Impact of nucleation temperature and hydroxyethyl starch on ice crystal growth: Implications for cell viability during extreme temperature fluctuations.

Extreme temperature fluctuations during routine handling and shipping of cryopreserved cell products significantly compromise product quality in ways that extend beyond the duration and peak temperature of the fluctuation. The type of cryoprotectant used and the initial ice nucleation temperature influence ice crystal growth during rewarming events, in turn impacting cell survival. Using a cryomicroscope together with temperature profiles recorded in cord-blood units, ice crystal growth was tracked through five transient-warming events (TWEs) that peaked at -30 °C, -20 °C, or -10 °C.

Innovations in red blood cell preservation.

The global infrastructure supporting nearly 100 million transfusions annually relies on the ability to store red cell concentrates (RCCs) for up to 42 days at hypothermic temperatures or indefinitely at low sub-zero temperatures. While these methods are generally effective, there is both an opportunity and, in specific settings, a need to refine storage techniques that have remained largely unchanged since the 1980s. Recent research has identified ways to address limitations that were not fully understood when these methods were first implemented in blood banks, with much of it focusing on modifying conventional storage strategies, while some studies explore alternative approaches.

Supercooled storage of red blood cells slows down the metabolic storage lesion.

Red blood cell (RBC) transfusion, a life-saving intervention, is limited by reduced RBC potency over time. Cold storage at +4 °C for up to 42 days can reduce transfusion efficacy due to alterations termed the "storage lesion." Strategies to mitigate the storage lesion include alkaline additive solutions and supercooled storage to extend storage by reducing metabolic stresses.

Cold storage surpasses the impact of biological age and donor characteristics on red blood cell morphology classified by deep machine learning.

Assessment of the morphology of red blood cells (RBCs) can improve clinical benefits following blood transfusion. Deep machine learning surpasses traditional microscopy-based classification methods, offering more accurate and consistent results while reducing time and labor intensity. RBCs from teenage males, teenage females, senior males, and senior females were biologically age-profiled or density-separated into dense/old RBCs (O-RBCs) and less-dense/young (Y-RBCs) throughout hypothermic storage and assessed using image flow cytometry with deep machine learning analysis.

Deglycerolization of manually glycerolized, frozen red cell concentrates using a closed system cell processor.

Background: Historically, red cell concentrates (RCCs) have been manually glycerolized and deglycerolized using an open system (COBE 2991, Terumo). Implementation of a closed system cell processor (ACP-215, Haemonetics) for glycerolization and deglycerolization of RCCs creates a challenge for management of the historic cryopreserved RCC inventory. A study was undertaken to determine whether manually glycerolized frozen RCCs could be deglycerolized using the closed system processor, as the open system processors are being discontinued.

Impact of input volume on red cell quality in deglycerolized RBCs using a modified ACP-215 protocol.

Background: The ACP 215 automated cell processor is used to glycerolize and deglycerolize red cell concentrates (RCCs). Its primary advantage over the COBE 2991, previously used to cryopreserve RCCs, is that it maintains a closed system enabling extended post-thaw expiry. However, it was observed that post-deglycerolization hematocrits (Hct) of units processed with the LN236 kit are markedly lower than those processed using the COBE 2991.

RBC subpopulations in RCCs affected by donor factors.

Understanding red blood cell (RBC) subpopulations is crucial for comprehending donor variability and enhancing transfusion outcomes. This review highlights the significance of RBC subpopulations, focusing on the properties of biologically young and old RBCs and underscores how donor variability impacts transfusion outcomes. The role of senescent RBCs in adverse transfusion reactions and the emerging significance of circulating erythroid cells (CECs) is discussed.

Shipment of Glycerolized RBC Segments for Red Cell Concentrate Compatibility Testing.

Red cell concentrate (RCC) cryopreservation allows for long-term storage of RCCs with rare phenotypes. Currently, tubing segments are not produced for these frozen units. Pre-transfusion compatibility testing therefore requires thawing and deglycerolization of the whole unit.

Extended supercooled storage of red blood cells.

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.

A perspective on exogenous redox regulation mediated by transfused RBCs subject to the storage lesion.

Granted with a potent ability to interact with and tolerate oxidative stressors, RBCs scavenge most reactive oxygen and nitrogen species (RONS) generated in circulation. This essential non-canonical function, however, renders RBCs susceptible to damage when vascular RONS are generated in excess, making vascular redox imbalance a common etiology of anemia, and thus a common indication for transfusion. This accentuates the relevance of impairments in redox metabolism during hypothermic storage, as the exposure to chronic oxidative stressors upon transfusion could be exceedingly deleterious to stored RBCs.

Towards the crux of sex-dependent variability in red cell concentrates.

Donor sex can alter the RBC 'storage lesion' progression, contributing to dissimilarities in blood product quality, and thus adverse post-transfusion reactions. The mechanisms underlying the reduced sensitivity of female RBCs to storage-induced stress are partially ascribed to the differential effects of testosterone, progesterone, and estrogen on hemolytic propensity. Contributing to this is the increased proportion of more robust, biologically 'young' subpopulations of RBCs in females.

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.

Comparing two extracellular additives to facilitate extended storage of red blood cells in a supercooled state.

Adenosine triphosphate (ATP) levels guide many aspects of the red blood cell (RBC) hypothermic storage lesions. As a result, efforts to improve the quality of hypothermic-stored red cell concentrates (RCCs) have largely centered around designing storage solutions to promote ATP retention. Considering reduced temperatures alone would diminish metabolism, and thereby enhance ATP retention, we evaluated: (a) whether the quality of stored blood is improved at -4°C relative to conventional 4°C storage, and (b) whether the addition of trehalose and PEG400 can enhance these improvements.

