Relevance of lower airway bacterial colonization, airway inflammation, and pulmonary function in the stable stage of chronic obstructive pulmonary disease

The objective of this investigation was to verify the hypothesis that the presence of lower airway bacterial colonization (LABC) can be a stimulating factor of airway inflammation, more frequent exacerbation, and impact on pulmonary function, independent of current tobacco smoking in the stable phase of chronic obstructive pulmonary disease (COPD). A total of 46 ex-smokers with moderate to severe COPD, 19 healthy non-smokers, and 17 ex-smokers without COPD were included in this study. Their sputum specimens were collected at the first baseline visit and at the second visit after a follow-up of one year. The samples were analyzed for bacterial growth by culture, and the levels of interleukin (IL)-6, IL-8, and tumor necrosis factor alpha (TNF-α) were measured by enzyme-linked immunosorbent assay (ELISA). The frequencies of exacerbations and pulmonary function were compared at visit 2. At visit 1, 37.0% (17/46) were found to have LABC with bacterial loads ≥10⁶ CFU/ml in their sputum specimens. Haemophilus influenzae was the predominant pathogenic organism isolated. IL-8, IL-6, and TNF-α in these patients’ sputum were significantly higher than those without LABC (p < 0.05). It was the presence of LABC that contributed to the significantly elevated IL-8 and IL-6 at the 1-year period (p < 0.05). LABC was also associated with significantly increased frequencies of exacerbations and declined forced expiratory volume in 1 s (FEV₁) (p < 0.05). LABC was documented in a subpopulation of stable COPD patients; it may be responsible for the deterioration of pulmonary function of COPD patients by promoting airway inflammation and/or increased frequency of exacerbations independently of tobacco smoking.     

Association between peripheral blood WBCs C3aR mRNA level and plasma C3a,C3aR,IL-1β concentrations and acute exacerbation of chronic obstructive pulmonary disease

BackgroundThe relationship between C3a-C3aR, IL-1β, and the acute exacerbation of chronic obstructive pulmonary disease is still unclear. This study aims to explore the expression levels of C3aR in peripheral blood WBCs and the concentrations of C3a, C3aR, and IL-1β in plasma in healthy controls and patients with chronic obstructive pulmonary disease (COPD).MethodsWBCs C3aR level in the peripheral blood, the concentrations of C3a, C3aR, and IL-1β in plasma were measured in 60 patients with acute exacerbation of COPD (AECOPD), 30 patients with stable COPD (SCOPD), and 30 healthy controls. The baseline characteristics and clinical data collected from enrolled patients, including age, gender, laboratory indicators, and lung function. We analyzed the correlation between C3a, C3aR, IL-1β, and lung function indicators (forced expiratory volume in the first second as a percentage of predicted value, FEV₁%pred) in the AECOPD group.ResultsThe white blood cell count (WBC), neutrophil/lymphocyte ratio (NLR), and C-reactive protein (CRP) of patients in COPD were higher than in healthy controls (P < 0.05). The peripheral blood WBCs C3aR mRNA and plasma C3a, C3aR, and IL-1β in AECOPD were higher than in SCOPD and healthy controls (P < 0.05). The peripheral blood WBCs C3aR mRNA and plasma C3aR, and IL-1β in AECOPD combined with respiratory failure were higher than in the non-respiratory failure group (P < 0.05). The peripheral blood WBCs C3aR mRNA and plasma C3a, C3aR, and IL-1β in AECOPD with high-risk were higher than in the low-risk group (P < 0.05). The peripheral blood WBCs C3aR mRNA and plasma C3a, C3aR, and IL-1β in AECOPD were negatively correlated with FEV₁pred%. The peripheral blood WBCs C3aR mRNA, the plasma C3a and C3aR in AECOPD were positively correlated with IL-1β.ConclusionThe peripheral blood WBCs C3aR mRNA and plasma C3a, C3aR, and IL-1β in COPD patients were significantly related to the risk of disease deterioration. The C3a-C3aR axis may be involved in airway inflammation in patients with COPD.     

