Lung Tissue Protein Research Articles

Anti-apoptosis effect of heme oxygenase-1 on lung injury after cardiopulmonary bypass.

BackgroundThis study aimed to investigate the anti-apoptosis effects of heme oxygenase-1 (HO-1) on lung injury (LI) after cardiopulmonary bypass (CPB) and its probable mechanisms.MethodsOne hundred and forty-four male Wistar rats were divided into 3 groups randomly: group A (control group), group B (cobalt protoporphyrin, CoPP), and group C [CoPP plus zinc protoporphyrin (ZnPP)]. Lung tissues were harvested at different time: before CPB (T0), 0 min after CPB (T1), 2 h after CPB (T2), 6 h (T3), 12 h (T4), and 24 h (T5).ResultsThe HO-1 protein expressions in lung tissue in group B were higher than those in group A and group C in any given time, and the same as HO-1 activity (P<0.05). The expressions of Bcl-2 protein in group B at all time point after bypass were higher than those in group A and group C, and the difference was statistically significant (P<0.05). Apoptosis index (AI) in group B at any time point after bypass were lower than those in group A and group C (P<0.05).ConclusionsCoPP can significantly increase the expression of HO-1 protein in lung tissue. HO-1 is still highly expressed after CPB, so as to play an important part in anti-apoptosis, and reduce LI.

Glyburide attenuates ozone-induced pulmonary inflammation and injury by blocking the NLRP3 inflammasome.

Glyburide is a classic antidiabetic drug that is dominant in inflammation regulation, but its specific role in ozone-induced lung inflammation and injury remains unclear. In order to investigate whether glyburide prevents ozone-induced pulmonary inflammation and its mechanism, C57BL/6 mice were intratracheally pre-instilled with glyburide or the vehicle 1 hour before ozone (1 ppm, 3 hours) or filtered air exposure. After 24 hours, the total inflammatory cells and total protein in bronchoalveolar lavage fluid (BALF) were detected. The pathological alternations in lung tissues were evaluated by HE staining. The expression of NLRP3, interleukin-1β (IL-1β), and IL-18 protein in lung tissues was detected by immunohistochemistry. Western blotting was used to examine the levels of caspase-1 p10 and active IL-1β protein. Levels of IL-1β and IL-18 in BALF were measured using ELISA kits. Glyburide treatment decreased the total cells in BALF, the inflammatory score, and the mean linear intercept induced by ozone in lung tissues. In addition, glyburide inhibited the expression of NLRP3, IL-18, and IL-1β protein in lung tissues, and also suppressed NLRP3 inflammasome activation, including caspase-1 p10, active IL-1β protein in lung tissues, IL-1β, and IL-18 in BALF. These results demonstrate that glyburide effectively attenuates ozone-induced pulmonary inflammation and injury via blocking the NLRP3 inflammasome.

CCR5 attenuates neutrophilic airway inflammation exacerbated by infection with rhinovirus

Human rhinovirus (hRV) is the most common cause of asthma exacerbation characterized by clinical and pathophysiological heterogeneity. Steroid-sensitive, Th2 type-eosinophilic asthma has been somewhat studied, but hRV-induced neutrophilic asthma exacerbation is poorly understood. Here, CCR5 was found to play a role in attenuating neutrophilic airway inflammation in hRV-induced asthma exacerbation using chicken ovalbumin (OVA)-based model. CCR5 deficiency resulted in exacerbated neutrophilic asthmatic responses in airways following hRV infection. CCR5-deficient mice showed enhanced mucus expression and altered expression of tight junction proteins in lung tissues. CCR5-deficient mice were also manifested with influx of CD45+CD11b+Siglec-F+Gr-1+ neutrophils, along with enhanced production of IL-17A, IFN-γ, IL-6, IL-1β cytokines in inflamed tissues. In contrast, CCR5-deficient mice elicited down-regulation of Th2-related cytokine proteins following hRV infection. More interestingly, the lack of CCR5 altered the equilibrium of CD4+FoxP3+ Tregs and IL-17+CD4+ Th17 in inflamed tissues. CCR5-deficient mice showed increased frequency and absolute number of IL-17-producing CD4+ Th17 cells in lung tissues compared to wild-type mice, whereas the reduced infiltration of CD4+FoxP3+ Treg cells was observed. CCR5 deficiency resulted in the skewed production of Th17 and Th1 cytokines in lymph nodes and lungs upon OVA stimulation. Likewise, CCR5-deficient mice showed enhanced expression of Th17-inducing cytokines (IL-1β, IL-6, and TNF-α) in lung tissues. These results imply that CCR5 deficiency facilitates Th17 airway inflammation during hRV-induced asthma exacerbation, along with suppressing Th2 responses. Furthermore, our results suggest that CCR5 attenuates hRV-induced neutrophilic airway inflammation through conserving the equilibrium of CD4+Foxp3+ Treg cells and IL-17+CD4+ Th17 cells in hRV-induced asthma exacerbation.

