川芎嗪对脓毒症诱导的急性肺损伤小鼠血管内皮生长因子水平变化的影响

目的研究川芎嗪(Tetramethylpyrazine,TMP)对脓毒症诱导的小鼠急性肺损伤(Acute lung injury,ALI)的保护作用以及对血管内皮生长因子(Vascular endothelial growth factor,VEGF)水平变化的影响。方法将小鼠随机分为假手术组(Sham组)、脓毒症组(Sep组)、治疗对照组(NS组)、TMP-A组和TMP-B组,Sep组、NS组、TMP-A组和TMP-B组采用盲肠结扎穿刺术(Cecal ligation and puncture,CLP)复制脓毒症相关性ALI模型,造模成功后,TMP-A组和TMP-B组分别经腹腔注射100 mg/kg和40 mg/kg TMP,Sham组和NS组经腹腔注射等量生理盐水,Sep组不作处理。分别于造模后0、12、24和48 h取材,HE染色观察小鼠肺组织病理学变化;ELISA检测小鼠血浆中VEGF含量变化;Western blot检测小鼠肺组织VEGF蛋白水平。结果 TMP组小鼠在造模后12 h时肺组织病理变化与Sep组相比有所减轻,24 h时肺泡水肿、肺出血及中性粒浸润均明显减轻,48 h时TMP作用最理想;TMP组小鼠血浆VEGF水平造模后24 h开始低于Sep组(P<0.05),48 h持续下降,已接近Sham组水平;TMP组小鼠肺组织VEGF蛋白水平在造模后24 h比Sep组明显升高,但低于Sham组(P<0.05);100 mg/kg TMP的效果优于40 mg/kg;小鼠肺组织VEGF水平与血浆VEGF水平呈负相关(P<0.05)。结论 TMP能够有效减轻脓毒症所致ALI,这一作用与降低血浆VEGF水平,恢复肺组织VEGF蛋白水平有关。     

Protective Role of Proton-Sensing TDAG8 in Lipopolysaccharide-Induced Acute Lung Injury

Acute lung injury is characterized by the infiltration of neutrophils into lungs and the subsequent impairment of lung function. Here we explored the role of TDAG8 in lung injury induced by lipopolysaccharide (LPS) administrated intratracheally. In this model, cytokines and chemokines released from resident macrophages are shown to cause neutrophilic inflammation in the lungs. We found that LPS treatment increased TDAG8 expression in the lungs and confirmed its expression in resident macrophages in bronchoalveolar lavage (BAL) fluids. LPS administration remarkably increased neutrophil accumulation without appreciable change in the resident macrophages, which was associated with increased penetration of blood proteins into BAL fluids, interstitial accumulation of inflammatory cells, and damage of the alveolar architecture. The LPS-induced neutrophil accumulation and the associated lung damage were enhanced in TDAG8-deficient mice as compared with those in wild-type mice. LPS also increased several mRNA and protein expressions of inflammatory cytokines and chemokines in the lungs or BAL fluids. Among these inflammatory mediators, mRNA and protein expression of KC (also known as CXCL1), a chemokine of neutrophils, were significantly enhanced by TDAG8 deficiency. We conclude that TDAG8 is a negative regulator for lung neutrophilic inflammation and injury, in part, through the inhibition of chemokine production.     

Methotrexate Ameliorates Systemic Inflammation and Septic Associated-Lung Damage in a Cecal Ligation and Puncture Septic Rat Model

Background: Sepsis is a serious, heterogeneous clinical entity produced by a severe and systemic host inflammatory response to infection. Methotrexate (MTX) is a folate-antagonist that induces the generation of adenosine and also inhibits JAK/STAT pathway; MTX it is widely used as an anti-inflammatory drug to control the immune system. Objective: The aim of this study was to assess the beneficial effects of a single and low dose of MTX in the systemic response and acute lung injury (ALI) induced by sepsis. As in the clinics, we treated our animals with antibiotics and fluids and performed the source control to mimic the current clinic treatment. Methods and main results: Sepsis was induced in rats by a cecal ligation puncture (CLP) procedure. Six hours after induction of sepsis, we proceeded to the source control; fluids and antibiotics were administered at 6 h and 24 h after CLP. MTX (2.5 mg/Kg) was administered 6 h after the first surgery in one CLP experimental group and to one Sham group. A protective effect of MTX was observed through a significant reduction of pro-inflammatory cytokines and a decrease infiltration of inflammatory cells in the lung. In addition, we found a regulation in adenosine receptor A2aR and the metalloproteinases by MTX. Conclusion: A single, low dose of MTX attenuates sepsis lung-associated damage by decreasing pro-inflammatory response, infiltration of pro-inflammatory cells and avoiding defective tissue lung remodeling.     

