Experimental Lung Research最新文献

Purpose of the study: The involvement of the IL-23/IL23R pathway is well known in the disease pathogenesis of sarcoidosis and other inflammatory diseases. To date, the pathogenic mechanism of IL-23 is most notably described on CD4⁺ Th17 lymphocytes. However, the function of the IL23R on myeloid cells in sarcoidosis is poorly understood. Thus, the aim of the study is to investigate the role of the IL23R on myeloid cell in pulmonary granuloma formation. Methods: We generated IL23RLysMCre mice lacking the IL23R gene in myeloid cells. The importance of IL23R in myeloid cells for the development of sarcoidosis was studied in a mouse model of inflammatory lung granuloma formation through embolization of PPD from Mycobacterium bovis-coated Sepharose beads into previously PPD-immunized mice. In addition the function of IL23R on myeloid cells was studied in LPS or IFNγ stimulated BMDMs and BMDCs. The mRNA and protein expression levels of relevant cytokines were analyzed by RT-PCR (TaqMan) and ELISA. The composition of immune cells in BALF was quantified by flow cytometry and alteration in granuloma sizes were observed by H&E stained lung sections. Results: Mycobacterium Ag-elicted pulmonary granulomas tend to be smaller in IL23RLysMCre mice and NF-κB dependent Th1 cytokines in the murine lungs are reduced compared to wildtype mice. In line, we observed that IL23R-deficient bone marrow-derived macrophages show a reduced production of Th1 cytokines after LPS stimulation. Conclusion: We here for the first time demonstrate a role for IL23R on myeloid cells in pulmonary inflammation and granuloma formation. Our findings provide essential insights in the pathogenesis of inflammatory lung diseases like sarcoidosis, which might be useful for the development of novel therapeutics targeting distinct immunological pathways like IL-23/IL23R.

Objective: Chronic obstructive pulmonary disease (COPD) is a respiratory disease with high morbidity and mortality worldwide, so far there is no ideal treatment method. Previous studies have shown that hydrogen (H₂) is involved in the treatment of COPD as an antioxidant. In this study, the effect of H₂ on M1/M2 polarization of alveolar macrophages in COPD rats was observed, and its anti-inflammatory mechanism was further elucidated. Methods: Twenty-four Sprague-Dawley rats were randomly divided into three groups including the control, COPD and H₂ group. A rat model of COPD was established by cigarette exposure combined with lipopolysaccharide (LPS) induction. H₂ therapy was administered 2 hours per day for 14 days. Lung function and pathology were assessed. The levels of interleukin (IL)-6, tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β1 and IL-10 in bronchoalveolar lavage fluid (BALF) and lung tissue were measured by enzyme-linked immunosorbent assay. The mRNA, protein expression and immunoreactivity of inducible nitric oxide synthase (iNOS) and arginase (Arg)-1 in lung were observed by quantitative real-time PCR, western blot and immunohistochemistry. Results: Compared with the control rats, there were a significant decline in lung function, a marked inflammatory infiltration and pulmonary parenchymal remodeling and the increases of IL-6, TNF-α and TGF-β1 levels in BALF and lung tissue, but a lower expression of IL-10 in COPD rats. The iNOS mRNA and protein expression, as well as its optical density (OD), were increased significantly in lung tissue, while those of Arg-1 decreased significantly. H₂ treatment improved the lung function and the parenchymal inflammation, reversed the increased levels of IL-6, TNF-α and TGF-β1, and the lower IL-10. Meanwhile, H₂ also down-regulated the expression of iNOS, but up-regulated expression of Arg-1 in lung tissue. Conclusion: H₂ reduces inflammation in the lung of COPD, which may be related to its inhibition of M1 type polarization and activation of M2 type polarization of alveolar macrophage.

Purpose: Asthma is associated with a T helper (Th)17/regulatory T (Treg) cells immune imbalance where the Notch signaling pathway contributes vitally. This study aimed to explore the role of Notch ligands Jagged1 and Delta4 in the Th17/Treg immune imbalance of chronic asthmatic mice.

