McQuinn Erin R, Smith Stephanie A, Viall Austin K, Wang Chong, LeVine Dana N
Department of Veterinary Clinical Sciences, Iowa State University, Ames, Iowa, USA.
Department of Biochemistry, College of Medicine, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
J Vet Intern Med. 2020 Sep;34(5):1894-1902. doi: 10.1111/jvim.15876. Epub 2020 Sep 3.
Neutrophil extracellular traps (NETs), webs of DNA and citrullinated histones extruded from activated neutrophils cause transfusion-related acute lung injury. Supernatants of stored red blood cell (RBC) units might promote NETosis in neutrophils from the units or from transfusion recipients.
(1) NETs form during storage of canine RBC, (2) leukoreduction (LR) before storage of RBC reduces NETosis, and (3) supernatant from stored, nonleukoreduced (NLR) RBC units induces NETosis in healthy canine neutrophils modeling transfusion recipients.
Six healthy purpose-bred research dogs were utilized for blood donation.
Prospective controlled study. RBC units were collected from each dog, aseptically divided into 2 equal subunits, 1 of which was leukoreduced, and stored for 42 days. Stored units were sampled biweekly for quantification of NET markers citrullinated histone H3 (Western blot) and cell-free DNA (cfDNA) (DNA dye binding). Unit supernatants were applied ex vivo to canine neutrophils and extracellular DNA release representing NETosis was assessed.
Markers of NETs increased during RBC storage (cfDNA P < .0001 and citrullinated H3 P = .0002) and were higher in NLR than LR units (day 42 LR cfDNA 0.34 ± 0.82 ng/mL vs day 42 NLR 1361.07 ± 741.00 ng/mL, P < .0001; day 42 LR citrullinated H3 0.19 ± 0.13 AU vs NLR 0.57 ± 0.34 AU, P = .007). Isolated neutrophils did not form NETs when exposed to stored canine RBC supernatant.
NETosis occurs in stored canine NLR RBC units, and is attenuated by LR before storage. NETs might be mediators of transfusion reactions.
中性粒细胞胞外陷阱（NETs）是由活化的中性粒细胞挤出的DNA和瓜氨酸化组蛋白构成的网络，可导致输血相关急性肺损伤。储存的红细胞（RBC）单位的上清液可能会促进来自这些单位或输血受者的中性粒细胞发生NETosis。
（1）犬RBC储存期间会形成NETs；（2）RBC储存前进行白细胞滤除（LR）可减少NETosis；（3）储存的未进行白细胞滤除（NLR）的RBC单位的上清液可在模拟输血受者的健康犬中性粒细胞中诱导NETosis。
使用6只健康的专门培育的研究犬进行献血。
前瞻性对照研究。从每只犬采集RBC单位，无菌分为2个相等的亚单位，其中1个进行白细胞滤除，并储存42天。每两周对储存的单位进行采样，以定量NET标志物瓜氨酸化组蛋白H3（蛋白质免疫印迹法）和游离DNA（cfDNA）（DNA染料结合法）。将单位上清液体外应用于犬中性粒细胞，并评估代表NETosis的细胞外DNA释放情况。
NETs的标志物在RBC储存期间增加（cfDNA P <.0001，瓜氨酸化H3 P =.0002），且NLR单位中的标志物高于LR单位（第42天LR的cfDNA为0.34±0.82 ng/mL，而第42天NLR的为1361.07±741.00 ng/mL，P <.0001；第42天LR的瓜氨酸化H3为0.19±0.13 AU，而NLR的为0.57±0.34 AU，P =.007）。分离的中性粒细胞在暴露于储存的犬RBC上清液时未形成NETs。
NETosis发生于储存的犬NLR RBC单位中，且在储存前通过LR可使其减弱。NETs可能是输血反应的介质。