Role of S100A8/A9 for Cytokine Secretion,Revealed in Neutrophils Derived from ER-Hoxb8 Progenitors

S100A9, a Ca²⁺-binding protein, is tightly associated to neutrophil pro-inflammatory functions when forming a heterodimer with its S100A8 partner. Upon secretion into the extracellular environment, these proteins behave like damage-associated molecular pattern molecules, which actively participate in the amplification of the inflammation process by recruitment and activation of pro-inflammatory cells. Intracellular functions have also been attributed to the S100A8/A9 complex, notably its ability to regulate nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation. However, the complete functional spectrum of S100A8/A9 at the intracellular level is far from being understood. In this context, we here investigated the possibility that the absence of intracellular S100A8/A9 is involved in cytokine secretion. To overcome the difficulty of genetically modifying neutrophils, we used murine neutrophils derived from wild-type and S100A9^−/− Hoxb8 immortalized myeloid progenitors. After confirming that differentiated Hoxb8 neutrophil-like cells are a suitable model to study neutrophil functions, our data show that absence of S100A8/A9 led to a dysregulation of cytokine secretion after lipopolysaccharide (LPS) stimulation. Furthermore, we demonstrate that S100A8/A9-induced cytokine secretion was regulated by the nuclear factor kappa B (NF-κB) pathway. These results were confirmed in human differentiated HL-60 cells, in which S100A9 was inhibited by shRNAs. Finally, our results indicate that the degranulation process could be involved in the regulation of cytokine secretion by S100A8/A9.     

Inhibition of Prostaglandin F2α Receptors Exaggerates HCl-Induced Lung Inflammation in Mice

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are severe respiratory disorders that are caused by aspiration, sepsis, trauma, and pneumonia. A clinical feature of ALI/ARDS is the acute onset of severe hypoxemia, and the mortality rate, which is estimated at 38–50%, remains high. Although prostaglandins (PGs) are detected in the bronchoalveolar lavage fluid of patients with ALI/ARDS, the role of PGF_2α in ALI remains unclear. We aimed to clarify the role of PGF_2α/PGF_2α receptor (FP) signaling in acid-induced ALI using an FP receptor antagonist, AL8810. Intratracheal injection of hydrochloric acid (HCl) increased neutrophil migration into the lungs, leading to respiratory dysfunction. Pre-administration of AL8810 further increased these features. Moreover, pre-treatment with AL8810 enhanced the HCl-induced expression of pro-inflammatory cytokines and neutrophil migratory factors in the lungs. Administration of HCl decreased the gene expression of lung surfactant proteins, which was further reduced by co-administration of AL8810. Administration of AL8810 also increased lung edema and reduced mRNA expression of epithelial sodium channel in the lungs, indicating that AL8810 reduced fluid clearance. Furthermore, AL8810 also increased lipopolysaccharide-induced expression of adhesion molecules such as intracellular adhesion molecule-1 and E-selectin in human umbilical vein endothelial cells. These results indicate that inhibition of FP receptors by AL8810 exacerbated HCl-induced ALI.     

Short-Term Blockade of Pro-Inflammatory Alarmin S100A9 Favorably Modulates Left Ventricle Proteome and Related Signaling Pathways Involved in Post-Myocardial Infarction Recovery

