One of the most common life-saving medical procedures is a red blood cell (RBC) transfusion. Unfortunately, RBCs for transfusion have a limited shelf life after donation due to detrimental storage effects on their morphological and biochemical properties. Inspired by nature, a biomimetics approach was developed to preserve RBCs for long-term storage using compounds found in animals with a natural propensity to survive in a frozen or desiccated state for decades. Trehalose was employed as a cryoprotective agent and added to the extracellular freezing solution of porcine RBCs. Slow cooling (-1 C min-1) resulted in almost complete hemolysis (1 ± 1 % RBC recovery), and rapid cooling rates had to be used to achieve satisfactory cryopreservation outcomes. After rapid cooling, the highest percentage of RBC recovery was obtained by plunging in liquid nitrogen and thawing at 55 C, using a cryopreservation solution containing 300 mM trehalose. Under these conditions, 88 ± 8 % of processed RBCs were recovered and retained hemoglobin (14 ± 2 % hemolysis). Hemoglobin’s oxygen-binding properties of cryopreserved RBCs were not significantly different to unfrozen controls and was allosterically regulated by 2,3-bisphosphoglycerate. These data indicate the feasibility of using trehalose instead of glycerol as a cryoprotective compound for RBCs. In contrast to glycerol, trehalose-preserved RBCs can potentially be transfused without time-consuming washing steps, which significantly facilitates the usage of cryopreserved transfusible units in trauma situations when time is of the essence.
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Assessment of stored red blood cells through lab-on-a-chip technologies for precision transfusion medicine
Transfusion of red blood cells (RBCs) is one of the most valuable and widespread treatments in modern medicine. Lifesaving RBC transfusions are facilitated by the cold storage of RBC units in blood banks worldwide. Currently, RBC storage and subsequent transfusion practices are performed using simplistic workflows. More specifically, most blood banks follow the “first-in-first-out” principle to avoid wastage, whereas most healthcare providers prefer the “last-in-first-out” approach simply favoring chronologically younger RBCs. Neither approach addresses recent advances through -omics showing that stored RBC quality is highly variable depending on donor-, time-, and processing-specific factors. Thus, it is time to rethink our workflows in transfusion medicine taking advantage of novel technologies to perform RBC quality assessment. We imagine a future where lab-on-a-chip technologies utilize novel predictive markers of RBC quality identified by -omics and machine learning to usher in a new era of safer and precise transfusion medicine.
more » « less- Award ID(s):
- 1941543
- NSF-PAR ID:
- 10471504
- Publisher / Repository:
- NSF
- Date Published:
- Journal Name:
- Proceedings of the National Academy of Sciences
- Volume:
- 120
- Issue:
- 32
- ISSN:
- 0027-8424
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Abstract Red blood cell (RBC) transfusions facilitate many life-saving acute and chronic interventions. Transfusions are enabled through the gold-standard hypothermic storage of RBCs. Today, the demand for RBC units is unfulfilled, partially due to the limited storage time, 6 weeks, in hypothermic storage. This time limit stems from high metabolism-driven storage lesions at +1-6 °C. A recent and promising alternative to hypothermic storage is the supercooled storage of RBCs at subzero temperatures, pioneered by our group. Here, we report on long-term supercooled storage of human RBCs at physiological hematocrit levels for up to 23 weeks. Specifically, we assess hypothermic RBC additive solutions for their ability to sustain supercooled storage. We find that a commercially formulated next-generation solution (Erythro-Sol 5) enables the best storage performance and can form the basis for further improvements to supercooled storage. Our analyses indicate that oxidative stress is a prominent time- and temperature-dependent injury during supercooled storage. Thus, we report on improved supercooled storage of RBCs at −5 °C by supplementing Erythro-Sol 5 with the exogenous antioxidants, resveratrol, serotonin, melatonin, and Trolox. Overall, this study shows the long-term preservation potential of supercooled storage of RBCs and establishes a foundation for further improvement toward clinical translation.
