Inflammaging increases susceptibility to cigarette smoke-induced COPD.

Chronic obstructive pulmonary disease (COPD) is related to an abnormal chronic inflammatory response of the lung to mainly cigarette smoke (CS) and the disease risk is increased in aged individuals. The source of this chronic inflammation is due to the repeated and progressive activation of immune cells. We hypothesize that in a chronic CS-induced mouse model, the predisposition to COPD pathogenesis in aged mice is characterized by an elevated immune response compared to young animals. We measured several characteristics of COPD in young and old mice (2 and 12 months of age) exposed to CS for 3 months. CS-exposed aged mice exhibited increased lung compliance (0.061 ± 0.008 vs. 0.055 ± 0.006 ml/cm H2O, p < 0.01), emphysema development (35.36 ± 0.71 vs. 25.31 ± 0.005 µm; p < 0.01) and airway remodeling (2.15 ± 0.37 vs. 1.09 ± 0.64 µm3/µm2; p < 0.01) compared to control animals, which was not seen in CS-exposed young mice. Quantification of lung tissue inflammation revealed a significantly greater volume of inducible bronchus-associated lymphoid tissue structures in aged mice after CS exposure (5.94 ± 2.89 vs. 2.37 ± 1.69 µm3/µm2; p < 0.01). Our results indicate that age-induced lung inflammation is further elevated after CS exposure in old mice, potentially via an age-induced change in immune cell susceptibility to CS thereby accelerating the pathophysiological hallmarks of COPD.

Secretory group V phospholipase A2 regulates acute lung injury and neutrophilic inflammation caused by LPS in mice.

We investigated the regulatory role of 14-kDa secretory group V phospholipase A(2) (gVPLA(2)) in the development of acute lung injury (ALI) and neutrophilic inflammation (NI) caused by intratracheal administration of LPS. Experiments were conducted in gVPLA(2) knockout (pla2g5(-/-)) mice, which lack the gene, and gVPLA(2) wild-type littermate control (pla2g5(+/+)) mice. Indices of pulmonary injury were evaluated 24 h after intratracheal administration of LPS. Expression of gVPLA(2) in microsections of airways and mRNA content in lung homogenates were increased substantially in pla2g5(+/+) mice after LPS-administered compared with saline-treated pla2g5(+/+) mice. By contrast, expression of gVPLA(2) was neither localized in LPS- nor saline-treated pla2g5(-/-) mice. LPS also caused 1) reduced transthoracic static compliance, 2) lung edema, 3) neutrophilic infiltration, and 4) increased neutrophil myeloperoxidase activity in pla2g5(+/+) mice. These events were attenuated in pla2g5(-/-) mice exposed to LPS or in pla2g5(+/+) mice receiving MCL-3G1, a neutralizing MAb directed against gVPLA(2), before LPS administration. Our data demonstrate that gVPLA(2) is an inducible protein in pla2g5(+/+) mice but not in pla2g5(-/-) mice within 24 h after LPS treatment. Specific inhibition of gVPLA(2) with MCL-3G1 or gene-targeted mice lacking gVPLA(2) blocks ALI and attenuates NI caused by LPS.

A comprehensive model of allergic rhinitis in guinea pigs.

INTRODUCTION: The economic and social impact of allergic rhinitis is substantial. The effectiveness of currently available medications is limited and therefore investigations for more effective drugs is essential. This study was intended to establish a model of allergic rhinitis in guinea pigs that can be utilized for further investigation of new drugs. METHODS: Male Dunkin Hartley guinea pigs were sensitized intranasally to, and challenged with, ovalbumin. Sneezing (SN) and nose rubbing (NR) response to allergen challenge were observed on day 21 post-initiation of sensitization in conscious guinea pigs. Nasal blockade (NB), leukocyte infiltration, and lung inflation pressure (LIP) were assessed in the same guinea pigs 23-28 days post-initiation of sensitization. A ventilator/flow method was used to measure NB and LIP. Leukocyte infiltration into nasal lavage fluid 60 min after challenge in the same animals was recorded as total and differential cell counts. RESULTS: Sensitized guinea pigs produced acute allergic responses after allergen challenge. This was characterized by increases in SN, NR, NB, and eosinophil infiltration. In addition, intranasal allergen challenge did not change lung inflation pressure. DISCUSSION: Allergen-induced rhinitis in guinea pigs resembles that in humans. The model reported in this study can be used to reflect the effectiveness of drugs currently used to treat allergic rhinitis and to investigate new potential drugs for the treatment of allergic rhinitis.