Prognosis after myocardial infarction (MI) varies greatly depending on the extent of damaged area and the management of biological processes during recovery. Reportedly, the inhibition of the pro-inflammatory S100A9 reduces myocardial damage after MI. We hypothesize that a S100A9 blockade induces changes of major signaling pathways implicated in post-MI healing. Mass spectrometry-based proteomics and gene analyses of infarcted mice left ventricle were performed. The S100A9 blocker (ABR-23890) was given for 3 days after coronary ligation. At 3 and 7 days post-MI, ventricle samples were analyzed versus control and Sham-operated mice. Blockade of S100A9 modulated the expressed proteins involved in five biological processes: leukocyte cell–cell adhesion, regulation of the muscle cell apoptotic process, regulation of the intrinsic apoptotic signaling pathway, sarcomere organization and cardiac muscle hypertrophy. The blocker induced regulation of 36 proteins interacting with or targeted by the cellular tumor antigen p53, prevented myocardial compensatory hypertrophy, and reduced cardiac markers of post-ischemic stress. The blockade effect was prominent at day 7 post-MI when the quantitative features of the ventricle proteome were closer to controls. Blockade of S100A9 restores key biological processes altered post-MI. These processes could be valuable new pharmacological targets for the treatment of ischemic heart. Mass spectrometry data are available via ProteomeXchange with identifier PXD033683.     

The Therapeutic Treatment with the GAG-Binding Chemokine Fragment CXCL9(74–103) Attenuates Neutrophilic Inflammation and Lung Dysfunction during Klebsiella pneumoniae Infection in Mice

Klebsiella pneumoniae is an important pathogen associated with hospital-acquired pneumonia (HAP). Bacterial pneumonia is characterized by a harmful inflammatory response with a massive influx of neutrophils, production of cytokines and chemokines, and consequent tissue damage and dysfunction. Targeted therapies to block neutrophil migration to avoid tissue damage while keeping the antimicrobial properties of tissue remains a challenge in the field. Here we tested the effect of the anti-inflammatory properties of the chemokine fragment CXCL9(74–103) in pneumonia induced by Klebsiella pneumoniae in mice. Mice were infected by intratracheal injection of Klebsiella pneumoniae and 6 h after infection were treated systemically with CXCL9(74–103). The recruitment of leukocytes, levels of cytokines and chemokines, colony-forming units (CFU), and lung function were evaluated. The treatment with CXCL9(74–103) decreased neutrophil migration to the airways and the production of the cytokine interleukin-1β (IL-1β) without affecting bacterial control. In addition, the therapeutic treatment improved lung function in infected mice. Our results indicated that the treatment with CXCL9(74–103) reduced inflammation and improved lung function in Klebsiella pneumoniae-induced pneumonia.     

Platelet Inhibition Prevents NLRP3 Inflammasome Activation and Sepsis-Induced Kidney Injury

Platelets, cellular mediators of thrombosis, are activated during sepsis and are increasingly recognized as mediators of the immune response. Platelet activation is significantly increased in sepsis patients compared to ICU control patients. Despite this correlation, the role of activated platelets in contributing to sepsis pathophysiology remains unclear. We previously demonstrated NOD-like receptor protein 3 inflammasome (NLRP3) inflammasome activation in sepsis-induced platelets from cecal-ligation puncture (CLP) rats. Activated platelets were associated with increased pulmonary edema and glomerular injury in CLP vs. SHAM controls. In this study, we investigated whether inhibition of platelet activation would attenuate NLRP3 activation and renal and pulmonary injury in response to CLP. CLP was performed in male and female Sprague Dawley (SD) rats (n = 10/group) to induce abdominal sepsis and SHAM rats served as controls. A subset of CLP animals was treated with Clopidogrel (10 mg/kg/day, CLP + CLOP) to inhibit platelet activation. At 72 h post-CLP, platelet activation and NLRP3 inflammasome assembly were evaluated, IL-1β and IL-18 were measured in plasma, and tissues, renal and pulmonary pathology, and renal function were assessed. Activated platelets were 7.8 ± 3.6% in Sham, 22 ± 6% in CLP and significantly decreased to 14.5 ± 0.6% in CLP + CLOP (n = 8–10/group, p < 0.05). NLRP3 inflammasome assembly was inhibited in platelets of CLP + CLOP animals vs. CLP. Significant increases in plasma and kidney IL-1β and IL-18 in response to CLP were decreased with Clopidogrel treatment. Renal injury, but not lung histology or renal function was improved in CLP + CLOP vs. CLP. These data provide evidence that activated platelets may contribute to sepsis-induced renal injury, possibly via NLRP3 activation in platelets. Platelets may be a therapeutic target to decrease renal injury in septic patients.     

