The Protective Role Of Perlfuorocarbon On Lipopolysaccharide-induced Impaired Function Of Epithelial Sodium Channelon Alveolar Epithelial Type â…¡ Cells

Objective: This study aimed to investigate the protective role of perfluorocarbonon lipopolysaccharide-induced impaired function of epithelial sodium channel onalveolar epithelial type II cell (ACE II).Methods:(1) injuries of A549cells induced by LPS (10μg/ml) were confirmed bydetecting mRNA expression of1L-1β, IL-6, TNF-α and ENaC-α mRNA, and proteinexpression of ENaC-α protein expression;(2) A549cells were divided into controlgroup, LPS group, PFC group and LPS/PFC group;(3)4h and6h after intervention,membrane and cytoplasm protein were extracted respectively, the leve of ENaC proteinon A549cytomembrane and cytoplasm were measured by western-blot;(4)4h afterafter intervention, whole-cell patch-clamp experiments were performedon A549cells torecording transmembrane current, transmembrane Na current through ENaC werecalculated by the change of transmembrane current recorded before and after addingENaCblocker amiloride(10-5M) to standard external solution;(5) A549cells were platedto snapwell filters, and cultured to form the tight monolayer epithelium.4h afterintervention, filters were mounted to ussing chamber for detecting short-circuit current(Isc), transepithelial Na current were define by the differences of Isc recorded beforeand after adding amiloride(10-5M) to K-H solution.Results:(1) LPS enhanced1L-1β, IL-6and TNF-α mRNA expression significantlyat4h, which were decreased but still higher than that of the control group at6h; LPSinhibited mRNA and protein expression of ENaC-α significantly at4h, which wereelevated but still lower than that of the control group at6group;(2) LPS inhibitedprotein level of ENaC-α on cytomembrane at both of4h and6h, and PFC improved that level inhibited by LPS respectively; while neither of LPS or PFC change the proteinlevel of ENaC-α in cytoplasm at4h and6h,respectively;(3) LPS reducedtransmembraneNa current through ENaC channel significantly4h after LPSintervention, while PFC can improve transmembrane sodium currentreduced by LPS;(4)LPS could significantly decreasedamiloride-sensitive Isc across A549monolayerepithelium4h after LPS intervention, while PFC can improve amiloride-sensitiveIscinhibited by LPS.Conclusions:LPS inhibitedNa transport on ACE II probaly by reducing the proteinlevel of ENaC on cytomenbrane, leading to damage to alveolar fluid clearance. PFCimprovedNa transport inhibited by LPS through increasing the protein level of ENaC oncytomenbrane, which suggested that PFC could enhanced the alveolar fluid clearance ofacute lung injury.