Exploratory Study of CD10low Polymorphonuclear Leukocytes Preceding and Correlating With Postsurgical Inflammation

Abstract

The lack of diagnostic and monitoring tools for postsurgical immunological complications such as systemic inflammation can contribute to a poor outcome despite modern intensive care. The need for reliable immune monitoring has been emphasised. Polymorphonuclear leukocytes (PMNs) play an important role in postsurgical inflammation. A subgroup of PMNs is particularly interesting, because they are released shortly after (iatrogenic) trauma: immature PMNs, characterised by, for example, their low CD10 expression. Therefore, we investigated the role of CD10^low PMNs in a non-interventional exploratory study by including patients undergoing scheduled, highly standardised cardiac surgery with extracorporeal circulation. We were able to demonstrate that the number of CD10^low PMNs released shortly after the beginning of surgery correlated with different fluid phase markers of inflammation and organ damage postsurgically. Among these parameters were CRP, IL-6, NGAL, CK-MB, and troponin-T. Noteworthy, the amount of CD10^low PMNs increased as early as 24 h before these well-established markers, suggesting superiority of CD10^low PMNs as an early diagnostic marker. Comparing CD10^low immature PMNs with CD10^high mature PMNs revealed potential involved mechanisms, including lower CD11b expression and a significant decrease in the formation of platelet-neutrophil complexes (PNCs) by CD10^low PMNs. In conclusion, we propose CD10^low PMNs as a potential early cellular biomarker to assess the postsurgical inflammatory response. In comparison to clinically established markers like CRP or IL-6 and scoring systems such as the SOFA-Score, CD10^low PMNs reflect a potential candidate for future immune monitoring to determine the risk of excessive inflammation and organ impairment more rapidly.

FIGURE 1

Perioperative numbers of CD10^low polymorphonuclear leukocytes (PMNs) in comparison to the total PMNs count. (A) CD10^low PMNs of total PMNs under unstimulated conditions before surgery (pre‐surgery), during surgery and 24, 48 and 120 h after surgery. (B) CD10^low PMNs numbers of total PMNs stimulated with cocktail (consisting of 10 μM fMLF, 1 μM PAF and 2.3 μM TNF) before surgery, during and 24, 48 and 120 h after surgery. Median ± interquartile range, n = 10. Kruskal–Wallis test with Dunn's post hoc correction when comparing to pre‐surgery with *p < 0.05, **p < 0.01, ****p < 0.0001.

FIGURE 2

Comparison of CD10^low polymorphonuclear leukocytes (PMNs) numbers perioperatively without stimulation (unstim) as well as with stimulation by an inflammatory cocktail (stim). (the cocktail consisted of10 μM fMLF, 1 μM PAF and 2.3 μM TNF). CD10^low PMNs are compared with established clinical diagnostic markers of inflammation like procalcitonin (PCT), interleukin‐6 (IL‐6) and C‐reactive protein (CRP) before, during, and 24, 48, and 120 h after surgery. Data normalised to 0% = pre‐surgery and 100% = maximal change. Median ± interquartile range, n = 10.

FIGURE 3

Simple linear regressions of CD10^low polymorphonuclear leukocytes (PMNs) of total PMNs during surgery and makers of inflammation and organ damage post‐surgery. (A, B) CD10^low PMNs under unstimulated conditions correlated with C‐reactive protein (CRP) at 48 h (A) and interleukin‐6 at 24 h (B) after surgery. (C, D) CD10^low PMNs under stimulation with an inflammatory cocktail (consisting of 10 μM fMLF, 1 μM PAF and 2.3 μM TNF) with troponin‐T at 48 h (C) and creatine kinase‐MB (CK‐MB) at 24 h (D). Spearman‐r correlation, n = 9–10.

FIGURE 4

Simple linear regressions of stimulated CD10^low polymorphonuclear leukocytes (PMNs) of the total PMNs numbers during surgery and makers of kidney damage post‐surgery. (A) CD10^low PMNs under unstimulated conditions (with buffer as ctrl) and plasma neutrophil gelatinase‐associated lipocalin (NGAL) at 24 h, plasma creatinine at 48 h (B), and urine creatinine at 24 h (C) post‐surgery. Spearman‐r correlation, n = 10.

FIGURE 5

Comparison of surface antigen expression of CD10^low polymorphonuclear leukocytes (PMNs) and CD10^high PMNs during surgery under unstimulated conditions (with buffer as ctrl) and after stimulation with an inflammatory cocktail (consisting of 10 μM fMLF, 1 μM PAF and 2.3 μM TNF). (A) CD11b and (B) CD62L from patients during surgery. Median ± interquartile range, n = 10. Mann–Whitney U Test with *p < 0.05, ****p < 0.0001.

FIGURE 6

Formation of platelet–neutrophil complexes (PNCs) of CD10^low polymorphonuclear leukocytes (PMNs) and CD10^high PMNs during surgery. (A) Representative PNC as detected by light microscopy (right). Platelets are marked as (P). (B) Flow cytometric analysis of PNC formation under unstimulated conditions (with buffer as ctrl), and (C) stimulated conditions with an inflammatory cocktail (consisting of 10 μM fMLF, 1 μM PAF and 2.3 μM TNF). Median ± interquartile range, n = 10. Mann–Whitney U test with **p < 0.01, ***p < 0.001.