Department of Surgery, University of Cincinnati, Cincinnati, Ohio.
Department of Surgery, University of Cincinnati, Cincinnati, Ohio.
J Surg Res. 2024 Nov;303:429-438. doi: 10.1016/j.jss.2024.09.042. Epub 2024 Oct 18.
Large-volume packed red blood cell (pRBC) transfusion is associated with lung injury and worsened outcomes. Amitriptyline reduces lung injury and inflammation in a murine sepsis model. We hypothesized that red cell microparticles (MP) activate endothelial cells, leading to lung injury and that treatment with amitriptyline would blunt the inflammatory response MPs through inhibition of acid sphingomyelinase (ASM).
Murine pRBCs were obtained from C57Bl/6 mice and stored in AS3 for 14 d. The MPs were isolated from pRBCs by serial centrifugation. Mouse lung endothelial cells (MLECs) were pretreated with amitriptyline (0, 2.5, 25, 27 μM, n = 5) for 30 min prior to MP treatment. Chemokine secretion and adhesion molecule shedding was assessed. ASM activity was measured from cell lysates.
MPs increased the secretion of chemokines and shedding of adhesion molecules in MLECs at both four and 24 h. Amitriptyline treatment of MLECs decreased ASM activity in the setting of MPs. Amitriptyline pretreatment decreased the secretion of chemokines and shedding of adhesion molecules in response to MPs at 4 h but did not decrease adhesion molecule shedding at 24 h CONCLUSIONS: Endothelial cell treatment with MPs induces secretion of chemokines responsible for chemotaxis (keratinocyte chemoattractant, regulated upon activation normal T cell expressed and presumably secreted, and G-granulocyte colony-stimulating factor) as well as many downstream proinflammatory effects (interleukin-6). Additionally, MPs induce adhesion molecule shedding (vascular cell adhesion molecule-1, intracellular adhesion molecule-1, P-selectin, and E-selectin), which has been shown to be associated with endothelial cell activation. Amitriptyline pretreatment decreases MLEC inflammatory response and ASM activity is decreased. These data suggest that ASM inhibition in MLECs is a potential strategy to blunt the inflammatory response to the red blood cell storage lesion.
大量输注浓缩红细胞（pRBC）与肺损伤和预后恶化有关。阿米替林可减轻小鼠脓毒症模型中的肺损伤和炎症。我们假设红细胞微粒（MP）激活内皮细胞，导致肺损伤，并且通过抑制酸性鞘磷脂酶（ASM），阿米替林治疗会减弱 MPs 的炎症反应。
从 C57Bl/6 小鼠中获得小鼠 pRBC，并在 AS3 中储存 14 天。通过连续离心从 pRBC 中分离 MP。用阿米替林（0、2.5、25、27μM，n=5）预处理小鼠肺内皮细胞（MLEC）30min，然后用 MP 处理。评估趋化因子分泌和黏附分子脱落。从细胞裂解物中测量 ASM 活性。
MP 在 4 和 24 小时均可增加 MLEC 中趋化因子的分泌和黏附分子的脱落。在 MPs 的情况下，阿米替林处理 MLEC 可降低 ASM 活性。阿米替林预处理可减少 4 小时时 MPs 引起的趋化因子分泌和黏附分子脱落，但不能减少 24 小时时黏附分子脱落。
内皮细胞用 MPs 处理会诱导趋化因子（角质细胞趋化因子、激活正常 T 细胞表达和假定分泌的调节因子、G-粒细胞集落刺激因子）的分泌，以及许多下游促炎作用（白细胞介素-6）。此外，MP 诱导黏附分子脱落（血管细胞黏附分子-1、细胞间黏附分子-1、P 选择素和 E 选择素），这与内皮细胞激活有关。阿米替林预处理可降低 MLEC 的炎症反应，并且 ASM 活性降低。这些数据表明，在 MLEC 中抑制 ASM 可能是减弱对红细胞储存损伤的炎症反应的一种策略。
