Nuclear factor-kappaB activation in neonatal mouse lung protects against lipopolysaccharide-induced inflammation

RATIONALE: Injurious agents often cause less severe injury in neonates as compared with adults. OBJECTIVE: We hypothesized that maturational differences in lung inflammation induced by lipopolysaccharide (LPS) may be related to the nature of the nuclear factor (NF)-kappaB complex activated, and the profile of target genes expressed. METHODS: Neonatal and adult mice were injected with intraperitoneal LPS. Lung inflammation was assessed by histology, and apoptosis was determined by TUNEL (terminal deoxynucleotidyl transferase UTP nick-end labeling). The expression of candidate inflammatory and apoptotic mediators was evaluated by quantitative real-time polymerase chain reaction and Western immunoblot. RESULTS: Neonates demonstrated reduced inflammation and apoptosis, 24 hours after LPS exposure, as compared with adults. This difference was associated with persistent activation of NF-kappaB p65p50 heterodimers in the neonates in contrast to early, transient activation of p65p50 followed by sustained activation of p50p50 in the adults. Adults had increased expression of a panel of inflammatory and proapoptotic genes, and repression of antiapoptotic targets, whereas no significant changes in these mediators were observed in the neonates. Inhibition of NF-kappaB activity in the neonates decreased apoptosis, but heightened inflammation, with increased expression of the same inflammatory genes elevated in the adults. In contrast, inhibition of NF-kappaB in the adults resulted in partial suppression of the inflammatory response. CONCLUSIONS: NF-kappaB activation in the neonatal lung is antiinflammatory, protecting against LPS-mediated lung inflammation by repressing similar inflammatory genes induced in the adult.     

Targeting the Linear Ubiquitin Assembly Complex to Modulate the Host Response and Improve Influenza A Virus Induced Lung Injury

Influenza virus infection is characterized by symptoms ranging from mild congestion and body aches to severe pulmonary edema and respiratory failure. While the majority of those exposed have minor symptoms and recover with little morbidity, an estimated 500,000 people succumb to IAV-related complications each year worldwide. In these severe cases, an exaggerated inflammatory response, known as “cytokine storm”, occurs which results in damage to the respiratory epithelial barrier and development of acute respiratory distress syndrome (ARDS). Data from retrospective human studies as well as experimental animal models of influenza virus infection highlight the fine line between an excessive and an inadequate immune response, where the host response must balance viral clearance with exuberant inflammation. Current pharmacological modulators of inflammation, including corticosteroids and statins, have not been successful in improving outcomes during influenza virus infection. We have reported that the amplitude of the inflammatory response is regulated by Linear Ubiquitin Assembly Complex (LUBAC) activity and that dampening of LUBAC activity is protective during severe influenza virus infection. Therapeutic modulation of LUBAC activity may be crucial to improve outcomes during severe influenza virus infection, as it functions as a molecular rheostat of the host response. Here we review the evidence for modulating inflammation to ameliorate influenza virus infection-induced lung injury, data on current anti-inflammatory strategies, and potential new avenues to target viral inflammation and improve outcomes.     

Host gene regulation during coxsackievirus B3 infection in mice: assessment by microarrays

Host genetic responses that characterize enteroviral myocarditis have not yet been determined. The injurious and inflammatory process in heart muscle may reflect host responses of benefit to the virus and ultimately result in congestive heart failure and dilated cardiomyopathy. On the other hand, host responses within the myocardium may secure the host against acute or protracted damage. To investigate the nature of modified gene expression in comparison with normal tissue, mRNA species were assessed in myocardium using cDNA microarray technology at days 3, 9, and 30 after infection. Of 7000 clones initially screened, 169 known genes had a level of expression significantly different at 1 or more postinfection time points as compared with baseline. The known regulated genes were sorted according to their functional groups and normalized expression patterns and, subsequently, interpreted in the context of viremic, inflammatory, and healing phases of the myocarditic process.     

