the vascular system. It is a global leader in heart valve replacement and heart valve cares. Mr. Shang is a senior technical specialist in the company’s R&D department. He introduced us to the current situation at Edwards, as well as the global heart valve market. We learned that traditional valves fall into two categories: 1) Biological valves 2) Mechanical valves. The issues with these are: 1) Biological valves are not strong enough and can easily tear 2) Mechanical valves have poor resistance to thrombosis and poor biocompatibility. Therefore, there's a need for a material with high mechanical strength and better biocompatibility. We believe that materials mainly composed of proteins from syn-bio can address the issue of biocompatibility. After conducting literature research, we found that fibers made up of Titin protein have mechanical strengths surpassing many traditional materials. We believe this material could become the next-generation heart valve material.
Figure 1. Edward Lifescience’s introduction to the commonly used heart valves in the current market.
Figure 2. Our Human Practices subgroup interviewed Mr. Shang from Edward Lifescience.
Activity 2. The cardiac surgeon’s perspective on new heart valve materials
[Stakeholder]: In-person Interview with Dr. Zhao at Peking Union Medical College Hospital [Team’s reflection]: We consulted a cardiac surgeon who affirmed the need for a new material. We had the opportunity to speak with Dr. Zhao, a renowned cardiac surgeon at the Peking Union Medical College Hospital, considered one of the premier institutions for heart surgery in Beijing. Dr. Zhao highlighted the pressing concern that heart valve diseases have led to a significantly high mortality rate in China. Factors like the number of valve leaflets, adaptability, and size are crucial considerations when addressing this medical condition. As it stands, China predominantly relies on international companies for heart valve supplies. Domestic entities, still in their nascent stages, often find their products priced steeply due to their limited experience in the market. Mechanical valves are priced at over 10,000 RMB, while biological ones can go beyond 20,000 RMB. When converted to USD, this is equivalent to 1-2 months of the average Beijing resident's income. Considering the vast population, millions potentially require heart valve surgeries, emphasizing the urgent need for affordable and efficient solutions.
Figure 3. Our team interviewed Dr. Zhao, a renowned cardiac surgeon at the Peking Union Medical College Hospital.
Activity 3. Investigation of the receptive attitude of the general public
[Stakeholder]: Survey the general public [Team’s reaction]: We confirmed that majority of users hold a positive view of this new material We designed a survey to study if a new heart valve material can be accepted by the general public. We conducted street interviews to gather public knowledge about heart valves and heart valve diseases, as well as their opinions on new valve materials. The results showed that the majority of the general public still has limited knowledge about heart valve diseases. Regarding the new materials, most people are open to our proposal, but a minority, especially the elderly, prefer the more established biological valves.
Figure 4. Survey results indicate that the younger generation is more receptive to the new heart valve material, whereas seniors tend to prefer the traditional option.
Stage II：Project Design and Experiments
Activity 4. To determine the synthesis and production conditions for Titin protein
[Stakeholder]：Online Interview with Mr. BoXiang Wang, PI of 19 Greatbay_SZ [Team’s reaction]: Learned experimental details, and re-designed experiment to examine Titin protein solubility. Mr. BoXiang Wang was the PI for the 2019 iGEM Team Greatbay-SZ. That year, the team researched the synthesis of spindron. Similar to our target, Titin, spindron is a protein that can be transformed into fibrous threads. We hoped to learn from him about the specifics of the experimental parameters. During our discussion, he shared details on protein purification using a nickel column, pointing out that this step is particularly challenging. Professor Wang emphasized that spindron is insoluble in water. Based on this insight, we designed a new experimental plan to test if we could achieve better solubility for the Titin protein under different conditions, which could impact the subsequent thread production. In conclusion, he offered advice on our project theme, highlighting that when designing for human implantation, it's crucial to consider the immunogenicity of the protein. He commended our choice of using the gene sequence from rabbitfish, which could effectively minimize the potential for cyclic reactions within the body.
Figure 5. Online Interview with Mr. BoXiang Wang
Activity 5. Improve experimental techniques to test protein synthesis.
[Stakeholder]：Interview with Prof. Yang Hu from Minzu University Dr. Hu is a professor at Minzu University specializing in biomedical engineering. He has extensive experience in examining protein expression using SDS-Page and Western Blot. In our experiments, the SDS-Page results indicated the presence of large proteins capable of forming fibers. However, due to our lack of expertise in conducting the SDS-Page, the results were somewhat ambiguous. Dr. Hu provided us with recommendations. We adjusted the experimental parameters, and while the results showed significant improvement, they still fell short of offering robust and solid evidence to prove the expression at a research level. After inducing our cultured bacteria, we sent the samples to Dr. Hu. He personally conducted the SDS-Page, producing clear evidence that our design was successful.
