Lung Wet-dry Weight Research Articles

Anti-inflammatory effect and pharmacokinetics of dehydroandrographolide, an active component of Andrographis paniculata, on Poly(I:C)-induced acute lung injury

Acute lung injury (ALI) is a common and critical respiratory disorder caused by various factors, with viral infection being the leading contributor. Dehydroandrographolide (DAP), a constituent of the Chinese herbal plant Andrographis paniculata, exhibits a range of activities including anti-inflammatory, in vitro antiviral and immune-enhancing effects. This study evaluated the anti-inflammatory effects and pharmacokinetics (PK) profile of DAP in ALI mice induced by intratracheal instillation of Poly(I:C) (PIC). The results showed that oral administration of DAP (10–40 mg/kg) effectively suppressed the increase in lung wet–dry weight ratio, total cells, total protein content, accumulation of immune cells, inflammatory cytokines and neutrophil elastase levels in bronchoalveolar lavage fluid of PIC-treated mice. DAP concentrations, determined by an LC-MS/MS method, in plasma after receiving DAP (20 mg/kg) were unchanged compared to those in normal mice. However, DAP concentrations and relative PK parameters in the lungs were significantly altered in PIC-treated mice, exhibiting a relatively higher maximum concentration, larger AUC, and longer elimination half-life than those in the lungs of normal mice. These results demonstrated that DAP could improve lung edema and inflammation in ALI mice, and suggested that lung injury might influence the PK properties of DAP, leading to increased lung distribution and residence. Our study provides evidence that DAP displays significant anti-inflammatory activity against viral lung injury and is more likely to distribute to damaged lung tissue.

Inhibition of S100A9 alleviates neurogenic pulmonary edema after subarachnoid hemorrhage

Background and PurposeNeurogenic pulmonary edema (NPE) frequently arises as a complication subsequent to subarachnoid hemorrhage (SAH). Heterodimers of S100A8 and S100A9 are commonly formed, thereby initiating an inflammatory reaction through receptor binding on the cell surface. Paquinimod serves as a specific inhibitor of S100A9. The objective of this investigation is to assess the impact of Paquinimod administration and S100A9 knockout on NPE following SAH. MethodsIn this study, SAH models of C57BL/6J wild-type (WT) and S100A9 knockout mice were established through intravascular perforation. These models were then divided into several groups, including the WT-sham group, S100A9-KO-sham group, WT-SAH group, WT-SAH + Paquinimod group, and S100A9-KO-SAH group. After 24 h of SAH induction, pulmonary edema was assessed using the lung wet-dry weight method and Hematoxylin and eosin (HE) staining. Additionally, the expression levels of various proteins, such as interleukin-1β (IL-1β), tumor necrosis factor α (TNF-α), occludin, claudin-3, Bax, Bcl-2, TLR4, MYD88, and pNF-κB, in lung tissue were analyzed using western blot and immunofluorescence staining. Lung tissue apoptosis was detected by TUNEL staining. ResultsFirstly, our findings indicate that the knockout of S100A9 has a protective effect on early brain injury following subarachnoid hemorrhage (SAH). Additionally, the reduction of brain injury after SAH can also alleviate neurogenic pulmonary edema (NPE). Immunofluorescence staining and western blot analysis revealed that compared to SAH mice with wild-type S100A9 expression (WT-SAH), the lungs of S100A9 knockout SAH mice (S100A9-KO-SAH) and mice treated with Paquinimod exhibited decreased levels of inflammatory molecules (IL-1β and TNF-α) and increased levels of tight junction proteins. Furthermore, the knockout of S100A9 resulted in upregulated expression of the apoptotic-associated protein Bax and down-regulated expression of Bcl-2. Furthermore, a decrease in TLR4, MYD88, and phosphorylated pNF-κB was noted in S100A9-KO-SAH and Paquinimod treated mice, indicating the potential involvement of the TLR4/MYD88/NF-κB signaling pathway in the inhibition of the protective effect of S100A9 on NPE following SAH. ConclusionThe knockout of S100A9 not only ameliorated initial cerebral injury following subarachnoid hemorrhage (SAH), but also mitigated SAH-associated neurogenic pulmonary edema (NPE). Additionally, Paquinimod was found to diminish NPE. These findings imply a correlation between the central nervous system and peripheral organs, highlighting the potential of safeguarding the brain to mitigate harm to peripheral organs.

Aspirin eugenol ester alleviates lipopolysaccharide-induced acute lung injury in rats while stabilizing serum metabolites levels.

