Resolution Of Alveolar Edema Research Articles

Hypercapnia Induces Inositol-Requiring Enzyme 1α-Driven Endoplasmic Reticulum-associated Degradation of the Na,K-ATPase β-Subunit.

Acute respiratory distress syndrome is often associated with elevated levels of CO2 (hypercapnia) and impaired alveolar fluid clearance. Misfolding of the Na,K-ATPase (NKA), a key molecule involved in both alveolar epithelial barrier tightness and resolution of alveolar edema, in the endoplasmic reticulum (ER) may decrease plasma membrane abundance of the transporter. Here, we investigated how hypercapnia affects the NKA β-subunit (NKA-β) in the ER. Exposing murine precision-cut lung slices and human alveolar epithelial A549 cells to elevated CO2 levels led to a rapid decrease of NKA-β abundance in the ER and at the cell surface. Knockdown of ER mannosidase α class 1B member 1 and ER degradation-enhancing α-mannosidase like protein 1 by siRNA or treatment with the mannosidase α class 1B member 1 inhibitor kifunensine rescued loss of NKA-β in the ER, suggesting ER-associated degradation (ERAD) of the enzyme. Furthermore, hypercapnia activated the unfolded protein response by promoting phosphorylation of inositol-requiring enzyme 1α (IRE1α), and treatment with an siRNA against IRE1α prevented the decrease of NKA-β in the ER. Of note, the hypercapnia-induced phosphorylation of IRE1α was triggered by a Ca2+-dependent mechanism. In addition, inhibition of the inositol trisphosphate receptor decreased phosphorylation levels of IRE1α in precision-cut lung slices and A549 cells, suggesting that Ca2+ efflux from the ER might be responsible for IRE1α activation and ERAD of NKA-β. In conclusion, here we provide evidence that hypercapnia attenuates maturation of the regulatory subunit of NKA by activating IRE1α and promoting ERAD, which may contribute to impaired alveolar epithelial integrity in patients with acute respiratory distress syndrome and hypercapnia.

Hypercapnia Impairs Na,K-ATPase Function by Inducing Endoplasmic Reticulum Retention of the β-Subunit of the Enzyme in Alveolar Epithelial Cells.

Alveolar edema, impaired alveolar fluid clearance, and elevated CO2 levels (hypercapnia) are hallmarks of the acute respiratory distress syndrome (ARDS). This study investigated how hypercapnia affects maturation of the Na,K-ATPase (NKA), a key membrane transporter, and a cell adhesion molecule involved in the resolution of alveolar edema in the endoplasmic reticulum (ER). Exposure of human alveolar epithelial cells to elevated CO2 concentrations caused a significant retention of NKA-β in the ER and, thus, decreased levels of the transporter in the Golgi apparatus. These effects were associated with a marked reduction of the plasma membrane (PM) abundance of the NKA-α/β complex as well as a decreased total and ouabain-sensitive ATPase activity. Furthermore, our study revealed that the ER-retained NKA-β subunits were only partially assembled with NKA α-subunits, which suggests that hypercapnia modifies the ER folding environment. Moreover, we observed that elevated CO2 levels decreased intracellular ATP production and increased ER protein and, particularly, NKA-β oxidation. Treatment with α-ketoglutaric acid (α-KG), which is a metabolite that has been shown to increase ATP levels and rescue mitochondrial function in hypercapnia-exposed cells, attenuated the deleterious effects of elevated CO2 concentrations and restored NKA PM abundance and function. Taken together, our findings provide new insights into the regulation of NKA in alveolar epithelial cells by elevated CO2 levels, which may lead to the development of new therapeutic approaches for patients with ARDS and hypercapnia.

A novel tumor necrosis factor-mediated mechanism of direct epithelial sodium channel activation.

Alveolar liquid clearance is regulated by Na(+) uptake through the apically expressed epithelial sodium channel (ENaC) and basolaterally localized Na(+)-K(+)-ATPase in type II alveolar epithelial cells. Dysfunction of these Na(+) transporters during pulmonary inflammation can contribute to pulmonary edema. In this study, we sought to determine the precise mechanism by which the TIP peptide, mimicking the lectin-like domain of tumor necrosis factor (TNF), stimulates Na(+) uptake in a homologous cell system in the presence or absence of the bacterial toxin pneumolysin (PLY). We used a combined biochemical, electrophysiological, and molecular biological in vitro approach and assessed the physiological relevance of the lectin-like domain of TNF in alveolar liquid clearance in vivo by generating triple-mutant TNF knock-in mice that express a mutant TNF with deficient Na(+) uptake stimulatory activity. TIP peptide directly activates ENaC, but not the Na(+)-K(+)-ATPase, upon binding to the carboxy-terminal domain of the α subunit of the channel. In the presence of PLY, a mediator of pneumococcal-induced pulmonary edema, this binding stabilizes the ENaC-PIP2-MARCKS complex, which is necessary for the open probability conformation of the channel and preserves ENaC-α protein expression, by means of blunting the protein kinase C-α pathway. Triple-mutant TNF knock-in mice are more prone than wild-type mice to develop edema with low-dose intratracheal PLY, correlating with reduced pulmonary ENaC-α subunit expression. These results demonstrate a novel TNF-mediated mechanism of direct ENaC activation and indicate a physiological role for the lectin-like domain of TNF in the resolution of alveolar edema during inflammation.

