TY - JOUR
T1 - Flow cytometric analysis of myeloid cells in human blood, bronchoalveolar lavage, and lung tissues
AU - Yu, Yen Rei A.
AU - Hotten, Danielle F.
AU - Malakhau, Yuryi
AU - Volker, Ellen
AU - Ghio, Andrew J.
AU - Noble, Paul W.
AU - Kraft, Monica
AU - Hollingsworth, John W.
AU - Gunn, Michael D.
AU - Tighe, Robert M.
N1 - Funding Information:
This work was supported by the Pulmonary Hypertension Association Proof of Concept Grant (Y.-R.A.Y.), the Mandel Foundation Fellowship Grant (Y.-R.A.Y.), P01 HL108793 (P.W.N.), R01 ES020350 (J.W.H.), and K08 HL105537 (R.M.T.).
Publisher Copyright:
Copyright © 2016 by the American Thoracic Society.
PY - 2016/1
Y1 - 2016/1
N2 - 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.
AB - 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.
KW - Alveolar macrophages
KW - Interstitial lung disease
KW - Interstitial macrophages
KW - Interstitial-associated macrophages
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U2 - 10.1165/rcmb.2015-0146OC
DO - 10.1165/rcmb.2015-0146OC
M3 - Article
C2 - 26267148
AN - SCOPUS:84963812591
VL - 54
SP - 13
EP - 24
JO - American Journal of Respiratory Cell and Molecular Biology
JF - American Journal of Respiratory Cell and Molecular Biology
SN - 1044-1549
IS - 1
ER -