Aspirin eugenol ester (AEE) was a novel drug compound with aspirin and eugenol esterified. AEE had various pharmacological activities, such as anti-inflammatory, antipyretic, analgesic, anti-oxidative stress and so on. In this study, it was aimed to investigate the effect of AEE on the acute lung injury (ALI) induced by lipopolysaccharide (LPS) in rats. In vitro experiments evaluated the protective effect of AEE on the LPS-induced A549 cells. The tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β) were measured in the cell supernatant. The Wistar rats were randomly divided into five groups (n = 8): control group, model group (LPS group), LPS + AEE group (AEE, 54 mg·kg−1), LPS + AEE group (AEE, 108 mg·kg−1), LPS + AEE group (AEE, 216 mg·kg−1). The lung wet-to-dry weight (W/D) ratio and immune organ index were calculated. WBCs were counted in bronchoalveolar lavage fluid (BALF) and total protein concentration was measured. Hematoxylin-Eosin (HE) staining of lung tissue was performed. Glutathione (GSH), glutathione peroxidase (GPx), catalase (CAT), antioxidant superoxide dismutase (SOD), total antioxidant capacity (T-AOC), lactate dehydrogenase (LDH), C-reactive protein (CRP), myeloperoxidase (MPO), malondialdehyde (MDA), macrophage mobility inhibitory factor (MIF), TNF-α, IL-6, and IL-1β activity were measured. The metabolomic analysis of rat serum was performed by UPLC-QTOF-MS/MS. From the results, compared with LPS group, AEE improved histopathological changes, reduced MDA, CRP, MPO, MDA, and MIF production, decreased WBC count and total protein content in BALF, pro-inflammatory cytokine levels, immune organ index and lung wet-dry weight (W/D), increased antioxidant enzyme activity, in a dose-dependent manner. The results of serum metabolomic analysis showed that the LPS-induced ALI caused metabolic disorders and oxidative stress in rats, while AEE could ameliorate it to some extent. Therefore, AEE could alleviate LPS-induced ALI in rats by regulating abnormal inflammatory responses, slowing down oxidative stress, and modulating energy metabolism.

Mechanism of curcumin inhibiting pyroptosis in infectious acute lung injury through NLRP3 inflammatory pathway

Purpose of this study the mechanism of curcumin inhibiting pyroptosis in infectious acute lung injury through NLRP3 inflammatory pathway. In this study, SPF SD male rats were randomly divided into blank control group, curcumin group, lipopolysaccharide (LPS) group, LPS + curcumin group, LPS + Nigerin treatment group, LPS + Nigerin + curcumin treatment group. Three in each group. Animal models were established according to the experimental design. Blood samples were collected after anesthesia, and the lung wet/dry weight ratio was calculated. TUNEL was used to detect pyroptosis of alveolar macrophages in left lower lobe, and Elisa was used to detect the expression levels of inflammatory factors IL-1β and IL-18 in left lung tissue. Western Blot was used to detect the expression of pyroptosis-related proteins such as NLRP3, Caspase1 and GSDMD in right lung alveolar lavage fluid. Compared with the blank control group, the lung wet weight and lung dry weight of the curcumin group were significantly increased (P < 0.01, P < 0.05). Compared with the LPS group, the lung wet weight and lung dry weight of the curcumin group There was a statistical difference in weight increase (P < 0.01). Compared with the blank control group, the apoptosis rate of lung tissue in the curcumin group, LPS group and LPS+ nigericin group was significantly increased (P < 0.01). Compared with the LPS + nigericin group, the lung tissue apoptosis rate in the LPS + curcumin + nigericin group was decreased, but there was no statistical difference (P > 0.05). LPS and LPS combined with Nigerian bacteriocin can cause pyroptosis in lung tissue. Curcumin can inhibit pyroptosis of lung tissue macrophages caused by LPS and LPS combined with Nigerian bacteriocin, and reduce inflammatory response.

Pro-Resolving Mediator Resolvin E1 Restores Alveolar Fluid Clearance in Acute Respiratory Distress Syndrome.

Acute respiratory distress syndrome (ARDS) is a life-threatening condition characterized by increased permeability of the alveolar-capillary barrier and impaired alveolar fluid clearance. Resolvin E1 (RvE1) is a specialized pro-resolving mediator derived endogenously from omega-3-polyunsaturated fatty acids. RvE1 (10 μg/kg i.v.) was injected to rats 6 h post-lipopolysaccharide (LPS) (14 mg/kg) induction. After another 3 h, alveolar fluid clearance was measured in live rats (n = 8-9). The primary Type II alveolar epithelial cell was isolated and treated by LPS (1 μg/mL) with or without RvE1 (250 nM). The expression of epithelial sodium channel (ENaC), Na+/K+-ATPase (NKA), AKT, serum- and glucocorticoid-induced kinase 1 (SGK1), and Nedd4-2 were detected. RvE1 improved survival rate (30% vs. 70%, P = 0.048), increased the clearance of alveolar fluid (13.34% vs. 18.73%, P < 0.001), reduced lung wet-dry weight ratio (5.01 vs. 4.63, P < 0.001), mitigated lung injury scores (13.38 vs. 7.0, P < 0.05) and inflammation in LPS-induced ARDS in rats. RvE1 upregulated alveolar ENaC and NKA expression in vivo and in vitro. In addition, RvE1 significantly increased the expression of phosphorylated AKT, SGK1, and phosphorylated Nedd4-2 in LPS-stimulated primary alveolar type II cells. The effects of RvE1 were abrogated by blocking phosphatidylinositide3'-kinase (PI3K) and SGK1 with LY294002 and GSK650394, respectively. In summary, RvE1 upregulated ENaC and NKA expression by activating PI3K/AKT/SGK1 pathway to promote alveolar fluid clearance, suggesting that RvE1 may be a potentially effective drug for ARDS treatment.