Alveolar macrophage activation is a key initiation signal for acute lung ischemia-reperfusion injury.

Lung ischemia-reperfusion (I/R) injury is a biphasic inflammatory process. Previous studies indicate that the later phase is neutrophil-dependent and that alveolar macrophages (AMs) likely contribute to the acute phase of lung I/R injury. However, the mechanism is unclear. AMs become activated and produce various cytokines and chemokines in many inflammatory responses, including transplantation. We hypothesize that AMs respond to I/R by producing key cytokines and chemokines and that depletion of AMs would reduce cytokine/chemokine expression and lung injury after I/R. To test this, using a buffer-perfused, isolated mouse lung model, we studied the impact of AM depletion by liposome-clodronate on I/R-induced lung dysfunction/injury and expression of cytokines/chemokines. I/R caused a significant increase in pulmonary artery pressure, wet-to-dry weight ratio, vascular permeability, tumor necrosis factor (TNF)-alpha, monocyte chemoattractant protein (MCP)-1, and macrophage inflammatory protein (MIP)-2 expression, as well as decreased pulmonary compliance, when compared with sham lungs. After AM depletion, the changes in each of these parameters between I/R and sham groups were significantly attenuated. Thus AM depletion protects the lungs from I/R-induced dysfunction and injury and significantly reduces cytokine/chemokine production. Protein expression of TNF-alpha and MCP-1 are positively correlated to I/R-induced lung injury, and AMs are a major producer/initiator of TNF-alpha, MCP-1, and MIP-2. We conclude that AMs are an essential player in the initiation of acute lung I/R injury.

Roles for early response cytokines during Escherichia coli pneumonia revealed by mice with combined deficiencies of all signaling receptors for TNF and IL-1.

During infection, inflammation is essential for host defense, but it can injure tissues and compromise organ function. TNF-alpha and IL-1 (alpha and beta) are early response cytokines that facilitate inflammation. To determine the roles of these cytokines with overlapping functions, we generated mice deficient in all of the three receptors mediating their effects (TNFR1, TNFR2, and IL-1RI). During Escherichia coli pneumonia, receptor deficiency decreased neutrophil recruitment and edema accumulation to half of the levels observed in wild-type mice. Thus these receptors contributed to maximal responses, but substantial inflammation progressed independently of them. Receptor deficiency compromised antibacterial efficacy for some infectious doses. Decreased ventilation during E. coli pneumonia was not affected by receptor deficiency. However, the loss of lung compliance during pneumonia was substantially attenuated by receptor deficiency. Thus during E. coli pneumonia in mice, the lack of signaling from TNF-alpha and IL-1 decreases inflammation and preserves lung compliance.

The C10/CCL6 chemokine and CCR1 play critical roles in the pathogenesis of IL-13-induced inflammation and remodeling.