Methods: The experimental animals were randomly assigned to the Saline, ovalbumin (OVA), and OVA + γ-secretase inhibitor (GSI) groups. A mouse model of chronic asthma was induced by OVA sensitization and challenge. GSI was injected intraperitoneally before the OVA challenge in the OVA + GSI group. Lung function, lung histopathology and immunohistochemistry to assess airway inflammation, enzyme-linked immunosorbent assay to measure cytokines levels, flow cytometry to measure the proportions of Th17 (Th17%) and Treg% in CD4⁺T cells, quantitative real-time polymerase chain reaction and western blot to measure mRNA and protein levels of Jagged1 and Delta4 in lung tissue, and correlation analysis were performed.

Results: Lung function and histopathology and IL-4, IL-13, and IFN-γ levels in the bronchoalveolar lavage fluid (BALF) of chronic asthmatic mice showed characteristic changes of asthma. The Th17%, Th17/Treg ratio, BALF and serum IL-17 levels, and IL-17/IL-10 ratio increased significantly in the OVA group, while the Treg% and IL-10 level significantly decreased. mRNA and protein expression levels of Jagged1 and Delta4 increased significantly. GSI could reduce the Th17%, Th17/Treg ratio, IL-17, IL-17/IL-10 ratio, and Jagged1 expression in chronic asthmatic mice. The mRNA and protein levels of Jagged1 and Delta4 were positively correlated with the Th17/Treg ratio in the OVA group, while only those of Jagged1 were positively correlated with the Th17/Treg ratio in the OVA + GSI group.

Conclusions: In chronic asthmatic mice, the Th17/Treg ratio increased, and the Notch ligands Jagged1 and Delta4 were overactive and positively regulated the Th17/Treg imbalance. GSI partially inhibited Jagged1 and relieved the Th17/Treg imbalance.

Purpose: Non-small lung (NSCLC) is the deadliest cancer, with survival measured in months. Earlier diagnosis using a robust biomarker would likely improve survival. This study aims to determine whether blood levels of the extracellular sulfatases (SULF1 and SULF2) and their bio-activity can serve as novel biomarkers for NSCLC early detection.

Materials and methods: Using human plasma specimens from NSCLC patients, nonmalignant COPD patients, and healthy individuals, we determined the association between plasma SULF levels and the presence of NSCLC. We assessed the plasma SULF levels as a function of sex and age. We also evaluated the plasma levels of heparin-binding factors potentially mobilized by the SULFs. To increase test specificity of blood SULF2 as a biomarker for the early diagnosis of NSCLC, we investigated the presence of a tumor-specific SULF2 isoform released in the blood, which could be used as a biomarker alone or in multiplex assays.

Results: The median level of plasma SULF2 was significantly elevated in NSCLC patients than in healthy controls (∼2 fold). However, these data were confounded by age. Surprisingly, COPD patients also showed a dramatically increased SULF2 plasma level. We showed a significant increase in the median plasma levels of several HSPG-binding factors in early-stage NSCLC patients compared to controls. Furthermore, we revealed a significant positive correlation of the SULF2 protein level with the plasma levels of two HSPG-binding factors IL6 and IL8. We demonstrated that NSCLC cancer cells and tissues overexpress a SULF2 splice variant. We determined the presence of a SULF2 splice variant form in NSCLC plasma, which was not detectable in COPD and control plasmas.

Conclusion: Our findings highlight the potential for the plasma levels of SULF2 protein and its bio-activity as novel blood biomarkers for early diagnosis of NSCLC.

Purpose: This study was prospectively designed to investigate the effects of different concentrations of mesenchymal stem cells treatment on respiratory mechanics, oxygenation, hemodynamics and inflammatory response in LPS-induced acute respiratory distress syndrome (ARDS) rat model. Methods: One hundred and twenty six LPS-induced ARDS model rats (weighted 200-220 g) were randomly divided into three groups: 1) Control group (N = 42); 2) low-dose hUC-MSC treatment group (MSC group 1, 1x10⁷ cell/kg, N = 42); 3) high-dose hUC-MSC treatment group (MSC group 2, 2x10⁷ cell/kg, N = 42), sham operation group as healthy group (N = 15). The rats were observed closely for 24 hours after hUC-MSC treatment, and the survival rate was calculated. At 24 hours, all rats were tested for hemodynamics, blood gas analysis, heart, lung, liver and kidney functions, inflammatory factors detection in blood samples and broncho-alveolar lavage fluid (BALF). The lung tissue of the rats was collected for HE staining analysis. Results: After LPS injection, ARDS was obvious in all LPS-infused rat groups, consistent with severe acute lung injury and high death rate. However, compared with the control group, a single intravenous injection hUC-MSC at dose of 1 × 10⁷ cells/kg (low dose group) and 2 × 10⁷ cells/kg (high dose group) reduced the mortality of rats with LPS-induced ARDS, as well as improving the lung function, increased the arterial oxygen pressure, improved the heart function, and reduced the levels of inflammatory factors including IL-1β, IL-6, and TNF-α. In addition, the high dose MSC group showed better lung injury therapeutic effects than the low dose MSC group. Data from this study demonstrated that injection of hUC-MSC had a significant therapeutic effect in treating the rat model of LPS-induced ARDS and multiple organ function injury.

