Presentation description

Hemorrhagic shock is a life-threatening condition resulting from severe blood loss, often leading to ischemia-reperfusion injury (IRI), multi-organ failure, and death. Mitochondria play a central role in this process, as their function is critically compromised during and after ischemia. Transplantation of healthy, functional mitochondria has emerged as a promising strategy to restore cellular bioenergetics and improve survival after IRIs. While mitochondrial transplantation is successful in controlled settings, adapting it for emergency use requires overcoming a significant limitation. Freshly isolated mitochondria have a fragile and short-lived nature. This project focuses on developing field-deployable mitochondrial therapies by evaluating whether mitochondria can be frozen or lyophilized for long-term storage without compromising function at the time of transfusion. My subproject specifically investigates whether our freezing protocol preserves mitochondrial membrane potential and oxygen consumption after thawing, two key indicators of mitochondrial integrity and viability. Freezing can damage mitochondria by forming ice crystals that physically disrupt the inner membrane, causing the leakage of soluble matrix enzymes, such as malate dehydrogenase and glutamate dehydrogenase, which are essential for NADH production and complex I-linked respiration. Previous studies suggest that swine mitochondria retain post-thaw function, but it remains unclear whether this resilience is species-specific or tissue-dependent.
Using mitochondria isolated from mouse pelvic leg muscle and pig abdominal wall muscle, we compare fresh and frozen/thawed samples (frozen at a controlled rate of −1°C/minute). Mitochondrial function is assessed using O2 flux measurements and Safranin-O fluorescence to evaluate membrane potential. By analyzing oxygen consumption and membrane potential simultaneously, we aim to determine whether freezing-induced loss of function is due to ETC disruption or substrate loss, and whether mitochondrial preservation varies across different species.