Lung Protein Expression Research Articles

Increased lethality of respiratory infection by Streptococcus pneumoniae in the Dp16 mouse model of Down syndrome.

We sought to investigate whether the Dp16 mouse model of Down syndrome (DS) is more susceptible to severe and lethal respiratory tract infection by Streptococcus pneumoniae. We infected controls and Dp16 mice with Streptococcus pneumoniae and measured survival rates. We compared cytokine production by primary lung cell cultures exposed to Streptococcus pneumoniae. We examined lung protein expression for interferon signaling related pathways. We characterized the histopathology and quantified the extent of bronchus-associated lymphoid tissue. Finally, we examined mouse tissues for the presence of oligomeric tau protein. We found that the Dp16 mouse model of DS displayed significantly higher susceptibility to lethal respiratory infection with Streptococcus pneumoniae compared to control mice. Lung cells cultured from Dp16 mice displayed unique secreted cytokine profiles compared to control mice. The Dp16 mouse lungs were characterized by profound lobar pneumonia with massive diffuse consolidation involving nearly the entire lobe. Marked red hepatization was noted, and Dp16 mice lungs contained numerous bronchus-associated lymphoid tissues that were highly follicularized. Compared to uninfected mice, both control mice and Dp16 mice infected with Streptococcus pneumoniae showed evidence of oligomeric tau aggregates. Increased susceptibility to severe respiratory tract infection with Streptococcus pneumoniae in Dp16 mice closely phenocopies infection in individuals with DS. The increase does not appear to be linked to overexpression of mouse interferon genes syntenic to human chromosome 21.

Naringin-Dextrin Nanocomposite Abates Diethylnitrosamine/Acetylaminofluorene-Induced Lung Carcinogenesis by Modulating Oxidative Stress, Inflammation, Apoptosis, and Cell Proliferation.

Nanotechnology has proven advantageous in numerous scientific applications, one being to enhance the delivery of chemotherapeutic agents. This present study aims to evaluate the mechanisms underlying the chemopreventive action of naringin-dextrin nanocomposites (Nar-Dx-NCs) against diethylnitrosamine (DEN)/2-acetylaminofluorene (2AAF)-induced lung carcinogenesis in male Wistar rats. DEN was administered intraperitoneally (i.p.) (150 mg/kg/week) for two weeks, followed by the oral administration of 2AAF (20 mg/kg) four times a week for three weeks. Rats receiving DEN/2AAF were concurrently treated with naringin or Nar-Dx-NCs orally at a dose of 10 mg/kg every other day for 24 weeks. Naringin and Nar-Dx-NCs treatments prevented the formation of tumorigenic cells within the alveoli of rats exposed to DEN/2AAF. These findings were associated with a significant decrease in lipid peroxidation, upregulation of antioxidant enzyme (glutathione peroxidase and superoxide dismutase) activity, and enhanced glutathione and nuclear factor erythroid 2-related factor 2 expression in the lungs. Naringin and Nar-Dx-NCs exerted anti-inflammatory actions manifested by a decrease in lung protein expression of tumor necrosis factor-α and interleukin-1β and mRNA expression of interleukin-6, interferon-γ, nuclear factor-κB, and inducible nitric oxide synthase, with a concurrent increase in interleukin-10 expression. The anti-inflammatory effect of Nar-Dx-NCs was more potent than naringin. Regarding the effect on apoptosis, both naringin and Nar-Dx-NCs significantly reduced Bcl-2 and increased Bax and P53 expressions. Moreover, naringin or Nar-Dx-NCs induced a significant decrease in the expression of the proliferator marker, Ki-67, and the effect of Nar-Dx-NCs was more marked. In conclusion, Nar-Dx-NCs improved naringin's preventive action against DEN/2AAF-induced lung cancer and exerted anticarcinogenic effects by suppressing oxidative stress and inflammation and improving apoptotic signal induction and propagation.

Protective effects of Qing-Re-Huo-Xue formula on bleomycin-induced pulmonary fibrosis through the p53/IGFBP3 pathway

