Influence Of Inhaled Nitric Oxide On Stored Red Blood Cell autotransfusion In Overweight Volunteers With Endothelial Dysfunction: Preliminary Results
Authors: Berra Lorenzo , Pinciroli Riccardo , Yu Binglan , Stowell Christopher P. , Bloch Kenneth D. , Zapol Warren M. 
Published: October 2019

Abstract

Introduction
Transfusion of stored packed erythrocytes is associated with excessive intra and extravascular hemolysis. High plasma cell-free hemoglobin levels rapidly scavenge endogenous nitric oxide (NO) leading in many species to systemic and pulmonary vasoconstriction, and a pro-inflammatory, pro-thrombotic state.

Methods
We conducted a crossover randomized interventional study of autologous stored blood transfusion. We enrolled fourteen overweight adults (BMI 28 to 40 Kg/m2) who also had evidence of impaired endothelial function.Endothelial dysfunction was determined by a reduced Reactive Hyperemia Index following 5 minutes of forearm tourniquet ischemia (RHI<1.8).
Over 8 months each volunteer returned to the blood bank on 3 occasions to donate a unit of blood. In a randomized scheme,autologous blood was leukoreduced and stored in AS-3, and re-infused after a) 3-days of storage (3-d), b) 40-days of storage (40-d), or c) 40-days of storage and breathing 80 parts per million (ppm) NO (40-d+NO).
Venous blood was sampled before and at 10min, 1h, 2h, 4h and 24h after autotransfusion.
Plasma hemoglobin measurement Plasma hemoglobin was measured with a QuantiChrom Hemoglobin Assay Kit (BioAssaySystems, Hayward, CA). These samples were collected using 16 G catheters avoiding negative aspirating pressures to minimize ex vivo hemolysis (1).
Plasma NO consumption The ability of plasma hemoglobin to scavenge
NO was measured with an NO consumption assay with a NO chemiluminescenceanalyzer (Sievers, Boulder, CO) (2)

Results
Eleven volunteers completed the entire study. Volunteer characteristics at baseline were (mean ± SD): age: 41± 13 years; sex: 9 males, 5 females; BMI 33.2 ± 4.9 Kg/m2; reactive hyperemia index 1.59±0.18, and plasma levels of C-Reactive Protein 4.75 ±3.44 md/L.
After autotransfusion the plasma hemoglobin and serum iron concentrations were increased from 1 to 4 hours after transfusing 40-days stored blood. No increase was detected after transfusing 3-days stored blood (p<0.01 value differs from b and c) (Figures 1a and 2).
The plasma NO consumption assay showed an increased level in the 40-d group (p<0.05 value differs from both b and c) (Figure 1b). Plasma concentrations of potassium, lactate dehydrogenase, haptoglobin, cytokines, and systemic blood pressure did not differ between a, b and c and all remained within the normal range.

Conclusions
Transfusion of autologous packed erythrocytes stored for 40 days in AS3 is associated with increased hemolysis and increased plasma NO consumption,the latter is prevented by breathing 80 ppm NO.
References
1. Berra L, Coppadoro A, Yu B, Lei C, Spagnolli E, Steinbicker AU, Bloch KD, Lin T, Sammy FY, Warren HS, Fernandez BO, Feelisch M, Dzik WH, Stowell CP, Zapol WM. Transfusion of stored autologous blood does not alter reactive hyperemia index in healthy volunteers. Anesthesiology. 2012;117(1):56-63.
2. Wang X, Tanus-Santos JE, Reiter CD, Dejam A, Shiva S, Smith RD, Hogg N, Gladwin MT. Biological activity of nitric oxide in the plasmatic compartment. ProcNatlAcadSci USA 2004;101:11477-11482.

Disclosures:
Off Label Use: Breathing Nitric Oxide during autologous stored blood transfusion. Bloch:MGH: Patents & Royalties. Zapol:IKARIA: Inhaled Nitric Oxide, Inhaled Nitric Oxide Patents & Royalties.