Anti-inflammatory mechanism of Crepis crocea based on NF-κB signaling pathway and ~1H-NMR metabonomics

Based on ~1H-NMR metabonomics technique and Western blot assay, the anti-inflammatory mechanism of Crepis crocea was discussed. In this study, male SD rats were treated with water extract(2.5 g·kg~(-1)) and dexamethasone acetate(6.25×10~(-4) g·kg~(-1)) for one week, and the inflammation model was induced by lipopolysaccharide(LPS). Then the counts of inflammatory cells white blood ceel(WBC), eosinophil(EO), lymphocyte(LY), basophils(BA) and neutrophils(NE) in whole blood of rats were observed. The levels of serum inflammatory factors tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), IL-6 and the expression of nuclear factor-κB(NF-κB) signaling pathway p65 and p-IκBα proteins in lung tissues were detected, and the change rules of serum endogenous metabolites were analyzed by ~1H-NMR metabonomics technique. The levels of TNF-α, IL-1β, IL-6 and NF-κB signaling pathway p65 and p-IκBα proteins were combined with ~1H-NMR metabonomics to study the anti-inflammatory mechanism of C. crocea. The results showed that the water extract of C. crocea significantly decreased the number of WBC, NE, EO, increased the number of BA and LY, decreased the levels of TNF-α, IL-1β, IL-6 and the expression of p65 and p-IκBα protein in NF-κB signaling pathway, and effectively alleviated the inflammatory symptoms. In the correlation analysis of differential metabolites regulated of C. crocea, four significant metabolites were obtained, including glycine, creatine, methionine and succinic acid. The anti-inflammatory mechanism of C. crocea may be related to the decrease of TNF-α, IL-1β, IL-6 levels and the protein expression of NF-κB signaling pathway, as well as the regulation of glycine, creatine, methionine and succinic acid metabolism.

Airway epithelial specific deletion of Jun-N-terminal kinase 1 attenuates pulmonary fibrosis in two independent mouse models.

The stress-induced kinase, c-Jun-N-terminal kinase 1 (JNK1) has previously been implicated in the pathogenesis of lung fibrosis. However, the exact cell type(s) wherein JNK1 exerts its pro-fibrotic role(s) remained enigmatic. Herein we demonstrate prominent activation of JNK in bronchial epithelia using the mouse models of bleomycin- or AdTGFβ1-induced fibrosis. Furthermore, in lung tissues of patients with idiopathic pulmonary fibrosis (IPF), active JNK was observed in various regions including type I and type II pneumocytes and fibroblasts. No JNK activity was observed in adjacent normal tissue or in normal control tissue. To address the role of epithelial JNK1, we ablated Jnk1 form bronchiolar and alveolar type II epithelial cells using CCSP-directed Cre recombinase-mediated ablation of LoxP-flanked Jnk1 alleles. Our results demonstrate that ablation of Jnk1 from airway epithelia resulted in a strong protection from bleomycin- or adenovirus expressing active transforming growth factor beta-1 (AdTGFβ1)-induced fibrosis. Ablation of the Jnk1 allele at a time when collagen increases were already present showed a reversal of existing increases in collagen content. Epithelial Jnk1 ablation resulted in attenuation of mesenchymal genes and proteins in lung tissue and preserved expression of epithelial genes. Collectively, these data suggest that epithelial JNK1 contributes to the pathogenesis of pulmonary fibrosis. Given the presence of active JNK in lungs from patients with IPF, targeting JNK1 in airway epithelia may represent a potential treatment strategy to combat this devastating disease.

Effects of dexmedetomidine on injury of lungs and CHOP protein expression in elderly patients with lung cancer during one-lung ventilation