Echinostoma caproni (Trematoda): differential in vivo mucin expression and glycosylation in high‐ and low‐compatible hosts

Enhanced mucus production and release appears to be a common mechanism for the clearance of intestinal helminths, and this expulsion is normally mediated by Th2‐type immune responses. To investigate the factors determining the expulsion of intestinal helminths, we have analysed in vivo expression of mucin genes at the site of infection in two host species displaying different compatibility with Echinostoma caproni (Trematoda). Surprisingly, a general down‐regulation on mucin mRNA expression was detected in low‐compatible hosts (rats) coinciding with the development of Th2/Th17 responses and the early rejection of the worms from the intestinal lumen. This suggests the existence of a mechanism by which the parasites can modulate the mucus barrier to favour their survival. In highly compatible hosts (mice), some mucin genes were found to be up‐regulated throughout the infection, probably, to protect the intestinal epithelium against the infection‐induced inflammation developed in this host species. Moreover, infection‐induced changes on mucin glycans were also studied by lectin histochemistry. Similar alterations were detected in the ileum of infected mice and rats, except with SNA lectin, indicating that sylated mucins might play an important role in determining the evolution of the infection in each host species.     

Microbiological and inflammatory factors associated with the development of pneumococcal pneumonia.

Pneumococcal pneumonia still is associated with a high mortality rate, despite appropriate antimicrobial therapy. Many gaps remain in the understanding of the pathogenesis of this deadly infection. The microbial and inflammatory events that characterize survival or death after intranasal inoculation of mice with an LD(50) inoculum of Streptococcus pneumoniae were investigated. Survival was associated with rapid bacterial clearance and low inflammation (surfactant and red blood cells in alveoli), but no neutrophil recruitment or lung tissue injury was noted. By contrast, death was preceded by strong bacterial growth that peaked 48 h after the infection and was associated with gradual increases in pulmonary levels of interleukin-6, macrophage inflammatory protein (MIP)-1alpha, MIP-2, monocyte chemoattractant protein-1, KC, and neutrophil recruitment. The injection of tumor necrosis factor-alpha or the addition of lipopolysaccharide or heat-killed S. pneumoniae to the inoculum enhanced early host response and survival. These observations may help develop appropriate markers of evolution of pneumonia, as well as new therapeutic strategies.     

The role of inflammation in respiratory impairment during Pneumocystis carinii pneumonia

Pneumocystis carinii remains an important cause of pneumonia in immunosuppressed hosts. Severe Pneumocystis pneumonia is characterized by an intense neutrophilic inflammatory response resulting in gas exchange abnormalities, diffuse alveolar damage, and respiratory failure. The inflammatory response directed against P. carinii involves a complex series of interactions between alveolar macrophages, CD4+ T lymphocytes, polymorphonuclear cells, and their various products. CD4+ T lymphocytes are crucial to host defense against P. carinii. Alveolar macrophages also provide essential functions that significantly enhance clearance of P. carinii infection. In addition, host proteins play an important role in augmenting the host inflammatory responses to this organism. Although essential for effective clearance of infection, excessive inflammatory responses also predispose the host to the development of lung injury and respiratory compromise. Understanding the complex processes involved in the host inflammatory response and its potential for causing lung injury may enable development of novel therapeutic approaches for this and other important fungal lung infections.     

Immunomodulatory therapies for cystic fibrosis.

Progressive pulmonary disease is the primary cause of morbidity and mortality in adult patients with cystic fibrosis (CF). The decrease in lung function associated with infection with Pseudomonas aeruginosa has been related to the severity of pulmonary inflammation. Thus therapies that reduce pulmonary inflammation may prove to be clinically efficacious. Therapeutic interventions that target pulmonary inflammation may be directed either at the infecting organism, especially P aeruginosa, or at host responses. Eradication of P aeruginosa from the airways of patients with CF has not been accomplished. However, reduction of the burden of P aeruginosa or modification of virulence factors are practical goals. Normalization of the host response ultimately depends on correction of the molecular defect. Until then, therapies are being investigated that may modulate pulmonary inflammation. These include therapies aimed at compensating for the defect in ion transport, down-regulating inflammatory cell responses, inhibiting host inflammatory products, or altering airway secretions. Preliminary data suggest that each of these approaches may have clinical efficacy. Large, multicenter trials addressing these issues are presently ongoing and hold the promise for continued improvement in the clinical course of patients with CF.     

Global analysis of gene expression in pulmonary fibrosis reveals distinct programs regulating lung inflammation and fibrosis

The molecular mechanisms of pulmonary fibrosis are poorly understood. We have used oligonucleotide arrays to analyze the gene expression programs that underlie pulmonary fibrosis in response to bleomycin, a drug that causes lung inflammation and fibrosis, in two strains of susceptible mice (129 and C57BL/6). We then compared the gene expression patterns in these mice with 129 mice carrying a null mutation in the epithelial-restricted integrin beta6 subunit (beta6(-/-)), which develop inflammation but are protected from pulmonary fibrosis. Cluster analysis identified two distinct groups of genes involved in the inflammatory and fibrotic responses. Analysis of gene expression at multiple time points after bleomycin administration revealed sequential induction of subsets of genes that characterize each response. The availability of this comprehensive data set should accelerate the development of more effective strategies for intervention at the various stages in the development of fibrotic diseases of the lungs and other organs.     