BackgroundIdiopathic pulmonary fibrosis (IPF) is a chronic, progressive fibrosing lung disease with high mortality. Inflammation and epithelial mesenchymal transformation (EMT) may play an important role in the occurrence and development of IPF. Qing-Re-Huo-Xue formula (QRHXF) has been used clinically by our team for half a century and has obvious therapeutic effects on lung disease. Nevertheless, the role and mechanism of QRHXF in the treatment of IPF have never been studied.MethodsA mouse pulmonary fibrosis model was established by intratracheal injection of BLM. The effects of QRHXF on the treatment of pulmonary fibrosis were studied by pulmonary function testing, imaging examination, pathological staining, transmission electron microscopy (TEM) observation and mRNA expression. Tandem mass tag (TMT)-based quantitative proteomics was carried out to analyse the lung protein expression profiles between the control (CTL), bleomycin (BLM) and QRHXF (BLM + QRHXF) groups. Immunohistochemistry and qRT-PCR were used to verify the possible existence of drug target proteins and signalling pathways.ResultsThe results of pulmonary function, lung pathology and imaging examinations showed that QRHXF could significantly alleviate BLM-induced pulmonary fibrosis in vivo. Additionally, inflammatory cell infiltration and EMT were markedly reduced in BLM-induced PF mice administered QRHXF. Proteomics detected a total of 35 proteins, of which 17 were upregulated and 18 were downregulated. A total of 19 differentially expressed proteins (DEPs) overlapped between the BLM versus CTL groups and the BLM + QRHXF versus BLM groups. The expression of p53 and IGFBP3 was reversed in the QRHXF intervention group, which was verified by immunohistochemistry and qRT-PCR.ConclusionsQRHXF attenuated BLM-induced pulmonary fibrosis, and regulation of the p53/IGFBP3 pathway might be associated with its efficacy, which holds promise as a novel treatment strategy for pulmonary fibrosis patients.Graphical

The synergistic effect and mechanisms of flavonoids and polysaccharides from Houttuynia cordata on H1N1-induced pneumonia in mice

Ethnopharmacological relevanceHouttuynia cordata Thunb. (HC, Saururaceae family) is a classical Traditional Chinese Medicine used to treat pneumonia clinically. The total flavonoids (HCF) and polysaccharides (HCP) are key medicinal components of H. cordata involved in its beneficial effect on viral pneumonia. Aim of the studyThe purpose of the study is to investigate the synergistic or complementary effects of combination of HCF and HCP on viral pneumonia as well as the mechanisms underlying. Materials and methodsHCF or HCP were administrated separately or combined in different proportions on influenza virus H1N1 - infected mice. The survival and lung weight of mice were recorded. The synergistic effect on HCF and HCP combination was calculated by Chou-Talalay method. H&E staining was performed to detect lung histomorphology. Western blot, immunohistochemistry and enzyme linked immunosorbent assay were done to analyze the representative protein expression in lung and intestine tissues. AB - PAS staining on intestine tissue sections was performed to evaluate the histopathology of intestines. Bacterial genomic DNA was extracted and sequenced for gut microbiota analysis. ResultsIn H1N1 lethally infected mice, the combined administration of HCF and HCP significantly increased the survival rate and prolonged the life span of mice, compared with mono-drug therapy. The viral pneumonia was remarkably improved by HCF and HCP combination reflected by lower lung index, more intact lung morphology, and less inflammatory cells and mediators. Furthermore, the combination of HCF and HCP regulated intestinal microbiota, significantly reduced the proportion of pathogenic Proteobacteria and the secretion of proinflammatory cytokine in gut. The combined HCF and HCP showed synergistic effect on reducing lung and intestine injury. The complementary interaction was also found in HCF and HCP combined therapy, as HCF provided the significant antiviral activity and HCP markedly improved intestinal physical barrier and increased the protein expression involving removal of edema. ConclusionsOur findings indicated that combination of HCF and HCP from H. cordata synergistically alleviated H1N1-induced viral pneumonia in mice via multimodal regulation of both pulmonary and intestinal homeostasis, which might imply novel therapeutic strategy for treating viral pneumonia.

Biochemical and genomic identification of novel biomarkers in progressive sarcoidosis: HBEGF, eNAMPT, and ANG-2.

Progressive pulmonary fibrosis is a serious complication in subjects with sarcoidosis. The absence of reliable, non-invasive biomarkers that detect early progression exacerbates the difficulty in predicting sarcoidosis severity. To potentially address this unmet need, we evaluated a panel of markers for an association with sarcoidosis progression (HBEGF, NAMPT, IL1-RA, IL-6, IL-8, ANG-2). This panel encompasses proteins related to inflammation, vascular injury, cell proliferation, and fibroblast mitogenesis processes. Plasma biomarker levels and biomarker protein expression in lung and lymph nodes tissues (immunohistochemical studies) from sarcoidosis subjects with limited disease and progressive (complicated) sarcoidosis were performed. Gene expression of the protein-coding genes included in this panel was analyzed using RNAseq in sarcoidosis granulomatous tissues from lung and lymph nodes. Except for IL-8, plasma levels of each biomarker-eNAMPT, IL-1RA, IL-6, ANG-2, and HBEGF-were significantly elevated in sarcoidosis subjects compared to controls. In addition, plasma levels of HBEGF were elevated in complicated sarcoidosis, while eNAMPT and ANG-2 were observed to serve as markers of lung fibrosis in a subgroup of complicated sarcoidosis. Genomic studies corroborated HBEGF and NAMPT among the top dysregulated genes and identified cytokine-related and fibrotic pathways in lung granulomatous tissues from sarcoidosis. These findings suggest HBEGF, eNAMPT, and ANG-2 may serve as potential novel indicators of the clinical severity of sarcoidosis disease.

