Ocalization of anti-CXCR2 (red) with RECA-1+ subepithelial blood vessels (green: 1006 original

Ocalization of anti-CXCR2 (red) with RECA-1+ subepithelial blood vessels (green: 1006 original magnification, and inset 6006 original magnification). doi:10.1371/journal.pone.0066432.gAcute Ischemia and CXC ChemokinesFigure 6. Effects of dexamethasone on parenchymal and airway cells and cytokines/receptors. (A) Summary of the average effect of dexamethasone on measured parameters reflecting changes within the alveolar space. Each bar represents the average decrease 6 h after LPAL in dexamethasone treated rats compared with vehicle treated rats (3?1 rats/group, *P,0.05). B . Average effects of dexamethasone treatment on CXCL1, CXCL2, (mRNA, protein), CXCR1 and CXCR2 (mRNA) expression in the left bronchus 6 h after LPAL. Dexamethasone has no significant effects on any of the variables measured in the bronchus (3? rats/group). doi:10.1371/journal.pone.0066432.gchemokines within both lung compartments. MedChemExpress SC66 However, the CXCL1 and CXCL2 modulation pattern was different within the two locations analyzed. CXCL1 release appears to be more generally associated with injury and anesthesia/surgery. Furthermore, anti-inflammatory treatment affected only the extent of lung injury, BAL CXCL1 level and BAL inflammatory cells. No changes were observed in the level of chemokine expression within 223488-57-1 site bronchial tissue or, importantly, the magnitude of angiogenesis. In conclusion, results suggest that early changes within the bronchial niche where arteriogenesis originates, contribute to subsequent neovascularization during pulmonary ischemia. Our initial experiments evaluated BAL content as an index of predominantly parenchymal responses. In this model, we hypothesized that alveolar fluid could redistribute and act as a conduit for growth factors, through mucociliary movement. We demonstrated that acute ischemia caused an early increase in BAL total protein suggesting acute lung injury. These permeability changes were substantial (3-fold increase), early, and seemed to be attenuated by24 h after the onset of ischemia. We had shown previously that permeability changes were again seen 14 d after LPAL [12,29]. In addition, the inflammatory cell response paralleled the protein leak and likely reflected resident cells because of the lack of pulmonary blood flow, and only the small bronchial vasculature at this time point. Specifically, the inflammatory cell profile showed a predominance of neutrophils and macrophages, which, in addition to epithelial cells, have the capacity to secrete both CXCL1 and CXCL2 chemokines [30]. Although CXCL1 showed a reproducible increase by 6 h after LPAL, changes in BAL CXCL2 were inconsistent. Measurements of CXCL1 were included since recent studies suggest that both chemokines can play a role in angiogenesis signaling [31,32]. However, given the pattern of protein and inflammatory cell changes at the three time points (0, 6 and 24 h), the appearance of CXCL1 seems to reflect injury. That we did not see robust early increases in CXCL2 was somewhat surprising since in our previous study, left lung homogenate showed a substantial increase between 4 and 24 hFigure 7. Changes in proliferating bronchial vessels. (A) Histologic section of airway demonstrating bronchial vessels by H E (left panel) and serial section stained for PCNA (right panel). Bronchial vessels show abundant PCNA+ 1676428 endothelial cells. (B) After LPAL (3 d), a significant increase in the fraction of PCNA+ bronchial vessels is observed. Treatment with dexamethasone had no sign.Ocalization of anti-CXCR2 (red) with RECA-1+ subepithelial blood vessels (green: 1006 original magnification, and inset 6006 original magnification). doi:10.1371/journal.pone.0066432.gAcute Ischemia and CXC ChemokinesFigure 6. Effects of dexamethasone on parenchymal and airway cells and cytokines/receptors. (A) Summary of the average effect of dexamethasone on measured parameters reflecting changes within the alveolar space. Each bar represents the average decrease 6 h after LPAL in dexamethasone treated rats compared with vehicle treated rats (3?1 rats/group, *P,0.05). B . Average effects of dexamethasone treatment on CXCL1, CXCL2, (mRNA, protein), CXCR1 and CXCR2 (mRNA) expression in the left bronchus 6 h after LPAL. Dexamethasone has no significant effects on any of the variables measured in the bronchus (3? rats/group). doi:10.1371/journal.pone.0066432.gchemokines within both lung compartments. However, the CXCL1 and CXCL2 modulation pattern was different within the two locations analyzed. CXCL1 release appears to be more generally associated with injury and anesthesia/surgery. Furthermore, anti-inflammatory treatment affected only the extent of lung injury, BAL CXCL1 level and BAL inflammatory cells. No changes were observed in the level of chemokine expression within bronchial tissue or, importantly, the magnitude of angiogenesis. In conclusion, results suggest that early changes within the bronchial niche where arteriogenesis originates, contribute to subsequent neovascularization during pulmonary ischemia. Our initial experiments evaluated BAL content as an index of predominantly parenchymal responses. In this model, we hypothesized that alveolar fluid could redistribute and act as a conduit for growth factors, through mucociliary movement. We demonstrated that acute ischemia caused an early increase in BAL total protein suggesting acute lung injury. These permeability changes were substantial (3-fold increase), early, and seemed to be attenuated by24 h after the onset of ischemia. We had shown previously that permeability changes were again seen 14 d after LPAL [12,29]. In addition, the inflammatory cell response paralleled the protein leak and likely reflected resident cells because of the lack of pulmonary blood flow, and only the small bronchial vasculature at this time point. Specifically, the inflammatory cell profile showed a predominance of neutrophils and macrophages, which, in addition to epithelial cells, have the capacity to secrete both CXCL1 and CXCL2 chemokines [30]. Although CXCL1 showed a reproducible increase by 6 h after LPAL, changes in BAL CXCL2 were inconsistent. Measurements of CXCL1 were included since recent studies suggest that both chemokines can play a role in angiogenesis signaling [31,32]. However, given the pattern of protein and inflammatory cell changes at the three time points (0, 6 and 24 h), the appearance of CXCL1 seems to reflect injury. That we did not see robust early increases in CXCL2 was somewhat surprising since in our previous study, left lung homogenate showed a substantial increase between 4 and 24 hFigure 7. Changes in proliferating bronchial vessels. (A) Histologic section of airway demonstrating bronchial vessels by H E (left panel) and serial section stained for PCNA (right panel). Bronchial vessels show abundant PCNA+ 1676428 endothelial cells. (B) After LPAL (3 d), a significant increase in the fraction of PCNA+ bronchial vessels is observed. Treatment with dexamethasone had no sign.

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