Anti-Septic Functions of Cornuside against HMGB1-Mediated Severe Inflammatory Responses

High mobility group box 1 (HMGB1) is acknowledged to have critical functions; therefore, targeting this protein may have therapeutic effects. One example is potential antiseptic activity obtained by suppressing HMGB1 secretion, leading to the recovery of vascular barrier integrity. Cornuside (CN), which is a product extracted from the fruit of Cornus officinalis Seib, is a natural bis-iridoid glycoside with the therapeutic effects of suppressing inflammation and regulating immune responses. However, the mechanism of action of CN and impact on sepsis is still unclear. We examined if CN could suppress HMGB1-induced excessive permeability and if the reduction of HMGB1 in response to LPS treatment increased the survival rate in a mouse model of sepsis. In human endothelial cells stimulated by LPS and mice with septic symptoms of cecal ligation and puncture (CLP), we examined levels of proinflammatory proteins and biomarkers as an index of tissue damage, along with decreased vascular permeability. In both LPS-treated human umbilical vein endothelial cells (HUVECs) and the CLP-treated mouse model of sepsis, we applied CN after the induction processes were over. CN suppressed excessive permeability and inhibited HMGB1 release, leading to the amelioration of vascular instability, reduced mortality, and improved histological conditions in the CLP-induced septic mouse model. Overall, we conclude that the suppressed release of HMGB1 and the increased survival rate of mice with CLP-induced sepsis caused by CN may be an effective pharmaceutical treatment for sepsis.     

RGS5 Determines Neutrophil Migration in the Acute Inflammatory Phase of Bleomycin-Induced Lung Injury

The regulator of G protein signaling (RGS) represents a widespread system of controllers of cellular responses. The activities of the R4 subfamily of RGSs have been elucidated in allergic pulmonary diseases. However, the R4 signaling in other inflammatory lung diseases, with a strong cellular immune response, remained unexplored. Thus, our study aimed to discern the functional relevance of the R4 family member, RGS5, as a potential modulating element in this context. Gene profiling of the R4 subfamily showed increased RGS5 expression in human fibrosing lung disease samples. In line with this, RGS5 was markedly increased in murine lungs following bleomycin injury. RGS knock-out mice (RGS-/-) had preserved lung function while control mice showed significant combined ventilatory disorders three days after bleomycin application as compared to untreated control mice. Loss of RGS5 was associated with a significantly reduced neutrophil influx and tissue myeloperoxidase expression. In the LPS lung injury model, RGS5-/- mice also failed to recruit neutrophils into the lung, which was accompanied by reduced tissue myeloperoxidase levels after 24 h. Our in-vitro assays showed impaired migration of RGS5-/- neutrophils towards chemokines despite preserved Ca²⁺ signaling. ERK dephosphorylation might play a role in reduced neutrophil migration in our model. As a conclusion, loss of RGS5 preserves lung function and attenuates hyperinflammation in the acute phase of bleomycin-induced pulmonary fibrosis and LPS-induced lung injury. Targeting RGS5 might alleviate the severity of exacerbations in interstitial lung diseases.     

Sinomenine Hydrochloride Protects against Polymicrobial Sepsis via Autophagy

Sepsis, a systemic inflammatory response to infection, is the major cause of death in intensive care units (ICUs). The mortality rate of sepsis remains high even though the treatment and understanding of sepsis both continue to improve. Sinomenine (SIN) is a natural alkaloid extracted from Chinese medicinal plant Sinomenium acutum, and its hydrochloride salt (Sinomenine hydrochloride, SIN-HCl) is widely used to treat rheumatoid arthritis (RA). However, its role in sepsis remains unclear. In the present study, we investigated the role of SIN-HCl in sepsis induced by cecal ligation and puncture (CLP) in BALB/c mice and the corresponding mechanism. SIN-HCl treatment improved the survival of BALB/c mice that were subjected to CLP and reduced multiple organ dysfunction and the release of systemic inflammatory mediators. Autophagy activities were examined using Western blotting. The results showed that CLP-induced autophagy was elevated, and SIN-HCl treatment further strengthened the autophagy activity. Autophagy blocker 3-methyladenine (3-MA) was used to investigate the mechanism of SIN-HCl in vitro. Autophagy activities were determined by examining the autophagosome formation, which was shown as microtubule-associated protein light chain 3 (LC3) puncta with green immunofluorescence. SIN-HCl reduced lipopolysaccharide (LPS)-induced inflammatory cytokine release and increased autophagy in peritoneal macrophages (PM). 3-MA significantly decreased autophagosome formation induced by LPS and SIN-HCl. The decrease of inflammatory cytokines caused by SIN-HCl was partially aggravated by 3-MA treatment. Taken together, our results indicated that SIN-HCl could improve survival, reduce organ damage, and attenuate the release of inflammatory cytokines induced by CLP, at least in part through regulating autophagy activities.     

