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Targeted induction of lung endothelial cell apoptosis causes emphysema-like changes in the mouse.

Giordano RJ, Lahdenranta J, Zhen L, Chukwueke U, Petrache I, Langley RR, Fidler IJ, Pasqualini R, Tuder RM, Arap W - J. Biol. Chem. (2008)

Bottom Line: As early as 4 days following peptide administration, mice developed air space enlargement associated with enhanced oxidative stress, influx of macrophages, and up-regulation of ceramide.Thus, our data enable the generation of a convenient mouse model of human emphysema.Finally, combinatorial screenings on immortalized cells followed by in vivo targeting establishes an experimental framework for discovery and validation of additional ligand-directed pharmacodelivery systems.

View Article: PubMed Central - PubMed

Affiliation: University of Texas M.D. Anderson Cancer Center, Houston, Texas 77030, USA.

ABSTRACT
Pulmonary gas exchange relies on a rich capillary network, which, together with alveolar epithelial type I and II cells, form alveolar septa, the functional units in the lung. Alveolar capillary endothelial cells are critical in maintaining alveolar structure, because disruption of endothelial cell integrity underlies several lung diseases. Here we show that targeted ablation of lung capillary endothelial cells recapitulates the cellular events involved in cigarette smoke-induced emphysema, one of the most prevalent nonneoplastic lung diseases. Based on phage library screening on an immortalized lung endothelial cell line, we identified a lung endothelial cell-binding peptide, which preferentially homes to lung blood vessels. This peptide fused to a proapoptotic motif specifically induced programmed cell death of lung endothelial cells in vitro as well as targeted apoptosis of the lung microcirculation in vivo. As early as 4 days following peptide administration, mice developed air space enlargement associated with enhanced oxidative stress, influx of macrophages, and up-regulation of ceramide. Given that these are all critical elements of the corresponding human emphysema caused by cigarette smoke, these data provide evidence for a central role for the alveolar endothelial cells in the maintenance of lung structure and of endothelial cell apoptosis in the pathogenesis of emphysema-like changes. Thus, our data enable the generation of a convenient mouse model of human emphysema. Finally, combinatorial screenings on immortalized cells followed by in vivo targeting establishes an experimental framework for discovery and validation of additional ligand-directed pharmacodelivery systems.

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Induction of morphological changes in mouse lungs after administration of CGSPGWVRC-GG-D(KLAKLAK)2. a, lung sections of mice 4 days after treatment with CGSPGWVRC-GG-D(KLAKLAK)2 peptide or with control peptides (CGSPGWVRC or D(KLAKLAK)2 peptides) were stained with hematoxylin and eosin. Lung sections from the CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated mice show increased air space enlargement when compared with the lung sections from the control-treated mice. Scale bar, 400 μm. b, quantification of the mean linear intercept (μm) in CGSPGWVRC-GG-D(KLAKLAK)2 and control peptide-treated lungs after 4 days of peptide treatment. CGSPGWVRC-GG-D(KLAKLAK)2 treatment shows a significant increase in the mean linear intercept values in versus control-treated mice. c, lung sections from mice treated for 21 days with CGSPGWVRC-GG-D(KLAKLAK)2, control peptides (CGSPGWVRC or D(KLAKLAK)2), or vehicle alone were stained with hematoxylin and eosin. Lung sections from the CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated mice show increased air space enlargement when compared with the lung sections from the control-treated mice. Scale bar, 200 μm. d, the mean linear intercept (μm) was measured from vehicle, CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated, and control peptide (CGSPGWVRC or D(KLAKLAK)2)-treated animals after 21 days of treatment. CGSPGWVRC-GG-D(KLAKLAK)2 treatment shows a significant increase in mean linear intercept values compared with control peptide-treated mice.
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fig4: Induction of morphological changes in mouse lungs after administration of CGSPGWVRC-GG-D(KLAKLAK)2. a, lung sections of mice 4 days after treatment with CGSPGWVRC-GG-D(KLAKLAK)2 peptide or with control peptides (CGSPGWVRC or D(KLAKLAK)2 peptides) were stained with hematoxylin and eosin. Lung sections from the CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated mice show increased air space enlargement when compared with the lung sections from the control-treated mice. Scale bar, 400 μm. b, quantification of the mean linear intercept (μm) in CGSPGWVRC-GG-D(KLAKLAK)2 and control peptide-treated lungs after 4 days of peptide treatment. CGSPGWVRC-GG-D(KLAKLAK)2 treatment shows a significant increase in the mean linear intercept values in versus control-treated mice. c, lung sections from mice treated for 21 days with CGSPGWVRC-GG-D(KLAKLAK)2, control peptides (CGSPGWVRC or D(KLAKLAK)2), or vehicle alone were stained with hematoxylin and eosin. Lung sections from the CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated mice show increased air space enlargement when compared with the lung sections from the control-treated mice. Scale bar, 200 μm. d, the mean linear intercept (μm) was measured from vehicle, CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated, and control peptide (CGSPGWVRC or D(KLAKLAK)2)-treated animals after 21 days of treatment. CGSPGWVRC-GG-D(KLAKLAK)2 treatment shows a significant increase in mean linear intercept values compared with control peptide-treated mice.