The Allelic Variant A391T of Metal Ion Transporter ZIP8 (SLC39A8) Leads to Hypotension and Enhanced Insulin Resistance.

The metal ion transporter ZIP8 (SLC39A8) mediates cellular uptake of vital divalent metal ions. Genome-wide association studies (GWAS) showed that the single-nucleotide polymorphism (SNP) variant A391T (rs13107325) is associated with numerous human traits, including reduced arterial blood pressure, increased body mass index and hyperlipidemia. We analyzed in vitro the transport properties of mutant ZIP8 A391T and investigated in vivo in mice the physiological effects of this polymorphism. In vitro, the intrinsic transport properties of mutant ZIP8 were similar to those of wild type ZIP8, but cellular uptake of zinc, cadmium and iron was attenuated due to reduced ZIP8 plasma membrane expression. We then generated the ZIP8 A393T mice (ZIP8KI) that carry the corresponding polymorphism and characterized their phenotype. We observed lower protein expression in lung and kidney membrane extracts in ZIP8KI mice. The ZIP8KI mice exhibited striking changes in metal ion composition of the tissues, including cobalt, palladium, mercury and platinum. In agreement with GWAS, ZIP8KI mice showed reduced arterial blood pressure. Body weight and plasma lipid composition remained unchanged, although these features were reported to be increased in GWAS. ZIP8KI mice also exhibited remarkable insulin resistance and were protected from elevated blood glucose when challenged by dietary sucrose supplementation. We showed that increased hepatic insulin receptor expression and decreased ZnT8 (slc30a8) metal ion transporter mRNA expression are associated with this phenotypic change. In conclusion, our data reveal that ZIP8 plays an important role in blood pressure regulation and glucose homeostasis.

The replacement of helper lipids with charged alternatives in lipid nanoparticles facilitates targeted mRNA delivery to the spleen and lungs.

The broad clinical application of mRNA therapeutics has been hampered by a lack of delivery vehicles that induce protein expression in extrahepatic organs and tissues. Recently, it was shown that mRNA delivery to the spleen or lungs is possible upon the addition of a charged lipid to a standard four-component lipid nanoparticle formulation. This approach, while effective, further complicates an already complex drug formulation and has the potential to slow regulatory approval and adversely impact manufacturing processes. We were thus motivated to maintain a four-component nanoparticle system while achieving shifts in tropism. To that end, we replaced the standard helper lipid in lipidoid nanoparticles, DOPE, with one of eight alternatives. These lipids included the neutral lipids, DOPC, sphingomyelin, and ceramide; the anionic lipids, phosphatidylserine (PS), phosphatidylglycerol, and phosphatidic acid; and the cationic lipids, DOTAP and ethyl phosphatidylcholine. While neutral helper lipids maintained protein expression in the liver, anionic and cationic lipids shifted protein expression to the spleen and lungs, respectively. For example, replacing DOPE with DOTAP increased positive LNP surface charge at pH7 by 5-fold and altered the ratio of liver to lung protein expression from 36:1 to 1:56. Similarly, replacing DOPE with PS reduced positive charge by half and altered the ratio of liver to spleen protein expression from 8:1 to 1:3. Effects were consistent across ionizable lipidoid chemistries. Regarding mechanism, nanoparticles formulated with neutral and anionic helper lipids best transfected epithelial and immune cells, respectively. Further, the lung-tropic effect of DOTAP was linked to reduced immune cell infiltration of the lungs compared to neutral or anionic lipids. Together, these data show that intravenous non-hepatocellular mRNA delivery is readily achievable while maintaining a four-component formulation with modified helper lipid chemistry.

Lack of Type I Interferon Signaling Ameliorates Respiratory Syncytial Virus-Induced Lung Inflammation and Restores Antioxidant Defenses.

Respiratory syncytial virus (RSV) infection in mouse and human lung is associated with pathogenic inflammation and oxidative injury. RSV impairs antioxidant responses by increasing the degradation of transcription factor NF-E2-related factor 2 (NRF2), which controls the expression of several antioxidant enzymes (AOEs). In addition to its protective effects, type I IFNs have been increasingly recognized as important mediators of host pathogenic responses during acute respiratory viral infections. We used a mouse model of RSV infection to investigate the effect of lack of type I interferon (IFN) receptor on viral-mediated clinical disease, airway inflammation, NRF2 expression, and antioxidant defenses. In the absence of type I IFN signaling, RSV-infected mice showed significantly less body weight loss and airway obstruction, as well as a significant reduction in cytokine and chemokine secretion and airway inflammation. Lack of type I IFN receptor was associated with greatly reduced virus-induced promyelocytic leukemia lung protein expression, which we showed to be necessary for virus-induced NRF2 degradation in a cell model of infection, resulting in restoration of NRF2 levels, AOE expression, and airway antioxidant capacity. Our data support the concept that modulation of type I IFN production and/or signaling could represent an important therapeutic strategy to ameliorate severity of RSV-induced lung disease.

