Chronic Obstructive Pulmonary Disease Model Group Research Articles

Electroacupuncture improves pulmonary function by reducing inflammatory reaction via inhibiting miR-19b-3p to regulate SOCS3/JAK1/STAT3 signaling pathway in mice with chronic obstructive pulmonary disease

To explore whether electroacupuncture(EA) can increase the expression of suppressor of cytokine signaling (SOCS)3 by affecting the expression of miR-19b-3p, inhibiting the continuous activation of janus kinase (JAK)1 /signal transducer and activator of transcription (STAT)3 signaling pathway, and improve pulmonary inflammation in chronic obstructive pulmonary disease (COPD) mice. Forty mice were randomly divided into normal, COPD model, COPD+EA, and COPD+miR-19b-3p agomir (agomir)+EA groups. The COPD model was simulated by cigarette smoke exposure for 1 h, twice a day for 3 months. After modeling, mice in the COPD+EA group and the COPD+agomir+EA group received EA stimulation of "Feishu" (BL13) and "Zusanli"(ST36) for 30 min, once every other day, for 14 days. The mice of the COPD+agomir+EA group received intranasal drops of miR-19b-3p agomir solution (50 μL) 24 h before every EA intervention. The pulmonary ventilation functions including forced vital capacity (FVC), forced expiratory volume (FEV) at the 0.05 s and 0.1 s (FEV0.05 and FEV0.1), FEV0.05/FVC and FEV0.1/FVC were detected using a pulmonary function analysis system. The pathological morphology of lung tissue was observed after H.E. staining. The contents of interleukin(IL)-6, tumor necrosis factor(TNF)-α and IL-1β in the bronchoalveolar lavage fluid were assayed using ELISA. And the expressions of SOCS3, JAK1, STAT3, phosphorylated(p)-JAK1 and p-STAT3 proteins in the lung tissue were detected using Western blot. The expressions of miR-19b-3p, JAK1, STAT3 and SOCS3 mRNA were detected using real-time fluorescence quantitative PCR. Compared with the normal group, the COPD model group had a significant decrease in the levels of FVC, FEV0.05, FEV0.1, FEV0.05/FVC and FEV0.1/FVC, and the expression levels of SOCS3 mRNA and protein (P<0.001) and a significant increase in the contents of IL-6, TNF-α and IL-1β, and the expression levels of miR-19b-3p, JAK1 and STAT1 mRNAs, and p-JAK/JAK1 and p-STAT3/STAT3 protein expression ratio (P<0.001). After EA intervention, the decreased and increased levels of all the indexes mentioned above were reversed in the COPD+EA group (P<0.001, P<0.01, P<0.05). The effect of the EA+agomir were significantly less than EA in up-regulating the levels of FVC, FEV0.05, FEV0.1, FEV0.05/FVC and FEV0.1/FVC, and the expression levels of SOCS3 protein and in down-regulating the contents of IL-6, IL-1β, TNF-α, and expressions of JAK1 and STAT3 mRNA, and p-STAT3/STAT3 (P<0.001, P<0.01, P<0.05), suggesting that the effects of EA were weakened after intranasal drops of miR-19b-3p agomir. H.E. staining showed thickened alveolar wall, obvious inflammatory cell infiltration, with some ruptured alveoli fused into large vesicles in the model group, slightly dilated alveoli and small amount of inflammatory cell infiltration in the COPD+EA group, and slight alveolar fusion, slight thickening of alveolar wall, and light inflammatory cell infiltration in the COPD+agomir+EA group. EA can inhibit the expression of miR-19b-3p, thereby up-regulating SOCS3 expression and inhibiting the overactivation of JAK1/STAT3 signaling, thus reducing lung inflammatory reaction to improve pulmonary function in mice with COPD.

Inhalation of Citrus Reticulata essential oil alleviates airway inflammation and emphysema in COPD rats through regulation of macrophages

Ethnopharmacological relevanceChronic obstructive pulmonary disease (COPD) is a chronic inflammatory respiratory disease. Citrus Reticulata peel, the dried ripe peel of Citrus Reticulata species, has been found to have anti-inflammatory and cough attenuation effects. However, the therapeutic effects and its precise underlying mechanisms of atomizing inhalation using Citrus Reticulata essential oil (CREO) have not yet been fully elucidated. Aim of the studyThe aim of this study was to assess the therapeutic effects of Citrus Reticulata essential oil and its associated anti-inflammatory mechanisms in COPD rat model. MethodsA total of 80 SD rats were randomized into four groups: control group (Con), COPD model group (COPD), COPD + ipratropium bromide (IB), and COPD + citrus reticulata essential oil (CREO). To induce COPD in rats, cigarette smoke (CS) exposure was used, while CREO and IB groups were administered through atomizing inhalation. The clinical signs, pathological lesions of the lung, percentages of antigen-presenting lung macrophages (CD11b/c+/CD86+ cells) and CD8+ T cells, and the content and mRNA expression of cytokines of the lung were analyzed. ResultsThe findings revealed that atomizing inhalation of Citrus reticulata essential oil had therapeutic effects on COPD rats. The treatment resulted in improvement in the body weight and mental status of COPD rats, reduced pathological injury of the lung, and increased proportion of CD11b/c+/CD86+ cells in lung macrophages, while also decreasing the number of CD8+ T cells. In addition, the Citrus Reticulata essential oil reduced the contents of IL-18, IL-17A, IL-12p70, and GM-CSF, downregulated the relative mRNA expression of IFN-γ, IL-4, and MMP-12, and upregulated the mRNA expression of IL-10. ConclusionsCitrus reticulata essential oil can alleviate histological injury of the lung and regulate macrophages and CD8+ T cells in COPD rats. The study suggests that citrus reticulata essential oil could be a potential therapeutic agent for COPD.

