Flow Cytometric Analysis of Myeloid Cells in Human Blood, Bronchoalveolar Lavage, and Lung Tissues.
Clear identification of specific cell populations by flow cytometry is important to understand functional roles. A well-defined flow cytometry panel for myeloid cells in human bronchoalveolar lavage (BAL) and lung tissue is currently lacking. The objective of this study was to develop a flow cytometry-based panel for human BAL and lung tissue. We obtained and performed flow cytometry/sorting on human BAL cells and lung tissue. Confocal images were obtained from lung tissue using antibodies for cluster of differentiation (CD)206, CD169, and E cadherin. We defined a multicolor flow panel for human BAL and lung tissue that identifies major leukocyte populations. These include macrophage (CD206(+)) subsets and other CD206(-) leukocytes. The CD206(-) cells include: (1) three monocyte (CD14(+)) subsets, (2) CD11c(+) dendritic cells (CD14(-), CD11c(+), HLA-DR(+)), (3) plasmacytoid dendritic cells (CD14(-), CD11c(-), HLA-DR(+), CD123(+)), and (4) other granulocytes (neutrophils, mast cells, eosinophils, and basophils). Using this panel on human lung tissue, we defined two populations of pulmonary macrophages: CD169(+) and CD169(-) macrophages. In lung tissue, CD169(-) macrophages were a prominent cell type. Using confocal microscopy, CD169(+) macrophages were located in the alveolar space/airway, defining them as alveolar macrophages. In contrast, CD169(-) macrophages were associated with airway/alveolar epithelium, consistent with interstitial-associated macrophages. We defined a flow cytometry panel in human BAL and lung tissue that allows identification of multiple immune cell types and delineates alveolar from interstitial-associated macrophages. This study has important implications for defining myeloid cells in human lung samples.
1 Department of Medicine, Duke University, Durham, North Carolina.
Yen-Rei A
YR
Yu
1 Department of Medicine, Duke University, Durham, North Carolina.
Danielle F
DF
Hotten
1 Department of Medicine, Duke University, Durham, North Carolina.
Yuryi
Y
Malakhau
1 Department of Medicine, Duke University, Durham, North Carolina.
Ellen
E
Volker
2 National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency, Chapel Hill, North Carolina.
Andrew J
AJ
Ghio
3 Department of Medicine, Cedar Sinai Medical Center, Los Angeles, California; and.
Paul W
PW
Noble
1 Department of Medicine, Duke University, Durham, North Carolina.
Monica
M
Kraft
4 Department of Medicine, Ohio State University Medical Center, Columbus, Ohio.
John W
JW
Hollingsworth
1 Department of Medicine, Duke University, Durham, North Carolina.
Michael D
MD
Gunn
1 Department of Medicine, Duke University, Durham, North Carolina.
Robert M
RM
Tighe
D000293:Adolescent; D000328:Adult; D015415:Biomarkers / Q000097:blood*; D001992:Bronchoalveolar Lavage Fluid / Q000166:cytology / Q000276:immunology*; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D016130:Immunophenotyping / Q000379:methods*; D008168:Lung / Q000166:cytology / Q000276:immunology*; D016676:Macrophages, Alveolar / Q000276:immunology; D008297:Male; D018613:Microscopy, Confocal; D022423:Myeloid Cells / Q000276:immunology*; D063267:Sialic Acid Binding Ig-like Lectin 1 / Q000097:blood; D055815:Young Adult
D015415:Biomarkers; C571166:SIGLEC1 protein, human; D063267:Sialic Acid Binding Ig-like Lectin 1
alveolar macrophages; interstitial lung disease; interstitial macrophages; interstitial-associated macrophages