Objective: To evaluate the effect of dexmedetomidine on lung injury and CCAAT/enhancer-binding protein homologous protein (CHOP) expression in elderly patients with lung cancer undergoing thoracoscopic lobectomy. Methods: A total of 120 elderly patients with lung cancer who underwent elective thoracoscopic lobectomy were selected from the Second Affiliated Hospital of Zhengzhou University from October 2018 to March 2019. The random phenotype was divided into dexmedetomidine group (group D, n=60) and control group (group C, n=60). The elapsed time before postoperative awake and spontaneous breathing recovery and agitation occurrence were recorded. The mean arterial pressure (MAP), heart rate (HR), oxygen saturation (SPO(2)), oxygen partial pressure (PaO(2)), alveolar-arterial oxygen partial pressure difference[P(A-a)O(2)], end-tidal carbon dioxide partial pressure (P(ET)CO(2)), and airway pressure peak (P(peak)) were recorded before the anesthetic induction (T(0)), immediately after tracheal intubation (T(1)), immediately after unilateral pulmonary ventilation(T(2)), 1 hour after unilateral pulmonary ventilation (T(3)), immediately after bilateral pulmonary ventilation (T(4)), and at the end of surgery(T(5)). The levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-8, IL-10, super oxide dismutase (SOD) and malondialdehyde (MDA)were compared at the time of T(0), T(1), T(2), T(3), T(4) and T(5) in two groups. The ratios of alveolar damage (IQA), apoptotic index (AI), and the expression of CHOP mRNA and their protein in lung tissue were detected. Results: The recovery time of conscious and spontaneous breathing in group D were significantly shorter than that in group C (P<0.05). The MAP and HR of the two groups at T(1)-T(5) were significantly lower than those at T(0)(P<0.05).The levels of P(peak) in group C at T(3)-T(5) was significantly higher than those in group D (P<0.05). The SpO(2) values in the group D at T(1)-T(5) were significantly higher than those in group C (P<0.05). The P(A-a)O(2) values in the group D at T(1)-T(5) was significantly lower than those in group C (P<0.05).The levels of IL-6, IL-8 and MDA in group D at T(1)-T(5) were significantly lower than those in group C(P<0.05). The levels of IL-10 and SOD in group D at T(1)-T(5) were significantly higher than those in group C (P<0.05). The values of IQA, AI, and the expression of CHOP mRNA and their protein in the two groups at T(4) were significantly higher than those at T(2) (P<0.05). The values of IQA, AI, and the expression of CHOP mRNA and their protein in group D at T(4) were significantly lower than those in group C (P<0.05). Conclusions: Dexmedetomidine could attenuate the extent of lung injury in elderly patients with lung cancer undergoing thoracoscopic lobectomy. The pulmonary protective mechanism could be related to the inhibition of inflammatory factors in dexmedetomidine, improving oxidative stress and reducing Chop-mediated apoptosis of lung tissue by the protein.

Ghrelin alleviates traumatic brain injury-induced acute lung injury through pyroptosis/NF-κB pathway

Acute lung injury (ALI) is one of the severe complications in patients with traumatic brain injury (TBI), contributing to the high mortality. Ghrelin has protective effects against various inflammatory diseases, but the effects of Ghrelin on TBI-induced ALI and its mechanisms remain unknown. In this study, Ghrelin administration was performed on the mice with TBI, then histological change in cortex and lung tissues, lung vascular permeability and macrophage number in bronchoalveolar lavage fluid (BALF) were examined, respectively. Simultaneously, the alterations of proinflammatory factors and pyroptosis-related proteins in lung tissues were detected. As a result, TBI-induced ALI was ameliorated after Ghrelin treatment, which was demonstrated by improved histology, reduced lung vascular permeability, and peripheral macrophage number. Furthermore, Ghrelin decreased the mRNA levels of proinflammatory factors (IL-1β, IL-6, TNF-α and IL-18), the protein levels of pyroptosis-related proteins (NLRP3, Caspase1-P20, HMGB1 and Gasdermin D), and the phosphorylation levels of NF-κB in lung tissues. These results showed that Ghrelin attenuating TBI-induced ALI might be via ameliorating inflammasome-induced pyroptosis by blocking NF-κB signal, which are important for the prevention and treatment of TBI-induced ALI.

Analysis of factors of pulmonary fungal infection in mice in respiratory medicine department based on logistic regression analysis model and Progranulin.

The objective of this research is to solve the current medical problems of a high incidence of fungal infections in the lungs, high misdiagnosis rate, and high mortality. In this study, firstly, the logistic regression model was used to conduct. Risk factors of pulmonary fungal infection in respiratory department were analyzed. Then a model of pulmonary fungal infection in mice was constructed, and the expression difference of Progranulin (PGRN) in serum was detected by enzyme-linked immuno sorbent assay (ELISA). The expression of PGRN in lung tissues of mice infected by pulmonary fungi was detected by Western bolt method and quantitative polymerase chain reaction (PCR). The PGRN protein and mRNA expression in the lung epithelial cells of mice were detected after the infection. Results logistic regression model was used to analyze the main risk factors affecting pulmonary infection in mice. The risk factors of pulmonary fungal infection were indent catheter, hypoproteinemia, long-term use of glucocorticoid and long-term use of antibiotics. The PGRN content in serum was obviously higher than that before pulmonary fungal infection (P < 0.01). The expression of PGRN mRNA and protein in lung tissue was obviously higher than that before infection (P < 0.01). The expression of PGRN mRNA and protein in lung tissues of the infected group was obviously higher than that of the non-infected group (P < 0.01). The expression of PGRN protein in the lung epithelial cells of mice was obviously higher at 24 h after infection than before infection (P < 0.01), and the expression of PGRN mRNA was obviously higher at 12 h after infection than before infection (P < 0.01), indicating that PGRN is highly expressed in fungal pulmonary infection and is involved in disease progression. Therefore, this study provides a new idea for the diagnosis and treatment of fungal pulmonary infection in the later stage and has a good guiding significance for the diagnosis and treatment of fungal pulmonary infection.