Baicalin Inhibits Inflammation in Rats with Chronic Obstructive Pulmonary Disease by the TLR2/MYD88/NF-κBp65 Signaling Pathway.

Objective Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory disease with a relatively high morbidity and death rate. This study aimed to investigate the inhibitory effect of baicalin (BA) on inflammation in COPD rats and its possible mechanism. Methods The experimental COPD of SD rats were induced by LPS, smoking, and cold stimulation, and they were randomly divided into the control group, COPD group, COPD + LB group, COPD + MB group, and COPD + HB group. The test of pulmonary function and the HE staining were carried out in COPD rats. The levels of TNF-α, IL-1β, IL-6, IL-10, and IL-8, as well as GSH, SOD, and MDA in serum, were detected by ELISA. The levels of TLR2, MYD88, TNF-α, and IL-1β mRNA in BALF were detected by qPCR. The expression of TLR2/MYD88/NF-κBp65 pathway-related proteins was also detected by the Western blot and immunohistochemistry assays. Results Compared to the COPD model group, BA treatment significantly improved the pulmonary function and pathologic changes, reduced the levels of TNF-α, IL-1β, IL-6, IL-10, IL-8, and MDA, and increased the levels of IL-10, SOD, and GSH in COPD rats. In addition, BA could also decrease the protein levels of MYD88, p–NF–κBp65/NF-κBp65, TLR2, and TLR4 but increase the protein level of p-IκBa/IκB in lung tissue of COPD rats. Conclusion BA ameliorated inflammatory response and oxidative stress in COPD rats by regulating the TLR2/MYD88/NF-κBp65 signaling pathway.

Effect of electroacupuncture on pulmonary neuroendocrine cells and secretion of neuroactive substances in lung of COPD rats

To observe the effect of electroacupuncture (EA) at "Zusanli" (ST36) and "Feishu" (BL13) on the activation and secretion of calcitonin gene-related peptide (CGRP) and 5-hydroxytryptamine (5-HT) of pulmonary neuroendocrine cells (PNECs) and inflammatory response in rats with chronic obstructive pulmonary disease (COPD), so as to explore its underlying mechanisms in treating COPD. Male SD rats were randomly divided into normal control, COPD model and EA groups, with 7 rats in each group. The COPD model was established by forced inhale of cigarette smoke for 1 h in a self-made box (1 m×1 m×1 m in volume), twice daily for 12 weeks. EA (4 Hz/20 Hz, 1-3 mA) was applied at bilateral ST36 and BL13 acupoints for 30 min, once a day for 14 consecutive days. The pulmonary function including the forced vital capacity (FVC), forced expiratory volume at 0.1 second (FEV0.1), FEV0.3, FEV0.1/FVC and FEV0.3/FVC was detected using a lung function analyzer for small animals. The lung tissue was sampled for observing histopathological changes by using H.E. staining, for observing expression and distribution of PNECs by Grimelius silver staining, and for detecting the immunoactivity (integrated optical density) of CGRP and 5-HT by using immunohistochemistry. The contents of CGRP, 5-HT, tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and transforming growth factor-β1 (TGF-β1) in the bronchoalveolar lavage fluid (BALF) and lung tissue were detected by ELISA, and the correlations between TNF-α and CGRP, IL-1β and CGRP, TNF-α and 5-HT, and IL-1β and 5-HT levels were analyzed. The mRNA and protein expression levels of nerve fiber markers of CGRP and purinergic receptor P2X ligand gated ion channel 3 (P2X3) which dominate PNECs in the lung tissue were detected by real-time fluorescence quantitative PCR and Western blot, respectively. Compared with the normal control group, the levels of FVC, FEV0.1, FEV0.3, and the ratios of FEV0.1/FVC and FEV0.3/FVC were significantly decreased (P<0.05, P<0.01), while the immunoactivity of PNECs, CGRP and 5-HT, the contents of CGRP, 5-HT, TNF-α, IL-1β and TGF-β1 in the BALF and lung tissue, and the expression levels of CGRP and P2X3 mRNAs and proteins in the lung tissue significantly increased in the COPD model group (P<0.01, P<0.05). Following EA intervention, both the increased and decreased levels of all the indexes mentioned above were reversed (P<0.05, P<0.01) except FEV0.3. H.E. staining showed severe deformed bronchial lumen with thickened wall and alveolar septum, and obvious inflammatory cell infiltration and reduced number of alveolar lumen fusion in the COPD model group, which was mild in the EA group. A positive correlation was found between TNF-α and CGRP, IL-1β and CGRP, TNF-α and 5-HT,IL-1β and 5-HT levels in both BALF and lung tissues (P<0.01). EA at ST36 and BL13 can improve lung function and reduce inflammatory response in COPD rats, which may be related to its function in inhibiting the activation of PNECs and release of neuroactive substances.