MEF2C alleviates acute lung injury in cecal ligation and puncture (CLP)-induced sepsis rats by up-regulating AQP1.

Sepsis is a systemic inflammatory response syndrome and leads to patient's death. Objective: To investigate the effect of myocyte enhancer factor 2 (MEF2C) on acute lung injury (ALI) with sepsis and its possible mechanism. The cecal ligation and puncture (CLP)-induced sepsis rat model was established. The lung injury was determined by lung wet-dry weight ratio, the concentration of inflammatory cytokines, including tumor necrosis factor-α (TNF-α), Interlukin (IL)-6, IL-1β, and IL-10, were measured by the enzyme-linked-immunosorbent serologic assay kit. The cell apoptosis was detected by TUNEL staining assay. Interestingly, MEF2C was down-regulated in this model. Moreover, adeno-associated virus (AAV)-MEF2C treatment markedly suppressed TNF-α, IL-1β, and IL-6 concentrations but promoted IL-10 concentration in serum in CLP-challenged rats. Besides, overexpression of MEF2C alleviates CLP-induced lung injury. Interestingly, AAV-MEF2C treatment was confirmed to suppress apoptosis in CLP-induced sepsis rats as well as promote aquaporin APQ1 expression. Mechanistically, the rescue experiments indicated that MEF2C alleviated CLP-induced lung inflammatory response and apoptosis via up-regulating AQP1. In summary, overexpression of MEF2C suppressed CLP-induced lung inflamma-tory response and apoptosis via up-regulating AQP1, providing a novel therapeutic target for sepsis-induced ALI.

Diammonium glycyrrhizinate lipid ligand ameliorates lipopolysaccharide-induced acute lung injury by modulating vascular endothelial barrier function.

The aim of the present study was to investigate the effects of diammonium glycyrrhizinate lipid ligand (DGLL) treatment on acute lung injury (ALI) and pulmonary edema induced by lipopolysaccharide (LPS) in Sprague-Dawley rats. Rats orally received 30, 60 and 120 mg/kg DGLL. After 1 h, the rat ALI model was established by LPS (10 mg/kg) intraperitoneal injection. After 6 h, lung injury was evaluated using hematoxylin and eosin staining techniques. Pulmonary edema was evaluated using lung wet-dry weight ratio, protein concentrations in the bronchoalveolar lavage fluid (BALF) and Evans blue (EB) extravasation in lung tissue. The expression levels of tumor necrosis factor (TNF)-α and interleukin (IL)-1β in lung tissues were measured using ELISA. Myeloperoxidase (MPO) expression levels were detected by immunohistochemical staining. Western blotting was used to measure the expression level changes of intercellular adhesion molecule (ICAM)-1, as well as adherent and tight junction proteins, including vascular endothelial (VE)-cadherin, zonula occludens (ZO)-1, occludin and junctional adhesion molecule (JAM)-1 that were associated with pulmonary inflammation and microvascular permeability. DGLL treatment significantly alleviated ALI induced by LPS, which was demonstrated by reduction of MPO-positive cells and expression levels of TNF-α, IL-1β and ICAM-1 in rat lung tissues. In addition, DGLL abrogated LPS-induced pulmonary edema, decreased the protein concentration in BALF and reduced EB extravasation. DGLL also reversed the reduced expression of VE-cadherin and tight junction proteins, including ZO-1, occludin and JAM-1 in the lung tissues caused by LPS. In conclusion, DGLL exhibits a protective effect on LPS-induced rat ALI, which is associated with the inhibition of inflammatory cell infiltration and microvascular barrier disruption. The present results provide a theoretical basis for the application of DGLL for the potential clinical treatment of ALI.

Partial liquid ventilation–induced mild hypothermia improves the lung function and alleviates the inflammatory response during acute respiratory distress syndrome in canines

