Sepsis-induced acute respiratory distress syndrome (ARDS) causes significant fatalities worldwide and lacks pharmacological intervention. Alveolar fluid clearance (AFC) plays a pivotal role in the remission of ARDS and is markedly impaired in the pathogenesis of ARDS. Here, we demonstrated that erythropoietin could effectively ameliorate lung injury manifestations and lethality, restore lung function and promote AFC in a rat model of lipopolysaccharide (LPS)-induced ARDS. Moreover, it was proven that EPO-induced restoration of AFC occurs through triggering the total protein expression of ENaC and Na,K-ATPase channels, enhancing their protein abundance in the membrane, and suppressing their ubiquitination for degeneration. Mechanistically, the data indicated the possible involvement of EPOR/JAK2/STAT3/SGK1/Nedd4–2 signaling in this process, and the pharmacological inhibition of the pathway markedly eliminated the stimulating effects of EPO on ENaC and Na,K-ATPase, and subsequently reversed the augmentation of AFC by EPO. Consistently, in vitro studies of alveolar epithelial cells paralleled with that EPO upregulated the expression of ENaC and Na,K-ATPase, and patch-clamp studies further demonstrated that EPO substantially strengthened sodium ion currents. Collectively, EPO could effectively promote AFC by improving ENaC and Na,K-ATPase protein expression and abundance in the membrane, dependent on inhibition of ENaC and Na,K-ATPase ubiquitination, and resulting in diminishing LPS-associated lung injuries.

Endothelial glycocalyx degradation, critical for increased pulmonary vascular permeability, is thought to facilitate the development of sepsis into the multiple organ failure. Maresin conjugates in tissue regeneration 1 (MCTR1), a macrophage-derived lipid mediator, which exhibits potentially beneficial effects via the regulation of bacterial phagocytosis, promotion of inflammation resolution, and regeneration of tissue. In this study, we show that MCTR1 (100 ng/mouse) enhances the survival of mice with lipopolysaccharide (LPS)-induced (15 mg/kg) sepsis. MCTR1 alleviates LPS (10 mg/kg)-induced lung dysfunction and lung tissue inflammatory response by decreasing inflammatory cytokines (tumor necrosis factor-alpha, interleukin-1 beta [IL-1 beta], and IL-6) expression in serum and reducing the serum levels of heparan sulfate (HS) and syndecan-1. In human umbilical vein endothelial cells (HUVECs) experiments, MCTR1 (100 nM) was added to the culture medium with LPS for 6 hr. MCTR1 treatment markedly inhibited HS degradation by downregulating heparanase (HPA) protein expression in vivo and in vitro. Further analyses indicated that MCTR1 upregulates sirtuin 1 (SIRT1) expression and decreases NF-kappa B p65 phosphorylation. In the presence of BOC-2 or EX527, the above effects of MCTR1 were abolished. These results suggest that MCTR1 protects against LPS-induced sepsis in mice by attenuating pulmonary endothelial glycocalyx injury via the ALX/SIRT1/NF-kappa B/HPA pathway.

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are common and devastating clinical disorders with high mortality and no specific therapy. An excessive inflammatory response results in the progression of ALI/ARDS, and the NLRP3 inflammasome is a key participant in inflammation. Erythropoietin (EPO), which is clinically used for anemia, reportedly exerts pleiotropic effects in ALI. However, whether EPO could protect against lipopolysaccharide (LPS)-induced ALI by regulating the NLRP3 inflammasome and its underlying mechanisms remain poorly elucidated. This study aimed to explore whether the therapeutic effects of EPO rely on the suppression of the NLRP3 inflammasome and the specific mechanisms in an LPS-induced ALI mouse model. ALI was induced in C57BL/6 mice by intraperitoneal (i.p.) injection of LPS (15 mg/kg). EPO was administered intraperitoneally at 5 U/g after LPS challenge. The mice were sacrificed 8 h later. Our findings indicated that application of EPO markedly diminished LPS-induced lung injury by restoring histopathological changes, lessened lung wet/dry (W/D) ratio, protein concentrations in bronchoalveolar lavage fluid (BALF) and myeloperoxidase (MPO) levels. Meanwhile, EPO evidently decreased interleukin-1β (IL-1β) and interleukin-18 (IL-18) secretion, the expression of NLRP3 inflammasome components including pro-IL-1β, NLRP3, and cleaved caspase-1 as well as phosphorylation of nuclear factor-κB (NF-κB) p65, which may be associated with activation of EPO receptor (EPOR), phosphorylation of Janus-tyrosine kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3). However, all the beneficial effects of EPO on ALI and modulation NLRP3 inflammasome were remarkably abrogated by the inhibition of EPOR/JAK2/STAT3 pathway and knockout (KO) of NLRP3 gene. Taken together, this study indicates that EPO can effectively attenuate LPS-induced lung injury in mice by suppressing the NLRP3 inflammasome, which is dependent upon activation of EPOR/JAK2/STAT3 signaling and inhibition of the NF-κB pathway.

