Toward Translational Impact of Low-Glucose Strategies on Red Blood Cell Storage Optimization.

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.

UPLC-MS/MS method for quantitative determination of the advanced glycation endproducts Nε-(carboxymethyl)lysine and Nε-(carboxyethyl)lysine.

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.

A 3D-printed, multi-modal microfluidic device for measuring nitric oxide and ATP release from flowing red blood cells.

In this paper, a 3D-printed multi-modal device was designed and fabricated to simultaneously detect nitric oxide (NO) and adenosine triphosphate (ATP) in red blood cell suspensions prepared from whole blood. Once a sample was injected into the device, NO was first detected (via amperometry) using a three-electrode, dual-opposed, electrode configuration with a platinum-black/Nafion coated gold working electrode. After in-line amperometric detection of NO, ATP was detected via a chemiluminescence reaction, with a luciferin/luciferase solution continuously pumped into an integrated mixing T and the resulting light being measured with a PMT underneath the channel. The device was optimized for mixing/reaction conditions, limits of detection (40 nM for NO and 30 nM for ATP), and sensitivity. This device was used to determine the basal (normoxic) levels of NO and ATP in red blood cells, as well as an increase in concentration of both analytes under hypoxic conditions. Finally, the effect of storing red blood cells in a commonly used storage solution was also investigated by monitoring the production of NO and ATP over a three-week storage time.

Sol-Gel-Derived Bioactive and Antibacterial Multi-Component Thin Films by the Spin-Coating Technique.

Although metallic alloys commonly used as prosthetics are durable and mechanically strong, they are often bioinert and lack antibacterial properties. Implementing a bioactive glass material with antibacterial properties as a coating on a metallic substrate provides mechanical strength and bioactivity, as well as antibacterial properties. Many coating methods have been extensively investigated; however, most of them can be expensive, are difficult to scale up, or do not form thin films, which could prevent their translation to clinical practice. The formation of thin films by spin-coating multi-component solution-gelation (sol-gel)-derived glass with antibacterial and bioactive properties has not been achieved previously. For this study, stainless steel 316L substrates were spin-coated with a sol-gel-derived bioactive and antibacterial glass coating in SiO2 58.3-P2O5 7.1-CaO 25.6-Al2O5 5.4-Ag2O 2.1-Na2O 1.5 wt% system (Ag-BG). A sol-gel processing condition that avoids elemental separation upon spin-coating when sintering happens at below the calcination temperature (500 °C) has been developed. This work demonstrates that silver reduction occurs when the concentrations of other cations such as Ca2+ and Na+ in the solution increase. Increasing the stirring duration time prior to the increase of cations, Ag+ ions are stabilized by aluminum tetrahedra, and their reduction to metallic silver does not occur. This study also shows that large dilution ratios (water:tetraethyl orthosilicate) greater than 25:1, accompanied by long stirring durations, produce morphologically homogeneous coatings. Using this strategy, thin films were formed with antibacterial properties against methicillin-resistant Staphylococcus aureus (MRSA) biofilm and biological responses that promote eukaryotic cell adhesion and proliferation. In total, the improved synthesis strategy opens new avenues for the development of novel bioactive and antibacterial thin-film coatings, as it reveals the processing characteristics that control the physicochemical and morphological properties of the formed films.