Background Acute respiratory distress syndrome (ARDS), which is the severest form of pulmonary injury, is the leading cause of death in critical care. At present, the mortality remains high in ARDS. Partial liquid ventilation (PLV) using perfluorocarbon (PFC) has been proven to improve gas exchange and respiratory dynamics of the lungs during ARDS. However, PLV has not been shown to reduce the mortality of ARDS. Some studies have shown that mild hypothermia therapy can reduce lung injuries in animal models of ARDS by reducing inflammatory cytokine levels in lung tissues. However, hypothermia cannot produce a lung protection effect alone, and it may have a synergistic effect with other protective measures. To explore the possible role of PLV combined with mild hypothermia in the treatment of ARDS, in this study, we used PFC liquid ventilation to induce mild hypothermia in dogs suffering from ARDS and analyzed the effects of PFC liquid ventilation-induced mild hypothermia on the levels of inflammatory factors and lung histopathology in dogs with ARDS. The experimental dogs were randomly divided into conventional mechanical ventilation (CMV), normal temperature PFC liquid ventilation (NPLV), hypothermic PFC liquid ventilation (HPLV), and mechanical ventilation (MV) groups. After induction of ARDS, the CMV group was treated with CMV for respiratory support, the HPLV group was treated with PLV-induced mild hypothermia using 15 °C PFC and maintained the rectal temperature at 34–36 °C, the NPLV group was treated with PLV using 36 °C PFC and maintained the rectal temperature at 36–38 °C. The MV group served as the control group. Analyses of the pulmonary pathology, partial pressure of oxygen in the blood, and lung wet-dry weight ratio (W/T) of each dog revealed that PLV-induced mild hypothermia significantly increased the PaO2 values and attenuated lung injury, and there were no adverse effects on hemodynamics. Furthermore, treatment with PLV-induced mild hypothermia significantly increased the expression of the anti-inflammatory factor IL-10 in bronchoalveolar lavage fluid (BALF) and attenuated the expression of interleukin (IL-6) and tumor necrosis factor-α (TNF-α) in peripheral blood and in lung BALF. Moreover, the results showed that the expression of myeloperoxidase (MPO) and NF-κB p65 in lung tissues was significantly decreased by PLV-induced mild hypothermia compared with NPLV and CMV. Our results indicated that PLV combined with mild hypothermia can provide protection against oleic acid-induced ARDS in dogs.

Effects and mechanism of fecal transplantation on acute lung injury induced by lipopolysaccharide in rats

Objective: To investigate the effect of fecal microbiota transplantation (FMT) on acute lung injury (ALI) induced by lipopolysaccharide (LPS) and its regulatory mechanism. Methods: Fifteen rats were divided into control group, LPS group and LPS+FMT group by random number table method. LPS group and LPS+FMT group were intraperitoneally injected with LPS to generate rat ALI model. After 24 h of modeling, feces (10 ml/kg) were given to the LPS+FMT group twice a day, and the control group and LPS group were given the same amount of normal saline. The intervention lasted for 2 days. After 24 h of the last fecal microbiota transplantation, arterial blood gas analysis was performed in each group. Then rats were sacrificed and enzyme-linked immunosorbent (ELISA) method was used to detect intercellular adhesion molecule 1 (ICAM-1) content in the serum and bronchoalveolar lavage fluid (BALF). The lung wet-dry weight ratio (W/D) was evaluated; HE staining and lung tissue pathology scoring, immunohistochemical detection of nuclear factor-kappa B (NF-κB) predominate nuclear expression and expression of ICAM-1 of alveolar epithelial cells were conducted; Western blot was used to detect the expression of proteins related to the intracellular phosphatidylinositol kinase (PI3K)/protein kinase (AKT) signaling pathway. Samples of rat feces were collected and DNA was extracted. Polymerase chain reaction (PCR) products of the V3 and V4 regions of the 16S ribosomal RNA gene (16SrDNA) were sequenced at high throughput, and bioinformatics analysis was conducted on the microbial community based on the operational classification unit. Results: The lung W/D and lung histopathological score of the LPS group were significantly higher than those of the control group, while the arterial partial oxygen pressure (PaO(2)) of the LPS group was significantly lower than that of the control group [(79.2±5.89 vs 95.2±2.77) mmHg, 1 mmHg=0.133 kPa](all P<0.05). The results of intestinal flora sequencing revealed that the diversity index of LPS group was significantly higher than that of the control group, while the lactobacillus of LPS group rats was significantly lower than that of the control group. The content of ICAM-1 in serum, BALF and its relative expression on the cell membrane in the LPS group was significantly higher than that in the control group [(8.64±0.87) vs (7.40±0.32) ng/L; (0.941±0.035) vs (0.739±0.079) ng/L; (0.250±0.010) vs (0.076±0.010)] (all P<0.05). Moreover, the relative expression levels of phosphorylated P65 (p-P65), p-PI3K and p-AKT nucleoprotein in the LPS group were significantly higher than those in the control group (4.89±0.27 vs 3.28±0.13, 0.265±0.030 vs 0.036±0.013 and 0.444±0.040 vs 0.109±0.016) (all P<0.05). The above injury effect was reduced after fecal fungus transplantation. The lung W/D and lung pathological score of LPS+FMT group were significantly lower than those of LPS group, and PaO(2) of LPS+FMT group was significantly higher than that of LPS group [(88.0±3.53) mmHg]. The results of intestinal flora sequencing revealed that the diversity index of LPS+FMT group was significantly lower than that of LPS group, and the lactobacillus genus of LPS+FMT group was significantly higher than that of LPS group. ICAM-1 in the blood serum ((7.44±0.46) ng/L), BALF (0.834±0.040) ng/L) and its relative expression on alveolar epithelial cell membrane (0.173±0.030), the relative expression of p-P65, p-PI3K and p-AKT protein of NF-κB in alveolar epithelial cells was down-regulated ((2.99±0.28, 0.090±0.013 and 0.206±0.018) in LPS+FMT group than those of LPS group, the differences were statistically significant (all P<0.05). Conclusion: Fecal transplantation can alleviate lipopolysaccharide-induced acute lung injury in rats, and its regulatory effect may be related to inhibiting the activation of PI3K/AKT/NF-κB signaling pathway and reducing the expression of inflammatory factor ICAM-1.

