Introduction Ex-vivo normothermic limb perfusion (EVNLP) has been proven to preserve limb viability better than standard cold storage. Perfusates containing packed red blood cells (pRBC) improve outcomes when compared to acellular perfusates. Limitations of pRBC-based perfusion include limited availability, need for cross match, mechanical hemolysis, and activation of pro-inflammatory proteins. Hemoglobin-based oxygen carrier (HBOC)-201 (Hemopure) is a solution of polymerized bovine hemoglobin, characterized by low immunogenicity, no risk of hemolytic reaction, and enhanced convective and diffusive oxygen delivery. This is a preliminary study on the feasibility of EVNLP using HBOC-201 as an oxygen carrier. Materials and Methods Three porcine forelimb perfusions were performed using an established EVNLP model and an HBOC-201-based perfusate. The perfusion circuit included a roller pump, oxygenator, heat exchanger, and reservoir. Electrolytes, limb temperature, weight, compartment pressure, nerve conduction, and perfusion indicated by indocyanine green angiography and infra-red thermography were monitored. Histological evaluation was performed with hematoxylin and eosin and electron microscopy. Results Three limbs were perfused for 21.3 ± 2.1 hours. Muscle contractility was preserved for 10.6 ± 2.4 hours. Better preservation of the mitochondrial ultrastructure was evident at 12 hours in contrast to crystallization and destruction features in the cold-storage controls. Conclusions An HBOC-201-EVNLP produced outcomes similar to RBC-EVNLP with preservation of muscle contractility and mitochondrial structure.
Original Clinical Science-General Background. Ischemia-reperfusion injury remains a primary concern in upper extremity transplantation. Ex vivo normothermic perfusion (EVNP) enables near-physiological organ preservation, avoiding the deleterious effects of hypoxia and cooling. We investigated the effectiveness of human limb EVNP compared with static cold storage (SCS). Methods. Twenty human upper extremities were procured. Ten were perfused at 38 °C with an oxygenated red blood cell-based solution, and contralateral limbs served as SCS control (4 °C). EVNP was terminated with systolic arterial pressure ≥115 mm Hg, compartment fullness, or a 20% decline in oxygen saturation. Weight, contractility, compartment pressure, tissue oxygen saturation, and uptake rates were assessed. Perfusate fluid dynamics, gases, electrolytes, and metabolites were measured. Myocyte injury scores and liquid chromatography-mass spectrometry analysis were performed. Results. EVNP duration was 41.6 ± 9.4 h. Vascular resistance averaged 173.0 ± 29.4 mm Hg × min/L. Weight change and compartment pressures were 0.4 ± 12.2% (P = 0.21) and 21.7 ± 15.58 mm Hg (P = 0.003), respectively. Arterial and venous carbon dioxide partial pressure, oxygen saturation, and pH were 509.5 ± 91.4 mm Hg, 15.7 ± 30.2 mm Hg, 87.4 ± 11.4%, and 7.3 ± 0.2, respectively. Oxygen uptake rates averaged 5.7 ± 2.8 mL/min/g. Lactate reached 20 mmol/L after 15 (interquartile range = 6) h. Limb contractility was preserved for 30.5 (interquartile range = 15.8) h (P < 0.001) and negatively correlated with perfusate potassium (ρ = -0.7, P < 0.001). Endpoint myocyte injury scores were 28.9 ± 11.5% (EVNP) and 90.2 ± 11.8% (SCS) (P < 0.001). A significant increase in taurine (P = 0.002) and decrease in tryptophan (P = 0.002) were detected. Infrared thermography and indocyanine green angiography confirmed the presence of peripheral perfusion. Conclusions. EVNP can overcome the limitations of cold preservation by extending preservation times, enabling limb quality assessment, and allowing limb reconditioning before transplantation.
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