Exposure to environmental tobacco smoke induces angiogenesis and leukocyte trafficking in lung microvessels

Abstract

Exposure to environmental tobacco smoke (ETS) is known to contribute to and exacerbate inflammatory diseases of the lung such as chronic obstructive pulmonary disease (COPD) and asthma. The effect of ETS on angiogenesis and leukocyte recruitment, both of which promote lung inflammation, was investigated using lung tissue from mice exposed to aged and diluted sidestream cigarette smoke or fresh air for 12 weeks and transplanted into dorsal skin-fold chambers in nude mice. Lung tissue from mice exposed to cigarette smoke for 12 weeks exhibited significantly increased vascular density (angiogenesis) associated with selectin-mediated increased intravascular leukocyte rolling and adhesion compared to fresh air-exposed lung tissue by intravital microscopy. Further, neutrophils from nicotine-exposed mice displayed significantly increased rolling and adhesion compared to control neutrophils in microvessels of nicotine-exposed lungs versus control lung microvessels, suggesting that nicotine in cigarette smoke can augment leukocyte-endothelial interactions. ETS-induced angiogenesis and leukocyte trafficking may play a key role in airway recruitment of inflammatory cells in ETS-associated disorders such as COPD bronchitis or asthma.

Figure 1

Exposure to ADSCS induces pulmonary angiogenesis. (A) Lung allografts obtained from FA-and ADSCS-exposed mice were transplanted into skin-fold chambers in nude mice and allowed to undergo revascularization. Representative photomicrographs of the microvasculature of the revascularized lung allografts from FA- and ADSCS-exposed mice observed by IVM on day 14 at magnifications of 40× (upper panels) and 100× (lower panels) are shown. (B) The density of the vascular network in revascularized lung allografts from ADSCS- and FA-exposed mice was determined from scanned images of the vascular network as described in Materials and Methods. Individual data of 4 different regions from 2 allografts each for FA and ADSCS is shown. The inset shows mean ± range of the data.

Figure 2

Exposure to ADSCS induces leukocyte rolling and adhesion within revascularized lung microvessels. (A) The interaction of acridine orange-labeled circulating leukocytes with the endothelium of the revascularized microvessels of lung allografts from ADSCS- and FA-exposed mice was evaluated by IVM and the number of rolling and adherent cells was determined by off-line analysis of recorded video images. Data represent mean ± SE. *P < .01 (B) Representative photomicrographs of acridine orange-labeled leukocytes within microvessels of lung allografts from ADSCS- (left panel) and FA- (right panel) exposed mice at magnification of 100× are shown.

Figure 3

ADSCS-induced rolling and adhesion of leukocytes in lung microvessels is selectin-mediated. The effect of anti-P- and anti-E-selectin mAb treatment on leukocyte rolling and adhesion in revascularized lung microvessels of ADSCS- and FA-exposed mice was investigated by IVM. Anti-selectin antibodies were used at a concentration of 2 mg/kg body weight. Rolling (upper panel) and adhesion (lower panel) of circulating acridine orange–labeled leukocytes was analyzed by off-line analysis of recorded video images. Data represent mean ± SE. *P < .05 versus rolling and adhesion in ADSCS-exposed microvessels.

Figure 4

Nicotine induces neutrophil rolling and adhesion in lung microvessels. (A) Rolling and adhesion of CFDA-labeled neutrophils from control and nicotine pellet-exposed mice in revascularized lung microvessels of allografts from control and nicotine-exposed mice was evaluated by IVM and off-line analysis of recorded video images. Data represent mean ± SE. *, **P < .01 versus rolling and adhesion in control lungs; §P < .05 versus rolling and adhesion of control neutrophils. (B) Representative photomicrographs of CFDA-labeled neutrophils from control and nicotine-exposed mice within microvessels of lung allografts from control and nicotine-exposed mice at a magnification of 100× are shown. Upper left panel: control neutrophils in control lung microvessels; lower left panel: neutrophils from nicotine-exposed mice in control lung microvessels; upper right panel: control neutrophils in nicotine-exposed lung microvessels; lower right panel: neutrophils from nicotine-exposed mice in nicotine-exposed lung microvessels.