Protective Effect and Mechanism of Alprostadil in Acute Respiratory Distress Syndrome Induced by Oleic Acid in Rats.

BackgroundThis study investigated the role and mechanism of alprostadil in acute respiratory distress syndrome (ARDS) induced by oleic acid (OA) in rats.Material/MethodsSprague-Dawley rats were randomly divided into control, OA model, and OA + Alprostadil (2.5, 5, and 10 μg/kg, respectively) groups. The ARDS model was induced by femoral vein injection of OA, and alprostadil was administrated immediately. Lung injury was evaluated by lung wet-dry weight ratio (W/D) and histological analyses. Expressions of ACE, inflammatory mediators, apoptotic-related proteins, and proteins in the MAPKs and NF-κB signaling pathways were determined by Western blot or immunohistochemical staining.ResultsCompared with the control group, the OA model group had significantly increased W/D, lung injury score, and collagen deposition at 3 h after OA injection. However, alprostadil (10 μg/kg) treatment significantly reduced OA-induced elevation of these indicators. Additionally, OA-induced expression of TNF-α and IL-1β were suppressed by alprostadil. The OA-induced activation of nuclear factor (NF) κB p65 was also reduced by alprostadil. Furthermore, we found that Alprostadil had an inhibitory effect on the phosphorylation of JNK, ERK1/2, and p38 MAPKs. Alprostadil inhibited Bax but increased Bcl-2, indicating a suppressive role in apoptosis. Remarkably increased expression of ACE in the OA model group was observed, which was decreased by alprostadil.ConclusionsAlprostadil has a protective effect on ARDS induced by OA in rats, possibly through inhibiting apoptosis, suppressing the activation of MAPKs and NF-κB signaling pathways, and decreasing ACE protein expression. Therefore, the use of alprostadil in clinical ARDS treatment is promising.

Long non-coding RNA CASC2 improved acute lung injury by regulating miR-144-3p/AQP1 axis to reduce lung epithelial cell apoptosis

Background and objectiveApoptosis of lung epithelial cell is implicated in the pathogenesis of acute lung injury (ALI). To study the protective effect and mechanism of cancer susceptibility candidate 2 (CASC2) on reducing lung epithelial cell apoptosis after LPS inducing acute lung injury in mice.Methods and resultsThe ALI mice model was performed by intratracheally instilling with lipopolysaccharide (LPS). The CASC2 expression detected by quantitative real-time polymerase chain reaction was significantly decreased in LPS-induced A549 cell and ALI mice model. LPS induced A549 cell apoptosis, while transfection with pcDNA-CASC2 reversed the increased cell apoptosis, suggesting overexpression of CASC2 inhibited LPS-induced A549 cell apoptosis. In addition, we found that miR-144-3p expression were opposite to CASC2, while Aquaporin-1 (AQP1) expression was opposite to miR-144-3p in LPS-induced A549 cell and ALI mice model. The RNA immunoprecipitation and RNA pull-down assay demonstrated that CASC2 could function as a miR-144-3p decoy. The luciferase reporter assay revealed that AQP1 was a target of miR-144-3p in A549 cell. And then, further in vitro studied showed that CASC2 controlled AQP1 expression by regulating miR-144-3p, and LPS induced A549 cell apoptosis by regulating CASC2/miR-144-3p/AQP1 axis. At last, after injection with lentivirus-expressing CASC2 or control lentivirus, the mice were intratracheally instilled with LPS. Comparing to the mice injected with pcDNA, the mice injected with pcDNA-CASC2 had a significantly reduced lung wet–dry weight ratio.ConclusionsLong non-coding RNA CASC2 improved acute lung injury by regulating miR-144-3p/AQP1 axis to reduce lung epithelial cell apoptosis.

3-Methyladenine and dexmedetomidine reverse lipopolysaccharide-induced acute lung injury through the inhibition of inflammation and autophagy.

The aim of the present study was to investigate the effects of 3-methyladenine (3-MA) and dexmedetomidine (DEX) pretreatment on lipopolysaccharide (LPS)-induced acute lung injury (ALI) and the potential mechanism underlying the effects. LPS was instilled into the trachea of BALB/c mice to induce the ALI model. Solutions of 3-MA or DEX were intravenously injected into the mice 1 h later to establish the 3-MA and DEX groups. On days 1, 3 and 5 after the injections, arterial blood gas analysis was conducted, and the lung wet-dry weight ratio (W/D) was determined. In addition, albumin, cytokine and myeloperoxidase (MPO) contents were evaluated using ELISAs, and hematoxylin and eosin (H&E) staining was conducted. Furthermore, western blot analysis was used to evaluate the protein expression levels of microtubule-associated protein 1A/1B-light chain 3 (LC3)-I, LC3-II, autophagy protein 5 (ATG5), Rab7 and lysosome-associated membrane protein 1 (LAMP1), and reverse transcription quantitative polymerase chain reaction (RT-qPCR) was used to detect the mRNA expression levels of nuclear factor-κB (NF-κB) and Toll-like receptor 4 (TLR4). Treatment with 3-MA or DEX increased the blood partial pressure of oxygen level compared with that in the model group, and restored the W/D and blood partial pressure of carbon dioxide to normal levels. The content of tumor necrosis factor-α, interleukin-6 and albumin in bronchoalveolar fluid and MPO in lung tissue was significantly decreased in the 3-MA and DEX groups compared with the model group (P<0.05). H&E staining demonstrated that 3-MA and DEX each reversed the ALI. In addition, 3-MA and DEX reduced the protein expression levels of LC3-I, LC3-II, ATG5, Rab7 and LAMP1. Also, RT-qPCR results revealed that NF-κB and TLR4 mRNA expression levels were clearly decreased in the 3-MA and DEX groups compared with the model group. In conclusion, LPS-induced ALI was effectively reversed by treatment with 3-MA and DEX through the reduction of inflammation and autophagy and inhibition of the TLR4-NF-κB pathway.