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Medical planning for space exploration is based on the “floating” blood bank model to store life-saving red blood cells (RBCs) for emergencies. The “floating” blood bank approach is not sufficient in cases where multiple crewmembers are affected by space anemia. In these situations, long-term preserved RBCs will be vital to guarantee the health and safety of crew members. Transfusable RBC units can only be refrigerated for 42 days or frozen at -80 C. However, storing frozen RBCs at -80 C is challenging during the confined condition of long-duration space flight. Freeze-dried, viable RBCs would be an appropriate alternative because they can be stored without cooling, are predicted to have a shelf-life of years, and could be transfused immediately after rehydration. This study explores if freeze-dried RBCs can be rehydrated and transfused in reduced gravity with similar outcomes in recovery as observed at Earth gravity. Experiments analyzing freeze-dried RBC recoveries, rehydration fluid dynamics, and transfusion flow rates were analyzed utilizing an experimental glovebox in simulated 0 g during parabolic flights. RBC recoveries and rehydration fluid dynamics for volumes of 5 mL and 10 mL were the same in simulated 0 g compared to results obtained at 1 g. A clinically acceptable range of flow rates for slow intravenous infusion and rapid fluid resuscitation was possible with the simple augmentation of a hand-pumped clinical pressure bag around a unit of rehydrated RBCs. The results demonstrate the potential feasibility of using freeze-dried cells for healthcare during deep-space exploration.more » « less
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Abstract In sickle cell disease (SCD), ‘disease severity’ associates with increased RBC adhesion to quiescent endothelium, but the impact on activated endothelium is not known. Increased concentrations of free heme result from intravascular hemolysis in SCD. Heme is essential for aerobic metabolism and plays an important role in numerous biological processes. Excess free heme induces reactive oxygen species generation and endothelial activation, which are associated with cardiovascular disorders including atherosclerosis, hypertension, and thrombosis. Here, we utilized an endothelialized microfluidic platform (Endothelium‐on‐a‐chip) to assess adhesion of sickle hemoglobin‐containing red blood cells (HbS RBCs), from adults with homozygous SCD, to heme‐activated human endothelial cells (EC)
in vitro . Confluent EC monolayers in microchannels were treated with pathophysiologically relevant levels of heme in order to simulate the highly hemolytic intravascular milieu seen in SCD. RBC adhesion to heme‐activated ECs varied from subject to subject, and was associated with plasma markers of hemolysis (LDH) and reticulocytosis, thereby linking those RBCs that are most likely to adhere with those that are most likely to hemolyze. These results re‐emphasize the critical contribution made by heterogeneous adhesive HbS RBCs to the pathophysiology of SCD. We found that adhesion of HbS RBCs to heme‐activated ECs varied amongst individuals in the study population, and associated with biomarkers of hemolysis and inflammation, age, and a recent history of transfusion. Importantly, the microfluidic approach described herein holds promise as a clinically feasible Endothelium‐on‐a‐chip platform with which to study complex heterocellular adhesive interactions in SCD. -
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m RBCs pass through 0.3 m-wide slits remains enigmatic. Does the slit caliber increase during RBC passage as sometimes suggested? Here, we elucidated the mechanisms that govern the RBC retention or passage dynamics in slits by combining multiscale modeling, live imaging, and microfluidic experiments on an original device with submicron-wide physiologically calibrated slits. We observed that healthy RBCs pass through 0.28 m-wide rigid slits at 37 °C. To achieve this feat, they must meet two requirements. Geometrically, their surface area-to-volume ratio must be compatible with a shape in two tether-connected equal spheres. Mechanically, the cells with a low surface area-to-volume ratio (28% of RBCs in a 0.4 m-wide slit) must locally unfold their spectrin cytoskeleton inside the slit. In contrast, activation of the mechanosensitive PIEZO1 channel is not required. The RBC transit time through the slits follows a 1 and 3 power law with in-slit pressure drop and slip width, respectively. This law is similar to that of a Newtonian fluid in a two-dimensional Poiseuille flow, showing that the dynamics of RBCs is controlled by their cytoplasmic viscosity. Altogether, our results show that filtration through submicron-wide slits is possible without further slit opening. Furthermore, our approach addresses the critical need for in vitro evaluation of splenic clearance of diseased or engineered RBCs for transfusion and drug delivery.
- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1073/pnas.2115616120