Effects of Maxingloushi decoction on immune inflammation and programmed death markers in mice with chronic obstructive pulmonary disease.

To investigate effects of Maxingloushi decoction on lung inflammation and programmed death markers (programmed death-1 [PD-1], programmed death-ligand 1 [PD-L1]) in the lung tissue, peripheral blood, and bronchoalveolar lavage fluid (BLF) in a mouse model of chronic obstructive pulmonary disease (COPD). Thirty-six mature male BALB/C mice were randomly divided into normal group (group A, n=6), COPD model group (group B, n=10), Maxingloushi decoction + COPD group (group C, n=10), and PD-1 inhibitor + COPD group (group D, n=10). The COPD model was established by smoke inhalation combined with lipopolysaccharide (LPS). Levels of PD-1 and PD-L1 in plasma and BLF were measured by enzyme-linked immunosorbent assay (ELISA). Histopathological techniques were used to semi-quantitatively analyze the immuno-fluorescence optical density (IOD) value of the lung tissue. In plasma and BLF, the expression of PD-1 in the group B was higher than that in the group A, and the expression of PD-L1 was lower than that in the group A. The expression of PD-1 and PD-L1 in the lung tissue was normalized in the group C in comparison with the group B (P<0.05) and the group D (P<0.05), and inflammatory cell infiltration in the lung tissue was also improved. These findings reveal that COPD causes an immune imbalance in the peripheral blood and lung tissue, and that both Maxingloushi decoction and PD-1 inhibitor treatment can mitigate lung inflammation in COPD by reducing PD-1 expression and increasing PD-L1 expression. The treatment effect of Maxingloushi decoction may be superior to that of PD-1 inhibitor.

Magnesium isoglycyrrhizinate inhibits airway inflammation in rats with chronic obstructive pulmonary disease

BackgroundChronic obstructive pulmonary disease (COPD) is a kind of chronic lung diseases with the characteristics of airway remodeling and airflow obstruction. Magnesium isoglycyrrhizinate (MgIG) is an anti-inflammatory glycyrrhizic acid preparation for treating hepatitis. However, whether MgIG can treat other diseases and its action mechanism is still obscure. In this study, we evaluated the anti-inflammatory effect of MgIG in rats with COPD and investigated the underlying mechanisms.MethodsRat model of COPD was constructed by endotracheal-atomized lipopolysaccharide exposure and cigarette smoke induction. Rats were randomly divided into 5 groups: control group, COPD model group, salmeterol fluticasone comparator group, low dose of MgIG group, and high dose of MgIG group. Except for normal control group, the other four groups received sensitization treatment by cigarette smoking and endotracheal-atomization of endotoxin lipopolysaccharide to construct COPD rats model. After model established successfully, the COPD rats in each group received corresponding dose of endotracheal-atomized normal saline, salmeterol fluticasone, and MgIG every day prior to exposure of cigarette smoke from days 30 to 45. Normal control group were treated with normal saline. Finally, All rats were euthanatized. Pulmonary function was measured. Cells in bronchoalveolar lavage fluid were classified, inflammatory factors IL-6 and TNF-α were determined, histopathological analysis was performed by HE staining, and expression of NLRP3 and cleaved caspase-1 in the lung tissue was also determined by Western blotting.ResultsIt showed that MgIG treatment (0.40 or 0.80 mg/kg/day) could recover the weight and the clinical symptoms of rats with COPD, accompanied with lung inflammation infiltration reduction, airway wall attenuation, bronchial mucus secretion reduction. Additionally, MgIG administration reduced inflammatory cells (white blood cells, neutrophils, lymphocytes and monocytes) accumulation in bronchoalveolar lavage fluid and decreased IL-6 and TNF-α production in the serum of COPD rats. Furthermore, MgIG treatment also reduced the expression level of NLRP3 and cleaved caspase-1.ConclusionIt indicate that MgIG might be an alternative for COPD treatment, and its mechanism of action might be related to the suppression of NLRP3 inflammasome.

Muscle metabolomics analysis reveals potential biomarkers of exercise‑dependent improvement of the diaphragm function in chronic obstructive pulmonary disease.

Decreased diaphragm function is a crucial factor leading to reduced ventilatory efficiency and worsening of quality of life in chronic obstructive pulmonary disease (COPD). Exercise training has been demonstrated to effectively improve the function of the diaphragm. However, the mechanism of this process has not been identified. The emergence of metabolomics has allowed the exploration of new ideas. The present study aimed to analyze the potential biomarkers of exercise-dependent enhancement of diaphragm function in COPD using metabolomics. Sprague Dawley rats were divided into three groups: COPD + exercise group (CEG); COPD model group (CMG); and control group (CG). The first two groups were exposed to cigarette smoke for 16 weeks to establish a COPD model. Then, the rats in the CEG underwent aerobic exercise training for 9 weeks. Following confirmation that exercise effectively improved the diaphragm function, a gas chromatography tandem time-of-flight mass spectrometry analysis system was used to detect the differential metabolites and associated pathways in the diaphragm muscles of the different groups. Following exercise intervention, the pulmonary function and diaphragm contractility of the CEG rats were significantly improved compared with those of the CMG rats. A total of 36 different metabolites were identified in the comparison between the CMG and the CG. Pathway enrichment analysis indicated that these different metabolites were involved in 17 pathways. A total of 29 different metabolites were identified in the comparison between the CMG and the CEG, which are involved in 14 pathways. Candidate biomarkers were selected, and the pathways analysis of these metabolites demonstrated that 2 types of metabolic pathways, the nicotinic acid and nicotinamide metabolism and arginine and proline metabolism pathways, were associated with exercise-induced pulmonary rehabilitation.

