AT₁ levels.

<p>(A) A semi-quantative measure of AT1 protein levels in AEC II by western blotting; AT1 levels were significantly increased by 1.67 fold after Ang II administration. *P<0.0001 as compared to control group. CT—Control. Ang II—Angiotensin II. AT1—Angiotensin-II receptor type 1. The bars represent mean ± SEM. (B) An immunohestochemical staining of AT1 receptor in AEC II cells treated or untreated with Ang II. The representative figure showing stronger staining of AT1 in the Ang II treated group compared to the control. CT—Control. Ang II—Angiotensin II.</p

A comparable scheme of AFC under normal vs. Ang II stimulated conditions.

<p>Under normal conditions Na+ is extruded out of the alveolar airspace by apical epithelial Na+ channels (ENaC), specifically highly selective cation channels composed of α, β and γ subunits (HSC) and basolateral Na,K-ATPase pump with water following osmoticaly, Whereas Ang II stimulation down regulated cAMP levels in AEC II, via AT1 receptors triggering, thus leading to the decrease of the two important AFC players; αNa,K-ATPase and the HSC, and an increase of the NSC (non-selective cation channels composed of α subunit alone), with a resultant impairment of sodium reabsorption and conceivable AFC decrease.</p

Effect of Ang II on AFC.

<p>(A) % Alveolar fluid clearance of the initial instilled volume was decreased in the Ang II groups in a dose dependent manner, from 8.6% ± 0.19 in control rats to 6.66% ± 0.13, 6.15% ± 0.11, 5.03% ± 0.31, 4.42% ± 0.29 and 5.25% ± 0.23 in Ang II (10⁻¹⁰ M, 10^-9M, 10⁻⁸ M, 10⁻⁷ M and 10⁻⁶ M) respectively. * P<0.001 As compared to control group; ** P<0.05 As compared to the rest of 10⁻¹⁰ M and 10⁻⁹ M Ang II treated groups. CT—Control. The bars represent mean ± SEM. (B) The albumin movement across the alveolar-capillary barrier did not differ significantly among the study groups indicating that the barrier was intact. CT—Control. The bars represent mean ± SEM.</p

Levels of cAMP.

<p>Levels of cAMP were decreased in Ang II treated AEC II cells by 0.17 folds compared to the control group. * P < 0.05 As compared to control group. CT—Control. Ang II—Angiotensin II. The bars represent mean ± SEM.</p

αNa,K-ATPase levels.

<p>(A) Na,K-ATPase α1 subunit levels in whole cell AEC II were measured by western blotting. Ang II administration decreased Na,K-ATPase levels by 0.65 folds compared to the control group. * P < 0.001 As compared to control group. CT—Control. Ang II—Angiotensin II. The bars represent mean ± SEM. (B) A representative Immunofluorescence staining of α1-Na,K-ATPase shows a shift of the protein membrane localization to internal organelles following Ang II administration. CT—Control. Ang II—Angiotensin II.</p

Different interventions effect on AFC.

<p>(A) Losartan restored Ang II effect on AFC from 5.25%±0.23 to 8.1%±0.13. AFC was not different in both losartan treated groups. * P<0.001 As compared to control group treated with Ang II. PD123319, AT₂ receptor antagonist, decreased AFC in both AngII treated (n = 4) and untreated groups (n = 4) (6.54%±0.2 and 6.51%±0.2 respectively). $ P<0.05 as compared to Ang II group, & P<0.001 as compared to control group. Ouabain, the Na,K-ATPase blocker, significantly inhibited AFC in both control and Ang II treated rat lungs (5.9% ± 0.4 and 4.4% ± 0.2 respectively). + P<0.05 as compared to control rat lungs treated with ouabain alone. Amiloride, the sodium channel blocker, significantly reduced AFC in both control and Ang II treated rats as compared to untreated lungs (a 5.6% ± 0.2 and 5.01 ± 0.2 respectively). However, AFC was similar in the two Amiloride treated groups. Activating the adrenergic pathway by norepinephrine 10^-6M increased the clearance percentage to 14.12% ± 1.8, when compared to control 8.6% ± 0.19. But when Ang II was also added, NE effect was abolished (7.3% ± 0.6). # P<0.05 as compared to control rat lungs treated with norepinephrine alone. ## P<0.0001 as compared to AngII group. CT—Control. Ang II—Angiotensin II. NE—Norepinephrine. The bars represent mean ± SEM. (B) The albumin movement across the alveolar-capillary barrier did not differ significantly among the study groups indicating that the barrier was intact. CT—Control. Ang II—Angiotensin II. Los—Losartan. PD—PD123319. Ouab—Ouabain. Amil—Amiloride. NE—Norepinephrine. The bars represent mean ± SEM.</p