Extracellular vesicles: effectors of transfusion-related acute lung injury.

Respiratory transfusion reactions represent some of the most severe adverse reactions related to receiving blood products. Of those, transfusion-related acute lung injury (TRALI) is associated with elevated morbidity and mortality. TRALI is characterized by severe lung injury associated with inflammation, pulmonary neutrophil infiltration, lung barrier leak, and increased interstitial and airspace edema that cause respiratory failure.

Engineered Compounds to Control Ice Nucleation and Recrystallization.

One of the greatest concerns in the subzero storage of cells, tissues, and organs is the ability to control the nucleation or recrystallization of ice. In nature, evidence of these processes, which aid in sustaining internal temperatures below the physiologic freezing point for extended periods of time, is apparent in freeze-avoidant and freeze-tolerant organisms. After decades of studying these proteins, we now have easily accessible compounds and materials capable of recapitulating the mechanisms seen in nature for biopreser-vation applications.

Donor sex, pre-donation hemoglobin, and manufacturing affect CD71 cells in red cell concentrates.

Background: Circulating CD71 red blood cells (RBCs) have been reported to play an immunomodulatory role in vivo, which may contribute to adverse donor-recipient sex-mismatched transfusion outcomes. However, it is not clear how CD71 RBC quantity in red cell concentrates (RCCs) is affected by manufacturing methods and donor factors such as donor sex, donor age, pre-donation hemoglobin (Hb), venous Hb (Hb ) levels, and donation frequency.

Methods: We determined CD71 RBCs and Hb levels in whole blood (WB) from healthy donors (42 male/38 female).

Preparing small-dose red cell concentrates (RCCs) for neonatal and pediatric transfusions: Impact of RCC volume, storage, and irradiation.

Background: Preparing small-dose red cell concentrates (RCCs) is a common practice for pediatric and neonatal transfusions. However, there is a lack of quality monitoring data to indicate that both the preparation and storage of small-dose RCCs does not alter in vitro red cell quality. The present study seeks to provide data to support this practice.

The response of a human haematopoietic cell line to trehalose-loaded liposomes and their effect on post-thaw membrane integrity.

Dimethyl sulphoxide (DMSO) used in haematopoietic stem cell (HSC) cryopreservation has been linked to an increased incidence of adverse reactions following transplantation. In the interest of reducing the required DMSO concentrations, we have evaluated the use of unilamellar liposomes to internalize the non-toxic, cell-impermeable disaccharide, trehalose into HSCs and characterized the cryoprotective efficacy of this strategy. A fluorescent marker, 5(6)-carboxyfluorescein (200 μmol/L), was used for trehalose internalization following a 5 h incubation at 37 °C with liposome concentrations ranging from 0.

The timing of gamma irradiation and its effect on cell-free and microvesicle-bound hemoglobin. The BEST collaborative study.

Background: Gamma irradiation of red cell concentrates (RCCs) is regularly used to prevent transfusion-associated graft-versus-host disease (TA-GvHD) in at-risk patients. While studies have indicated that irradiated RCCs exhibit increased hemolysis, there have been no efforts to differentiate between free- and microvesicle (MV)-bound hemoglobin (Hb). As an increase in the proportion of free-Hb in irradiated RCCs could alter vascular function, we sought to characterize differences in the state of extracellular Hb based on the timing of irradiation.

High sub-zero organ preservation: A paradigm of nature-inspired strategies.

In recent years there have been several advancements in organ preservation that have yet to see widespread clinical translation. While static cold storage (SCS) at 2 °C-4 °C continues to be the state-of-the-art strategy, it contributes to the current shortage of transplantable organs due to the limited preservation times it affords combined with the limited ability of marginal grafts to tolerate SCS. The era of optimizing storage solutions to minimize SCS-induced hypothermic injury has largely plateaued in its improvements, resulting in a shift towards the use of machine perfusion systems to provide continuous metabolic support, or the use of sub-zero storage temperatures to leverage the protection brought forth by a reduction in metabolic demand.

Transient loss of membrane integrity following intracellular ice formation in dimethyl sulfoxide-treated hepatocyte and endothelial cell monolayers.

Immediate post-thaw evaluation of membrane integrity has proven to yield overestimates of cell survival under conditions that preclude intracellular ice formation (IIF). However, prominent theories on the mechanisms of intracellular nucleation suggest a damaged membrane can reseal, prompting us to evaluate whether immediate post-thaw assessments of membrane integrity can in fact underestimate cell survival under conditions that promote IIF. HUVEC and HepG2 monolayers were treated with 1.

Cryoprotectant-dependent control of intracellular ice recrystallization in hepatocytes using small molecule carbohydrate derivatives.

To promote the recovery of cells that undergo intracellular ice formation (IIF), it is imperative that the recrystallization of intracellular ice is minimized. Hepatocytes are more prone to IIF than most mammalian cells, and thus we assessed the ability of novel small molecule carbohydrate-based ice recrystallization inhibitors (IRIs) to permeate and function within hepatocytes. HepG2 monolayers were treated with N-(4-chlorophenyl)-d-gluconamide (IRI 1), N-(2-fluorophenyl)-d-gluconamide (IRI 2), or para-methoxyphenyl-β-D-glycoside (IRI 3) and fluorescent cryomicroscopy was used for real time visualization of intracellular ice recrystallization.