Neutrophil elastase ameliorates matrix metalloproteinase-9 to promote lipopolysaccharide-induced acute lung injury in mice 1.

To investigate the regulatory roles of neutrophil elastase (NE) and matrix metalloproteinase-9 (MMP-9) in lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice. To construct LPS-induced ALI mouse models, wild-type C57BL/6 mice were administered 5.0 mg/kg of LPS through endotracheal, and/or 1.0 mg/kg of ONO-5046, and/or 20.0 mg/kg of chemically modified tetracycline-3 (CMT-3) by gavage. The levels of MMP-9, tissue inhibitor of metalloprotease-1, interleukin (IL)-6 were detected by real time RT-PCR at 6 h, 24 h and 48 h, and tumor necrosis factor (TNF), lung wet-dry weight ratio, white blood cell (WBC) count and polymorphonuclear (PMN) count in bronchoalveolar lavage fluid (BALF) were tested at 48 h after administration. The 5-day survival analysis of the ALI mice was also performed. Both ONO-5046 and CMT-3, regardless of being used individually or combined, significantly reduced the levels of MMP-9, IL-6, and TNF in lung tissue as well as in BALF, and the WBC and PMN count in BALF. Combined treatment with ONO-5046 and CMT-3 remarkably improved the survival rate of ALI mice. Neutrophil elastase synergizes with matrix metalloproteinase-9 to promote and regulate the release of inflammatory mediators and the infiltration of inflammatory cells, consequently affecting the survival of lipopolysaccharide-induced acute lung injury mice.

Recombinant human brain natriuretic peptide ameliorates trauma-induced acute lung injury via inhibiting JAK/STAT signaling pathway in rats.

JAK/STAT signal pathway plays an important role in the inflammation process of acute lung injury (ALI). This study aimed to investigate the correlation between recombinant human brain natriuretic peptide (rhBNP) and the JAK/STAT signaling pathway and to explore the protective mechanism of rhBNP against trauma-induced ALI. The arterial partial pressure in oxygen, lung wet-dry weight ratios, protein content in bronchoalveolar lavage fluid, the histopathologic of the lung, as well as the protein expressions of STAT1, JAK2, and STAT3 were detected. Sprague-Dawley rats were randomly divided into five groups: a control group, a sham-operated group, an ALI group, an ALI + rhBNP group, and an ALI + AG490 group. At 4 hours, 12 hours, 1 day, 3 days, and 7 days after injury, injured lung specimens were harvested. rhBNP pretreatment significantly ameliorated hypoxemia and histopathologic changes and alleviated pulmonary edema in trauma-induced ALI rats. rhBNP pretreatment reduced the phosphorylated protein and total protein level of STAT1. Similarly to JAK-specific inhibitor AG490, rhBNP was shown to significantly inhibit the phosphorylation of JAK2 and STAT3 in rats with trauma-induced ALI. Our experimental findings indicated that rhBNP can protect rats against trauma-induced ALI and that its underlying mechanism may be related to the inhibition of JAK/STAT signaling pathway activation.

Smoke inhalation injury repaired by a bone marrow-derived mesenchymal stem cell paracrine mechanism: Angiogenesis involving the Notch signaling pathway.

Smoke inhalation injury is an acute lung injury induced by smoke exposure characterized by vascular endothelial injury and increased permeability. Cell therapy is an attractive new therapeutic approach, although its underlying mechanism and signaling pathway remain poorly understood. We investigated the effect of systemic transplantation of preconditioned bone marrow-derived mesenchymal stem cells (BMSCs) on angiogenesis in rat model of smoke inhalation injury and explored the underlying mechanism and possible signaling pathway. After the establishment of a smoke inhalation injury rat model, the animals were further randomized into subgroups that received either a tail vein injection of 2 × 10(6) preconditioned or nonpreconditioned BMSCs in 5-mL phosphate-buffered saline to explore the characteristics of preconditioned BMSCs, pulmonary microvessel quantities in smoke inhalation injury, and its Notch1 expression. BMSCs preconditioned by 60Co γ-ray radiation at an appropriate dose were inhibited differentiation potential in vitro without significantly affecting the paracrine activity, the ability of cell proliferation, viability, and homing. Systemic preconditioned BMSC transplantation significantly increased the quantities of microvessels in rat with smoke inhalation injury, improved the lung wet-dry weight ratio, and alleviated lung injury simply through paracrine activity. Immunofluorescence staining and Western blot analysis confirmed that the expression level of Notch microvessel and Notch1 protein increased significantly after systemic transplantation. Our findings indicate that systemic transplantation of preconditioned BMSCs promotes angiogenesis through paracrine activity after smoke inhalation injury and that the Notch signaling pathway is involved in the angiogenesis process.