TLR2 Favors Allergic Airway Inflammation Through JNK Signaling Pathway with Activation of Autophagy

Background: Numerous studies indicate that TLR2 activation led to contradictory effects in asthma. In addition, the occurance of autophagy such an innate immune response in asthma pathogenesis is still incompletely understood. This study investigated the role of TLR2 and the underlying mechanisms in allergic airway inflammation (AAI) and autophagy activation. Methods: Wild-type (WT) and TLR2 knockout (TLR2-/-) mice were subjected to an ovalbumin (OVA)-immunized and airway-challenged model to investigate the role of TLR2 in AAI and autophagy activation. SP600125, a specific JNK inhibitor, was administered in mice to study its effects on AAI and autophagy activation. Findings: Our study showed that the expression of TLR2 was increased concomitantly with airway inflammation upon OVA challenge, and TLR2 deficiency was associated with a dramatical decrease in serum OVA-specific IgE, pulmonary inflammatory cells infiltration, goblet cell metaplasia and bronchoalveolar lavage fluid (BALF) inflammatory cytokine production. Meanwhile, the expression of inflammatory proteins in lung tissues, autophagy activation were induced in OVA-challenge mice, while such inductions were significantly weakened by TLR2 deficiency. Moreover, inhibition of JNK by SP600125 also down-regulated airway inflammation and autophagy activation in OVA-challenged WT mice. Interestingly, treating TLR2-/- mice with SP600125 upon OVA challenge showed similar pulmonary inflammation manifestations, lung inflammatory proteins expression as well as autophagy activation compared to that in vehicle-treated TLR2-/- mice. Interpretation: TLR2 might contribute to the maintenance of AAI through JNK signaling pathway accompanying with autophagy activation. These findings may provide a novel signal target for prevention of AAI. Funding Statement: This study was supported by the National Natural Science Foundation of China (NO. 81170030, 81270082) and Natural Science Foundation of Anhui Province (NO. 1908085QH311). Declaration of Interests: The authors declare no conflicts of interest. Ethics Approval Statement: All procedures were approved by the Animal Care and Use Committee of University of Science and Technology of China.

Down-regulation of long non-coding RNA SNHG14 protects against acute lung injury induced by lipopolysaccharide through microRNA-34c-3p-dependent inhibition of WISP1

BackgroundAccumulating evidence has shown the important roles of long non-coding RNAs (lncRNAs) in acute lung injury (ALI). This study aimed to investigate the potential role of lncRNA small nucleolar RNA host gene 14 (SNHG14) in lipopolysaccharides (LPS)-induced ALI.MethodsExpression of SNHG14, microRNA-34c-3p (miR-34c-3p) and Wnt1 inducible signaling pathway protein 1 (WISP1) in LPS-exposed mouse alveolar macrophages (MH-S) and lung tissues from mice with LPS-induced ALI was determined by reverse transcription quantitative polymerase chain reaction. The interactions among SNHG14, miR-34c-3p and WISP1 were analyzed by dual-luciferase reporter and RIP assays. Using gain-of-function or loss-of-function approaches, the contents of proinflammatory proteins were determined and MH-S cell viability was assessed to evaluate the in vitro functions of SNHG14, miR-34c-3p and WISP1, and wet/dry weight ratio and proinflammatory proteins in lung tissues were determined to assess their in vivo effects.ResultsSNHG14 and WISP1 expression was increased, while miR-34c-3p was decreased in ALI models. SNHG14 bound to miR-34c-3p, resulting in impaired miR-34c-3p-dependent down-regulation of WISP1. Both SNHG14 silencing and miR-34c-3p over-expression reduced the levels of proinflammatory proteins IL-18, IL-1β, TNF-α and IL-6 and inhibited MH-S cell viability. SNHG14 silencing or miR-34c-3p over-expression decreased the wet/dry weight ratio in lung tissues from ALI mice. The reductions induced by SNHG14 silencing or miR-34c-3p over-expression were rescued by WISP1 over-expression.ConclusionThis study demonstrated that lncRNA SNHG14 silencing alleviated inflammation in LPS-induced ALI through miR-34c-3p-mediated inhibition of WISP1. Our findings suggest that lncRNA SNHG14 may serve as a therapeutic target for ALI.