The association between the decreased expression levels of FOXJ1 and the activation of NF-κB pathway in interstitial lung disease of MRL/Lpr mice

Background: Pulmonary manifestations of systemic lupus erythematosus (SLE) are appearing in 4-5% of patients involving lung in almost half of the cases during the disease course. Objective: We compared the autoimmune pulmonary inflammation in the lung tissue of mice to determine the association between decreased expression levels of Forkhead Box J1 (FOXJ1) and the activation of the NF-κB pathway in autoimmune pulmonary inflammation of MRL/Lpr mice. Methods: The female BALB/c mice (n=6) and MRL/Lpr mice (n=30) were divided into 5 groups including a control group (BALB/c), and five MRL/Lpr mice groups (8W, 12W, 16W, 24W, and 32W). The infiltration of the inflammatory cells was determined in lung tissue by performing the histological analysis. The western blotting was used to examine the expression levels of the age-related FOXJ1, and p50 and p65 proteins in the lungs of MRL/Lpr mice. The expression levels of MMP2 and MMP9 were determined via immunohistochemistry and immunofluorescence. Results: There were severe infiltrates of lung cells with high levels of tracheal damage, perivascular injury and interstitial inflammatory cell infiltration when the MRL/Lpr mice from 16w to 32w comparing to the 8w old healthy MRL/Lpr mice in the control group (p <0.05). Moreover, the reduced expression levels of FOXJ1 were associated with the activation of the NF-κB pathway in interstitial lung disease of MRL/Lpr mice via the modulation of p50 and p65. In addition, the expression levels of MMP2 and MMP9 pro-inflammation factors increased in the lungs of the MRL/Lpr mice from 16w to 32w. Conclusions: The expression level of FOXJ1 might be an indicator of the degree of lung disease in lupus-prone mice.     

Evaluation of tool-like receptor-2 and 4 and interleukin-6 gene expressions in Turkish rheumatoid arthritis patients

Rheumatoid arthritis (RA) is a progressive inflammatory disease. Although the etiology and pathogenesis of RA are not known well, genetic and environmental factors are proposed to initiate an autoimmune process. We aimed to investigate mRNA expression levels of Toll-like receptor-2 (TLR-2), TLR-4, and interleukin-6 (IL-6) genes in RA disease. This study was conducted with 50 patients who were diagnosed with RA according to the American College of Rheumatology classification criteria for RA and 50 age-matched healthy control individuals who did not have any joint diseases and autoimmune diseases. We collected whole blood from all participants and analyzed expression of TLR-2, TLR-4, and IL-6 genes at mRNA level using real-time qPCR. TLR-2 expression was detected to increase 3.8-fold and IL-6 expression was detected to increase 6.8-fold in RA patients compared to healthy controls. No difference was found between patient and control groups with regard to TLR-4 expression. Overexpression of TLR-2 and IL-6 may be responsible for RA pathogenesis. Inhibition of both TLR and IL signaling pathways may prevent joint inflammation and destruction.     

IKK NBD peptide inhibits LPS induced pulmonary inflammation and alters sphingolipid metabolism in a murine model

Airway epithelial NF-κB is a key regulator of host defence in bacterial infections and has recently evolved as a target for therapeutical approaches. Evidence is accumulating that ceramide, generated by acid sphingomyelinase (aSMase), and sphingosine-1-phosphate (S1-P) are important mediators in host defence as well as in pathologic processes of acute lung injury. Little is known about the regulatory mechanisms of pulmonary sphingolipid metabolism in bacterial infections of the lung. The objective of this study was to evaluate the influence of NF-κB on sphingolipid metabolism in Pseudomonas aeruginosa LPS-induced pulmonary inflammation. In a murine acute lung injury model with intranasal Pseudomonas aeruginosa LPS we investigated TNF-α, KC (murine IL-8), IL-6, MCP-1 and neutrophilic infiltration next to aSMase activity and ceramide and S1-P lung tissue concentrations. Airway epithelial NF-κB was inhibited by topically applied IKK NBD, a cell penetrating NEMO binding peptide. This treatment resulted in significantly reduced inflammation and suppression of aSMase activity along with decreased ceramide and S1-P tissue concentrations down to levels observed in healthy animals. In conclusion our results confirm that changes in sphingolipid metabolim due to Pseudomonas aeruginosa LPS inhalation are regulated by NF-κB translocation. This confirms the critical role of airway epithelial NF-κB pathway for the inflammatory response to bacterial pathogens and underlines the impact of sphingolipids in inflammatory host defence mechanisms.     