Effects of dexmedetomidine on oxidative stress and inflammatory response in lungs during mechanical ventilation in COPD rats.

Effects of dexmedetomidine (Dex) on oxidative stress and inflammatory response in lungs during mechanical ventilation in chronic obstructive pulmonary disease (COPD) rats were investigated. Eleven out of 38 SD rats were randomly selected as the blank control group, and the other 27 rats were subjected to modeling. After the modeling, 11 rats in the blank control group and 11 rats randomly selected from the model group received non-invasive test for lung function. Three rats from the blank control group and 3 rats from the model group were selected for hematoxylin and eosin (HE) staining to confirm successful modeling, and the other 24 rats were randomly divided into 3 groups, 8 rats in each group, including model control, Dex low-dose and Dex high-dose group. A COPD rat model was established by passive cigarette smoking and intratracheal instillation of lipopolysaccharide. Each group underwent mechanical ventilation for 2 h. The Dex low-dose group and Dex high-dose group were intravenously administered at 1.0 µg/kg/h and 5.0 µg/kg/h of Dex, and the other two groups received intravenous drip of the same amount of normal saline. Blood gas analysis was performed to calculate carbon dioxide partial pressure (PaCO2), oxygen partial pressure (PaO2) and blood pH. HE staining was performed to analyze pulmonary pathological features of COPD rat model. Serum inflammatory factors interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-α) and malondialdehyde (MDA) were detected by ELISA, and the levels of antioxidant enzymes superoxide dismutase 2 (SOD2) and catalase were analyzed by western blot analysis. After 28 days of modeling, TV, PEF, FEV0.3 and FEV0.3/FVC decreased significantly in the COPD model group. HE staining showed that in the model group, the alveolar cells became larger, the alveolar wall became thinner, and some alveolar walls were even broken. The lung lobule showed obvious cell degeneration, necrosis and shedding, and the interstitial inflammatory cell infiltration, suggesting that the COPD rat model was successfully established. After 2 h of mechanical ventilation and Dex intravenous infusion, PaCO2 decreased, PaO2 increased, and blood pH value increased (p<0.05). Inflammatory factors IL-8 and TNF-α decreased (p<0.05). Oxidative stress index MDA also decreased (p<0.05), antioxidant enzymes SOD2 and catalase increased (p<0.05). Dexmedetomidine can improve the oxidative stress response during mechanical ventilation in rats with COPD, and can reduce the inflammation of lung tissue, thus protecting the lung tissue of COPD rats.

Berberine Attenuates Cigarette Smoke Extract-induced Airway Inflammation in Mice: Involvement of TGF-β1/Smads Signaling Pathway.

Although several studies confirmed that berberine may attenuate airway inflammation in mice with chronic obstructive pulmonary disease (COPD), its underlying mechanisms were not clear until now. We aimed to establish an experiment mouse model for COPD and to investigate the effects of berberine on airway inflammation and its possible mechanism in COPD model mice induced by cigarette smoke extract (CSE). Twenty SPF C57BL/6 mice were randomly divided into PBS control group, COPD model group, low-dose berberine group and high-dose berberine group, 5 mice in each group. The neutrophils and macrophages were examined by Wright's staining. The levels of inflammatory cytokines TNF-α and IL-6 in bronchoalveolar lavage fluid (BALF) were determined by enzyme-linked immunosorbent assay. The expression levels of TGF-β1, Smad2 and Smad3 mRNA and proteins in lung tissues were respectively detected by quantitative real-time polymerase chain reaction and Western blotting. It was found that CSE increased the number of inflammation cells in BALF, elevated lung inflammation scores, and enhanced the TGF-β1/Smads signaling activity in mice. High-dose berberine restrained the alterations in the COPD mice induced by CSE. It was concluded that high-dose berberine ameliorated CSE-induced airway inflammation in COPD mice. TGF-β1/Smads signaling pathway might be involved in the mechanism. These findings suggested a therapeutic potential of high-dose berberine on the CSE-induced airway inflammation.

Allyl isothiocyanate may reverse the expression of MRP1 in COPD rats via the Notch1 signaling pathway.

In the present study, the roles of AITC in up-regulating the MRP1 expression and its relationship with the activation of the Notch1 signaling pathway were investigated by combining the in vivo and in vitro experiments. AITC was administered to the COPD model rats and normal rats to explore the association between Notch1 and MRP1. The human bronchial epithelial cells were treated with DAPT, the Notch1 signaling pathway inhibitor, to verify the effect of Notch1 on the expression of AITC-induced MRP1. Compared with the control group, the expressions of Notch1, Hes1 (the target gene of Notch1) and MRP1 in the lung tissue of the COPD model group were significantly inhibited. In contrast to the COPD model group, the expressions of MRP1, Hes1 and Notch1 dramatically up-regulated following the treatment with Low/High doses of AITC. The expression of MRP1 in the 16 HBE cells was down-regulated by the inhibition of Notch in a DAPT concentration-dependent manner. Additionally, the AITC-induced up-regulation of the MRP1 expression was markedly impaired following the inhibition of Notch1. The above results indicated that the pulmonary function and the expression of MRP1 in COPD rats could be improved by AITC, which was partly dependent on the Notch1 signaling pathway. Therefore, targeting the Notch signaling pathway may present as an effective therapeutic strategy for COPD treatment.