Mentions: As assessed by lung morphometry, C57Bl6/6J mice that received intraperitoneal injections of 240 μg of CGSPGWVRCGG-D(KLAKLAK)2 peptide or control peptides every other day for a total of either 4 days or 21 days developed emphysema (Fig. 4). Alveolar destruction was already evident after only 4 days (two doses) of treatment (Fig. 4, a and b), and it became more prominent at 21 days (10 doses) of treatment with the peptide CGSPGWVRC-GG-D(KLAKLAK)2 (Fig. 4c). By the end of the 21-day experiment (n = 5 mice in each group), CGSPGWVRCGG-D(KLAKLAK)2-treated lungs exhibited an approximately 20% increase from a mean linear intercept of 59 ± 0.84 μmin CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated mice compared with 49.7 ± 0.9 μm in control-treated mice with CGSPGWVRC peptide, 49.5 ± 1.0 μm for untargeted D(KLAKLAK)2-treated mice and 49 ± 0.6 μm for vehicle-treated lungs (Fig. 4d). Hematoxylin and eosin-stained tissue sections of several control organs (kidney, heart, liver, and bone marrow) did not reveal any detectable histological changes in CGSPGWVRCGG-D(KLAKLAK)2-treated mice (Fig. S1).


Targeted induction of lung endothelial cell apoptosis causes emphysema-like changes in the mouse.

Giordano RJ, Lahdenranta J, Zhen L, Chukwueke U, Petrache I, Langley RR, Fidler IJ, Pasqualini R, Tuder RM, Arap W - J. Biol. Chem. (2008)