IL-13 is a potent stimulator of inflammation and tissue remodeling that plays a key role in the pathogenesis of a wide variety of human disorders. To further understand these responses, studies were undertaken to define the role(s) of the chemokine C10/CCL6 in the pathogenesis of IL-13-induced alterations in the murine lung. IL-13 was a very potent stimulator of C10/CCL6 mRNA and protein, and IL-13-induced inflammation, alveolar remodeling, and compliance alterations were markedly ameliorated after C10/CCL6 neutralization. Treatment with anti-C10/CCL6 decreased the levels of mRNA encoding matrix metalloproteinase-2 (MMP-2), MMP-9, and tissue inhibitor of metalloproteinase-4 (TIMP-4) in lungs from wild-type mice. C10/CCL6 neutralization also decreased the ability of IL-13 to stimulate the production of monocyte chemoattractant protein-1, macrophage inflammatory protein-1alpha, MMP-2, MMP-9, and cathepsins-K, -L, and -S and the ability of IL-13 to inhibit alpha1-antitrypsin. In accord with these findings, a targeted null mutation of CCR1, a putative C10/CCL6 receptor, also decreased IL-13-induced inflammation and alveolar remodeling and caused alterations in chemokines, proteases, and antiproteases comparable to those seen after C10/CCL6 neutralization. These C10/CCL6 and CCR1 manipulations did not alter the production of transgenic IL-13. These studies demonstrate that IL-13 is a potent stimulator of C10/CCL6 and highlight the importance of C10/CCL6 and signaling via CCR1 in the pathogenesis of the IL-13-induced pulmonary phenotype. They also describe a C10/CCL6 target gene cascade in which C10/CCL6 induction is required for optimal IL-13 stimulation of selected chemokines (monocyte chemoattractant protein-1 and MIP-1alpha) and proteases (MMP-2, MMP-9, and cathepsins-K, -L, and -S) and the inhibition of alpha1-antitrypsin.

Increased airway responsiveness of a common fragrance component, 3-carene, after skin sensitisation--a study in isolated guinea pig lungs.

Lungs from skin-sensitised and non-sensitised guinea pigs were exposed via the airways to 3-carene (1900 mg/m3) and perfused with buffer containing either autologous plasma or lymphocytes. The experiments were performed in order to investigate the importance of blood components for the increased lung responsiveness seen in skin-sensitised animals. A reduction in lung function was noted in all lungs during 3-carene exposure. There was no difference in the 3-carene response between lungs from skin-sensitised animals versus lungs from non-sensitised animals when the perfusion buffer contained lymphocytes. However, when plasma diluted with buffer was used as perfusion medium, there was a significant enhancement in the response in lungs from sensitised versus lungs from non-sensitised animals. This implies that skin sensitisation increases lung responses to inhaled 3-carene and those components in plasma, and not the lymphocyte fraction, contributes to the observed increased lung responsiveness.

[Effects of vaccae on airway contraction and inflammation in asthmatic guinea pigs].

OBJECTIVE: To study the effects of Mycobacterium vaccae (M. vaccae) on the lung function, airway hyperresponsiveness and airway inflammation in a guinea pig model of asthma. METHODS: Asthma was induced with ovalbumin (OVA) in 71 guinea pigs. The changes of lung resistance (R(L)) and dynamic lung compliance (C(dyn)), the accumulation of inflammatory cells in bronchoalveolar lavage fluids (BALF), and the contraction reaction curve of tracheal smooth muscles to carbachol in vitro were determined. RESULTS: Pretreatment with M. vaccae as a single dosage by intramuscular injection (i.m) inhibited the early phase of asthma in the sensitized guinea pigs in a dose-dependent manner. R(L) (1 min to 15 min) increased 46.4% in the 2.5 micro g group, 29.6% in the 7.5 micro g group, and 20.8% in the 22.5 micro g group, which were significantly lower than that in the model control group (95.3%) (P < 0.05 approximately 0.01), respectively. The mean decrease of C(dyn) was 26.8% in the 2.5 micro g group, 23.5% in the 7.5 micro g group, and 21.5% in the 22.5 micro g group, which were significantly lower than that in the model control group (38.7%) (P < 0.05), respectively. Pretreatment with M. vaccae in a dosage of 2.5 micro g, 7.5 micro g and 22.5 micro g (per guinea pig) i.m also inhibited the late phase of asthma. The total white blood cells in BALF was (16.2 +/- 3.2) x 10(8)/L in the 2.5 micro g group, (14.6 +/- 3.4) x 10(8)/L in the 7.5 micro g group, and (15.4 +/- 2.5) x 10(8)/L in the 22.5 micro g group, which were significantly lower than that in the model control group (22.3 +/- 2.2) x 10(8)/L (P < 0.01 approximately 0.001), respectively. BAL eosinophil count was (11.6 +/- 3.0) x 10(8)/L in the 2.5 micro g group, (6.0 +/- 1.5) x 10(8)/L in the 7.5 micro g group, and (2.2 +/- 1.7) x 10(8)/L in the 22.5 micro g group, which were significantly lower than that in the model control group (15.9 +/- 5.2) x 10(8)/L (P < 0.01 approximately 0.001), respectively. M. vaccae significantly inhibited carbachol-induced airway hyperresponsiveness in the tracheal smooth muscle (TSM) from the sensitized guinea pigs. EC(50) (the concentration of carbachol required to increase TSM contraction value by 50% from the baseline) was 0.140 micro mol/L in the 2.5 micro g group, 0.179 micro mol/L in the 7.5 micro g group and 0.236 micro mol/L in the 22.5 micro g group, which were higher than that in the model control group (0.038 micro mol/L). CONCLUSIONS: M. vaccae was found to improve lung function, inhibit airway inflammation and airway hyperresponsiveness in this animal model, which supports its application in the treatment of asthma.