Resolution of pulmonary edema. Thirty years of progress.

In the last 30 years, we have learned much about the molecular, cellular, and physiological mechanisms that regulate the resolution of pulmonary edema in both the normal and the injured lung. Although the physiological mechanisms responsible for the formation of pulmonary edema were identified by 1980, the mechanisms that explain the resolution of pulmonary edema were not well understood at that time. However, in the 1980s several investigators provided novel evidence that the primary mechanism for removal of alveolar edema fluid depended on active ion transport across the alveolar epithelium. Sodium enters through apical channels, primarily the epithelial sodium channel, and is pumped into the lung interstitium by basolaterally located Na/K-ATPase, thus creating a local osmotic gradient to reabsorb the water fraction of the edema fluid from the airspaces of the lungs. The resolution of alveolar edema across the normally tight epithelial barrier can be up-regulated by cyclic adenosine monophosphate (cAMP)-dependent mechanisms through adrenergic or dopamine receptor stimulation, and by several cAMP-independent mechanisms, including glucocorticoids, thyroid hormone, dopamine, and growth factors. Whereas resolution of alveolar edema in cardiogenic pulmonary edema can be rapid, the rate of edema resolution in most patients with acute respiratory distress syndrome (ARDS) is markedly impaired, a finding that correlates with higher mortality. Several mechanisms impair the resolution of alveolar edema in ARDS, including cell injury from unfavorable ventilator strategies or pathogens, hypoxia, cytokines, and oxidative stress. In patients with severe ARDS, alveolar epithelial cell death is a major mechanism that prevents the resolution of lung edema.

Fluid management in acute respiratory distress syndrome

Fluid management is one of the most important measures shown to impact acute respiratory distress syndrome (ARDS) outcomes. This review summarizes the current strategies aimed at evaluating and modulating lung fluid balance. Multiple recent studies have shown that a conservative fluid management in ARDS patients had beneficial effects on morbidity and mortality. These findings were replicated also in different patient populations assumed to have potential deleterious effects from this approach. So far, only one retrospective study raised the possibility of impaired cognitive function in ARDS patients managed with a conservative fluid strategy. Thermodilution methods and serum biomarkers can be used to monitor lung fluid balance and guide therapy. Recent evidence has indicated significant detrimental effects associated with beta-2 agonists use in ARDS, despite a putative beneficial role in the resolution of alveolar edema seen in preliminary studies. Dynamic monitoring of lung fluid balance needs to be implemented to guide fluid therapy in ARDS patients. A conservative fluid strategy seems safe and yields overall good clinical outcomes, but its impact on cognitive function needs to be evaluated in further studies. The role of colloids and other pharmacological agents deserves further investigation.

Variations in ENaC subunit composition may determine amiloride sensitivity and β-adrenergic stimulation of lung fluid absorption

reabsorption of fetal lung fluid at birth and resolution of alveolar edema in the adult lung are driven by active sodium and chloride transport across the alveolar epithelial barrier (for review see Ref. [8][1]). The rate of fetal lung fluid absorption and alveolar edema removal can be increased by

Thrombin Impairs Alveolar Fluid Clearance by Promoting Endocytosis of Na+,K+-ATPase

Coagulation is an emerging area of interest in the pathogenesis and treatment of acute lung injury. Concentrations of the edemagenic coagulation protease thrombin are elevated in plasma and lavage fluids from afflicted patients. We explored the impact of thrombin on the formation and resolution of alveolar edema. Intravascularly applied thrombin inhibited active transepithelial 22Na transport in intact rabbit lungs, suppressing alveolar fluid clearance. Epithelial permeability was unaffected, whereas endothelial permeability was increased. In A549 human lung epithelial cells and in mouse primary alveolar type II cells, thrombin blocked ouabain-sensitive Na+,K+-ATPase-mediated 86Rb+ uptake, without altering amiloride-sensitive sodium currents. Furthermore, thrombin downregulated cell-surface expression of Na+,K+-ATPase, but not ENaC alpha and beta subunits. The endocytosis inhibitor phalloidin oleate blocked all thrombin-induced effects on sodium transport activity. Similarly, diphenyleneiodonium chloride, an inhibitor of reactive oxygen radical production, as well as a protein kinase C-zeta inhibitor, prevented these thrombin-induced effects. Thus, thrombin signaling via reactive oxygen species and protein kinase C-zeta promotes Na+,K+-ATPase endocytosis, resulting in loss of function. We propose here a dual role for thrombin in mediating disturbances to fluid balance in the lung: thrombin concomitantly provokes edema formation by increasing endothelial permeability, and inhibits alveolar edema resolution by blocking Na+,K+-ATPase function.

