NETosis in pulmonary pleomorphic carcinoma

Abstract

Pulmonary pleomorphic carcinoma (PC) is a rare non-small-cell lung carcinoma (NSCLC) with a poor prognosis, characterized by tumor necrosis (TN). NETosis is a form of neutrophil-specific cell death, which is morphologically characterized by prominent neutrophil infiltration and cell detritus in the necrotic foci. Seventy-six patients with pulmonary PC who underwent complete resection were enrolled. Tumor necrosis was evaluated using digitally scanned resected specimens. The regions of NETosis were quantified using citrullinated histone H3 (citH3)- and myeloperoxidase-positive regions. We examined the association between the NETosis area and the prognostic outcomes and assessed the correlation between the NETosis area and systemic inflammation. Tumor necrosis was observed in 70 patients (92%). In all the cases, the TN region was accompanied by a citH3-positive region. The patients with high NETosis area (n = 54) had significantly shorter overall survival than those with low NETosis area (n = 16) (p = 0.013). Furthermore, a high NETosis area was an independent poor prognostic factor in the multivariate analyses. Systemic inflammatory markers, including C-reactive protein (CRP), CRP-to-albumin ratio, and neutrophil-to-lymphocyte ratio, were significantly higher in patients with high NETosis area than in those with low NETosis area. Furthermore, the levels of these inflammatory markers were significantly decreased postsurgery. This study shows that in surgically resected pulmonary PC, patients with high NETosis areas have higher systemic inflammation and worse prognosis.

Keywords: NET; NETosis; dirty necrosis; pulmonary pleomorphic carcinoma; systemic inflammation.

FIGURE 1

Dirty necrosis (DN) (+) and DN (−) tumor necrosis (TN), and area of TN in pulmonary pleomorphic carcinoma (PC). (A) DN (+) with abundant neutrophils and cell detritus in pulmonary PC. Scale bars: 500 μm (left), 250 μm (middle), 50 μm (right). (B) DN (−) with only tumor cells showing coagulation necrosis in pulmonary PC. Scale bars: 500 μm (left), 125 μm (middle), 50 μm (right). (C) Area of the TN in pulmonary PC. The x‐ and y‐axis represent the patient number and area of TN, respectively.

FIGURE 2

Immunohistochemical analysis of citrullinated histone H3 (citH3) and myeloperoxidase (MPO), and area of citH3 and MPO. (A) Immunohistochemical expression of citH3 and MPO in NETosis (+). Scale bars, 250 μm. (B) Immunohistochemical expression of citH3 and MPO in NETosis (−). Scale bars, 250 μm. (C) Area of citH3 in pulmonary PC. The x‐ and y‐axis represent patient number and area of citH3, respectively. (D) Area of MPO in pulmonary PC. The x‐ and y‐axis represent patient number and area of MPO, respectively.

FIGURE 3

Overall survival (OS) and relapse‐free survival (RFS) curves for patients with tumor necrosis (TN) and NETosis. (A) OS curves for patients with TN. Black line represents the group with a low TN; red line represents the group with a high TN. (B) RFS curves for patients with TN. Black line represents the group with a low TN; red line represents the group with a high TN. (C) OS curves of patients with NETosis. Black line shows the group with low NETosis areas; red line shows the group with high NETosis areas. (D) RFS curves of patients with NETosis. Black line shows the group with low NETosis areas; red line shows the group with high NETosis areas.