Surfactant protein A is a required mediator of keratinocyte growth factor after experimental marrow transplantation.

We reported an association between the ability of recombinant human keratinocyte growth factor (rHuKGF) to upregulate the expression of surfactant protein A (SP-A) and to downregulate pulmonary inflammation that occurs after allogeneic bone marrow transplantation (BMT). To establish a causal relationship, rHuKGF (5 mg/kg) was administered subcutaneously for three consecutive days before irradiation to SP-A-sufficient and -deficient [SP-A(+/+) and SP-A(-/-), respectively] mice given inflammation-inducing allogeneic spleen T cells at the time of BMT. In contrast with SP-A(+/+) mice, rHuKGF failed to suppress the high levels of TNF-alpha, IFN-gamma, and nitric oxide contained in bronchoalveolar lavage fluids collected on day 7 after BMT from SP-A(-/-) mice. Early post-BMT weight loss was attenuated by rHuKGF in both SP-A(+/+) and SP-A(-/-) recipients. In the absence of supportive respiratory care, however, SP-A deficiency eventually abolished the ability of rHuKGF to prevent weight loss and to improve survival monitored for 1 mo after allogeneic BMT. In further experiments, the addition of cyclophosphamide (which is known to cause severe injury to the alveolar epithelium in donor T cell-recipient mice) to the conditioning regimen prevented rHuKGF-induced upregulation of SP-A and suppression of lung inflammation in both SP-A(+/+) and SP-A(-/-) mice. We conclude that endogenous baseline SP-A levels and optimal upregulation of SP-A are required for the anti-inflammatory protective effects of KGF after allogeneic transplantation.

Increased airway responsiveness after skin sensitisation to 3-carene, studied in isolated guinea pig lungs.

Inhalation of 3-carene has been shown to induce bronchoconstriction in concentrations not far from the threshold limit value. In this study, one group of guinea-pigs were sensitised by dermal exposure to 3-carene according to the modified Cumulative Contact Enhancement Test protocol and another group of animals was used as controls. Lungs from the skin-sensitised and control guinea-pigs were perfused with diluted autologous blood (13 ml blood/87 ml buffer) and exposed to 3-carene at an air concentration of 3000 mg/m(3). In both groups there was a reduction in compliance and conductance but this reduction was significantly (P<0.05) more pronounced (2.5-3 times) in lungs obtained from sensitised animals than from control animals. In a previous study with similar design, but with plain buffer instead of diluted autologous blood as perfusate, we found no statistically significant difference in lung bronchoconstriction. Thus, it is concluded that skin sensitisation can increase lung reactivity to 3-carene and that important mediators of this effect seem to be present in the blood.

Lung function and ventilation inhomogeneity in rat lungs after allergen challenge.