Model difference in the effect of cilostazol on the development of experimental pulmonary hypertension in rats

BackgroundPreventing pulmonary vascular remodeling is a key strategy for pulmonary hypertension (PH). Causes of PH include pulmonary vasoconstriction and inflammation. This study aimed to determine whether cilostazol (CLZ), a phosphodiesterase-3 inhibitor, prevents monocrotaline (MCT)- and chronic hypoxia (CH)-induced PH development in rats.MethodsFifty-one male Sprague–Dawley rats were fed rat chow with (0.3% CLZ) or without CLZ for 21 days after a single injection of MCT (60 mg/kg) or saline. Forty-eight rats were fed rat chow with and without CLZ for 14 days under ambient or hypobaric (air at 380 mmHg) CH exposure. The mean pulmonary artery pressure (mPAP), the right ventricle weight-to-left ventricle + septum weight ratio (RV/LV + S), percentages of muscularized peripheral pulmonary arteries (%Muscularization) and medial wall thickness of small muscular arteries (%MWT) were assessed. Levels of the endothelial nitric oxide synthase (eNOS), phosphorylated eNOS (peNOS), AKT, pAKT and IκB proteins in lung tissue were measured using Western blotting. Monocyte chemotactic protein (MCP)-1 mRNA in lung tissue was also assessed.ResultsmPAP [35.1 ± 1.7 mmHg (MCT) (n = 9) vs. 16.6 ± 0.7 (control) (n = 9) (P < 0.05); 29.1 ± 1.5 mmHg (CH) (n = 10) vs. 17.5 ± 0.5 (control) (n = 10) (P < 0.05)], RV/LV + S [0.40 ± 0.01 (MCT) (n = 18) vs. 0.24 ± 0.01 (control) (n = 10) (P < 0.05); 0.41 ± 0.03 (CH) (n = 13) vs. 0.27 ± 0.06 (control) (n = 10) (P < 0.05)], and %Muscularization and %MWT were increased by MCT injection and CH exposure. CLZ significantly attenuated these changes in the MCT model [mPAP 25.1 ± 1.1 mmHg (n = 11) (P < 0.05), RV/LV + S 0.30 ± 0.01 (n = 14) (P < 0.05)]. In contrast, these CLZ effects were not observed in the CH model. Lung eNOS protein expression was unchanged in the MCT model and increased in the CH model. Lung protein expression of AKT, phosphorylated AKT, and IκB was downregulated by MCT, which was attenuated by CLZ; the CH model did not change these proteins. Lung MCP-1 mRNA levels were increased in MCT rats but not CH rats.ConclusionsWe found model differences in the effect of CLZ on PH development. CLZ might exert a preventive effect on PH development in an inflammatory PH model but not in a vascular structural change model of PH preceded by vasoconstriction. Thus, the preventive effect of CLZ on PH development might depend on the PH etiology.

Hyperoxia-induced S1P1 signaling reduced angiogenesis by suppression of TIE-2 leading to experimental bronchopulmonary dysplasia.

We have earlier shown that hyperoxia (HO)-induced sphingosine kinase 1 (SPHK1)/sphingosine-1-phosphate (S1P) signaling contribute to bronchopulmonary dysplasia (BPD). S1P acts through G protein-coupled receptors, S1P1 through S1P5. Further, we noted that heterozygous deletion of S1pr1 ameliorated the HO-induced BPD in the murine model. The mechanism by which S1P1 signaling contributes to HO-induced BPD was explored. S1pr1+/+ and S1pr1+/- mice pups were exposed to either room air (RA) or HO (75% oxygen) for 7 days from PN 1-7. Lung injury and alveolar simplification was evaluated. Lung protein expression was determined by Western blotting and immunohistochemistry (IHC). In vitro experiments were performed using human lung microvascular endothelial cells (HLMVECs) with S1P1 inhibitor, NIBR0213 to interrogate the S1P1 signaling pathway. HO increased the expression of S1pr1 gene as well as S1P1 protein in both neonatal lungs and HLMVECs. The S1pr1+/- neonatal mice showed significant protection against HO-induced BPD which was accompanied by reduced inflammation markers in the bronchoalveolar lavage fluid. HO-induced reduction in ANG-1, TIE-2, and VEGF was rescued in S1pr1+/- mouse, accompanied by an improvement in the number of arterioles in the lung. HLMVECs exposed to HO increased the expression of KLF-2 accompanied by reduced expression of TIE-2, which was reversed with S1P1 inhibition. HO induces S1P1 followed by reduced expression of angiogenic factors. Reduction of S1P1 signaling restores ANG-1/ TIE-2 signaling leading to improved angiogenesis and alveolarization thus protecting against HO-induced neonatal lung injury.

Glucagon-like peptide-1 receptor agonist inhibits aeroallergen-induced activation of ILC2 and neutrophilic airway inflammation in obese mice.