Assessment of bacterial exposure on phagocytic capability and surface marker expression of sputum macrophages and neutrophils in COPD patients

Defective phagocytosis has been shown in chronic obstructive pulmonary disease (COPD) bronchoalveolar lavage and blood monocyte‐derived macrophages. Phagocytic capabilities of sputum macrophages and neutrophils in COPD are unknown. We investigated phagocytosis in these cells from COPD patients and controls. Phagocytosis of Streptococcus pneumoniae or fluorescently labelled non‐typeable Haemophilus influenzae (NTHi) by sputum macrophages and neutrophils was determined by gentamycin protection assay (COPD; n = 5) or flow cytometry in 14 COPD patients, 8 healthy smokers (HS) and 9 healthy never‐smokers (HNS). Sputum macrophages and neutrophils were differentiated by adherence for the gentamycin protection assay or receptor expression (CD206 and CD66b, respectively), by flow cytometry. The effects of NTHi on macrophage expression of CD206 and CD14 and neutrophil expression of CD16 were determined by flow cytometry. There was greater uptake of S. pneumoniae [~10‐fold more colony‐forming units (CFU)/ml] by sputum neutrophils compared to macrophages in COPD patients. Flow cytometry showed greater NTHi uptake by neutrophils compared to macrophages in COPD (67 versus 38%, respectively) and HS (61 versus 31%, respectively). NTHi uptake by macrophages was lower in HS (31%, p = 0.019) and COPD patients (38%, p = 0.069) compared to HNS (57%). NTHi uptake by neutrophils was similar between groups. NTHi exposure reduced CD206 and CD14 expression on macrophages and CD16 expression on neutrophils. Sputum neutrophils showed more phagocytic activity than macrophages. There was some evidence that bacterial phagocytosis was impaired in HS sputum macrophages, but no impairment of neutrophils was observed in HS or COPD patients. These results highlight the relative contributions of neutrophils and macrophages to bacterial clearance in COPD.     

Neutrophil CD64 as a Marker of Bacterial Infection in Acute Exacerbations of Chronic Obstructive Pulmonary Disease

Acute exacerbations of chronic obstructive pulmonary disease (AECOPD) are responsible for most mortality in patients with chronic obstructive pulmonary disease (COPD) and are caused mainly by bacterial infection. We analyzed and compared neutrophil CD64 expression (using the ratio of CD64 level in neutrophils to that in lymphocytes as an index), serum C-reactive protein (CRP), procalcitonin (PCT) levels, white blood cell (WBC) count, and neutrophil percentage among healthy subjects and patients with stable COPD or AECOPD. Compared with patients with COPD and healthy subjects, patients with AECOPD demonstrated significantly increased CD64 index, CRP, PCT, WBC count, and neutrophil percentage. Interestingly, CD64 index and PCT were both significantly higher in patients with AECOPD with positive bacterial sputum culture than those with negative culture. Furthermore, CD64 index and PCT were positively correlated in AECOPD, and there was also correlation between CD64 index and CRP, WBC, and neutrophil percentage. These data suggest that CD64 index is a relevant marker of bacterial infection in AECOPD. We divided patients with AECOPD into CD64-guided group and conventional treatment group. In CD64-guided group, clinicians prescribed antibiotics based on CD64 index; while in the conventional treatment group, clinicians relied on experience and clinical symptoms to determine the necessity for antibiotics. We found that the efficacy of antibiotic treatment in CD64-guided group was significantly improved compared with the conventional treatment group, including reduction of hospital stays and cost and shortened antibiotic treatment duration. Thus, the CD64 index has important diagnostic and therapeutic implications for antibiotic treatment of patients with AECOPD.     