Carbocisteine inhibits the expression of Muc5b in COPD mouse model.

BackgroundCigarette smoke (CS) results in chronic mucus hypersecretion and airway inflammation, contributing to COPD pathogenesis. Mucin 5B (MUC5B) and mucin 5 AC (MUC5AC) are major mucins implicated in COPD pathogenesis. Carbocisteine can reduce mucus viscosity and elasticity. Although carbocisteine decreased human elastase-induced MUC5AC expression in vitro and reduced MUC5AC expression that alleviated bacteria adhesion and improved mucus clearance in vivo, the roles of carbocisteine in inducing MUC5B expression in COPD remain unclear.MethodsTo investigate the Muc5b/Muc5ac ratio and the gene and protein levels of Muc5b in COPD and carbocisteine intervention models. C57B6J mice were used to develop COPD model by instilling intratracheally with lipopolysaccharide on days 1 and 14 and were exposed to CS for 2 hr twice a day for 12 weeks. Low and high doses of carbocisteine 112.5 and 225 mg/kg/d, respectively, given by gavage administration were applied for the treatment in COPD models for the same duration, and carboxymethylcellulose was used as control. Carbocisteine significantly attenuated inflammation in bronchoalveolar lavage fluid and pulmonary tissue, improved pulmonary function and protected against emphysema.ResultsHigh-dose carbocisteine significantly decreased the overproduction of Muc5b (P<0.01) and Muc5ac (P<0.001), and restored Muc5b/Muc5ac ratio in COPD model group (P<0.001). Moreover, the Muc5b/Muc5ac ratio negatively correlated with pro-inflammatory cytokines such as IL-6 and keratinocyte-derived cytokine, mean linear intercept, functional residual capacity and airway resistance, but positively correlated with dynamic compliance.ConclusionsThese findings suggest that carbocisteine attenuated Muc5b and Muc5ac secretion and restored Muc5b protein levels, which may improve mucus clearance in COPD.

Effects of neuromuscular electrical stimulation on muscle atrophy and muscle fiber composition in quadriceps muscles of chronic obstructive pulmonary disease rats

Objective To investigate the effect of neuromuscular electrical stimulation(NMES)on muscle atrophy and muscle fiber type conversion in quadriceps muscles of rats with chronic obstructive pulmonary disease(COPD). Methods Forty-eight male SPF Wistar rats were randomly divided into three groups: the healthy control group, COPD model group and NMES treatment group, with sixteen rats in each group.Rats in the NMES treatment group were given alternating electrical stimulation at 100 Hz and 2 Hz, with each stimulation time of 30 min, 7 days per week, for 4 weeks.The control group and the COPD model group were only treated with electrode sheets.After the intervention, the endurance running distance measurement, the pathological changes of rat quadriceps muscle detected by HE staining, the mRNA and protein content of MSTN, MyHC(Myosin Heavy Chain)-Ⅰ, MyHC-Ⅱa and MyHC-Ⅱx in quadriceps muscles determined by Realtime PCR and Western Blot were performed. Results Compared with the healthy control group, the cross-sectional area, endurance running distance and MyHC-Ⅰ mRNA and protein expressions in the quadriceps muscles were decreased[(654.7±53.7) μm2vs.(1079.8±117.2) μm2, (396.7±42.4) m vs.(607.4±56.3) m, 0.407±0.054 vs.0.997±0.069, 0.884±0.102 vs.1.723±0.156, t=4.642, 3.785, 24.723 and 18.640, P=0.008, 0.033, 0.000 and 0.000, respectively], while MSTN, MyHC-Ⅱa and MyHC-Ⅱx mRNA and protein expressions were increased in the COPD model group(all P<0.01). Compared with COPD model, the cross-sectional area(997.5±92.4 μm2), endurance running distance(597.2±65.8 m)and the expression levels of MyHC-ⅠmRNA(0.884±0.097)and protein(1.534±0.182)in the quadriceps muscles were increased(t=3.734, 3.602, 19.988 and 13.666, P=0.035, 0.043, 0.000, 0.000), and the mRNA and protein expression levels of MSTN, MyHC-Ⅱa and MyHC-Ⅱx were decreased in the NMES treatment group(all P<0.01). Further correlation analysis found that the protein expression of MSTN in the quadriceps muscles was negatively correlated with muscle cross-sectional area, MyHC-Ⅰprotein levels and endurance running distance(r=-0.724, -0.426 and -0.536, P=0.002, 0.036 and 0.007, respectively), and positively correlated with the protein expressions of MyHC-Ⅱa and MyHC-Ⅱx(r=0.408 and 0.392, P=0.042 and 0.048). Conclusions The expression of MSTN is increased in quadriceps muscle of COPD rats, and NMES can inhibit the expression of MSTN in quadriceps muscles.MSTN is associated with muscle atrophy and muscle fiber conversion in COPD.Inhibition of MSTN expression can improve muscle atrophy and reverse muscle fiber transformation. Key words: Pulmonary disease, chronic obstructive; Electric stimulation therapy; Muscular disorders, atrophic