BackgroundObesity is a risk factor for the development of asthma. However, pharmacologic therapeutic strategies that specifically target obese asthmatics have not been identified. We hypothesize that glucagon‐like peptide‐1 receptor agonist (GLP‐1RA) treatment inhibits aeroallergen‐induced early innate airway inflammation in a mouse model of asthma in the setting of obesity.MethodsSWR (lean) and TALLYHO (obese) mice were challenged intranasally with Alternaria alternata extract (Alt‐Ext) or PBS for 4 consecutive days concurrent with GLP‐1RA or vehicle treatment.ResultsTALLYHO mice had greater Alt‐Ext‐induced airway neutrophilia and lung protein expression of IL‐5, IL‐13, CCL11, CXCL1, and CXCL5, in addition to ICAM‐1 expression on lung epithelial cells compared with SWR mice, and all endpoints were reduced by GLP‐1RA treatment. Alt‐Ext significantly increased BALF IL‐33 in both TALLYHO and SWR mice compared to PBS challenge, but there was no difference in the BALF IL‐33 levels between these two strains. However, TALLYHO, but not SWR, mice had significantly higher airway TSLP in BALF following Alt‐Ext challenge compared to PBS, and BALF TSLP was significantly greater in TALLYHO mice compared to SWR mice following airway Alt‐Ext challenge. GLP‐1RA treatment significantly decreased the Alt‐Ext‐induced TSLP and IL‐33 release in TALLYHO mice. While TSLP or ST2 inhibition with a neutralizing antibody decreased airway eosinophils, they did not reduce airway neutrophils in TALLYHO mice.ConclusionsThese results suggest that GLP‐1RA treatment may be a novel pharmacologic therapeutic strategy for obese persons with asthma by inhibiting aeroallergen‐induced neutrophilia, a feature not seen with either TSLP or ST2 inhibition.

L‐Citrulline Decreases LPS‐Induced Inflammation and Oxidative Stress in Newborn Rat Lungs

Introduction Extremely low birth weight infants have immature lungs at birth as well as low levels of anti-inflammatory and antioxidative defences, placing them at high risk of developing bronchopulmonary dysplasia (BPD). Adiponectin (APN) is produced by adipocytes and has potent anti-inflammatory and antioxidative properties, and anti-proliferative effects; APN levels are quite low in premature infants. We showed that exogenous APN markedly attenuated lipopolysaccharide (LPS)-induced lung injury (Ivanovska et al. 2020) in newborn rats. Recombinant APN is not available for human use; however, L-citrulline (L-CIT) increases APN gene expression in adult rats (Joffin et al., 2015) and upregulates expression of peroxisome proliferator-activated receptor-gamma coactivator alpha (PGC-1α). PGC-1α is a master coactivator of genes involved in mitochondrial biogenesis, inflammation and oxidative stress. L-CIT levels are also low in premature infants. Thus, we sought to determine the effect of exogenous L-CIT on LPS-induced changes in protein expression of PGC-1α, APN, APN receptors (AdipoR1/2), cytokines, and superoxide dismutase (SOD) in lung and pulmonary artery smooth muscle cells (PASMC) homogenates, and on LPS-induced changes of intracellular ROS levels in PASMC. Methods in-vivo: Sprague Dawley rat pups were treated with L-CIT (2.5 g/kg) or saline intraperitoneally from postnatal day (PND) 1-5. On PND 5, intra-pharyngeal LPS (5mg/kg) or saline was given to L-CIT (n=4-6) or saline (n=4-6) treated animals; pups were euthanized 24 h after LPS treatment and lungs were removed and processed for western blot. In-vitro: PASMCisolated from PND 4 rats were treated with L-CIT (4 mM/ml), LPS (5μg/ml) and a combination of the two, for 3 h; cells were harvested and processed for protein expression (n=4 replicate experiments). PASMC were assayed for the level of LPS-induced intracellular ROS with fluorescence dye in the presence and absence of blocker for PGC-1α (SR 18292) Results In vivo, L-CIT, in the presence of LPS, increased lung protein expression of APN (1.36-fold), AdipoR1/2 (2-fold), PGC-1α (1.8-fold), phosphorylated AMP-kinase (2.1-fold), SOD1 (2.3-fold), SOD2 (1.7-fold), SOD3 (1.43-fold), and decreased tumor necrosis factor α (1.2-fold), and interleukin-8 (1.28-fold) compared to saline and LPS treated animals (p<0.05 L-CIT+LPS vs saline+LPS for all proteins). In vitro, in the presence of LPS, L-CIT increased PASMC protein expression of PGC-1α (1.55-fold), SOD1 (2-fold), SOD2 (1.57-fold), and SOD3 (2.2-fold) compared to cells only exposed to LPS (p<0.05; L-CIT+LPS vs medium+LPS). L-CIT decreased LPS-induced ROS production in PASMC, an effect that was inhibited in the presence of PGC-1α blocker. Conclusion L-CIT upregulates APN signaling in presence of LPS and decreases LPS-induced lung inflammation and oxidative stress in newborn rats through activation of PGC1-α. The translational significance of these findings warrants further clinical studies of the L-CIT potential preventative/curative benefit in BPD.