Expression of nicotinamide adenine dinucleotide phosphate-reduced oxidase-4/reactive oxygen species and cystathionine-γ-lyase/hydrogen sulfide in patients with chronic obstructive pulmonary disease-related pulmonary hypertension

Objective: To observe the levels of serum reactive oxygen species (ROS) and hydrogen sulfide(H(2)S) in patients with acute exacerbation of chronic obstructive pulmonary disease (AECOPD), and nicotinamide adenine dinucleotide phosphate-reduced (NADPH) oxidase 4 (NOX4) and cystathionine-γ-lyase (CSE) in lung tissue of patients with stable chronic obstructive pulmonary disease (COPD). Methods: (1) A total of 60 patients with AECOPD admitted to the Department of Respiratory Medicine at Ningxia Hui People's Hospital from November 2015 to December 2016 were recruited. According to the results of pulmonary function and echocardiography, the participants were divided into AECOPD-related pulmonary hypertension (PH) group(A) and AECOPD non-PH group (B).Other 30 healthy subjects were selected as the control group (C).Serum ROS and H(2)S of group A, B and C were detected by enzyme-linked immunosorbent assay (ELISA).(2)The lung tissues of patients undergoing lobectomy for lung cancer from November 2012 to April 2017 were collected, who were divided into COPD-related PH group (D), COPD non-PH group (E) and negative control (F). The expression of NOX4 and CSE protein in lung tissue was detected by immunohistochemistry and the thickness of pulmonary arteriole wall was measured. Results: (1)The serum ROS level in group A was higher than group B and C which were (613.52±69.66)IU/ml,(565.76±71.33)IU/ml, (294.63±60.39)IU/ml, respectively with that in group B higher than that in group C (P<0.05). Serum H(2)S level in group A was lower than group B and C, with that in group B lower than group C [(18.59±5.50) nmol/ml, (20.49±4.97) nmol/ml, (38.03±4.43) nmol/ml, respectively P<0.05]. ROS level was positively correlated with pulmonary systolic pressure (PASP) (r=0.59, P<0.05), H(2)S level was negatively correlated with PASP(r=-0.62, P<0.05).(2)The lung tissue expression of NOX4 in group D was higher than group E and F (P<0.05), which were 0.08±0.01,0.06±0.01,0.03±0.01, respectively,while the level of NOX4 in group E was higher than group F (P<0.05). The expression of CSE between group D, E and F were all significantly different (P<0.05),which were 0.03±0.01, 0.07±0.02,0.12±0.02, respectively.(3)Smooth muscle thickness of pulmonary arterioles as a percentage of vascular diameter (WT%) between group D, E and F was all different(P<0.05), which were (40.58±6.63)%,(36.87±5.60)%,(31.27±6.24)%, respectively; so was smooth muscle area of pulmonary arterioles as a percentage of total vascular area(WA%) with (32.33±6.27)%, (30.20±5.28)%, (25.20±4.31)%, respectively (P<0.05). (4)The expression of NOX4 was positively correlated with WT% and WA%, r was 0.81 and 0.66, respectively (P<0.05). The expression of CSE was negatively correlated with WT% and WA%, r was -0.55 and -0.39 respectively (P<0.05). Conclusions: NOX4/ROS and CSE/H(2)S signaling pathways may play an important role in the pathogenesis of COPD related PH.

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.

Regulation of calpain 2 expression by miR-223 and miR-145

Calpain 2 (CAPN2) is a Ca2+-dependent cysteine-protease that is involved in different cellular processes. Despite its important role, little is known about how CAPN2 expression is regulated. This study addressed the potential regulation of CAPN2 by microRNAs (miRNAs) in human endothelial cells. Two miRNAs were found to regulate CAPN2 expression by two distinct mechanisms, one direct and the other indirect. MiR-223 directly targeted CAPN2 by binding to the CAPN2 3′-untranslated region. Mir-223 overexpression decreased CAPN2 protein levels in cultured cells and in mice miR-223 antagonism led to an increase in CAPN2 protein in lung tissue. MiR-145 overexpression also decreased CAPN2 expression but did not affect a CAPN2 luciferase construct, indicating that the effect was indirect. MiR-145 targets histone deacetylase (HDAC) 2, and HDAC inhibition transcriptionally regulated CAPN2 expression by hyperacetylation of the promoter of CAPN2 gene and a subsequent decrease in polymerase 2 binding. Indeed, down regulation of HDAC2 by miR-145 not only decreased CAPN2 protein expression and calpain activity, but also protected paxillin against calpain-dependent degradation. Thus, protein levels of CAPN2 are regulated by miR-223, acting directly on the 3′-untranslated region as well as by miR-145, which acts via an increase in HDAC2. EnzymesCalpain 2 (EC 3.4.22.53), histone deacetylase 2 (EC 3.5.1.98).