Induction of morphological changes in mouse lungs after administration of CGSPGWVRC-GG-D(KLAKLAK)2. a, lung sections of mice 4 days after treatment with CGSPGWVRC-GG-D(KLAKLAK)2 peptide or with control peptides (CGSPGWVRC or D(KLAKLAK)2 peptides) were stained with hematoxylin and eosin. Lung sections from the CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated mice show increased air space enlargement when compared with the lung sections from the control-treated mice. Scale bar, 400 μm. b, quantification of the mean linear intercept (μm) in CGSPGWVRC-GG-D(KLAKLAK)2 and control peptide-treated lungs after 4 days of peptide treatment. CGSPGWVRC-GG-D(KLAKLAK)2 treatment shows a significant increase in the mean linear intercept values in versus control-treated mice. c, lung sections from mice treated for 21 days with CGSPGWVRC-GG-D(KLAKLAK)2, control peptides (CGSPGWVRC or D(KLAKLAK)2), or vehicle alone were stained with hematoxylin and eosin. Lung sections from the CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated mice show increased air space enlargement when compared with the lung sections from the control-treated mice. Scale bar, 200 μm. d, the mean linear intercept (μm) was measured from vehicle, CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated, and control peptide (CGSPGWVRC or D(KLAKLAK)2)-treated animals after 21 days of treatment. CGSPGWVRC-GG-D(KLAKLAK)2 treatment shows a significant increase in mean linear intercept values compared with control peptide-treated mice.
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fig4: Induction of morphological changes in mouse lungs after administration of CGSPGWVRC-GG-D(KLAKLAK)2. a, lung sections of mice 4 days after treatment with CGSPGWVRC-GG-D(KLAKLAK)2 peptide or with control peptides (CGSPGWVRC or D(KLAKLAK)2 peptides) were stained with hematoxylin and eosin. Lung sections from the CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated mice show increased air space enlargement when compared with the lung sections from the control-treated mice. Scale bar, 400 μm. b, quantification of the mean linear intercept (μm) in CGSPGWVRC-GG-D(KLAKLAK)2 and control peptide-treated lungs after 4 days of peptide treatment. CGSPGWVRC-GG-D(KLAKLAK)2 treatment shows a significant increase in the mean linear intercept values in versus control-treated mice. c, lung sections from mice treated for 21 days with CGSPGWVRC-GG-D(KLAKLAK)2, control peptides (CGSPGWVRC or D(KLAKLAK)2), or vehicle alone were stained with hematoxylin and eosin. Lung sections from the CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated mice show increased air space enlargement when compared with the lung sections from the control-treated mice. Scale bar, 200 μm. d, the mean linear intercept (μm) was measured from vehicle, CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated, and control peptide (CGSPGWVRC or D(KLAKLAK)2)-treated animals after 21 days of treatment. CGSPGWVRC-GG-D(KLAKLAK)2 treatment shows a significant increase in mean linear intercept values compared with control peptide-treated mice.
Mentions: As assessed by lung morphometry, C57Bl6/6J mice that received intraperitoneal injections of 240 μg of CGSPGWVRCGG-D(KLAKLAK)2 peptide or control peptides every other day for a total of either 4 days or 21 days developed emphysema (Fig. 4). Alveolar destruction was already evident after only 4 days (two doses) of treatment (Fig. 4, a and b), and it became more prominent at 21 days (10 doses) of treatment with the peptide CGSPGWVRC-GG-D(KLAKLAK)2 (Fig. 4c). By the end of the 21-day experiment (n = 5 mice in each group), CGSPGWVRCGG-D(KLAKLAK)2-treated lungs exhibited an approximately 20% increase from a mean linear intercept of 59 ± 0.84 μmin CGSPGWVRC-GG-D(KLAKLAK)2 peptide-treated mice compared with 49.7 ± 0.9 μm in control-treated mice with CGSPGWVRC peptide, 49.5 ± 1.0 μm for untargeted D(KLAKLAK)2-treated mice and 49 ± 0.6 μm for vehicle-treated lungs (Fig. 4d). Hematoxylin and eosin-stained tissue sections of several control organs (kidney, heart, liver, and bone marrow) did not reveal any detectable histological changes in CGSPGWVRCGG-D(KLAKLAK)2-treated mice (Fig. S1).

Bottom Line: As early as 4 days following peptide administration, mice developed air space enlargement associated with enhanced oxidative stress, influx of macrophages, and up-regulation of ceramide.Thus, our data enable the generation of a convenient mouse model of human emphysema.Finally, combinatorial screenings on immortalized cells followed by in vivo targeting establishes an experimental framework for discovery and validation of additional ligand-directed pharmacodelivery systems.

View Article: PubMed Central - PubMed

Affiliation: University of Texas M.D. Anderson Cancer Center, Houston, Texas 77030, USA.

ABSTRACT
Pulmonary gas exchange relies on a rich capillary network, which, together with alveolar epithelial type I and II cells, form alveolar septa, the functional units in the lung. Alveolar capillary endothelial cells are critical in maintaining alveolar structure, because disruption of endothelial cell integrity underlies several lung diseases. Here we show that targeted ablation of lung capillary endothelial cells recapitulates the cellular events involved in cigarette smoke-induced emphysema, one of the most prevalent nonneoplastic lung diseases. Based on phage library screening on an immortalized lung endothelial cell line, we identified a lung endothelial cell-binding peptide, which preferentially homes to lung blood vessels. This peptide fused to a proapoptotic motif specifically induced programmed cell death of lung endothelial cells in vitro as well as targeted apoptosis of the lung microcirculation in vivo. As early as 4 days following peptide administration, mice developed air space enlargement associated with enhanced oxidative stress, influx of macrophages, and up-regulation of ceramide. Given that these are all critical elements of the corresponding human emphysema caused by cigarette smoke, these data provide evidence for a central role for the alveolar endothelial cells in the maintenance of lung structure and of endothelial cell apoptosis in the pathogenesis of emphysema-like changes. Thus, our data enable the generation of a convenient mouse model of human emphysema. Finally, combinatorial screenings on immortalized cells followed by in vivo targeting establishes an experimental framework for discovery and validation of additional ligand-directed pharmacodelivery systems.

Show MeSH
Related in: MedlinePlus