Absence of Vaccine-enhanced Disease With Unexpected Positive Protection Against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by Inactivated Vaccine Given Within 3 Days of Virus Challenge in Syrian Hamster Model.

BackgroundMass vaccination against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is ongoing amidst widespread transmission during the coronavirus disease-2019 (COVID-19) pandemic. Disease phenotypes of SARS-CoV-2 exposure occurring around the time of vaccine administration have not been described.MethodsTwo-dose (14 days apart) vaccination regimen with formalin-inactivated whole virion SARS-CoV-2 in golden Syrian hamster model was established. To investigate the disease phenotypes of a 1-dose regimen given 3 days prior (D-3), 1 (D1) or 2 (D2) days after, or on the day (D0) of virus challenge, we monitored the serial clinical severity, tissue histopathology, virus burden, and antibody response of the vaccinated hamsters.ResultsThe 1-dose vaccinated hamsters had significantly lower clinical disease severity score, body weight loss, lung histology score, nucleocapsid protein expression in lung, infectious virus titers in the lung and nasal turbinate, inflammatory changes in intestines, and a higher serum neutralizing antibody or IgG titer against the spike receptor-binding domain or nucleocapsid protein when compared to unvaccinated controls. These improvements were particularly noticeable in D-3, but also in D0, D1, and even D2 vaccinated hamsters to varying degrees. No increased eosinophilic infiltration was found in the nasal turbinate, lung, and intestine after virus challenge. Significantly higher serum titer of fluorescent foci microneutralization inhibition antibody was detected in D1 and D2 vaccinated hamsters at day 4 post-challenge compared to controls despite undetectable neutralizing antibody titer.ConclusionsVaccination just before or soon after exposure to SARS-CoV-2 does not worsen disease phenotypes and may even ameliorate infection.

Sleeve gastrectomy ameliorates alveolar structures and surfactant protein expression in lungs of obese and diabetic rats.

Bariatric surgeries have been shown to be effective in reversing damaged pulmonary function in individuals suffering from obesity and type 2 diabetes mellitus, whereas its underlying mechanisms remain largely unknown. Sleeve gastrectomy (SG) was performed on obese and diabetic Wistar rats, and their pulmonary function and lung tissues were compared to sham-operated (SH) obese and diabetic rats, and age-matched healthy controls (C) to explore the improvements in microstructures and expression of surfactant protein (SP)-A and -C at postoperative 4th, 8th, and 12th week. Apart from the profound metabolic changes and improvement in pulmonary function, lung volume was restored along with an improved diffusion capacity noted by thinned capillary basement membrane and decreased harmonic mean length of diffusion barrier in SG rats. The digital slices of light microscope showed the general changes brought on by the SG, including normalized basic structures, ameliorated inflammatory status, as well as reduced lipid deposition, where the hydroxyproline (HYP), triglyceride (TG) assays, and electron microscope further suggested that the improvement in alveolar structures lies in reduced collagen fibers, lipids and septal tissues, increased capillary blood, and normalized alveolar type 2 (AT2) cells. Besides, disrupted SP-A and SP-C expression were also normalized after SG. The improvement of lung function after SG is related to the ameliorated alveolar structures, and surface protein expression induced by weight loss and improved glucose metabolism.

TSLP and IL-33 reciprocally promote each other's lung protein expression and ILC2 receptor expression to enhance innate type-2 airway inflammation.

BackgroundThe epithelial cell‐derived danger signal mediators thymic stromal lymphopoietin (TSLP) and IL‐33 are consistently associated with adaptive Th2 immune responses in asthma. In addition, TSLP and IL‐33 synergistically promoted group 2 innate lymphoid cell (ILC2) activation to induce innate allergic inflammation. However, the mechanism of this synergistic ILC2 activation is unknown.MethodsBALB/c WT and TSLP receptor‐deficient (TSLPR−/−) mice were challenged intranasally with Alternaria extract (Alt‐Ext) or PBS for 4 consecutive days to evaluate innate airway allergic inflammation. WT mice pre‐administered with rTSLP or vehicle, TSLPR−/− mice, and IL‐33 receptor‐deficient (ST2−/−) mice were challenged intranasally with Alt‐Ext or vehicle once or twice to evaluate IL‐33 release and TSLP expression in the lung. TSLPR and ST2 expression on lung ILC2 were measured by flow cytometry after treatment of rTSLP, rIL‐33, rTSLP + rIL‐33, or vehicle.ResultsThymic stromal lymphopoietin receptor deficient mice had significantly decreased the number of lung ILC2 expressing IL‐5 and IL‐13 following Alt‐Ext‐challenge compared to WT mice. Further, eosinophilia, protein level of lung IL‐4, IL‐5, and IL‐13, and airway mucus score were also significantly decreased in TSLPR−/− mice compared to WT mice. Endogenous and exogenous TSLP increased Alt‐Ext‐induced IL‐33 release into BALF, and ST2 deficiency decreased Alt‐Ext‐induced TSLP expression in the lung. Further, rTSLP and rIL‐33 treatment reciprocally increased each other's receptor expression on lung ILC2 in vivo and in vitro.ConclusionThymic stromal lymphopoietin and IL‐33 signaling reciprocally enhanced each other's protein release and expression in the lung following Alt‐Ext‐challenge and each other's receptor expression on lung ILC2 to enhance ILC2 activation.