Effects of Xinjiaxiangruyin on the TLR7 pathway in influenza virus-infected lungs of mice housed in a hygrothermal environment

BackgroundTo investigate the effects and immunological mechanisms of the traditional Chinese medicine Xinjiaxiangruyin on controlling influenza virus (FM1 strain) infection in mice housed in a hygrothermal environment.MethodsMice were housed in normal and hygrothermal environments, and intranasally infected with influenza virus (FM1). A high-performance liquid chromatography fingerprint of Xinjiaxiangruyin was used to provide an analytical method for quality control. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to measure messenger RNA expression of Toll-like receptor 7 (TLR7), myeloid differentiation primary response 88 (MyD88), and nuclear factor-kappa B (NF-κB) p65 in the TLR7 signaling pathway and virus replication in the lungs. Western blotting was used to measure the expression levels of TLR7, MyD88, and NF-κB p65 proteins. Flow cytometry was used to detect the proportion of Th17/T-regulatory cells.ResultsXinjiaxiangruyin effectively alleviated lung inflammation in C57BL/6 mice in hot and humid environments. Guizhimahuanggebantang significantly reduced lung inflammation in C57BL/6 mice. The expression of TLR7, MyD88, and NF-κB p65 mRNA in lung tissue of WT mice in the normal environment, GZMHGBT group was significantly lower than that in the model group (P < 0.05). In WT mice exposed to the hot and humid environment, the expression levels of TLR7, MyD88, and NF-κB p65 mRNA in the XJXRY group were significantly different from those in the virus group. The expression levels of TLR7, MyD88, and NF-κB p65 protein in lung tissue of WT mice exposed to the normal environment, GZMHGBT group was significantly lower than those in the model group. In WT mice exposed to hot and humid environments, the expression levels of TLR7, MyD88, and NF-κB p65 protein in XJXRY group were significantly different from those in the virus group.ConclusionGuizhimahuanggebantang demonstrated a satisfactory therapeutic effect on mice infected with the influenza A virus (FM1 strain) in a normal environment, and Xinjiaxiangruyin demonstrated a clear therapeutic effect in damp and hot environments and may play a protective role against influenza through downregulation of the TLR7 signal pathway.

Is NLRP3 or NLRP6 inflammasome activation associated with inflammation-related lung tumorigenesis induced by benzo(a)pyrene and lipopolysaccharide?

Chronic inflammation has been shown to play a vital role in lung tumorigenesis. Recently, we have successfully developed a C57BL/6 mouse model of inflammation-related lung tumorigenesis induced by benzo(a)pyrene [B(a)p] and lipopolysaccharide (LPS), which will contribute to better understand the association between pulmonary inflammation and cancer. In this study, we aim to explore the role of NLRP3 and NLRP6 inflammasome in lung tumorigenesis in the animal model that we set up previously. Levels of NLRP3, NLRP6, interleukin-1β (IL-1β) and IL-18 protein in lung tissues were detected by using immunohistochemistry. The co-localization of NLRP3 or NLRP6 with caspase-1 was examined using immunofluorescence and confocal. Western blotting was used to evaluate the levels of caspase-1 p10 and cleaved-IL-1β protein. The expression of IL-18 in bronchoalveolar lavage fluid (BALF) was measured using ELISA kit. The expression of NLRP3, NLRP6 and IL-18 protein in the lung tissues of mice exposed to B(a)p plus LPS was upregulated significantly compared with those in Vehicle control group. Immunofluorescent results indicated the co-localization of NLRP3 with caspase-1 was increased in the lung tissues of LPS-, B(a)p- or B(a)p plus LPS-exposed mice than that in Vehicle control group, but no co-localization of NLRP6 with caspase-1. Additionally, caspase-1 activation was induced, cleaved-IL-1β in lung tissues and IL-18 protein in BALF were increased in B(a)p plus LPS-exposed mice compared with those in B(a)p group. In conclusion, our results from this study demonstrate that NLRP3 inflammasome, not NLRP6 inflammasome, activation is involved in B(a)p plus LPS-induced inflammation-related lung tumorigenesis in mice, but the mechanisms of NLRP6 participate in the development of lung cancer should be further investigated.