姜黄素对脓毒症小鼠急性肺损伤的保护作用及对细胞间黏附分子-1和肿瘤坏死因子-α表达的影响

目的 研究姜黄素对脓毒症小鼠急性肺损伤(Acute lung injury,ALI)的保护作用及对细胞间黏附分子-1(Intercel-lular adhesion molecule-1,ICAM-1)和肿瘤坏死因子-α(Tumor necrosis factor-α,TNF-α)表达的影响。方法将SD小鼠随机分为假手术组(Sham组)、脓毒症组(Sep组)、二甲基亚砜组(DMSO组)和姜黄素组(Cur组)。采用盲肠结扎穿刺术(Cecal ligation andpuncture,CLP)复制脓毒症相关性ALI模型,造模24 h后,Cur组给予200 mg/(kg.d)姜黄素,Sham和Sep组给予等量生理盐水,DMSO组给予等量DMSO,均经腹腔注射给药。HE染色观察小鼠肺组织病理形态学变化;ELISA法检测小鼠血浆中ICAM-1和TNF-α含量的变化;Western blot分析小鼠肺组织中ICAM-1和TNF-α蛋白的表达。结果 Cur组小鼠在给药后12 h肺组织病理变化与Sep组相比有所减轻,48 h姜黄素作用达最强,且各种病理改变明显减轻,部分肺组织已恢复到正常形态;Cur组小鼠血浆中lCAM-1的含量在给药后6、12、24和48 h均明显低于Sep组(P<0.05),Cur组小鼠血浆中TNF-α的含量在给药后24 h明显低于Sep组(P<0.05);给药后24 h,Cur组小鼠肺组织中ICAM-1和TNF-α蛋白的表达水平与Sep组相比明显降低(P<0.05);DMSO组与Sep组各项指标差异均无统计学意义(P>0.05)。结论姜黄素能够有效减轻脓毒症所致ALI,这一作用与抑制ICAM-1和TNF-α的过度表达有关。     

FoxO3a基因缺失致小鼠脾脏进行性慢性炎性反应的观察

目的观察FoxO3a基因缺失致小鼠脾脏进行性慢性炎性反应。方法分别于16周、24周和12个月称取FoxO3a基因敲除小鼠和对照小鼠体重;于16、24和38周处死各组小鼠,取脾脏,肉眼观察脾脏变化,并制备脾细胞悬液,进行细胞计数,同时取各组小鼠卵巢、肺及皮下等组织,观察其炎性变化。流式细胞术检测24周的FoxO3a基因敲除小鼠和对照小鼠脾细胞中T、B淋巴细胞和Mac-1+细胞数量及百分率;Real-time PCR检测脾细胞中炎性细胞因子S100A8和S100A9基因mRNA的表达水平。结果与对照小鼠相比,FoxO3a基因敲除小鼠脾脏明显肥大,且随年龄增加更加显著,卵巢、肺及皮下等组织内可见大量炎性细胞浸润,脾细胞数明显升高;基因敲除组老年小鼠的体重明显低于对照小鼠;FoxO3a基因敲除小鼠的脾细胞中T、B淋巴细胞数增加明显,但所占脾细胞的百分率无明显增加,Mac-1+细胞数和百分率均明显增加;S100A8和S100A9基因mRNA的表达水平明显高于对照小鼠。结论 FoxO3a基因缺失致小鼠脾脏进行性慢性炎性反应。     

The Emerging Role of Neutrophils in the Pathogenesis of Thrombosis in COVID-19

Previous studies have shown that COVID-19 leads to thrombotic complications, which have been associated with high morbidity and mortality rates. Neutrophils are the largest population of white blood cells and play a pivotal role in innate immunity. During an infection, neutrophils migrate from circulation to the infection site, contributing to killing pathogens. This mechanism is regulated by chemokines such as IL-8. Moreover, it was shown that neutrophils play an important role in thromboinflammation. Through a diverse repertoire of mechanisms, neutrophils, apart from directly killing pathogens, are able to activate the formation of thrombi. In COVID-19 patients, neutrophil activation promotes neutrophil extracellular trap (NET) formation, platelet aggregation, and cell damage. Furthermore, neutrophils participate in the pathogenesis of endothelitis. Overall, this review summarizes recent progress in research on the pathogenesis of COVID-19, highlighting the role of the prothrombotic action of neutrophils in NET formation.     

