At Harvard Medical School and Massachusetts General Hospital, Dr. Rasha Al-attar’s research is at the forefront of advancing heart regeneration and organ transplantation. Initially focusing on stem cell-derived cardiomyocyte (hPSC-CM) transplantation to repair infarcted heart tissue, Dr. Al-attar faced a significant challenge: while immature hPSC-CMs survive freezing, they often induce arrhythmias post-transplantation, limiting their clinical application. To address this, Dr. Al-attar transitioned to her postdoctoral work in Dr. Shannon N. Tessier's lab, where she applies nature-inspired techniques to advance cryopreservation methods. Inspired by freeze-tolerant frogs, her research includes developing techniques to preserve hearts at normothermic levels ex vivo, enabling real-time monitoring of arrhythmias and improving transplantation outcomes. Her work aims to overcome gaps in cryopreservation and organ preservation, enhancing the clinical feasibility of stem cell therapies and addressing key issues in heart regeneration.

Early Inspirations

Dr. Al-attar’s interest in cryogenics began with the wood frog (Rana sylvatica), known for its ability to endure freezing temperatures and transition from suspended animation to full functionality. This curiosity led her to pursue a Master’s degree in Dr. Ken Storey’s lab at Carleton University, where she investigated the molecular mechanisms of cryobiology. Her research there fueled a broader interest in freeze-tolerant and hibernating species. Her passion grew during her Ph.D. studies, where she explored survival strategies of hibernating animals like bears, bats and squirrels.

“Towards the end of my Ph.D., I sought to apply my understanding of cryobiology to clinically relevant models, which led me to join Dr. Michael Laflamme’s lab at the McEwen Stem Cell Institute,” she explains. In Laflamme’s lab, she worked on hPSC-CMs aimed at repairing damaged hearts. Despite advancements, arrhythmias remained a significant challenge. Dr. Al-attar noted, “These arrhythmias are partly attributed to the immature characteristics of hPSC-CMs, which differ from those of mature adult cardiomyocytes.” Efforts to mature the cells through electrical stimulation or metabolic supplementation compromised their freezing tolerance, highlighting the need for better cryopreservation techniques.

Challenges and Innovations in Cryopreservation

To address the issues associated with arrhythmias, Dr. Al-attar moved to Dr. Tessier’s lab, where her research focuses on improving organ preservation and developing protocols for freezing complex organoids. “By learning from nature’s strategies used by freeze-tolerant frogs and hibernating mammals,” she details, “I aim to enhance organ preservation techniques.” This work could improve the viability of transplantable organs, addressing the problem of discarded human organs due to short ex vivo viability periods.

Dr. Al-attar’s early postdoctoral work significantly shaped her focus on cryopreservation and stem cell transplantation. During this time, she studied the electrophysiology of hPSC-CMs and their arrhythmias. Despite a 60-70% viability rate post-thaw using conventional methods, immature hPSC-CMs presented challenges, including arrhythmias. She recalls, “As we explored the literature, it became evident that metabolically mature hPSC-CMs offered superior functional properties compared to their immature counterparts for both in vitro testing and transplantation.” However, mature cells had lower viability after thawing, prompting her to investigate cryopreservation techniques for these samples. “Recognizing the need for deeper insights into cryopreservation techniques for medically relevant samples,” she notes, “I decided to join a lab dedicated to this specialty.”

In Laflamme’s lab, Dr. Al-attar learned that while progress had been made with hPSC-CMs, gaps remained in understanding cryopreservation’s impact on these cells. “Most existing studies provide inconsistent results regarding their viability and functionality post-thaw,” she explains. Moving to Dr. Tessier’s lab allowed her to apply insights from her prior work to tailor solutions for organ preservation and stem cell research.

Dr. Al-attar’s work on hPSC-CMs highlights critical challenges, such as mechanical displacement due to the heart’s pulsatile nature, which impacts cell integration and functionality. “Injecting hPSC-CMs into a contracting heart presents notable difficulties due to the heart’s pulsatile nature, which can lead to mechanical displacement of the cells from their targeted injection site,” she notes. This issue, combined with acute ischemia causing a 90% cell loss within the first 24 hours, underscores the need for improved techniques.

