Department of Surgery, University of Chicago, Chicago, IL, United States.
Biological Sciences Division, University of Chicago, Chicago, IL, United States.
Front Immunol. 2022 Jun 2;13:788935. doi: 10.3389/fimmu.2022.788935. eCollection 2022.
Normothermic machine perfusion (NMP) allows viability assessment and potential resuscitation of donor livers prior to transplantation. The immunological effect of NMP on liver allografts is undetermined, with potential implications on allograft function, rejection outcomes and overall survival. In this study we define the changes in immune profile of human livers during NMP.
Six human livers were placed on a NMP device. Tissue and perfusate samples were obtained during cold storage prior to perfusion and at 1, 3, and 6 hours of perfusion. Flow cytometry, immunohistochemistry, and bead-based immunoassays were used to measure leukocyte composition and cytokines in the perfusate and within the liver tissue. Mean values between baseline and time points were compared by Student's t-test.
Within circulating perfusate, significantly increased frequencies of CD4 T cells, B cells and eosinophils were detectable by 1 hour of NMP and continued to increase at 6 hours of perfusion. On the other hand, NK cell frequency significantly decreased by 1 hour of NMP and remained decreased for the duration of perfusion. Within the liver tissue there was significantly increased B cell frequency but decreased neutrophils detectable at 6 hours of NMP. A transient decrease in intermediate monocyte frequency was detectable in liver tissue during the middle of the perfusion run. Overall, no significant differences were detectable in tissue resident T regulatory cells during NMP. Significantly increased levels of pro-inflammatory and anti-inflammatory cytokines were seen following initiation of NMP that continued to rise throughout duration of perfusion.
Time-dependent dynamic changes are seen in individual leukocyte cell-types within both perfusate and tissue compartments of donor livers during NMP. This suggests a potential role of NMP in altering the immunogenicity of donor livers prior to transplant. These data also provide insights for future work to recondition the intrinsic immune profile of donor livers during NMP prior to transplantation.
常温机器灌注（NMP）允许在移植前对供体肝脏进行活力评估和潜在的复苏。NMP 对肝移植物的免疫影响尚不确定，这可能对移植物功能、排斥反应结果和总体存活率产生影响。在这项研究中，我们定义了 NMP 过程中人类肝脏免疫谱的变化。
将 6 个供体肝脏置于 NMP 设备上。在灌注前的冷保存期间以及灌注 1、3 和 6 小时时获取组织和灌流液样本。使用流式细胞术、免疫组织化学和基于珠子的免疫分析来测量灌流液和肝组织内的白细胞组成和细胞因子。通过 Student's t 检验比较基线和时间点之间的平均值。
在循环灌流液中，在 NMP 开始 1 小时后可检测到 CD4 T 细胞、B 细胞和嗜酸性粒细胞的频率显著增加，并在灌注 6 小时时继续增加。另一方面，NK 细胞频率在 NMP 开始 1 小时后显著降低，并且在灌注期间持续降低。在肝组织中，在 NMP 开始 6 小时后可检测到 B 细胞频率显著增加，但中性粒细胞减少。在灌注过程中的中间阶段，可检测到中间单核细胞频率的短暂下降。总体而言，在 NMP 期间，组织驻留 T 调节细胞中未检测到明显差异。在 NMP 开始后，可检测到促炎和抗炎细胞因子的水平显著增加，并在整个灌注过程中持续升高。
在 NMP 过程中，供体肝脏的灌流液和组织部分中的单个白细胞细胞类型都出现了时间依赖性的动态变化。这表明 NMP 在移植前可能改变供体肝脏的免疫原性。这些数据还为未来的工作提供了见解，即在移植前的 NMP 期间对供体肝脏的固有免疫谱进行调理。