The anti-inflammatory effect of BML-111 on COPD may be mediated by regulating NLRP3 inflammasome activation and ROS production

The aim of this study is to investigate whether the lipoxin receptor agonist BML-111 exerts a protective effect against inflammation in a mouse model of chronic obstructive pulmonary disease (COPD) by regulating NLRP3 inflammasome activation and reactive oxygen species (ROS) production. In this study, mice were randomly divided into the following five groups: control group (Control), COPD model group (Model), BML-111 low-dose group (Low-BML), BML-111 high-dose group (High-BML) and Dexamethasone group (Dex). NLRP3 involvement and oxidative stress were evaluated. Differential cell counts in the BALF were calculated to obtain a reliable enumeration of each cell type, and the levels of TGF-β, TNF-α, IL-1β, and IL-10 in BALF were evaluated using ELISA. We found that the white blood cell and lymphocyte numbers in the BALF were significantly lower in the High-BML group than in the Model group. ELISA of the BALF showed that BML-111 reduced TGF-β and IL-1β levels to some extent. HE staining showed various degrees of reduction in inflammatory cell infiltration in the bronchopulmonary tissue and blood vessels of the Low-BML, High-BML and Dex groups. Measurement of oxidative stress showed that SOD activity was significantly upregulated and that the increase in MDA content was prevented in the High-BML and Dex groups. According to the Western blotting analysis, the levels of NLRP3, Cleaved-IL-1β and Cleaved-caspase-1 were decreased and Nrf-2 was increased to various extents in the Low-BML, High-BML and Dex groups. Based on these findings, BML-111 may prevent NLRP3 inflammasome activation and inhibit ROS production via upregulation of Nrf-2, thereby exerting an anti-inflammatory effect on COPD model mice.

The Function of Ophiocordyceps sinensis in Airway Epithelial Cell Senescence in a Rat COPD Model.

Ophiocordyceps sinensis (O. sinensis) seems to be able to alleviate airway epithelial cell senescence in chronic obstructive pulmonary disease (COPD). The objective of the study is to evaluate the effect of O. sinensis on airway epithelial senescence in the COPD model both in vitro and in vivo. We observed the expression of P16 and P21 in the airway epithelia of 30 patients with COPD. The optimal concentration of O. sinensis and exposure time of the cigarette smoke extract (CSE) were determined in vitro, and senescence-associated β-galactosidase (SA-β-gal) and 5-bromodeoxyuridine (BrdU) were used to evaluate the senescence and proliferation of human bronchial epithelial (16HBE) cells pretreated with O. sinensis by staining kits. COPD model rats were treated with O. sinensis at various concentrations to determine the changes in P16 and P21 expression in airway epithelial tissues. It was found that the expression levels of P16 and P21 were higher in the airway epithelia of COPD patients than those in the control group based on immunohistochemical staining, real-time quantitative PCR, and western blotting. The CSE could induce 16HBE cell senescence, and O. sinensis could alleviate CSE-induced senescence and promote the proliferation of 16HBE cells. The expression levels of P16 and P21 were also higher in the airway epithelia of COPD model rats; however, the levels of P16 and P21 in the groups treated with all concentrations of O. sinensis were obviously lower than those in the COPD model group based on real-time quantitative PCR and western blotting. In conclusion, the CSE can induce airway epithelium senescence, and O. sinensis can inhibit CSE-induced cellular senescence, both in vitro and in vivo.

MicroRNA-21 aggravates chronic obstructive pulmonary disease by promoting autophagy

ABSTRACTMicroRNAs and autophagy play important roles in chronic obstructive pulmonary disease (COPD). This study was designed to explore the role of microRNA-21 (miR-21) induced autophagy in COPD. Using the C57BL/6, miR-21−/− mice and human bronchial epithelial (16HBE) cell line, we found that in the lung tissues of mice, the level of autophagy in the COPD model group was significantly higher than that in the control group. However, compared to the COPD model, the level of autophagy was significantly lower in the miR-21−/− CSE+CS group. In the COPD model, miR-21 was overexpressed. Moreover, in human bronchial epithelial (16HBE) cells exposed to cigarette smoke extract (CSE), miR-21 expression was upregulated and autophagy was notably increased. In addition, pretreatment of 16HBE cells with miR-21 inhibitor significantly inhibited autophagy activity and decreased apoptosis, indicating that miR-21 is involved in CSE-induced autophagy and apoptosis. The results showed that miR-21 could increase autophagy and promote the apoptosis of 16HBE cells in COPD. This information contributes to our further understanding of COPD.

Effects of 1,25-Dihydroxyvitamin D3 on the Prevention of Chronic Obstructive Pulmonary Disease (COPD) in Rats Exposed to Air Pollutant Particles Less than 2.5 Micrometers in Diameter (PM2.5).

