COPD lung contains a sub-population of steroid-insensitive macrophages

In COPD, alveolar macrophages (AM) increase, release more inflammatory mediators but respond poorly to glucocorticosteroids. Different macrophage phenotypes are identified in animals based on density but no definitive studies on human lung macrophages exist. Cells were isolated from resected human lung tissue from non-smokers (NS, n=5) smokers (S, n=11) and COPD (n=7) patients. Cells were separated into three viable fractions using Percoll density gradients (A: 30-40%, B: 40-50% C: 50-60%). Responses to budesonide after stimulation with lipopolysaccharide (LPS) were investigated by measuring TNFα, CXCL8 and IL-10 release by ELISA. Baseline and LPS-stimulated release of TNFα, CXCL8, and IL-10 did not differ between cell fractions or subjects. LPS-stimulated TNFα release by fraction A from NS and S were responsive to budesonide (EC 50 NS: 0.5±0.04nM vs S: 1.8±1.1nM), with inhibition at 10 -6 M being ∼80% (NS) and ∼60% (S). However, COPD cells were unresponsive. Budesonide (10 -6 M) inhibited LPS-stimulated CXCL8 release from fraction A similarly in NS and S cells but less effectively in COPD cells (∼30%, p 50 values 0.6±0.1nM (NS), 1.0±0.3nM (S) and 2.4±0.9nM (COPD) cells. This apparent steroid insensitivity of COPD macrophages from fraction A was selective, as budesonide inhibited LPS-stimulated IL-10 release by ∼55% in fractions A-C from S and COPD patients (EC 50 S=2.0±0.71nM vs COPD 1.7±1.16nM) but by 80% (EC 50 =0.9±0.2nM) in NS cells. TNFα and CXCL8 responses of cells from fractions B and C did not differ between subjects. Fraction A COPD macrophages were less responsive to budesonide and may represent AM. Identifying selective fraction A markers will allow development of directed therapies.