Endothelial Progenitor Cells Attenuate Ventilator-Induced Lung Injury with Large-Volume Ventilation

Ventilator-induced lung injury (VILI) is a common complication that results from treatment with mechanical ventilation (MV) in acute respiratory distress syndrome (ARDS) patients. The present study investigated the effect of endothelial progenitor cell (EPC) transplantation on VILI. Wistar rats were divided into three groups (n = 8): sham (S), VILI model (V) induced by tidal volume ventilation (17 mL/kg), and VILI plus EPC transplantation (VE) groups. The lung PaO2/FiO2 ratio, pulmonary wet-to-dry (W/D) weight ratio, number of neutrophils, total protein, neutrophil elastase level, and inflammatory cytokines in bronchoalveolar lavage fluid (BALF) and serum were examined. Furthermore, the histological and apoptotic analysis, and lung tissue protein expression analysis of Bax, Bcl-2, cleaved caspase-3, matrix metalloproteinase (MMP)-9, total nuclear factor kappa B (total-NF-κB), phosphorylated NF-κB (phospho-NF-κB) and myosin light chain (MLC) were performed. The ventilation-induced decrease in PaO2/FiO2 ratio, and the increase in W/D ratio and total protein concentration were prevented by the EPC transplantation. The EPC transplantation (VE group) significantly attenuated the VILI-induced increased expression of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-8, MMP-9, phospho-NF-κB and MLC, neutrophil elastase levels and neutrophil counts in BALF. In addition, the anti-inflammatory factor IL-10 increased in the VE group. Furthermore, pulmonary histological injury and apoptosis (TUNEL-positive cells, increase in Bax and cleaved caspase-3) were considerably diminished by the EPC transplantation. The EPC transplantation ameliorated the VILI. The mechanism may be primarily through the improvement of epithelial permeability, inhibition of local and systemic inflammation, and reduction in apoptosis.

Lung tissue extracellular matrix-derived hydrogels protect against radiation-induced lung injury by suppressing epithelial-mesenchymal transition.

This study aimed to examine whether lung tissue extracellular matrix (ECM) hydrogels have protective effects on radiation-induced lung injury (RILI). The cytocompatibility and histocompatibility were tested for the obtained ECM-derived hydrogel. Sprague-Dawley rats were randomly divided into three groups (n = 18): control group (control); rats receiving irradiation and intratracheal injection of normal saline (IR + NS); and rats receiving irradiation and intratracheal injection of lung ECM-derived hydrogel (IR + ECM). The wet/dry weight ratio was used to evaluate the congestion and edema of the lungs. Histopathological analysis of lung tissues was performed using hemotoxylin and eosin staining and Masson's trichrome staining. Immunohistochemical staining and western blot analyses were carried out to determine the expression of epithelial-mesenchymal transition (EMT)-related proteins in lung tissues (E-cadherin, α-smooth muscle actin [α-SMA], and vimentin). In addition, tumor necrosis factor-α (TNF-α), transforming growth factor-β1 (TGF-β1) and interleukin-6 (IL-6), hydroxyproline, malondialdehyde (MDA), and superoxide dismutase (SOD) levels were also evaluated. The ECM-derived hydrogels had good cytocompatibility and histocompatibility. ECM-derived hydrogel treatment improved lung histopathology injury and pulmonary edema. Higher expression of E-cadherin and lower expression of vimentin and α-SMA were found in the IR + ECM group compared with those in the IR + NS group. Hydroxyproline levels were reduced by ECM-derived hydrogel treatment compared with those in the IR + NS group. Obvious increases of TNF-α, IL-6, and TGF-β1 were identified following irradiation. Marked reductions in MDA content and increases in SOD were induced by ECM-derived hydrogel treatment in rats after radiation. ECM-derived hydrogels were shown to protect against RILI, potentially by reducing EMT, inflammation, and oxidative damage.

Expression of CD63 in Lung Tissue of Guinea Pigs Dying of Anaphylactic Shock.