Figure 6. Meeting Dr. Hu in the lab and we learned about more experiment techniques for a better SDS-Page run.
Figure 7. Our SDS-Page results were ambiguous in proving the Titin protein synthesis. We then cultured the bacteria and induced the protein expression under exactly the same condition. The sample was sent to Dr. Hu for an SDS-Page run. The results show clear bands at the expected positions. (+iPTG -1, -2, -3 are repetitions with exactly same experiment conditions.)
Activity 6. Adjusting the coding sequence design and validating through experiments
[Stakeholder]：Interview with Dr. Hu from Minzu University Dr. Hu suggested that we experiment with varying the number of repeated sequences in the fibrous protein's sequence to investigate which design would be optimal for fiber production. We adjusted our initial design from 4XT to 8XT.
Figure 8. Sequence design of 8XT-NC to test production efficiency.
Our team carried out experiments and found that the 8XT-NC design did not exhibit satisfactory expression. Dr. Hu proposed using a more sensitive method, the Western Blot, for further verification. Due to limitations in our experimental conditions and techniques, we were unable to conduct this advanced verification. We then handed our samples over to Dr. Hu. Upon conducting the Western Blot test, Dr. Hu was able to confirm that the 8XT-NC design did not achieve successful large protein production. This led us to conclude that our original design, 4XT-NC, was indeed more stable.
Figure 9. SDS-Page results and Western Blog results (From Dr. Hu) to verify 8XT-NC design was not as stable as the 4XT-NC.
Stage III：Project Hardware
With the successful synthesis of the Titin protein, we turned our attention to converting this protein into fibers. We discovered that we could employ a technique called wet spinning to transform the protein into fibers interconnected by β-sheets. After this transformation, we can utilize plain weaving technology to craft these protein fibers into a fabric. This fabric can then serve as the intermediary layer in a 'sandwich structure.
Activity 7. Learning the fundamentals of Wet Spinning
[Stakeholder]：Online Interview with Prof. Wang from Donghua University To verify the feasibility of our approach, we consulted Prof. Wang Jun from Donghua University's School of Textiles. Prof. Wang is an expert in thread and textile machinery at Donghua University, located in Shanghai, China, which is renowned for its expertise in the textile field. Prof. Wang initiated our discussion by introducing us to the basics of weaving and confirming the methods for weaving both long and short threads. He expressed his support to provide us with technical advice and help us optimize our experimental steps. He was enthusiastic about our ideas and current conceptualizations. We inquired about general protein fiber weaving techniques and the specifics of the necessary equipment. The professor illustrated his explanations using the most common example: soy protein fiber. He further detailed the specific parameters one should consider in protein fiber weaving, such as viscosity, temperature, pushing capacity, advancing speed, and other relevant data.
Figure 10. Online interview with Dr. Wang Jun
Subsequently, we delved into the procedures of wet spinning and learned how to implement the mechanical structures required for the wet spinning process. We acquired a detailed understanding of how to use stepper motors and ball screw mechanisms to control a needle, allowing for a consistent rate of protein solution extrusion into a coagulation bath to form fibers. Armed with this knowledge, we drafted a preliminary design diagram and further discussed our design ideas with Prof. Wang. Ultimately, we successfully fabricated a hardware system that allows for adjustable and controllable extrusion rates.
Figure 11. Our hand-drawn design of a combination of the stepper motor and frame. We constructed the frame with customized metal bars and brackets, and 3D-printed the connector parts, and eventually assemble the injection drive.