Therapeutic Effect of Intravenous Infusion of Perfluorocarbon Emulsion on LPS-Induced Acute Lung Injury in Rats

Acute lung injury (ALI) and its more severe form, acute respiratory distress syndrome (ARDS) are the leading causes of death in critical care. Despite extensive efforts in research and clinical medicine, mortality remains high in these diseases. Perfluorocarbon (PFC), a chemical compound known as liquid ventilation medium, is capable of dissolving large amounts of physiologically important gases (mainly oxygen and carbon dioxide). In this study we aimed to investigate the effect of intravenous infusion of PFC emulsion on lipopolysaccharide (LPS) induced ALI in rats and elucidate its mechanism of action. Forty two Wistar rats were randomly divided into three groups: 6 rats were treated with saline solution by intratracheal instillation (control group), 18 rats were treated with LPS by intratracheal instillation (LPS group) and the other 18 rats received PFC through femoral vein prior to LPS instillation (LPS+PFC group). The rats in the control group were sacrificed 6 hours later after saline instillation. At 2, 4 and 6 hours of exposure to LPS, 6 rats in the LPS group and 6 rats in LPS+PFC group were sacrificed at each time point. By analyzing pulmonary pathology, partial pressure of oxygen in the blood (PaO2) and lung wet-dry weight ratio (W/D) of each rat, we found that intravenous infusion of PFC significantly alleviated acute lung injury induced by LPS. Moreover, we showed that the expression of pulmonary myeloperoxidase (MPO), intercellular adhesion molecule-1 (ICAM-1) of endothelial cells and CD11b of polymorphonuclear neutrophils (PMN) induced by LPS were significantly decreased by PFC treatment in vivo. Our results indicate that intravenous infusion of PFC inhibits the infiltration of PMNs into lung tissue, which has been shown as the core pathogenesis of ALI/ARDS. Thus, our study provides a theoretical foundation for using intravenous infusion of PFC to prevent and treat ALI/ARDS in clinical practice.

Left Ventricular Failure Produces Profound Lung Remodeling and Pulmonary Hypertension in Mice

Chronic left ventricular failure causes pulmonary congestion with increased lung weight and type 2 pulmonary hypertension. Understanding the molecular mechanisms for type 2 pulmonary hypertension and the development of novel treatments for this condition requires a robust experimental animal model and a good understanding of the nature of the resultant pulmonary remodeling. Here we demonstrate that chronic transverse aortic constriction causes massive pulmonary fibrosis and remodeling, as well as type 2 pulmonary hypertension, in mice. Thus, aortic constriction-induced left ventricular dysfunction and increased left ventricular end-diastolic pressure are associated with a ≤5.3-fold increase in lung wet weight and dry weight, pulmonary hypertension, and right ventricular hypertrophy. Interestingly, the aortic constriction-induced increase in lung weight was not associated with pulmonary edema but resulted from profound pulmonary remodeling with a dramatic increase in the percentage of fully muscularized lung vessels, marked vascular and lung fibrosis, myofibroblast proliferation, and leukocyte infiltration. The aortic constriction-induced left ventricular dysfunction was also associated with right ventricular hypertrophy, increased right ventricular end-diastolic pressure, and right atrial hypertrophy. The massive lung fibrosis, leukocyte infiltration, and pulmonary hypertension in mice after transverse aortic constriction clearly indicate that congestive heart failure also causes severe lung disease. The lung fibrosis and leukocyte infiltration may be important mechanisms in the poor clinical outcome in patients with end-stage heart failure. Thus, the effective treatment of left ventricular failure may require additional efforts to reduce lung fibrosis and the inflammatory response.

Effects of cholinergic anti-inflammatory pathway on ventilator-induced lung injury in rats