The PDE4 Inhibitor Tanimilast Restrains the Tissue-Damaging Properties of Human Neutrophils

Neutrophils, the most abundant subset of leukocytes in the blood, play a pivotal role in host response against invading pathogens. However, in respiratory diseases, excessive infiltration and activation of neutrophils can lead to tissue damage. Tanimilast-international non-proprietary name of CHF6001—is a novel inhaled phosphodiesterase 4 (PDE4) inhibitor in advanced clinical development for the treatment of chronic obstructive pulmonary disease (COPD), a chronic inflammatory lung disease where neutrophilic inflammation plays a key pathological role. Human neutrophils from healthy donors were exposed to pro-inflammatory stimuli in the presence or absence of tanimilast and budesonide—a typical inhaled corticosteroid drug-to investigate the modulation of effector functions including adherence to endothelial cells, granule protein exocytosis, release of extracellular DNA traps, cytokine secretion, and cell survival. Tanimilast significantly decreased neutrophil-endothelium adhesion, degranulation, extracellular DNA traps casting, and cytokine secretion. In contrast, it promoted neutrophil survival by decreasing both spontaneous apoptosis and cell death in the presence of pro-survival factors. The present work suggests that tanimilast can alleviate the severe tissue damage caused by massive recruitment and activation of neutrophils in inflammatory diseases such as COPD.     

Usefulness of Intratracheal Instillation Studies for Estimating Nanoparticle-Induced Pulmonary Toxicity

Inhalation studies are the gold standard for the estimation of the harmful effects of respirable chemical substances, while there is limited evidence of the harmful effects of chemical substances by intratracheal instillation. We reviewed the effectiveness of intratracheal instillation studies for estimating the hazards of nanoparticles, mainly using papers in which both inhalation and intratracheal instillation studies were performed using the same nanoparticles. Compared to inhalation studies, there is a tendency in intratracheal instillation studies that pulmonary inflammation lasted longer in the lungs. A difference in pulmonary inflammation between high and low toxicity nanoparticles was observed in the intratracheal instillation studies, as in the inhalation studies. Among the endpoints of pulmonary toxicity, the kinetics of neutrophil counts, percentage of neutrophils, and chemokines for neutrophils and macrophages, heme oxygenase-1 (HO-1) in bronchoalveolar lavage fluid (BALF), reflected pulmonary inflammation, suggesting that these markers may be considered the predictive markers of pulmonary toxicity in both types of study. When comparing pulmonary inflammation between intratracheal instillation and inhalation studies under the same initial lung burden, there is a tendency that the inflammatory response following the intratracheal instillation of nanoparticles is greater than or equal to that following the inhalation of nanoparticles. If the difference in clearance in both studies is not large, the estimations of pulmonary toxicity are close. We suggest that intratracheal instillation studies can be useful for ranking the hazard of nanoparticles through pulmonary inflammation.     

Induced Pluripotent Stem Cell-Derived Conditioned Medium Promotes Endogenous Leukemia Inhibitory Factor to Attenuate Endotoxin-Induced Acute Lung Injury

The conditioned medium of induced pluripotent stem cells (iPSC-CM) can attenuate neutrophil recruitment and endothelial leakage of lipopolysaccharide (LPS)-induced acute lung injury (ALI). Therefore, we investigated the mechanisms by which iPSC-CM regulate the interaction between neutrophils and the endothelium in ALI. Murine iPSCs (miPSCs) were delivered intravenously to male C57BL/6 mice (8–12 weeks old) 4 h after intratracheal LPS injection. A miPSC-derived conditioned medium (miPSC-CM) was delivered intravenously to mice after intratracheal LPS injection. DMSO-induced HL-60 cells (D-HL-60, neutrophil-like cells) and human umbilical vein endothelial cells (HUVECs) were used as in vitro models to assess the interaction of neutrophils and endothelial cells. miPSC-CM diminished the histopathological changes in the lungs and the neutrophil count in bronchoalveolar lavage fluids of ALI mice. miPSC-CM attenuated the expression of adhesion molecules in the lungs of ALI mice. Human iPSC conditioned medium (hiPSC-CM) reduced the expression of adhesion molecules in a HUVEC and D-HL-60 co-culture after LPS stimulation, which decreased the transendothelial migration (TEM) of D-HL-60. A human angiogenesis factors protein array revealed that leukemia inhibitory factor (LIF) was not detected in the absence of D-HL-60 and hiPSC-CM groups. hiPSC-CM significantly promoted the production of endogenous LIF in in vitro models. Administration of an anti-LIF antibody not only reversed the effect of iPSC-CM in ALI mice, but also blocked the effect of iPSC-CM on neutrophils TEM in in vitro models. However, a controlled IgG had no such effect. Our study demonstrated that iPSC-CM promoted endogenous LIF to inhibit neutrophils TEM and attenuate the severity of sepsis-induced ALI.     

