| Objective Acute lung injury (ALI) is characterized by uncontrolledinflammatory reponse, increased permeablility pulmonary edema, alveolarfilling and respiratory failure. Angiotensin II (Ang II) is the main effector ofrennin-angiotensin system (RAS), and exerts its effect in maintainingsodium balance and extracellular fluid volume. Ang II has beendemonstrated a pro-inflammatory effect in acute lung injury, however,studies of the effect of Ang II on the formation of pulmonary edema andalveolar filling remains unclear. Epithelial sodium channel (ENaC) is therate-limiting step for sodium absorption in alveoli, which in turn regulatesthe alveolar fluid volume. There is mounting evidence form renal andcolonic studies demonstrating that Ang II regulates ENaC expression andits electrogenous activity. However, the effect of Ang II on ENaCexpression in lung is still unknown. Therefore, in this study was verified theregulation of alveolar fluid clearance (AFC) and the expression of epithelialsodium channel (ENaC) by endogenous and exogenous Ang II. The effect ofAng II on ENaC expression was still verified in A549cells.Methods In study on endogenous Ang II effects, SD rats wereanesthetized and intratracheally injected with1mg/kg lipopolysaccharide (LPS), whereas control rats received only saline vehicle. Angiotensin type1(AT1) receptor antagonist ZD7155(10mg/kg) was injected intraperitoneally30min before administration of LPS. Then lungs were isolated en bloc. Theleft lungs were separated to measure lung water volume and bronchoalveolarlavage fluid (BALF). The right lungs were prepared to assess alveolar fluidclearance. The lungs were processed histological analysis andimmunological studies with HE stain and immunocytochemistry. Ang IIlevels in plasma and lung tissue were determined by ELISA. The proteinexpression of ENaC was detected by Western blot. In study on exogenousAng II effects, SD rats were anesthetized and were given Ang II withincreasing doses (1,10and100μg/kg-1·min-1) via osmotic minipumps,whereas control rats received only saline vehicle. AT1receptor antagonistZD7155(10mg/kg) and inhibitor of cAMP degeneration rolipram (1mg/kg)were injected intraperitoneally30min before administration of Ang II. Thenlungs were isolated en bloc. The left lungs were separated to measure lungwater volume and bronchoalveolar lavage fluid (BALF). The right lungswere prepared to assess alveolar fluid clearance. The lungs were processedhistological analysis and immunological studies with HE stain andimmunocytochemistry. TNF-α, IL-1β and cAMP levels in lung tissue weredetermined by ELISA and RIA, respectively. The mRNA and proteinexpression of ENaC were detected by RT-PCR and Western blot. A549cellswere treated with different doses (10-7M and10-5M) of Ang II and the change in protein level caused by Ang II was determined by Western blot.Results In study on endogenous Ang II effects, ALI induced by LPScaused an increase in Ang II levels in plasma and lung tissue, but adecrease in alveolar fluid clearance. The time-course of the Ang II levelsparalleled that of the alveolar fluid clearance (In plasma: ALI2h2.53±0.4μg/l, ALI4h3.78±1.15μg/l, ALI6h4.91±1.1μg/l; In lung: ALI2h190.2±38.23μg/l, ALI4h305±72.99μg/l, ALI6h580.4±129.16μg/l).Pretreatment with ZD7155prevented ALI-induced reduction of alveolarfluid clearance (7.8±1.61%vs4.4±1.14%). ZD7155also reversed theALI-induced reduction of the abundance of β and γ-ENaC, whereas furtherdecreased the abundance of α-ENaC (P<0.05), and ameliorated interstitialedema and inflammatory cell infiltration induced by ALI. In study onexogenous Ang II effects, Exposure to higher doses of Ang II reduced AFCin a dose-dependent manner and resulted in a non-coordinate regulation ofα-ENaC versus the regulation of β-and γ-ENaC, however Ang II type1(AT1) receptor antagonist ZD7155prevented the Ang II-induced inhibitionof fluid clearance (10.8±1.483%vs6.2±2.683%) and dysregulation ofENaC expression (P<0.05). The similar non-coordinate regulation ofα-ENaC versusβ-and γ-ENaC by Ang II was still observed in A549cells,and Ang II dysregulated ENaC expression in a concentration-dependentmanner (P<0.05). In addition, exposure to inhibitor of cAMP degradationrolipram blunted the Ang II-induced inhibition of fluid clearance (9.8± 3.033%vs6.2±2.683%).Conclusions Endogenous and exogenous Ang II promotes pulmonaryedema and alveolar filling by inhibition of alveolar fluid clearance throughactivation of AT1receptor. Ang II regulates α-ENaC expresson versus β-and γ-ENaC expression in a non-coordinate manner, which may lead tochanges in proportion of HSC channels and NSC channels, and results inreduction of AFC. AT1receptor antagonist and inhibitor of cAMPdegeneration can protect from Ang II-induced reduction of AFC.
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