Alveolar Epithelium: Role in Lung Fluid Balance and Acute Lung Injury

The resolution of alveolar edema is regulated by active sodium and chloride transport across the pulmonary epithelium, including alveolar epithelial type I and II cells as well as distal airway epithelia. Catecholamine-dependent mechanisms can markedly upregulate alveolar fluid clearance even under pathological conditions, an effect that is mediated by both epithelial sodium channel (ENaC) and cystic fibrosis transmembrane conductance regulator (CFTR). Under pathological conditions, impaired alveolar fluid clearance is associated with worse survival in patients with acute lung injury. However, there is some experimental and clinical evidence that cAMP stimulation could accelerate the resolution of pulmonary edema in the presence of acute lung injury. Clinical trials are needed to test this potential therapeutic strategy in patients with acute lung injury.

Integrated control of lung fluid balance

This review summarizes the highlights of the EB2004 symposium that dealt with the integrated aspects of the lung fluid balance. It is apparent that maintenance of lung fluid balance requires the proper functioning of vascular endothelial and alveolar epithelial barriers. Under physiological conditions, the transcytotic pathway requiring repeated fission-fusion events of the caveolar membrane with other caveolae solely transports albumin. Caveolin-1, which forms caveolae, and albumin-binding proteins play a central role in signaling the transcytosis of albumin. Signals responsible for increasing endothelial permeability in lung microvessels in response to inflammatory mediators were also described. These studies in gene knockout mouse models revealed the importance of Ca(2+) signaling via store-operated transient receptor channel 4 and the activation of endothelial myosin light chain kinase isoform in mediating the increase in microvessel permeability. Increases in the cytosolic Ca(2+) in situ in microvessel endothelia can occur by mitochondria-dependent as well as mitochondria-independent pathways (such as the endoplasmic reticulum). Both these pathways, by triggering endothelial cell activation, may result in lung microvascular injury. The resolution of alveolar edema, requiring clearance of fluid from the air space, is another area of intense investigation in animal models. Although beta-adrenergic agonists can activate alveolar fluid clearance, signaling pathways regulating these events in intact alveoli remain to be established. Development of mouse models in which the function of regulatory proteins (identified in cell culture studies) can be systematically analyzed will provide a better and more integrated picture of lung fluid balance. In vivo veritas!

Functional identification of the alveolar edema reabsorption activity of murine tumor necrosis factor-alpha.

Tumor necrosis factor-alpha (TNF-alpha) activates sodium channels in Type II alveolar epithelial cells, an important mechanism for the reported fluid resorption capacity of the cytokine. Both TNF-alpha receptor-dependent and -independent effects were proposed for this activity in vitro, the latter mechanism mediated by the lectin-like domain of the molecule. In this study, the relative contribution of the receptor-dependent versus receptor-independent activities was investigated in an in situ mouse lung model and an ex vivo rat lung model. Fluid resorption due to murine TNF-alpha (mTNF-alpha) was functional in mice that were genetically deficient in both types of mTNF-alpha receptor, establishing the importance of mTNF-alpha receptor-independent effects in this species. In addition, we assessed the capacity of an mTNF-alpha-derived peptide (mLtip), which activates sodium transport by a receptor-independent mechanism, to reduce lung water content in an isolated, ventilated, autologous blood-perfused rat lung model. The results show that in this model, mLtip, in contrast to mTNF-alpha, produced a progressive recovery of dynamic lung compliance and airway resistance after alveolar flooding. There was also a significant reduction in lung water. These results indicate that the receptor-independent lectin-like domain of mTNF-alpha has a potential physiological role in the resolution of alveolar edema in rats and mice.

Regulation of ion and fluid transport across the distal pulmonary epithelia: new insights

this issue of the American Journal of Physiology-Lung Cellular and Molecular Physiology features nine articles that provide new insights into the mechanisms and the cellular pathways that regulate ion transport across the distal pulmonary epithelia in both the adult and fetal lung. Although much has

Recent developments in pulmonary edema.

Research on lung fluid balance and pulmonary edema has increased during the last decade. New approaches have led to insights into the role of each component of the alveolar-capillary barrier. The capillary endothelium is the first line of defense against lung fluid accumulation. The interstitium may play a more important role in lung fluid balance than previously appreciated. Active and passive transport properties of alveolar epithelium may be important in the pathogenesis and resolution of alveolar edema. New methods for the determination of epithelial permeability and lung water are being evaluated. The developments reviewed here may have an impact on the institution of new diagnostic and therapeutic approaches to pulmonary edema during the next decade.