A new approach for analyzing cellular infiltration during allergic airway inflammation.
A mouse model for allergic airway inflammation involving ovalbumin (OVA) sensitization and challenge has been developed that reproduces hallmark features of human asthma and has provided valuable insight into the mechanisms by which this disease occurs. Cellular infiltrate in lungs of mice used in this model have conventionally been evaluated using histological examination of tissue sections and light microscopic analysis of lung lavage samples. As an alternative or complementary approach for characterizing cellular infiltrate, we developed a multicolor fluorescence-activated cell sorter (FACS) method involving the simultaneous detection of seven different markers on lung cell suspensions: CD4, CD8, B220, CD11b, Gr-1, CD49b, and FcepsilonRI. Only some of these cell types increased in OVA-challenged mice compared to PBS controls, including the CD4(+), B220(+), CD11b(+), and FcepsilonRI(+) groups. We also examined subpopulations of cells for coexpression of these markers and dissected heterogeneous populations as further evaluation procedures to characterize the cellular infiltrate resulting from OVA challenge. Finally, we combined FACS with real-time PCR to analyze certain cell types in terms of mRNA levels for factors involved in asthma, including GATA-3 and IL-1beta. Overall, these FACS-based techniques provide a powerful approach for analyzing cellular profiles in lung tissue from mice used in the mouse model of asthma and may also prove valuable in evaluating cellular infiltrates for other models of inflammation and immune responses.
Hoffmann
Peter R
PR
Department of Cell and Molecular Biology, University of Hawai'i, Honolulu, HI 96813, USA. peterh@pbrc.hawaii.edu
Gurary
Alexandra
A
Hoffmann
Fukun W
FW
Jourdan-Le Saux
Claude
C
Teeters
Kelsa
K
Hashimoto
Ann C
AC
Tam
Elizabeth K
EK
Berry
Marla J
MJ
D000485:Allergens / Q000276:immunology; D000818:Animals; D001992:Bronchoalveolar Lavage Fluid / Q000166:cytology; D005434:Flow Cytometry / Q000379:methods*; D016131:Lymphocyte Subsets / Q000166:cytology*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010047:Ovalbumin / Q000276:immunology; D012333:RNA, Messenger / Q000032:analysis; D012130:Respiratory Hypersensitivity / Q000276:immunology*; D020133:Reverse Transcriptase Polymerase Chain Reaction