Background and objectives: Idiopathic pulmonary fibrosis (IPF) is a progressive and lethal lung disease. An increased expression of somatostatin receptor subtype 2 in patients with IPF was identified and lung fibroblasts expressed somatostatin receptors in vitro. In addition, somatostatin analogue inhibits the expression of transforming growth factor-β, insulin-like growth factor (IGF) -1, platelet-derived growth factor, and basic fibroblast growth factor. Therefore, we examined the effects of somatostatin analogue on bleomycin-induced pulmonary fibrosis in mice. In a similar model, it has been reported that administration of high-dose somatostatin analogs suppressed acute inflammation and subsequent pulmonary fibrosis. However, it was clarified that the same effect can be obtained even at the dose used in clinical practice.

Methods: C57BL/6 mice received a single tracheal instillation of bleomycin. After randomly allocated, mice were treated with subcutaneous injection of either normal saline or somatostatin analogue.

Results: Somatostatin analogue reduced the number of neutrophils and lymphocytes in bronchoalveolar lavage (BAL) and IGF-1 level in serum and BAL fluid and attenuated weight loss. The hydroxyproline content of the lung homogenates in somatostatin analogue treatment group was significantly lower than in that of normal saline treatment group.

Conclusions: These results suggest that somatostatin analogue may attenuate pulmonary fibrosis after bleomycin treatment at the dose used in clinical practice.

Purpose: To delve into the related molecular mechanism of ACTL6A on non-small cell lung cancer (NSCLC) cell growth and apoptosis.Methods: Quantitative real-time polymerase chain reaction, immunohistochemical staining, and western blot assays were employed to examine ACTL6A mRNA and protein expression in four NSCLC cell line (NCI-H2170, LTEP-s, NCI-H1703, and PC-9) and normal lung cell line (BEAS-2B). CCK-8 cell viability assays and clone formation assay were applied to verify the cell proliferation of NCI-H2170 cell line after knockdown of ACTL6A. Flow cytometry assays were applied to check the role of ACTL6A in the apoptosis of NSCLC cells. The western blot assays were employed to examine the protein expression of WWC1, YAP, TAZ, and CYR61 in NCI-H2170 after knockdown of ACTL6A. Finally, xenograft tumor was taken out and checked the tumor volumes and weight. Immunohistochemical staining and western blot assays were employed to examine cell proliferation and apoptosis of NSCLC in vivo.Results: In this study, the results showed that the mRNA and protein expression level of ACTL6A was higher in four NSCLC cell line than normal lung cell line, respectively. Suppression of ACTL6A inhibited the growth and promoted apoptosis of NSCLC cells. Meanwhile, ACTL6A promotes tumor growth and inhibits apoptosis of NSCLC in vivo via Hippo/YAP signaling pathway.Conclusion: ACTL6A promotes the proliferation in NSCLC by regulating Hippo/YAP pathway.