BackgroundThis study aimed to investigate the effects of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) on airway changes in chronic obstructive pulmonary disease (COPD) rats exposed to air pollutant particles less than 2.5 micrometers in diameter (PM2.5), and to evaluate the mechanisms.Material/MethodsThree groups were included in this study: a normal group, a COPD model group, and a COPD with 1,25(OH)2D3 treatment group. In each group, the rats were divided into four subgroups: control and different doses of PM2.5 (1.6, 8 and 40 mg/kg body weight). Apoptosis in lung tissue was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL). The expression of c-Jun N-terminal kinase 1 (JNK1) and mucin 5AC (MUC5AC) were detected by real-time polymerase chain reaction (RT-PCR), Western blotting and immunofluorescence staining.ResultsCompared with corresponding subgroups in normal group, the apoptotic rates in COPD group were significantly increased. By contrast, 1,25(OH)2D3 treatment group significantly reduced COPD-induced apoptosis in lung tissue. Upon the dose increase of PM2.5, the apoptotic rate was also elevated in each group. Compared with the corresponding control in each group, PM2.5 increased apoptosis in a dose-dependent manner. Importantly, 1,25(OH)2D3 also prevented apoptosis in COPD rats exposed to PM2.5. Mechanically, the expression of MUC5AC and JNK1 in COPD group was significantly upregulated, compared with corresponding subgroups in the normal group. Treatment with 1,25(OH)2D3 reduced expression of MUC5AC and JNK1 in COPD rats. It was found that the expression of MUC5AC and JNK1 was elevated with the dose increase of PM2.5 in each group. Consistently, 1,25(OH)2D3 also reduced the expression of MUC5AC and JNK1 in COPD rats exposed to PM2.5.Conclusions1,25(OH)2D3 prevented lung injury in COPD rats with or without PM2.5 exposure. Our results suggest that 1,25(OH)2D3 is useful to mitigate the injury caused by COPD.

Tumor necrosis factor-like weak inducer of apoptosis regulates quadriceps muscle atrophy and fiber-type alteration in a rat model of chronic obstructive pulmonary disease

BackgroundIn chronic obstructive pulmonary disease (COPD), weakness and muscle mass loss of the quadriceps muscle has been demonstrated to predict survival and mortality rates of patients. Tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK), as a member of the TNF superfamily, has recently been identified as a key regulator of skeletal muscle wasting and metabolic dysfunction. So our aim was to study the role of TWEAK during quadriceps muscle atrophy and fiber-type transformation in COPD model rats and its possible pathway.MethodsForty-four healthy male adult Wistar rats were randomly divided into two groups: A normal control group (n = 16) and a COPD model group (n = 28). The COPD group was exposed to cigarette smoke for 90 d and injected with porcine pancreatic elastase on day 15, whereas the control group was injected with saline alone. Following treatment, weights of the quadriceps muscles were measured and hematoxylin and eosin staining was performed to identify structural changes in lung and quadriceps muscle tissue. Immunohistochemical staining was also conducted to determine the localization of TWEAK, nuclear factor (NF)-κB, muscle ring finger (MuRF)-1 and proliferator-activated coactivator (PGC)-1a proteins in the quadriceps muscle, and western blotting was used to assess the level of protein expression.ResultsCompared with controls, COPD model rats exhibited significantly lower quadriceps muscle weight (P < 0.05) accompanied by fiber atrophy and disordered fiber arrangement, a wide gap between adjacent muscle fibers, a significant reduction in nuclear number (P < 0.05) and an uneven size distribution. The proportion of fiber types was also significantly altered (P < 0.05). In addition, TWEAK expression in the quadriceps muscle of COPD model rats was significantly higher than that in control rats (P < 0.05), and was significantly associated with quadriceps atrophy and fiber-type alteration (P < 0.05). Levels of NF-κB, MuRF1 and PGC-1α expression also significantly differed between the two groups (P < 0.05).ConclusionsCollectively these data suggest that increased levels of TWEAK may lead to skeletal muscle atrophy and fiber-type alteration, which in turn may be associated with activation of the ubiquitin-proteasome pathway, involving NF-κB, MuRF1 and PGC-1α as potential regulatory factors. These preliminary results in rats suggest that TWEAK may be a therapeutic target for the treatment of muscle atrophy in COPD.

Effects of mTOR-Cdc42 signaling pathway on phagocytosis of alveolar macrophages in chronic obstructive pulmonary disease mice

