Janicova Andrea, Becker Nils, Xu Baolin, Simic Marija, Noack Laurens, Wagner Nils, Müller Andreas J, Bertrand Jessica, Marzi Ingo, Relja Borna
Experimental Radiology, Department of Radiology and Nuclear Medicine, Otto-von-Guericke-University Magdeburg, Leipziger Straße 44, 39120 Magdeburg, Germany.
Department of Trauma, Hand and Reconstructive Surgery, Goethe University, Theodor-Stern-Kai 7, 60590 Frankfurt am Main, Germany.
J Clin Med. 2021 Sep 14;10(18):4139. doi: 10.3390/jcm10184139.
Severe traumatic injury has been associated with high susceptibility for the development of secondary complications caused by dysbalanced immune response. As the first line of the cellular immune response, neutrophils and monocytes recruited to the site of tissue damage and/or infection, are divided into three different subsets according to their CD16/CD62L and CD16/CD14 expression, respectively. Their differential functions have not yet been clearly understood. Thus, we evaluated the phenotypic changes of neutrophil and monocyte subsets among their functionality regarding oxidative burst and the phagocytic capacity in severely traumatized patients.
Peripheral blood was withdrawn from severely injured trauma patients (TP; = 15, ISS ≥ 16) within the first 12 h post-trauma and from healthy volunteers (HV; = 15) and stimulated with fMLP and PMA. CD16CD62L (immature), CD16CD62L (mature) and CD16CD62L (CD62L) neutrophil subsets and CD14CD16 (classical), CD14CD16 (intermediate) and CD14CD16 (non-classical) monocyte subsets of HV and TP were either directly analyzed by flow cytometry or the examined subsets of HV were sorted first by fluorescence-activated cell sorting and subsequently analyzed. Subset-specific generation of reactive oxygen species (ROS) and of bioparticle phagocytosis were evaluated.
In TP, the counts of immature neutrophils were significantly increased vs. HV. The numbers of mature and CD62L neutrophils remained unchanged but the production of ROS was significantly enhanced in TP vs. HV and the stimulation with fMLP significantly increased the generation of ROS in the mature and CD62L neutrophils of HV. The counts of phagocyting neutrophils did not change but the mean phagocytic capacity showed an increasing trend in TP. In TP, the monocytes shifted toward the intermediate phenotype, whereas the classical and non-classical monocytes became less abundant. ROS generation was significantly increased in all monocyte subsets in TP vs. HV and PMA stimulation significantly increased those level in both, HV and TP. However, the PMA-induced mean ROS generation was significantly lower in intermediate monocytes of TP vs. HV. Sorting of monocyte and neutrophil subsets revealed a significant increase of ROS and decrease of phagocytic capacity vs. whole blood analysis.
Neutrophils and monocytes display a phenotypic shift following severe injury. The increased functional abnormalities of certain subsets may contribute to the dysbalanced immune response and attenuate the antimicrobial function and thus, may represent a potential therapeutic target. Further studies on isolated subsets are necessary for evaluation of their physiological role after severe traumatic injury.
严重创伤性损伤与免疫反应失衡导致的继发性并发症高易感性相关。作为细胞免疫反应的第一线，招募到组织损伤和/或感染部位的中性粒细胞和单核细胞，分别根据其CD16/CD62L和CD16/CD14表达分为三个不同的亚群。它们的不同功能尚未完全清楚。因此，我们评估了严重创伤患者中性粒细胞和单核细胞亚群在氧化爆发和吞噬能力方面的表型变化及其功能。
在创伤后12小时内，从严重受伤的创伤患者（TP；n = 15，损伤严重程度评分[ISS]≥16）和健康志愿者（HV；n = 15）中采集外周血，并用N-甲酰甲硫氨酰-亮氨酰-苯丙氨酸（fMLP）和佛波酯（PMA）刺激。通过流式细胞术直接分析HV和TP的CD16⁺CD62L⁻（未成熟）、CD16⁺CD62L⁺（成熟）和CD16⁻CD62L⁺（CD62L⁻）中性粒细胞亚群以及CD14⁺CD16⁻（经典）、CD14⁺CD16⁺（中间）和CD14⁻CD16⁺（非经典）单核细胞亚群，或者先通过荧光激活细胞分选对HV的检测亚群进行分选，随后进行分析。评估亚群特异性活性氧（ROS）生成和生物颗粒吞噬情况。
在TP中，未成熟中性粒细胞计数相对于HV显著增加。成熟和CD62L⁻中性粒细胞数量保持不变，但与HV相比，TP中ROS生成显著增强，并且用fMLP刺激显著增加了HV中成熟和CD62L⁻中性粒细胞的ROS生成。吞噬性中性粒细胞计数没有变化，但TP中的平均吞噬能力呈增加趋势。在TP中，单核细胞向中间表型转变，而经典和非经典单核细胞数量减少。与HV相比，TP中所有单核细胞亚群的ROS生成均显著增加，PMA刺激显著增加了HV和TP中的ROS水平。然而，与HV相比，TP中间单核细胞中PMA诱导的平均ROS生成显著降低。单核细胞和中性粒细胞亚群的分选显示，与全血分析相比，ROS显著增加，吞噬能力降低。
严重损伤后中性粒细胞和单核细胞表现出表型转变。某些亚群功能异常增加可能导致免疫反应失衡并削弱抗菌功能，因此可能是一个潜在的治疗靶点。对分离亚群进行进一步研究对于评估其在严重创伤性损伤后的生理作用是必要的。
