Compartment‐Specific Effects of Alveolar Heparan Suflate Shedding after Intratracheal Lipopolysaccharide

Abstract

Acute lung injury can occur as a consequence of direct (pulmonary) insults that target the alveolar epithelium or indirect (extrapulmonary) insults that injure the alveolar endothelium. During indirect lung injury, the endothelial glycocalyx is degraded, releasing highly‐sulfated heparan sulfate (HS) octasaccharides into the vasculature. In contrast to indirect lung injury, it is unknown whether direct lung injury induces release of HS from the apical surface of the damaged alveolar epithelium or the luminal endothelial glycocalyx. We hypothesized that during direct lung injury HS would be shed from injured epithelium, and aimed to determine the function of the shed HS.To determine if HS was shed from the alveolar epithelium into the airspace or from the luminal endothelium into the vasculature, we intratracheally (IT) instilled mice with lipopolysaccharide (LPS) and performed mass spectrometry for HS on bronchoalveolar lavage fluid (BALF) and plasma collected from these mice. We measured increased BALF HS in mice treated with IT LPS vs PBS on day 2 (24.5 vs 6.5ng/ml) and day 4 (18.8 vs 7.5ng/ml) after injury. There was also a trend of increased plasma HS in mice treated with IT LPS vs PBS 4 days after injury (106.8 vs 63.9ng/ml, p = 0.10). We next aimed to determine the function of the shed HS within both the airspace and the vasculature by supplementing mice with exogenous IT or IV heparin octasaccharides (a heavily sulfated HS fragment) on days 2 and 4 after IT LPS injury. Mice supplemented with IV heparin octasaccharides vs PBS on day 2 after injury were protected from injury, exhibiting decreased lung permeability (BALF protein: 0.34 vs 0.40mg/ml) and inflammation (BALF total cells: 3.14 vs 5.63×106). In contrast, animals treated with IT heparin octasaccharides on day 4 after injury exhibited a trend of increased lung permeability (BALF protein: 0.23 vs 0.20mg/ml, p=0.07). Our data indicate that during direct lung injury HS is largely shed from the damaged epithelium, but may also be shed from the endothelium. While intravascular HS is pro‐reparative and decreases injury, airway HS surprisingly appears to exacerbate injury and reduce repair. We are currently investigating the mechanisms by which IT and IV heparin affect lung injury.Support or Funding InformationNHLBI RO1 HL125371NHLBI F30 HL132424