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Endothelial cells isolated from pulmonary arteries (RPAEC) and microcirculation (RPMVEC) of rat lungs were grown to confluence on porous filters and mounted on an Ussing-type chamber. Transmembrane pressure (ΔP) was controlled by the reservoir height, and the filtration rate corrected for surface area ( Jv/ A) was measured by timing fluid movement in a calibrated micropipette. These parameters were used to calculate hydraulic conductance (Lp) by using linear regression of Jv/ A on ΔP. Mean Lp values for newly confluent RPAEC monolayers were 22 times higher than those for RPMVEC monolayers (28.6 ± 5.6 vs. 1.30 ± 0.50 × 10−7 cm·s−1·cmH2O−1; P ≤ 0.01). After confluence was reached, electrical resistance and Lp remained stable in RPAEC but continued to change in RPMVEC with days in culture. Both phenotypes exhibited an initial time-dependent sealing response, but Lp also had an inverse relationship to ΔP in RPMVEC monolayers ≥4 days postconfluence that was attributed to cell overgrowth rather than junctional length. In a comparison of the cadherin contents, E-cadherin was predominant in RPMVEC, but VE-cadherin was predominant in RPAEC. At a constant ΔP of 40–45 cmH2O for 2 h, Jv/ A increased 225% in RPAEC monolayers but did not change significantly in RPMVEC monolayers. Significant decreases in Lp were obtained after treatment with 5% albumin, GdCl3, or isoproterenol plus rolipram in both phenotypes. Thus lung microvascular endothelial cells exhibited a significantly lower Lp than conduit vessel endothelium, which would limit alveolar flooding relative to perivascular edema cuff formation during increased pulmonary vascular pressures.

We compared the transport of three proteins with different hydrodynamic radii with ultrastructural changes in lungs of intact mice ventilated at peak inflation pressures (PIP) of 15, 35, 45, and 55 cmH2O for 2 h and PIP of 55 cmH2O for 0.5 and 1 h. After 2 h of ventilation, significant increases were observed in plasma Clara cell secretory protein (1.9 nm radius) at 35 cmH2O PIP and in bronchoalveolar lavage fluid albumin (3.6 nm radius) at 45 cmH2O PIP and IgG (5.6 nm radius) at 55 cmH2O PIP. Increased concentrations of all three proteins and lung wet-to-dry weight ratios were significantly correlated with PIP and ventilation time. Clara cell secretory protein and albumin increased significantly after 0.5 h of 55 cmH2O PIP, but IgG increased only after 2 h. Separation of endothelium or epithelium to form blebs was apparent only in small vessels (15-30 μm diameter) at 45 cmH2O PIP and after 0.5 h at 55 cmH2O PIP but became extensive after 2 h of ventilation at 55 cmH2O PIP. Junctional gaps between cells were rarely observed. Ultrastructural lung injury and protein clearances across the air-blood barrier were related to ventilation time and PIP levels. Protein clearances increased in relation to molecular size, consistent with increasing dimensions and frequency of transmembrane aqueous pathways.

Lung injury is a broad descriptor that can be applied to conditions ranging from mild interstitial edema without cellular injury to massive and fatal destruction of the lung. This review addresses those methods that can be readily applied to rats and mice whose small size limits the techniques that can be practically used to assess injury. The methodologies employed range from nonspecific measurement of edema formation to techniques for calculating values of specific permeability coefficient for the microvascular membrane in lung. Accumulation of pulmonary edema can be easily and quantitatively measured using gravimetric methods and indicates an imbalance in filtration forces or restrictive properties of the microvascular barrier. Lung compliance can be continuously measured, and light and electron microscopy can be used regardless of lung size to detect edema and structural damage. Increases in fluid and/or protein flux due to increased permeability must also be separated from those due to increased filtration pressure for mechanistic interpretation. Although an increase in the initial lung albumin clearance compared with controls matched for size and filtration pressure is a reliable indicator of endothelial dysfunction, calculated alterations in capillary filtration coefficient Kf,c, reflection coefficient σ, and permeability-surface area product PS are the most accurate indicators of increased permeability. Generally, PS and Kf,cwill increase and σ will decrease with vascular injury, but derecruitment of microvascular surface area may attenuate the affect on PS and Kf,cwithout altering measurements of σ.