Myeloid Phenotypes in Tracheostomy‐Associated Granulation Tissue

Abstract

Objective(s)Tracheostomy‐associated granulation tissue is a common, recurrent problem occurring secondary to chronic mucosal irritation. Although granulation tissue is composed of predominantly innate immune cells, the phenotype of monocytes and macrophages in tracheostomy‐associated granulation tissue is unknown. This study aims to define the myeloid cell population in granulation tissue secondary to tracheostomy.MethodsGranulation tissue biopsies were obtained from 8 patients with tracheostomy secondary to laryngotracheal stenosis. Cell type analysis was performed by flow cytometry and gene expression was measured by quantitative real‐time polymerase chain reaction. These methods and immunohistochemistry were used to define the monocyte/macrophage population in granulation tissue and were compared to tracheal autopsy control specimens.ResultsFlow cytometry demonstrated macrophages (CD45+CD11b+) and monocytes (CD45+FSClowSSClow) represent 23.2 ± 6% of the granulation tissue cell population. The M2 phenotype (CD206) is present in 77 ± 11% of the macrophage population and increased compared to the M1 phenotype (p = 0.012). Classical monocytes (CD45+CD14highCD16low) were increased in granulation tissue compared to controls (61.2 ± 7% and 30 ± 8.5%, p = 0.038). Eighty‐five percent of macrophages expressed pro‐inflammatory S100A8/A9 and 36 ± 4% of macrophages co‐localized CD169, associated with tissue‐resident macrophages. M2 gene expression (Arg1/CD206) was increased in granulation tissue (3.7 ± 0.4, p = 0.035 and 3.5 ± 0.5, p = 0.047) whereas M1 gene expression (CD80/CD86) was similar to controls (p = 0.64, p = 0.3). Immunohistochemistry of granulation tissue demonstrated increased cells co‐localizing CD11b and CD206.ConclusionsM2 macrophages are the dominant macrophage phenotype in tracheostomy‐associated granulation tissue. The role of this cell type in promoting ongoing inflammation warrants future investigation to identify potential treatments for granulation tissue secondary to tracheostomy.Level of Evidence3 Laryngoscope, 133:2346–2356, 2023

The Activation of M1 Macrophages is Associated with the JNK-m6A-p38 Axis in Chronic Obstructive Pulmonary Disease

“… 18 The gating strategy for peripheral blood of C57 mice was as follows: SSC and CD45 were used to set a gate to circle the macrophage population, then SSC and F4/80 were utilized to circle the macrophage population again and F4/80 and CD11C were used to locate the M1 macrophages. 19 , 20 A total of 100 µL of peripheral blood was inserted into the flow tube and 1 uL of antibody was added to the negative control sample without any antibody. The following antibodies were used: PE/Dazzle 594 anti-human CD14 (Clone 63D3, BioLegend), Alexa Fluor 488 anti-human CD86 (Clone 2331 (FUN-1), BD), PE/Cyanine7 anti-human CD68 (Clone Y1/82A (RUO), BD), PE/Dazzle 594 anti-mouse F4/80 (Clone BMB, BioLegend), PE/Cyanine5 anti-mouse CD11c (Clone N418, BioLegend) and FITC anti-mouse CD45 (Clone 30-F11, BioLegend).…”