Cryopreservation introduces additional complexities. Dr. Al-attar notes. “Immature hPSC-CMs generally exhibit better tolerance to ischemic conditions compared to their metabolically mature counterparts.” While immature cells have higher post-thaw viability, Dr. Al-attar notes, they can cause arrhythmias, presenting a Catch-22 scenario where increased ischemia tolerance comes with a risk of arrhythmia.” Mature hPSC-CMs, though more susceptible to ischemia, have heightened metabolic demands. The challenge is balancing ischemia tolerance with arrhythmia risk.

To address these issues, Dr. Al-attar emphasizes enhancing hPSC-CMs’ maturation before transplantation. Techniques such as electrical stimulation, mechanical stretch, gene editing and pharmacological interventions are being explored. Cardiac patches and microtissues, though promising, currently cannot be cryopreserved, presenting practical challenges.

Dr. Tessier’s lab utilizes nature-inspired methods, drawing from organisms like the wood frog. Dr. Al-attar elaborates, “Dr. Tessier’s lab has developed a novel preservation protocol incorporating cryoprotectants such as glucose, inspired by the wood frog’s ability to survive freezing.” This approach has extended liver tissue viability, offering new possibilities for organ preservation.

Further, the lab’s work on zebrafish larvae exemplifies these techniques. “By preserving zebrafish embryos or larvae, researchers can maintain the wealth of transgenic lines available, facilitating long-term storage and distribution for research,” Dr. Al-attar explains. This method benefits genetic diversity and provides insights into preserving metabolically active cells like hPSC-CMs.

Preserving hearts at normothermic levels offers notable advantages. As Dr. Al-attar describes, “At normothermic temperatures, the heart remains in a state that closely mirrors physiological conditions, which is vital given the strong connection between metabolic fitness and arrhythmias.” This allows for real-time monitoring of electrical propagation and precise arrhythmia management. “Real-time monitoring enables researchers to pinpoint specific areas of damage and identify the origins of arrhythmias,” she notes, enhancing therapeutic interventions.

Future Directions and Impact on Regenerative Medicine

Looking forward, Dr. Al-attar’s research could revolutionize cryopreservation and transplantation. “By harnessing the power of hibernator biology, I aim to revolutionize the field of cryopreservation and organ transplantation,” she states. Her future work includes developing stress-resistant hPSC-CMs and extending human organ preservation times, advancing regenerative medicine. Dr. Al-attar’s interdisciplinary approach, integrating cryogenics, stem cell biology and organ preservation, is crucial in addressing complex issues. She highlights, “Combining perspectives from different scientific fields has been instrumental in addressing complex issues that might be overlooked when using a single approach.” Collaborations with experts, including Dr. Rohil Jain’s work on Raman spectroscopy and Dr. Michael Garton’s expertise in genome engineering, exemplify the value of cross-disciplinary
efforts.

Dr. Al-attar’s work, blending fundamental research with practical applications, aims to overcome key challenges in cryopreservation and regenerative medicine. As she progresses toward securing a faculty position, she envisions using nature-inspired methods to enhance cryopreservation and transplantation. She advises aspiring scientists to “stay curious, adaptable and committed,” emphasizing the importance of interdisciplinary collaboration and mentorship.

Dr. Al-Attar's long-term goals include improving cryopreservation techniques, developing ischemia-resistant hPSC-CMs and extending organ preservation times for both ex vivo testing and transplantation, all with the goal of revolutionizing regenerative medicine and transplantation. Cold Facts will continue to follow Dr. Al-attar’s research. hms.harvard.edu

Image: Dr. Rasha Al-attar is advancing heart regeneration by developing nature-inspired cryopreservation techniques to improve stem cell-derived cardiomyocytes and organ preservation. Credit: Harvard Medical School and the Massachusetts General Hospital