RESUMO
Transfusion of stored red blood cells (RBCs) to patients is a critical component of human healthcare. Following purification from whole blood, RBCs are stored in one of many media known as additive solutions for up to 42 days. However, during the storage period, the RBCs undergo adverse chemical and physical changes that are often collectively known as the RBC storage lesion. Storage of RBCs in additive solutions modified to contain physiological levels of glucose, as opposed to hyperglycemic levels currently used in most cases, reduces certain markers of the storage lesion, although intermittent doses of glucose are required to maintain normoglycemic conditions. Here, we describe an electrically actuated valving system to dispense small volumes of glucose into 100 mL PVC storage bags containing packed RBCs from human donors. The RBCs were stored in a conventional additive solution (AS-1) or a normoglycemic version of AS-1 (AS-1N) and common markers of stored RBC health were measured at multiple time points throughout storage. The automated feeding device delivered precise and predictable volumes of concentrated glucose to maintain physiological glucose levels for up to 37 days. Hemolysis, lactate accumulation, and pH values of RBCs stored in AS-1N were statistically equivalent to values measured in AS-1, while significant reductions in osmotic fragility and intracellular sorbitol levels were measured in AS-1N. The reduction of osmotic fragility and oxidative stress markers in a closed system may lead to improved transfusion outcomes for an important procedure affecting millions of people each year.
RESUMO
During blood storage, red blood cells (RBCs) undergo physical, chemical, and metabolic changes that may contribute to post-transfusion complications. Due to the hyperglycemic environment of typical solutions used for RBC storage, the formation of advanced glycation endproducts (AGEs) on the stored RBCs has been implicated as a detrimental chemical change during storage. Unfortunately, there are limited studies involving quantitative determination and differentiation of carboxymethyl-lysine (CML) and carboxyethyl-lysine (CEL), two commonly formed AGEs, and no reported studies comparing these AGEs in experimental storage solutions. In this study, CML and CEL were identified and quantified on freshly drawn blood samples in two types of storage solutions, standard additive solution 1 (AS-1) and a normoglycemic version of AS-1 (AS-1N). To facilitate detection of the AGEs, a novel method was developed to reliably extract AGEs from RBCs, provide Food and Drug Administration (FDA) bioanalytical guidance criteria, and enable acceptable selectivity for these analytes. Ultra-performance liquid chromatography with tandem mass spectrometry (UPLC-MS/MS) was utilized to identify and quantify the AGEs. Results show this method is accurate, precise, has minimal interferences or matrix effects, and overcomes the issue of detecting AGE byproducts. Importantly, AGEs can be detected and quantified in both types of blood storage solutions (AS-1 and AS-1N), thereby enabling long-term (6 weeks) blood storage related studies.