Enhancement of tumor suppressor RAR-β protein expression by cationic liposomal-ATRA treatment in benzo(a)pyrene-induced lung cancer mice model.

Targeting the specific molecular proteins or genes which are responsible for the suppression of cancer growth is currently an emerging molecular method to treat cancer. ATRA treatment is now considered as a molecular targeted therapy for many cancers. As ATRA exhibits its therapeutic effect through its receptors, this study was focused to investigate the effect and action of liposomal-ATRA treatment on the expression of RAR-β protein which is also a tumor suppressor. The liposomal-ATRA was developed with cationic DOTAP and cholesterol by thin-film formation method. The benzo(a)pyrene(50mg/kg b.wt)-induced mice were treated with free and liposomal-ATRA(0.60mg/kg b.wt). The RAR-β protein expression in lung and liver tissue samples were analyzed by immunohistochemistry (IHC) and western blotting (WB) on the 30th and 120th days. Almost nil expression of RAR-β protein was observed in B(a)P cancer control and liposome alone-treated groups. A comparatively elevated expression was seen in the free ATRA-treated group (IHC score-2+ in lung on the 120th day with band density of 14.46 ± 1.24% in WB). Interestingly, the liposomal-ATRA treatment demonstrated a significantly (p ≤ 0.01) higher RAR-β expression in lung (35.20 ± 3.398% band intensity and score 4+ in the 120th day) than that of in ATRA alone treatment. This study results indicate that the RAR-β protein expression was suppressed by B(a)P during cancer induction even on the 30th day itself. The treatment could reactivate the suppression and the lipo-ATRA treatment could show significantly higher RAR-β expression on the 120th day, which implies that it accumulated more ATRA in target site and sustained it for enhanced action.

Impact of nitrate therapy on the expression of caveolin-1 and its phosphorylated isoform in lungs in the model of monocrotaline induced pulmonary hypertension

Abstract Aim: Nitric oxide signalling pathway showed to be one of the crucial factors in the treatment and pathogenesis of pulmonary arterial hypertension. The aim of this study was to determine the effect of administration of inorganic nitrate, NaNO3, on the expression of caveolin-1 and its phosphorylated isoform (pTyr14Cav-1) in lungs in the experimental model of monocrotaline induced pulmonary hypertension. Methods: 10 weeks old male Wistar rats were subcutaneously injected with 60 mg/kg dose of monocrotaline (MCT) or vehicle (CON). Twelve days after the injection, part of the MCT group was receiving 0.3 mM NaNO3 (MCT+N0.3) daily in the drinking water and rest was receiving 0.08% NaCl solution. Four weeks after MCT administration, the rats were sacrificed in CO2. Protein expression in lungs was determined by western blot. Results: We observed a significant decrease in the caveolin-1 expression and a significant shift towards the expression of pTyr14Cav-1 in the group treated with nitrate (p &lt; 0.05). Conclusion: NaNO3 administration affected the expression of caveolin-1 and the ratio of its active (phosphorylated) isoform increased.

Overexpression of APC11 predicts worse survival in lung adenocarcinoma.

BackgroundAnaphase-promoting complex subunit 11 (APC11) plays an important role in gathering E2 and catalyzing ubiquitin-chain formation to support ubiquitination of substrates by acting as a catalytic core subunit of anaphase-promoting complex (APC/C). However, whether APC11 is implicated in the tumorigenesis of lung cancer is never known.Materials and methodsIn this study, we used an online survival analysis software to estimate the prognostic value of APC11 mRNA expression level for lung cancer. Cell Counting Kit-8 assay, colony-forming assay, and transwell assay were used to assess the biological functions of APC11 in lung cancer cells. Then, 107 lung cancer patient tissues were collected to examine the expression level of APC11 by immunohistochemistry staining. Kaplan–Meier method and univariate Cox regression analysis were performed to reveal the prognostic value of APC11 protein expression in lung cancer.ResultsHigher mRNA level of APC11 was significantly associated with worse survival for lung adenocarcinoma, but not for lung squamous cell carcinoma. Knockdown of APC11 by siRNA apparently inhibited cell proliferation and colon formation in both H1299 and H358 cells. In addition, silencing of APC11 decreased cell migrative and invasive abilities. Moreover, immunohistochemical analysis showed that APC11 was highly expressed in lung cancer tissues, and multivariate analysis suggested that APC11 overexpression was an independent prognostic factor in lung adenocarcinoma.ConclusionWe suggest that APC11 could serve as a prognostic biomarker and a novel target in treating lung adenocarcinoma.