We studied the early response to ovalbumin challenge in sensitized Brown-Norway rats through its effect on N(2), He, and SF(6) phase III slopes of the single-breath washout and on indexes of lung function. Sensitized rats showed varying degrees of response in terms of pulmonary pressure (PL), with increases ranging between 125 and 225% of baseline. The sensitized rats presented decreased quasistatic compliance, forced vital capacity, and end-expiratory flow, with all three lung function indexes showing a significant negative correlation with corresponding PL values. They also showed significant positive correlations of PL with the N(2), He, and SF(6) phase III slopes, reflecting diffusion-convection-dependent inhomogeneities generated by conformation changes throughout the entire rat lung. In addition, the rats showing the most marked PL increases (>150% baseline PL) also revealed a reversal of the SF(6)-He slope difference because of a more marked SF(6) than He slope increase. This latter finding suggests that the degree of structural heterogeneity during early response is even more marked in the most peripheral rat lung generations.

Intratracheal anti-tumor necrosis factor-alpha antibody attenuates ventilator-induced lung injury in rabbits.

To evaluate the role of tumor necrosis factor (TNF)-alpha in the pathogenesis of ventilator-induced lung injury, we 1) measured TNF-alpha production in the lung caused by conventional mechanical ventilation (CMV) and 2) evaluated the protective effect of anti-TNF-alpha antibody (Ab) in saline-lavaged rabbit lungs. After they received saline lung lavage, rabbits were intratracheally instilled with 1 mg/kg of polyclonal anti-TNF-alpha Ab in the high-dose group (n = 6), 0.2 mg/kg of anti-TNF-alpha Ab in the low-dose group (n = 6), serum IgG fraction in the Ab control group (n = 6), and saline in the saline control group (n = 7). Animals then underwent CMV for 4 h. Levels of TNF-alpha in lung lavage fluid were significantly higher after CMV than before in both control groups. Pretreatment with intratracheal instillation of high and low doses of anti-TNF-alpha Ab improved oxygenation and respiratory compliance, reduced the infiltration of leukocytes, and ameliorated pathological findings. CMV led to TNF-alpha production in the lungs, and intratracheal instillation of anti-TNF-alpha Ab attenuated CMV-induced lung injury in this model.

Development of eosinophilic airway inflammation and airway hyperresponsiveness in mast cell-deficient mice.

Mast cells are the main effector cells of immediate hypersensitivity and anaphylaxis. Their role in the development of allergen-induced airway hyperresponsiveness (AHR) is controversial and based on indirect evidence. To address these issues, mast cell-deficient mice (W/W v) and their congenic littermates were sensitized to ovalbumin (OVA) by intraperitoneal injection and subsequently challenged with OVA via the airways. Comparison of OVA-specific immunoglobulin E (IgE) levels in the serum and numbers of eosinophils in bronchoalveolar lavage fluid or lung digests showed no differences between the two groups of mice. Further, measurements of airway resistance and dynamic compliance at baseline and after inhalation of methacholine were similar. These data indicate that mast cells or IgE-mast cell activation is not required for the development of eosinophilic inflammation and AHR in mice sensitized to allergen via the intraperitoneal route and challenged via the airways.

Effect of antigen provocation of IL-5 transgenic mice on eosinophil mobilization and bronchial hyperresponsiveness.

We investigated whether allergen-induced eosinophil recruitment into mouse airways modifies the in vivo bronchopulmonary responses to standard agonists, and adaptation of a technique described for larger animals. Swiss, CBA, and IL-5 transgenic mice were immunized with ovalbumin and challenged intranasally after 14 days. Immunization alone was followed by increased eosinophil counts in bone marrow and blood, whereas antigenic challenge induced eosinophil infiltration in lungs and bronchoalveolar lavage fluid, which was suppressed by dexamethasone. Despite the high eosinophil counts, no bronchopulmonary hyperreactivity to methacholine or serotonin was detected 3 to 96 hours after antigenic provocation. Our results demonstrate that immunization augments the production of eosinophils by mice, which is further increased by antigenic challenge, but that eosinophil overproduction and lung infiltration, per se, are not sufficient to induce bronchopulmonary hyperreactivity, even in constitutively hypereosinophilic IL-5 transgenic mice.