In vitro-transfusional model for red-blood-cell study: the advantage of lowering hematocrit

The quality of red blood cells (RBCs) stored in red cell concentrates (RCCs) is influenced by processing, storage and donor characteristics, and can have a clinical impact on transfused patients. To evaluate RBC properties and their potential impact in a transfusion setting, a simple in vitro-transfusional model has been developed.

Transfusion was simulated by mixing a washed RBC pool from two male-derived RCCs stored at 4°C with a pool of 15 male-derived fresh frozen plasma (FFP) units, representing the recipient, at a hematocrit (HCT) of 30% ("control" setting) or 5% (alternative model). The mixtures were incubated at 37°C, 5% of CO₂ up to 48 h. Different metabolites, hemolysis and microvesicles (MVs) were quantified at several incubation times and RBC-morphology changes and deformability after incubation. For each model, biological triplicates have been investigated with RCCs at storage days 2 and 43.

The 5%-HCT model restored the 2,3-DPG level and maintained the ATP level. Furthermore, glucose consumption and corresponding lactate production were increased in the 5%- vs the 30%-HCT condition. Lower hemolysis was observed with 5%-HCT, but only at day 2. However, morphological analysis by digital holographic microscopy (DHM) revealed a decreased fraction of discocytes at 5% rather than at 30% of HCT at storage day 2 but at day 43, the trend was inverted. Concordantly, RBCs incubated at 5% of HCT were more deformable than at 30% at day 43 (p<0.0001).

Higher metabolic activity of RBCs in the 5%-HCT condition was promoted by a higher glucose availability and limited cell-waste accumulation. The conditions of the new proposed model thus enabled rejuvenation of RBCs and maintained them in a physiological-close state in contrast to the 30%-HCT model. It may be used as a first approach to evaluate e.g., the impact of donor and recipient characteristics on RBC properties.