Corpuscular Fragility and Metabolic Aspects of Freshly Drawn Beta-Thalassemia Minor RBCs Impact Their Physiology and Performance Post Transfusion: A Triangular Correlation Analysis In Vitro and In Vivo

Abstract

The clarification of donor variation effects upon red blood cell (RBC) storage lesion and transfusion efficacy may open new ways for donor-recipient matching optimization. We hereby propose a "triangular" strategy for studying the links comprising the transfusion chain-donor, blood product, recipient-as exemplified in two cohorts of control and beta-thalassemia minor (βThal⁺) donors (n = 18 each). It was unraveled that RBC osmotic fragility and caspase-like proteasomal activity can link both donor cohorts to post-storage states. In the case of heterozygotes, the geometry, size and intrinsic low RBC fragility might be lying behind their higher post-storage resistance to lysis and recovery in mice. Moreover, energy-related molecules (e.g., phosphocreatine) and purine metabolism factors (IMP, hypoxanthine) were specifically linked to lower post-storage hemolysis and phosphatidylserine exposure. The latter was also ameliorated by antioxidants, such as urate. Finally, higher proteasomal conservation across the transfusion chain was observed in heterozygotes compared to control donors. The proposed "triangularity model" can be (a) expanded to additional donor/recipient backgrounds, (b) enriched by big data, especially in the post-transfusion state and (c) fuel targeted experiments in order to discover new quality biomarkers and design more personalized transfusion medicine schemes.

Keywords: biomarkers; donor variation; energy metabolism; fragility; purine metabolism; red blood cells; transfusion.

Figure 1

Study design. Proposed routes for the linkage between donor parameters and post-storage/post-transfusion variables.

Figure 2

Statistically significant correlations between freshly drawn blood, stored, and reconstituted RBCs from control donors. The correlation triangles focus on osmotic fragility (MCF) and caspase-like (CASP-like) proteasomal activity. All connections represent repeatable (at every recipient plasma and storage time-point measured) significant correlations. The R-values concern late storage and thalassemic plasma.

Figure 3

Statistically significant hemolysis correlations between freshly drawn blood, stored, and reconstituted RBCs from beta-thalassemia trait (βThal⁺) donors. The correlation triangles focus on (A) osmotic, (B) mechanical and (C) spontaneous hemolysis of reconstituted samples. All connections represent repeatable (at every recipient plasma and storage time-point measured) significant correlations. The R-values concern late storage and thalassemic plasma. * SPTA1 was used as an example of an array of structural components that correlate with MFI (e.g., ankyrin-1, glycophorin C and 4.2 protein).

Figure 4

Statistically significant redox-related correlations between freshly drawn blood, stored, and reconstituted RBCs from beta-thalassemia trait (βThal⁺) donors. The correlation triangles focus on (A) PS-exposing RBCs and extracellular vesicles (EVs), (B) proteasome activity and (C) diamide-induced intracellular ROS of reconstituted samples. All connections represent repeatable (at every recipient plasma and storage time-point measured) significant correlations. The R-values concern late storage and thalassemic plasma. Antioxidant capacities of (A) are extracellular, while urate is the intracellular metabolite.

Figure 5

Statistically significant correlations of parameters of freshly drawn blood with the post-reconstitution physiology and 24 h recovery of beta-thalassemia trait (βThal⁺) donated RBCs. (A) Baseline levels of metabolites correlating with physiological parameters of reconstituted RBCs. (B) Baseline fragility indices correlating with post-transfusion recovery in mice. The selected scatterplots concern late storage, thalassemic plasma (A) and C57BL/6J mice (B), but significant connections (with slightly different R² values) were evident at every condition tested (i.e., early/late storage, control/thalassemic plasma, immunodeficient/sufficient mice), with the exception of ATP and hemolysis, a correlation evident only in early storage. PE: phosphoethanolamine; 2,3-BPG: 2,3-bisphosphoglycerate; PS: phosphatidylserine; EVs: extracellular vesicles.