Activity 8. Design of Stretching Machinery for Fiber
[Stakeholder]：Second Interview with Prof. Wang from Donghua University In our second interview, Professor Wang commended our design and offered further suggestions to improve it. He proposed incorporating two sets of stretching devices, ensuring that the protein thread is elongated before entering the coagulation bath. This measure would mitigate the tendency of the protein solution to undergo diameter dispersion in the bath, ensuring the consistent diameter of the threads. He explained that one stretching device consists of an upper rubber cylinder paired with a lower metal cylinder. The active rotation of the rubber cylinder would drive the passive rotation of the metal cylinder, facilitating the elongation of the protein thread as it passes between the two cylinders. The rotation speed near the needle head would be faster than the injection speed of the needle itself, and the speed further away from the needle head, closer to the aramid, would be even faster than the first rotation speed. Moreover, the stretching device is designed to move vertically, which allows for swift and precise placement of the protein fibers. In conclusion, Professor Wang believed that once the fibers are successfully manufactured, transitioning to a plain weaving process to produce fabric would be entirely feasible. He also expressed his affirmation and high expectations for our future endeavors. Additionally, he emphasized the importance of the diameter of the injection needle. The needle's diameter plays a pivotal role in ensuring that the final product is a continuous thread.
Figure 12. We purchased the roller device to study the design.
Figure 13. We tested needles with various diameters.
Activity 9. Testing Wet Spinning Injection Parameters
[Stakeholder]：Online Interview with Prof. Wang from Donghua University In another discussion, aiming to produce finer fibers, the professor emphasized the importance of extensive testing and adjusting the injection speeds. Recognizing that our protein production is still experimental and not yet scalable enough to generate vast quantities needed for repeated hardware testing, he suggested using generic protein fibers as a stand-in to evaluate the spinning equipment's performance. This approach would enable us to refine the equipment settings and ensure the resulting fibers meet the desired specifications. By using a readily available protein as a proxy, we can gain insights into the behavior of our specialized protein under similar conditions without depleting our limited resources. Furthermore, the use of a common protein can also provide a baseline for performance, making it easier to identify any unique challenges or advantages presented by our proprietary protein when it's eventually used.
Video: Protein solution injecting into cold coagulation bath, showcasing the fiber spinning test.
Figure 14. Testing wet spinning and injection conditions, such as temperature, solution viscosity, and injection diameter/speed.
Stage IV：Project Implementation
Activity 10. Designing the Shape of the Heart Valve
[Stakeholder]：Interview with Dr. Xiong Wei, Chief Surgeon of Cardiovascular Surgery We consulted Dr. Xiong Wei regarding the implementation of Titin material into a marketable a heart valve. Dr. Xiong pointed out that the subpar anti-thrombotic resistance of mechanical heart valves arises due to certain fluid dynamic issues. Mechanical valves tend to induce turbulent flow, whereas an ideal artificial synthetic valve should promote laminar flow. He believes that a tricuspid valve design would be more effective for our purpose, prompting us to evolve our initial bicuspid design into a tricuspid one. Additionally, he mentioned that each flap of the heart valve should approximate the shape of a trapezoid.
Figure 15: Online Interview with Dr. Xiong
Figure 16. The new proposed shape design of heart valve using titin protein fiber.
Activity 11. Acquiring more suggestions on the product design
[Stakeholder]: Presenting the Titin Heart Valve project and Heart Valve Design at international conference IEEE 3M-Nano, Chengdu, China At the IEEE-3M Nano International Conference on Nanomaterials held in Chengdu, three of our team members represented our group to showcase the project design. Professors in attendance highlighted that many materials lead to thrombosis due to the uneven surface of the material. They suggested considering adding smooth layers to both the top and bottom of the fabric woven from Titin protein fibers. Of course, it's imperative that this coating is biocompatible. After a thorough literature review, we discovered a material that fits this description: TPU (Thermoplastic Polyurethane). As a result, we updated the design of our heart valve product.
Activity 12. Exploring Other Applications of Titin Fibers
[Stakeholder]: Dr. Tong Dedi, a Microsurgery Specialist; Dr. Zhu Shan, a Cosmetic Surgery Expert We hope to broaden the applications of the Titin we produce, so we consulted with doctors who frequently use surgical sutures. Both experts emphasized that surgical sutures must possess three key characteristics: strength, elasticity, and fineness. They believe our Titin fibers, due to their high mechanical strength and biocompatibility, are ideal for creating single-strand sutures. These sutures, given their smooth surface, pass easily through tissues, leading to reduced inflammation and minimal rejection. This makes them especially suitable for use in microsurgery and cosmetic surgery procedures. Regarding the diameter of the sutures, the doctors noted that the thickest sutures currently available on the market are 0.3-0.4mm, while the finest measure 0.01mm. After reviewing relevant literature, our team found that we can produce Titin fibers as thin as 0.01mm. The doctors' primary suggestion for our venture into surgical sutures was to test the degradation time of Titin. They also highlighted several advantages of Titin: its wide availability, its tissue origin closely resembling the human body, and its impressive elasticity, strength, and resilience.