Aim of the study: Obstructive sleep apnea, which is characterized by intermittent hypoxia (IH), is a common respiratory disease. The aim of the present study was to explore the relationship between hypoxia and endothelial progenitor cell (EPC) function, and explain the role of IH in endothelial repair.Materials and methods: Peripheral blood mononuclear cells (PBMCs) were isolated from a mouse model of IH. The number of CD133⁺ kinase insert domain receptor (KDR)⁺, CD133⁺CD34⁺, CD34⁺KDR⁺ and ALDH^lowCD34⁺KDR⁺ EPCs was determined by flow cytometry. HIF-1α, stromal-derived factor-1 (SDF-1) α and VEGF were measured by ELISA. The proliferative ability of PBMCs was determined. EPC migration was assessed by Transwell assay and surface proteins by western blot analysis. EPCs were co-cultured with mouse brain endothelial cells and their angiogenic ability was analyzed.Results: The number of CD133⁺KDR⁺, CD133⁺CD34⁺ and CD34⁺KDR⁺ EPCs increased with IH ingravescence. The number of ALDH^lowCD34⁺KDR⁺ EPCs with mild IH stimulation was higher and gradually decreased in the moderate and severe IH groups. The release of HIF-1α, SDF-1α and VEGF in the serum increased with the increase in the degree of IH. In the mild IH treatment, the migration and angiogenesis of EPCs, as well as the expression of vascular endothelial growth factor receptor 2 and cysteine-X-cysteine receptor 4, were higher than those in the control group, but progressively decreased in the groups with moderate and severe IH.Conclusion: Increased levels of IH accelerated the increase in vasoactive factors in peripheral blood, thereby mobilizing a large number of EPCs. Increasing of IH diminished the mobilization, chemotactic and angiogenetic ability of EPCs.

Objectives: The purpose of this study was to analyze the α-SiO₂ content, composition, dispersion, morphology, and free radical content of dust between the alveolar and the workplace, to explore the possible changes in the properties (especially the pathogenicity) of dust after it enters the lung.

Methods: We collected the dust in the workplace in HANDAN Coal mine. They were selected by a 400 mesh sieve and was made a suspension of 50 mg/ml, which would be used to perfuse into the trachea of rats. When one week, four weeks, eight weeks, fourteen weeks, twenty weeks after perfusing, we harvested dust in rats alveolar through lung lavage for further processing.

Results: In the animal test, typical fibrous nodules appeared 20 weeks after dust exposure. No inflammatory reaction was observed in the saline group. The results of animal experiments showed that there was no significant difference in the content of α-SiO₂ between dust in the workplace and the lung lavage (P > 0.05). The content of the Fe element gradually increased with dust exposure time. The 12 elements of Al, Mg, Si, Pb, Mn, Ni, Zn, Cu, Cr, Sb, Cd, and AS were reduced in the experiment group compared with the workplace group. The shape of the dust in the workplace was mostly spherical. The shape of the dust extracted from the lung lavage fluid was mostly blocky and angular, and a few dust edges were sharp, and more than 80% of the particle size was smaller than 5 μm, while less than 1% of the particle size was larger than 10 μm. The amount of hydroxyl radical released by lung lavage dust in phosphate buffer was higher than that of the workplace dust.

Conclusions: After the dust entered the alveoli, the content of α-SiO₂ in the dust did not change with dust exposure time, while the content of elements in the dust, the morphology, and dispersion of the dust changed. The ability of dust in alveoli to produce hydroxyl radicals in phosphate buffer was higher than that in the workplace.

Lung adenocarcinoma (LUAD), one of the most common cancers, is a major threat to people's health due to its high mortality, and the survival of most patients suffering LUAD remains poor. This study aimed to explore the mechanism of Deleted in Liver Cancer 1 (DLC1) as a tumor suppressor underlying the occurrence and progression of LUAD. As revealed by bioinformatics analysis and qRT-PCR, DLC1 was significantly down-regulated in LUAD tumor tissue and cells. A series of cellular experiments including CCK-8, wound healing and Transwell assays were performed to detect the effect of DLC1 on the biological function of LUAD cells. It was found that overexpressing DLC1 significantly inhibited LUAD cell proliferative, migratory and invasive abilities, while knockdown of DLC1 promoted these abilities. Gene Set Enrichment Analysis (GSEA) and dual-luciferase assay were used to explore the downstream signaling pathway of DLC1, finding that DLC1 could remarkably inhibit the activity of mitogen-activated protein kinase (MAPK) signaling pathway. Western blot implemented for MAPK signaling pathway-related proteins further identified that DLC1 restrained the activation of MAPK/ERK signaling pathway. Furthermore, rescue experiments suggested that DLC1 inhibited LUAD cell proliferation and invasion by suppressing the MAPK/ERK signaling pathway. Overall, our study discussed the DLC1-dependent mechanism involved in LUAD. We found that the up-regulation of DLC1 may inhibit the malignant progression of LUAD by suppressing MAPK signaling pathway, which supports the view that DLC1 may serve as a molecular target for the targeted therapy of LUAD patients.