Serial characterisation of monocyte and neutrophil function after lung resection

Abstract

Objectives: The primary aim of this prospective study was to perform a comprehensive serial characterisation of monocyte and neutrophil function, circulating monocyte subsets, and bronchoalveolar lavage (BAL) fluid after lung resection. A secondary aim was to perform a pilot, hypothesis-generating evaluation of whether innate immune parameters were associated with postoperative pneumonia.

Methods: Forty patients undergoing lung resection were studied in detail. Blood monocytes and neutrophils were isolated preoperatively and at 6, 24 and 48 h postoperatively. BAL was performed preoperatively and immediately postoperatively. Monocyte subsets, monocyte responsiveness to lipopolysaccharide (LPS) and neutrophil phagocytic capacity were quantified at all time points. Differential cell count, protein and cytokine concentrations were measured in BAL. Pneumonia evaluation at 72 h was assessed using predefined criteria.

Results: After surgery, circulating subsets of classical and intermediate monocytes increased significantly. LPS-induced release of proinflammatory cytokines from monocytes increased significantly and by 48 h a more proinflammatory profile was found. Neutrophil phagocytosis demonstrated a small but significant fall. Factors associated with postoperative pneumonia were: increased release of specific proinflammatory and anti-inflammatory cytokines from monocytes; preoperative neutrophilia; and preoperative BAL cell count.

Conclusions: We conclude that postoperative lung inflammation is associated with specific changes in the cellular innate immune response, a better understanding of which may improve patient selection and prediction of complications in the future.

Keywords: Innate Immunity; Thoracic Surgery.

Figure 1

Postoperative changes in blood monocyte subsets. (A) Serial flow cytometry plots from a single representative patient, illustrating the quantification strategy (as outlined in the Methods section). Mononuclear cells were obtained from whole blood subjected to dextran sedimentation and discontinuous percoll gradients. The monocyte population was identified by gating on characteristic forward scatter and side scatter appearances. Within this gate, HLA-DR expressing cells were identified, and in this population staining for CD14 and CD16 defined ‘classical’ CD14++CD16− monocytes (Q3, bottom right quadrant), ‘intermediate’ CD14++CD16+ monocytes (Q2, top right quadrant) and ‘non-classical’ CD14+CD16++ monocytes (Q1, top left quadrant). As an additional quality control, expression of CCR2 and CX₃CR1 was assessed, to confirm that the expected expression pattern of these receptors within each monocyte subset was observed. (B) Serial monocyte subset counts in blood. Median values are shown with the IQR in brackets. Paired t tests were used to determine postoperative change, n=39 preoperatively, n=38 at 24 h and n=27 at 48 h.