Figure 18，Interview with Dr Tong and Dr. Shan.
Activity 13. Policy Issues on Application of Titin Fibers as Surgical Sutures
[Stakeholder]: Mr. Han, Senior Technical Specialist from a Surgical Suture Company We introduced our idea of using Titin as surgical sutures to Mr. Han. Based on our description, he believes that our Titin qualifies as a non-absorbable material. He noted the advantages of Titin, emphasizing its minimal rejection response, high strength, and great resilience. Mr. Han also detailed how the absorption period of surgical sutures is typically tested. He suggested placing the Titin-made threads in flowing saline solution or air and then using a tensile tester to measure the strength over different time intervals. The strength of surgical sutures is generally compared to that of standard threads, and they are deemed ineffective when their tensile strength drops below 23-25% of their original capacity. Furthermore, we inquired about policies related to surgical sutures. Mr. Han mentioned that the domestic government is highly supportive of the development of surgical sutures, especially since there has been limited original development in surgical sutures within China.
Figure 19，Interview with Mr. Han.
Stage V: Public Awareness
Activity 14. Developing a Comprehensive Health Awareness Campaign: The Heart of the Matter
In collaboration with a dedicated team of medical professionals, we've crafted an informative and engaging poster to enhance public understanding of heart health. Displayed in the hallways of prominent hospitals with proper hospital staff review, our posters are positioned to reach a wide audience, from patients to their visiting family and friends. Our poster begins with a visual and descriptive overview of the heart’s function and diseases. We delve into common risk factors for heart diseases and offer simple, everyday steps to mitigate these risks, from dietary choices to exercise routines. We proudly introduce the innovative Titin protein material to promote public awareness and to collect feedbacks. We plan to analyze and reflect on the feedbacks to address any safety or healthcare concerns, as well as optimizing product designs. Using layman’s terms and illustrative visuals, we explain its potential as a superior alternative for heart valves, highlighting its biocompatibility, durability, and overall benefits for cardiac patients.
Figure 20，Poster to promote knowledge of cardiovascular and heart valve issues.
Activity 15. Cardiovascular and Heart Valve Health Education
After recognizing the knowledge gap in the general public regarding heart-related information, we designed a series of educational activities aimed at reaching a wider audience. Through survey investigations, we have found that the current understanding of heart knowledge is not comprehensive and widespread. Conducting educational outreach to students will help expand social awareness to some extent. In order to better promote our project to all age groups, our team members have recorded an approximately eight-minute educational video. It includes basic knowledge about the heart, a brief introduction to our iGEM project, heart valve diseases, and related preventive measures.
Figure 21，Model of a heart we purchased for educational lectures.
Within our capabilities, we have pursued offline explanations to achieve better promotional results. Through communication with middle school teachers, we have conducted project promotions to multiple classes in middle schools. In order to connect the project content with middle school knowledge while promoting the project, we have specifically selected simple and easy-to-understand illustrations similar to middle school textbooks, as well as interesting and imaginative metaphors to facilitate middle school students' understanding of the project.
Figure 22，Education lectures delivered by our team members
Similarly, in our high school synthetic biology club, we also conducted relevant promotional presentations. However, unlike with middle school students, when facing high school students from international schools, we focused our explanations on promoting and explaining the iGEM project, building upon their existing knowledge of synthetic biology. To achieve better promotional results, we opted for more authentic and detailed English illustrations to enhance the effectiveness of our presentations.
Figure 23，Education lectures delivered by our team members
In offline lectures, whenever we finish the explanations, we conduct a Q&A session, creating a perfect feedback loop. During this session, middle school students had their doubts addressed through our careful explanations, and some expressed interest in biology, looking forward to further study in high school. High school students, on the other hand, provided interesting ideas for our project design, allowing us to further develop in the future. To better promote our project across all age groups, our team members recorded an approximately eight-minute promotional video for better outreach. In situations where face-to-face communication was not possible, we chose to send our video as a supplementary means of promotion. For adults who showed interest in our project, we approached them and promoted our team's project, gaining recognition as well. Targeting our main audience, the elderly, we contacted nursing homes and presented our lectures in the form of videos. In these lectures, we focused on popular science and prevention of heart valve diseases, making the project more relevant to the needs of the elderly. Finally, we uploaded our video on the Bilibili platform to reach a wider audience. It has gained a good number of views and received affirmation from many online users.