To investigate the effects of cholinergic anti-inflammatory pathway on ventilator-induced lung injury (VILI) in rats. Thirty-six healthy Sprague-Dawley (SD) rats were randomly divided into three groups: control group, in which rats did not receive ventilation; high-tidal volume (HVT) ventilation group; nicotine treatment (HVT+nicotine) group, in which rats received intraperitoneal injection of nicotine (2 mg/kg) 10 minutes before HVT ventilation; equal amount of normal saline was given to rats in other two groups. A rat model of VILI was reproduced by volume-controlled mechanical ventilation with HVT. Hemodynamic parameters were measured throughout the study period. Arterial blood gases were measured every 1 hour. After maintaining ventilation for 2 hours, rats were sacrificed and lung tissue specimens were harvested. Lung wet-dry weight ratio (W/D) and myeloperoxidase (MPO) activity were measured. Interleukin-8 (IL-8) level in bronchoalveolar lavage fluid (BALF) and intercellular adhesion molecule-1 (ICAM-1) level in lung tissue homogenate were measured by enzyme-linked immunosorbent assay (ELISA), respectively. After hematoxylin and eosin (HE) staining, pathological examination of lung tissue was performed, and diffuse alveolar damage (DAD) score was estimated. Mean pH of arterial blood in HVT group and HVT+nicotine group tended to be higher than the baseline value during the ventilation. Mean partial pressure of oxygen in arterial blood (PaO2), mean partial pressure of carbon dioxide in arterial blood (PaCO2), and mean arterial pressure (MAP) in HVT group and HVT+nicotine group were lower than baseline value during the ventilation. Mean PaO2 (mm Hg, 1 mm Hg=0.133 kPa) in HVT+nicotine group was significantly higher than that in HVT group after 2 hours of ventilation (85+/-4 vs. 76+/-3, P<0.05). Mean W/D ratio and mean MPO activity in HVT group were significantly higher than those in control group [W/D ratio: 5.66+/-0.33 vs. 4.53+/-0.21, P<0.01; MPO (U/g): 1.73+/-0.50 vs. 0.89+/-0.17, P<0.05]. Mean W/D ratio (5.02+/-0.37) and mean MPO activity (1.11+/-0.33) in HVT+nicotine group were significantly lower than those in HVT group (both P<0.05). Compared with control group, DAD scores in HVT group and HVT+nicotine group (10.40+/-1.85, 7.90+/-1.67 vs. 1.60+/-1.20), IL-8 concentration (ng/L: 1 625.3+/-271.7, 965.5+/-310.5 vs. 428.5+/-120.6) and ICAM-1 concentration (microg/L: 589.4+/-87.5, 452.5+/-89.3 vs. 247.5+/-73.7) were significantly higher (all P<0.01). But DAD score, IL-8 concentration, ICAM-1 concentration in HVT+nicotine group were significantly lower than those in HVT group (P<0.05 or P<0.01). Activation of cholinergic anti-inflammatory pathway can protect the lung against VILI by suppressing IL-8 and ICAM-1 expression, inhibiting neutrophil aggregation and infiltration.

Effects of intravenous 2-chloroadenosine on endotoxin-induced acute lung injury

To assess the effects of 2-chloroadenosine (2CA) on acute lung injury caused by endotoxin (lipopolysaccharide), guinea pigs were given 2CA intravenously. Three groups were used: saline control, endotoxin control and 2CA+ endotoxin. In the endotoxin and 2CA+ endotoxin groups, neutrophils accumulated in bronchoalveolar lavage fluid and in lung tissue. However, neutrophil accumulation did not differ significantly between the endotoxin and the 2CA+ endotoxin groups. The number of macrophages in bronchoalveolar lavage fluid was significantly higher in the endotoxin group than in the saline control group, but the difference between the saline control and the 2CA+ endotoxin groups was not significant. The lung wet-dry weight ratio and 125I-albumin lung tissue-plasma ratio, which were used to measure acute lung injury, were significantly higher in the endotoxin group than in the 2CA+ endotoxin and the saline control groups. However, these ratios did not differ between the 2CA+ endotoxin and the saline control groups. These results suggest that 2CA attenuated endotoxin induced acute lung injury in guinea pigs.

Granulocyte colony-stimulating factor exacerbates acute lung injury induced by intratracheal endotoxin in guinea pigs.

The effects of recombinant human granulocyte colony-stimulating factor (rG-CSF) on the lung injury induced by intratracheal endotoxin were studied using guinea pigs. Animals were divided into four groups: (1) saline control, (2) endotoxin alone, (3) cyclophosphamide (CPA)+endotoxin, and (4) CPA+rG-CSF+endotoxin. CPA was injected intraperitoneally to suppress hematopoietic function 7 d before the study. rG-CSF at a dose of 100 micrograms/kg was administered subcutaneously twice a day for 5 consecutive d beginning 2 d after the CPA pretreatment. Saline or 0.2 mg/kg of endotoxin was administered via the airway, and the animals were observed for 4 h. 99mTc-labeled macroaggregated albumin was mixed with saline or endotoxin to obtain a lobar distribution. Lung injury was assessed by the concentration ratio of 125I-labeled albumin in lung tissue to plasma (T/P) and lung wet-dry weight ratio (W/D). We also counted the number of neutrophils in bronchoalveolar lavage (BAL) fluid and fixed lung tissues. T/P, but not W/D, increased in endotoxin-alone and CPA+endotoxin groups compared with the saline control group (p < 0.01). Both T/P and W/D of the CPA+rG-CSF+endotoxin group were significantly higher than those of the endotoxin-alone and CPA+endotoxin groups (p < 0.01). In the CPA+rG-CSF+endotoxin group, histopathologic examination of the lung sections showed neutrophil recruitment into the lung, and neutrophil counts in BAL fluid were elevated. In conclusion, pretreatment with rG-CSF increased sequestration of neutrophils into the lung and exacerbated the lung injury induced by intratracheal endotoxin in CPA-treated guinea pigs.