Therapeutic CFTR Correction Normalizes Systemic and Lung-Specific S1P Level Alterations Associated with Heart Failure

Heart failure (HF) is among the main causes of death worldwide. Alterations of sphingosine-1-phosphate (S1P) signaling have been linked to HF as well as to target organ damage that is often associated with HF. S1P’s availability is controlled by the cystic fibrosis transmembrane regulator (CFTR), which acts as a critical bottleneck for intracellular S1P degradation. HF induces CFTR downregulation in cells, tissues and organs, including the lung. Whether CFTR alterations during HF also affect systemic and tissue-specific S1P concentrations has not been investigated. Here, we set out to study the relationship between S1P and CFTR expression in the HF lung. Mice with HF, induced by myocardial infarction, were treated with the CFTR corrector compound C18 starting ten weeks post-myocardial infarction for two consecutive weeks. CFTR expression, S1P concentrations, and immune cell frequencies were determined in vehicle- and C18-treated HF mice and sham controls using Western blotting, flow cytometry, mass spectrometry, and qPCR. HF led to decreased pulmonary CFTR expression, which was accompanied by elevated S1P concentrations and a pro-inflammatory state in the lungs. Systemically, HF associated with higher S1P plasma levels compared to sham-operated controls and presented with higher S1P receptor 1-positive immune cells in the spleen. CFTR correction with C18 attenuated the HF-associated alterations in pulmonary CFTR expression and, hence, led to lower pulmonary S1P levels, which was accompanied by reduced lung inflammation. Collectively, these data suggest an important role for the CFTR-S1P axis in HF-mediated systemic and pulmonary inflammation.     

n-3 Polyunsaturated Fatty Acids Improve Inflammation via Inhibiting Sphingosine Kinase 1 in a Rat Model of Parenteral Nutrition and CLP-Induced Sepsis

The sphingosine kinase 1 (SphK1)/sphingosine‐1‐phosphate (S1P) pathway plays a key role in inflammation. Parenteral nutrition containing n‐3 polyunsaturated fatty acids (n‐3 PUFA) may regulate inflammatory reactions. The aim of this study is to determine whether n‐3 PUFA may improve inflammatory responses by neutralizing SphK1 signaling. Rat models of parenteral nutrition, cecal ligation and puncture (CLP)‐induced sepsis were generated. Male Sprague–Dawley rats were operated for CLP on day 2 after venous catheterization. The rats were randomized to receive normal saline (NS; n = 20), parenteral nutrition (PN; n = 20), or PN + fish oil (FO; n = 20) for 5 days. The daily intake of fish oil (1.25–2.82 g EPA and 1.44–3.09 g DHA per 100 ml) in the FO group was approximately 1.8 g/kg body weight/day. Rats in the control group (n = 10) were subjected to sham operation and received a chow diet. Spleen tissues were collected for SphK1 and S1P receptor expression analysis. Our data showed that n‐3 PUFA ameliorated the survival rate. SphK1 expression and its enzymatic activity were significantly upregulated in sepsis rats. Furthermore, mRNA and protein levels of S1PR3, but not S1PR1, were also facilitated after CLP. However, PN + FO dramatically decreased SphK1 mRNA level and its enzymatic activity. S1PR3 expression was also attenuated by FO addition. In conclusion, the anti‐inflammatory effect of n‐3 PUFA may be linked to the inhibition of the SphK1/S1P pathway in a rat model of parenteral nutrition and CLP‐induced sepsis.