The effects of methylprednisolone on NLRP3 inflammasome in rats with acute lung injury Induced by Phosgene

Objective: To investigate the effects of methylprednisolone on NOD-like receptor hot protein domain-associated protein 3 (NLRP3) inflammasome in phosgene-induced acute lung injury. Methods: Rats were randomly divided into four groups, 10 rats in Air group (inhalation of air of the same volume as the phosgene group) , 10 rats in Phosgene group (inhalation of 8.33 mg/L with 100% purity phosgene for 5 min) , 10 rats in Saline control group (inhalation of the same dose of phosgene and 2 mg/kg saline via tail vein injection one hour later) , 10 rats in MP group (inhalation of the same dose of phosgene and 2 mg/kg MP via tail vein injection one hour later) . The specimens of serum, bronchoalveolar lavage fluid (BALF) and lung tissue were collected after 6h. Morphological changes were observed by HE staining. The expression of NLRP3 in the lung of four groups was detected by immunohistochemistry. NLRP3、ASC and caspase-1 expression in the lung tissue was quantified by Western blot. Reverse transcription-polymerase chain reaction (RT-PCR) were used to detect the expression of NLRP3、ASC and caspase-1 mRNA in the lung tissue. The concentrations of IL-1β、IL-18 and IL-33 in the serum and BALF were measured by enzyme-linked immunosorbent assay. Results: We successfully replicated the model of phosgene-induced ALI in rats. Morphological of HE staining after phosgene exposure to 6 h observed inflammatory cell infiltration in lung tissue in Phosgene group. Immunohistochemical staining results showed that there were many NLRP3 positive cells in lung tissue in Phosgene group. The levels of NLRP3, caspase-1 mRNA and protein expression in lung were significantly increased (P<0.05) in Phosgene group compared with Air group; compared with Phosgene group, The levels of NLRP3 and caspase-1 mRNA and protein expression in MP group were significantly decreased (P<0.05) . Compared with Air group, The levels IL-1β、IL-18 and IL-33 mRNA protein expression in the serum and BALF were significantly increased (P<0.05) in Phosgene group. Compared with Phosgene group, The levels IL-1β、IL-18 and IL-33 mRNA protein expression in the serum and BALF were significantly decreased (P<0.05) in MP group. Conclusion: Methylprednisolone can effectively protect the rats from phosgene-induced acute lung injury by inhibiting the expression of the NLRP3 inflammasome and reducing the release of inflammatory factors such as interleukin-1β (IL-1β) mediated by it.

Changes of apoE protein expression in lung of mice with hypoxic pulmonary arterial hypertension

To observe the changes of apolipoprotein E (apoE) protein expression of pulmonary tissue in mice with pulmonary hypertension induced by hypoxia. The animal model of hypoxic pulmonary hypertension was established by exposing the mice to isobaric hypoxic chamber for 3 weeks (23 h/d, regular chow feed).Twenty male wild type (WT) C57BL/6 mice and twenty apoE gene knockout (apoE-KO) mice were randomly divided into normoxia group and hypoxia group. The plasma concentrations of low density lipoprotein (LDL), high density lipoprotein (HDL) and total cholesterol were detected by ELISA method. The protein expression of apoE in lung and liver, and peroxisome proliferators-activated receptor gamma (PPARγ) in lung were measured by Western blot. ①In WT mice, the right ventricular systolic pressure (RVSP) and the weight ratio of right ventricle (RV) to left ventricle plus septum (LV+S) of hypoxia group were significantly higher than those of normoxia group by 68% and 59% (P<0.05), respectively. The plasma concentration of HDL and HDL/LDL of hypoxia group were significantly lower than those of normoxia group by 17% and 40% (P<0.05), respectively.The protein expression of apoE in lung and in liver of hypoxia group were significantly down-regulated than those of normoxia group by 48% and 52% (P<0.05), respectively.The protein expression of PPARγ in lung was significantly down-regulated than that of normoxia group by 37%(P<0.05).RVSP were significantly negative correlated with the protein levels of apoE and PPARγ in lung (P<0.01).② In apoE-KO mice, RVSP and the weight ratio of RV to LV+S of hypoxia group were significantly higher than those of normoxia group by 96% and 86% (P<0.05), respectively.RVSP and RV to (LV+S) of hypoxia group in apoE-KO mice were significantly higher than those of hypoxia group in WT mice by 29% and 24% (P<0.05), respectively. Down-regulated expression of apoE in lung tissue participates in the pathological proceeding of pulmonary hypertension induced by hypoxia.