Objective: To investigate effects of mammalian target of rapamycin (mTOR)- cell division cycle 42 (Cdc42) signaling pathway on phagocytosis of alveolar macrophages (AMs) in chronic obstructive pulmonary disease (COPD) mice. Methods: Forty mice were randomly divided into control group and model group. Each group contained 20 mice. COPD model group were established by cigarette smoking exposure. AMs were isolated from lung tissue by discontinuous density gradient centrifugation. AMs from control group were divided into health control group and rapamycin control group while AMs from model group were divided into COPD group and rapamycin COPD group. The AMs from rapamycin control group and rapamycin COPD group were incubated with a final concentration of 10 nmol/L rapamycin for 24 hours. Mean fluorescence intensity (MFI) and the positive percent of alveolar macrophage engulfed flurescein isothiocyanate-labeled Escherichina coli (FITC-E.coli) AM (AM%) were detected by flow cytometry. Real time PCR (RT-PCR) and Western blot were applied to detect mRNA and protein. The activity of Cdc42 was detected by G-LISA Small GTPase Activation Assays (G-LISA) Cdc42 Kit. The cytoskeleton structure of AM was observed by laser scanning confocal microscopy. Results: MFI and AM% in COPD group were decreased than those in health control group[4 060±590 vs 9 190±988 and (28.65±1.26)% vs (67.50±4.56)%]; Compared with COPD group, MFI and AM% in rapamycin COPD group[4 856±762, (38.31±1.71)%]were increased (all P<0.05). The expression of mRNA, protein and activity of mTOR in COPD group were increased than those in health control group[(2.62±0.46, 1.30±0.52, 1.46±0.43) vs (1.00±0.00, 0.48±0.27, 0.58±0.26)]; compared with COPD group, the expression of mRNA, protein and activity of mTOR in rapamycin COPD group (1.40±0.36, 0.90±0.66, 0.92±0.28) were decreased (all P<0.05). The Cdc42 mRNA, protein and activity in COPD group were higher than those in health control group[(2.56±0.50, 1.61±0.37, 0.46±0.09) vs (1.00±0.00, 0.67±0.22, 0.30±0.07)](all P<0.01); compared with COPD group, the expression of mRNA, protein and activity of Cdc42 in rapamycin COPD group (1.38±0.34, 0.91±0.48, 0.36±0.06) were decreased (all P<0.01). Filopodia protruding can not be seen in the cytoskeleton of AMs from health control group and rapamycin control group; some filopodia protruding can be seen in AM from COPD group; some long filopodia protruding can be seen in AM from rapamycin COPD group. Negative correlations were existed between the mRNA, protein and activity of mTOR, Cdc42 and MFI in all group. Conclusions: mTOR-Cdc42 signaling pathway is activated and related to phagocytosis deficiency of AM in COPD. It can be inferred that the pathway is involved in the pathogenesis of COPD.

Effect of Radix Stemonae concentrated decoction on the lung tissue pathology and inflammatory mediators in COPD rats.

BackgroundChronic obstructive pulmonary disease (COPD) is a common and frequently occurring respiratory disease. At present, western medicine treatment of COPD mainly focuses on symptomatic treatment. Using Chinese medicines or integrated Chinese and Western medicines to treat stable COPD has significant efficacy. In this study, we aimed to observe the effect of Radix Stemonae concentrated decoction on the lung tissue pathology and inflammatory mediators in COPD rats and explore its possible mechanism.MethodsSD rats were randomized into blank group, COPD model group and Radix Stemonae group, 10 cases in each group. Rats were fed for 112 days. Before the rats were sacrificed, lung function of the animals was tested. The right lower lung was fixed for morphologic observation. The inflammatory mediators in serum were determined using enzyme-linked immuno sorbent assay.ResultsBody weight of animals in the model group was significantly decreased compared with blank group (P < 0.05). After gavage therapy with Radix Stemonae, body weight was significantly increased (P < 0.05). Compared with the blank group, pulmonary functions of rats in the model group were significantly abnormal (P < 0.05), while in Radix Stemonae group, these indicators turned much better than model group (P < 0.05). As for pathological changes in lungs, airway inflammation in the model group was aggravated. In the Radix Stemonae group, inflammation and emphysema were much milder. The concentrations of TNF-α, IL-8 and LTB4 in both model group and Radix Stemonae group were increased significantly (P < 0.05). But the levels in Radix Stemonae group were decreased significantly than model group (P < 0.05).ConclusionRadix Stemonae concentrated decoction may mitigate and improve airway rebuilding in the lungs of COPD rats by inhibiting the release of inflammatory mediators.

Curcumin ameliorates alveolar epithelial injury in a rat model of chronic obstructive pulmonary disease

AimsTo investigate the effects of curcumin on alveolar epithelial injury in a rat model of chronic obstructive pulmonary disease (COPD) and its potential mechanism. Main methodsThe rat COPD model was established by cigarette smoke exposure combined with intratracheal administration of lipopolysaccharide. Thirty-eight male Sprague-Dawley rats were randomly divided into four groups: control, COPD model, COPD with curcumin and COPD with solvent groups. Neutrophil and macrophage infiltration in bronchoalveolar lavage fluid (BALF) was evaluated, and the levels of IL-6, IL-8 and TNF-α in BALF and serum were determined by ELISA. Histopathological examination and TUNEL staining were used to assess the alveolar epithelial injury. The protein expression of p66Shc and p-p66Shc in the lung tissues was determined by immunohistochemistry and western blot. Key findingsCurcumin significantly decreased the numbers of total cells, neutrophils and macrophages in BALF from COPD rats. In addition, the levels of IL-6, IL-8 and TNF-α in BALF and serum of COPD rats were significantly decreased after treatment with curcumin. Moreover, curcumin ameliorated emphysema and ultrastructural damage of alveolar epithelial cells in COPD rats. The apoptosis index of alveolar epithelial cells in the COPD with curcumin group was significantly lower than that in the COPD model group. Furthermore, the protein expression of p66Shc and p-p66Shc in alveolar epithelia was significantly decreased in the COPD with curcumin group compared with COPD model group. SignificanceCurcumin attenuates alveolar epithelial injury in COPD rats, which may be partially due to the down-regulation of p66Shc.