Objective To study the protein expression of cluster of differentiation 63 （CD63） in lung tissues of guinea pigs that died of anaphylactic shock and discuss the diagnostic value of CD63 for death from anaphylactic shock. Methods Twenty guinea pigs were randomly divided into control group, anaphylactic shock immediate death group, cold storage group （4 ℃ for 48 h） and frozen group （-20 ℃ for 7 d）. The animal model of guinea pigs that died of anaphylactic shock was established with human mixed serum injection. The expression changes of CD63 protein and CD63 mRNA in lung tissues were detected by hematoxylin-eosin （HE） staining, immunohistochemical staining, Western blotting, enzyme-linked immunosorbent assay （ELISA） and real-time RT-PCR. Results HE staining results showed congestion, and edema of lung tissues, and eosinophil infiltration in the anaphylactic shock groups. Western blotting analysis results showed that the expression of CD63 protein in the lung tissues of guinea pigs that died of anaphylactic shock was significantly higher than that in the control group （P<0.05）. Comparison between the anaphylactic shock groups was made, and the differences had no statistical significance. The results of immunohistochemical staining and real-time RT-PCR were consistent with that of Western blotting. ELISA results showed that CD63 protein expression in the immediate death group was higher than that in the control group （P<0.05）. Conclusion The expression of CD63 protein and CD63 mRNA in the lung tissues of guinea pigs that died of anaphylactic shock is significantly enhanced. Animal carcasses which were put in cold storage for 48 h and frozen for 7 d do not affect the examination of the above indicators. CD63 protein is expected to become an auxiliary diagnostic indicator of death from anaphylactic shock.

Alda-1 Prevents Pulmonary Epithelial Barrier Dysfunction following Severe Hemorrhagic Shock through Clearance of Reactive Aldehydes

Severe hemorrhagic shock and resuscitation (HS/R) can lead to lung injury, resulting in respiratory insufficiency. We investigated whether treatment with Alda-1, an ALDH2 activator, decreased lung injury induced by severe HS/R in a rat model. Male Sprague-Dawley rats were randomized into three groups, hemorrhagic shock + placebo, hemorrhagic shock + Alda-1, and sham. All animals were heparinized, and then 50% of the total calculated blood volume was collected over 60 minutes. After 40 minutes of hemorrhagic shock, animals were reinfused with the shed blood over 40 minutes and then observed for an additional 2 hours. Concentrations of 4-HNE, TNF-α, IL-6, and ALDH2 activity were detected; lung injury and lung wet-to-dry weight ratios were assessed. Expression of occludin and ZO-1 proteins in lung tissues was also determined. At 2 hours after resuscitation, lung injury was significantly reduced and the wet-to-dry weight ratio was notably decreased in the Alda-1 group compared with placebo (P<0.05). Alda-1 treatment also significantly increased the activity of ALDH2 and decreased the levels of toxic 4-HNE (P<0.05). In the Alda-1 group, IL-6 and TNF-α were dramatically decreased compared with placebo-treated animals (P<0.05). Expression of occludin and ZO-1 proteins was significantly decreased in the placebo group compared with the Alda-1 group (P<0.05). Thus, in a rat model of severe HS/R, treatment with Alda-1 increased the activity of ALDH2, significantly accelerated the clearance of reactive aldehydes, and concomitantly alleviated lung injury through improvement of pulmonary epithelial barrier integrity resulting in decreased alveolar epithelial tissue permeability, lung edema, and diffuse infiltration of inflammatory cells.

Surfactant Protein D Is Altered in Experimental Malaria-Associated Acute Lung Injury/Acute Respiratory Distress Syndrome.

Surfactant protein D (SP-D) is in the collectin family of C-type lectins and plays an important role in the regulation of inflammation and the innate immune defense against pathogens. This protein has been proposed as a biomarker for acute lung injury. However, the expression of SP-D in the lung and the circulating levels of SP-D during malaria infection have received limited attention. Therefore, the aim of this study was to determine the location and expression of the SP-D protein in lung tissue and to measure the plasma level of SP-D in experimental malaria-associated acute lung injury/acute respiratory distress syndrome (ALI/ARDS). Malaria-infected mice induced by Plasmodium berghei ANKA were classified into two groups, namely, the ALI/ARDS and non-ALI/ARDS groups, according to lung histopathology. The lungs of uninfected mice were used as a control group. The location and expression of SP-D in the lung tissues were investigated by immunohistochemical staining and Western blot analysis. In addition, the level of SP-D in plasma and lung homogenate was measured by an enzyme-linked immunosorbent assay. Immunohistochemical staining of SP-D was significantly increased in the lung tissues of the malaria-infected mice in the ALI/ARDS group compared with that in the malaria-infected mice in the non-ALI/ARDS group and the mice in the control group (p < 0.05). The levels of SP-D in the plasma and lung homogenate were significantly increased in the malaria-infected mice in the ALI/ARDS group compared with those in the malaria-infected mice in the non-ALI/ARDS group and the mice in the control group (p < 0.05). There was a significant positive correlation between SP-D in the plasma and SP-D in the lung homogenate (rs = 0.900, p = 0.037). In conclusion, this study demonstrated increased expression levels of SP-D in the lung tissue and high levels of plasma SP-D in the malaria-infected mice with ALI/ARDS compared with those in the mice in the other groups. The current study supports that the elevation of the plasma SP-D level may provide useful biological confirmation of the diagnosis of ALI/ARDS during malaria infection.