Maresin Conjugates in Tissue Regeneration 1 (MCTR1) is a newly identified macrophage-derived sulfido-conjugated mediator that stimulates the resolution of inflammation. This study assessed the role of MCTR1 in alveolar fluid clearance (AFC) in a rat model of acute lung injury (ALI) induced by lipopolysaccharide (LPS). Rats were intravenously injected with MCTR1 at a dose of 200 ng/rat, 8 hours after administration of 14 mg/kg LPS. The level of AFC was then determined in live rats. Primary rat ATII (Alveolar Type II) epithelial cells were also treated with MCTR1 (100 nmol/L) in a culture medium containing LPS for 8 hours. MCTR1 treatment improved AFC (18.85 +/- 2.07 vs 10.11 +/- 1.08, P < .0001) and ameliorated ALI in rats. MCTR1 also significantly promoted AFC by up-regulating epithelial sodium channel (ENaC) and Na+-K+-adenosine triphosphatase (Na, K-ATPase) expressions in vivo. MCTR1 also activated Na, K-ATPase and elevated phosphorylated-Akt (P-Akt) by up-regulating the expression of phosphorylated Nedd4-2 (P-Nedd4-2) in vivo and in vitro. However, BOC-2 (ALX inhibitor), KH7 (cAMP inhibitor) and LY294002 (PI3K inhibitor) abrogated the improved AFC induced by MCTR1. Based on the findings of this study, MCTR1 may be a novel therapeutic approach to improve reabsorption of pulmonary oedema during ALI/acute respiratory distress syndrome (ARDS).

Gram-negative bacterial infection causes an excessive inflammatory response and acute organ damage or dysfunction due to its outer membrane component, lipopolysaccharide (LPS). Protectin conjugates in tissue regeneration 1 (PCTR1), an endogenous lipid mediator, exerts fundamental anti-inflammation and pro-resolution during infection. In the present study, we examined the properties of PCTR1 on the systemic inflammatory response, organic morphological damage and dysfunction, and serum metabolic biomarkers in an LPS-induced acute inflammatory mouse model. The results show that PCTR1 reduced serum inflammatory factors and ameliorated morphological damage and dysfunction of the lung, liver, kidney, and ultimately improved the survival rate of LPS-induced acute inflammation in mice. In addition, metabolomics analysis and high performance liquid chromatography-mass spectrometry revealed that LPS-stimulated serum linoleic acid (LA), arachidonic acid (AA), and prostaglandin E2 (PGE2) levels were significantly altered by PCTR1. Moreover, PCTR1 upregulated LPS-inhibited fatty acid desaturase 1 (FADS1), fatty acid desaturase 2 (FADS2), and elongase of very long chain fatty acids 2 (ELOVL2) expression, and downregulated LPS-stimulated phospholipase A2 (PLA2) expression to increase the intrahepatic content of AA. However, these effects of PCTR1 were partially abrogated by a lipoxin A4 receptor (ALX) antagonist (BOC-2). In summary, via the activation of ALX, PCTR1 promotes the conversion of LA to AA through upregulation of FADS1, FADS2, and ELOVL2 expression, and inhibits the conversion of bound AA into free AA through downregulation of PLA2 expression to decrease the serum AA and PGE2 levels.

Acute respiratory distress syndrome (ARDS) is a lethal clinical syndrome characterized by damage of the epithelial barriers and accumulation of pulmonary edema fluid. Protectin conjugates in tissue regeneration 1 (PCTR1), an endogenously produced lipid mediator, are believed to exert anti‐inflammatory and pro‐resolution effects. PCTR1 (1 µg/kg) was injected at 8 hr after lipopolysaccharide (LPS; 14 mg/kg) administration, and the rate of pulmonary fluid clearance was measured in live rats at 1 hr after PCTR1 treatment. The primary type II alveolar epithelial cells were cultured with PCTR1 (10 nmol/ml) and LPS (1 μg/ml) for 8 hr. PCTR1 effectively improved pulmonary fluid clearance and ameliorated morphological damage and reduced inflammation of lung tissue, as well as improved the survival rate in the LPS‐induced acute lung injury (ALI) model. Moreover, PCTR1 markedly increased sodium channel expression as well as Na, K‐ATPase expression and activity in vivo and in vitro. In addition, PCTR1i also upregulated the expression of LYVE‐1 in vivo. Besides that, BOC‐2, HK7, and LY294002 blocked the promoted effect of PCTR1 on pulmonary fluid clearance. Taken together, PCTR1 upregulates sodium channels' expression via activating the ALX/cAMP/P‐Akt/Nedd4‐2 pathway and increases Na, K‐ATPase expression and activity to promote alveolar fluid clearance. Moreover, PCTR1 also promotes the expression of LYVE‐1 to recover the lymphatic drainage resulting in the increase of lung interstitial fluid clearance. In summary, these results highlight a novel systematic mechanism for PCTR1 in pulmonary edema fluid clearance after ALI/ARDS, suggesting its potential role in a therapeutic approach for ALI/ARDS.

Maresin1 (MaR1), a new anti-inflammatory and proresolving lipid mediator, has been proven to exert organ-protective effects in septic animal models. However, the potential mechanisms are still not fully elucidated. In this study, we sought to explore the impact of MaR1 on metabolic dysfunction in cecal ligation and puncture- (CLP-) induced septic mice. We found that MaR1 significantly increased the overall survival rate and attenuated lung and liver injuries in septic mice. In addition, MaR1 markedly reduced the levels of proinflammatory cytokines (TNF-α and IL-6) and alleviated mitochondrial damage. Based on a 1H NMR-based metabolomics analysis, CLP-induced septic mice had increased levels of acetate, pyruvate, and lactate in serum and decreased levels of alanine, aspartate, glutamate, and fumarate in lungs. However, these metabolic disorders, mainly involving energy and amino acid metabolism, can be recovered by MaR1 treatment. Therefore, our results suggest that the protective effects of MaR1 on sepsis could be related to the recovery of metabolic dysfunction and the alleviation of inflammation and mitochondrial damage.