Hypercapnia via reduced rate and tidal volume contributes to lipopolysaccharide-induced lung injury

Appreciating that CO2 modifies the chemical reactivity of nitric oxide (NO)-derived inflammatory oxidants, we investigated whether hypercapnia would modulate pulmonary inflammatory responses. Rabbits (n = 72) were ventilated with approximately 7-ml/kg tidal volume for 6 hours. Animals were randomized to one of the following conditions: eucapnia (Pa(CO2) at approximately 35-40 mm Hg), eucapnia + lipopolysaccharide (LPS), eucapnia + LPS + inhaled NO (iNO delivered at approximately 20 ppm), hypercapnia (Pa(CO2) at approximately 60 mm Hg), hypercapnia + LPS, and hypercapnia + LPS + iNO. The hypercapnia + LPS groups compared with groups exposed to eucapnia + LPS displayed significantly increased bronchoalveolar lavage fluid protein concentrations (p < 0.05), lung wet-to-dry ratios (p < 0.05), bronchoalveolar lavage fluid cell counts (p < 0.05), and lung histologic alterations consistent with greater injury. Furthermore, expression of inducible nitric oxide synthase (p < 0.05), tissue myeloperoxidase content (p < 0.05), and formation of lung protein 3-nitrotyrosine derivatives (p < 0.05) was greatest under conditions of hypercapnia + LPS. Groups exposed to hypercapnic conditions without LPS did not manifest these changes. The inhalation of iNO attenuated selected indices of lung injury. We conclude that hypercapnia induced by means of reduced rate and tidal volume amplifies pulmonary inflammatory responses.     

慢性阻塞性肺疾病组蛋白去乙酰化酶与糖皮质激素抵抗

慢性阻塞性肺疾病(chronic obstructive pulmonary disease,COPD)是一种慢性气道炎症,炎症基因的表达受组蛋白去乙酰化酶(histone deacetylase,HDAC)和组蛋白乙酰化酶(histone acetyltransferase,HAT)调控,通常组蛋白乙酰化增高促进基因转录,降低则抑制基因转录.糖皮质激素(glucocorticoid,GC)的一个重要作用是募集HDAC2到基因表达位点,使基因组蛋白乙酰化程度降低,从而抑制炎症基因表达.由吸烟引起的COPD患者,香烟烟雾氧化应激使得HDAC2减少,降低了GC的抗炎作用,导致GC抵抗,而GC抵抗是COPD抗炎治疗的主要障碍.本文讨论GC抵抗的一些分子机制,为COPD患者治疗方案的制定及改善预后提供理论参考依据.     

Impact of cigarette smoke on clearance and inflammation after Pseudomonas aeruginosa infection

The object of this study was to investigate the impact of cigarette smoke on bacterial clearance and immune inflammatory parameters after infection with Pseudomonas aeruginosa in mice. We observed a delayed rate of bacterial clearance in smoke-exposed compared with sham-exposed mice. This was associated with increased inflammation characterized by greater numbers of neutrophils and mononuclear cells in the bronchoalveolar lavage. After infection, we observed increased levels of proinflammatory cytokines (tumor necrosis factor-alpha, interleukin-1beta, and interleukin-6) and chemokines (monocyte chemoattractant protein-1 [MCP-1] and macrophage inflammatory protein-2 [MIP-2]) as well as myeloperoxidase and proteolytic activity in the lungs of smoke-exposed compared with sham-exposed animals. Delayed clearance was associated with increased morbidity and greater weight loss of smoke-exposed mice. After delivery of inactivated bacteria, we observed a similar inflammatory response, clinical score, and tumor necrosis factor-alpha expression in smoke- and sham-exposed animals, suggesting that increased inflammation and altered clinical presentation are due to the delayed rate of bacterial clearance. Our findings suggest that cigarette smoke affects respiratory immune-inflammatory responses elicited by bacteria. We postulate that altered respiratory host defense may be implicated in smoking-related diseases such as chronic obstructive pulmonary disease.