Airway Microbiome Dynamics in Exacerbations of Chronic Obstructive Pulmonary Disease

Specific bacterial species are implicated in the pathogenesis of exacerbations of chronic obstructive pulmonary disease (COPD). However, recent studies of clinically stable COPD patients have demonstrated a greater diversity of airway microbiota, whose role in acute exacerbations is unclear. In this study, temporal changes in the airway microbiome before, at the onset of, and after an acute exacerbation were examined in 60 sputum samples collected from subjects enrolled in a longitudinal study of bacterial infection in COPD. Microbiome composition and predicted functions were examined using 16S rRNA-based culture-independent profiling methods. Shifts in the abundance (≥2-fold, P < 0.05) of many taxa at exacerbation and after treatment were observed. Microbiota members that were increased at exacerbation were primarily of the Proteobacteria phylum, including nontypical COPD pathogens. Changes in the bacterial composition after treatment for an exacerbation differed significantly among the therapy regimens clinically prescribed (antibiotics only, oral corticosteroids only, or both). Treatment with antibiotics alone primarily decreased the abundance of Proteobacteria, with the prolonged suppression of some microbiota members being observed. In contrast, treatment with corticosteroids alone led to enrichment for Proteobacteria and members of other phyla. Predicted metagenomes of particular microbiota members involved in these compositional shifts indicated exacerbation-associated loss of functions involved in the synthesis of antimicrobial and anti-inflammatory products, alongside enrichment in functions related to pathogen-elicited inflammation. These trends reversed upon clinical recovery. Further larger studies will be necessary to determine whether specific compositional or functional changes detected in the airway microbiome could be useful indicators of exacerbation development or outcome.     

Predicting Sputum Eosinophilia in Exacerbations of COPD Using Exhaled Nitric Oxide

Fractional exhaled nitric oxide (FENO) may be a pulmonary biomarker in chronic obstructive pulmonary disease (COPD). In this prospective study, the relationship between FENO and airway inflammation was assessed in COPD exacerbations. FENO and lung function were measured, and sputum was collected from 49 ex-smoking COPD patients, first at the time of hospital admission and again at discharge following treatment. There was a significant positive correlation between the percentage of sputum eosinophils and FENO concentrations, both at exacerbation (r?=?0.593, p?<?0.001) and discharge (r?=?0.337, p?=?0.044). The increase in forced expiratory volume in one second (FEV₁) after treatment was greater in patients with sputum eosinophilia (ΔFEV₁ 0.35?±?0.12 vs. 0.13?±?0.04 L, p?=?0.046), and FENO was a strong predictor of sputum eosinophilia (area under the receiver operating characteristic curve, 0.89). The optimum cut point was 19 parts per billion (sensitivity: 90 %; specificity: 74 %). Our data suggest that FENO is a good surrogate marker of eosinophilic inflammation in COPD patients with exacerbations.     

HMGB1 and RAGE levels in induced sputum correlate with asthma severity and neutrophil percentage

Previous work indicated that high mobility group box-1 (HMGB1) protein may be involved in neutrophilic asthma. Here, we sought to investigate the correlation between HMGB1 and one of its receptors, receptor for advanced glycosylation end products (RAGE), with the severity of bronchial asthma. Compared to the control group (30 healthy individuals), patients in the asthma group (n = 72) exhibited a higher percentage of neutrophils and higher HMGB1 and RAGE levels in induced sputum samples (P < 0.05). Concurrently, FEV₁% was significantly lower in the asthma group (P < 0.05). Further, compared to mild and moderate asthma, in patients with severe asthma ACQ scores, the percentage of neutrophils, and HMGB1 levels were significantly higher, while FEV₁% was significantly lower (P < 0.05). The percentage of neutrophils and HMGB1 and RAGE levels were lower after treatment than before treatment (P < 0.05). Finally, negative correlations were observed between HMGB1 or RAGE levels and FEV₁% (r = −0.777 and r = −0.291, P < 0.05), and positive correlations were detected between HMGB1 or RAGE levels and percentage of neutrophils (r = 0.803 and r = 0.326, P < 0.05). Additionally, positive correlations were observed between HMGB1 and RAGE levels within the asthma group (r = 0.306, P < 0.05). Therefore, HMGB1 protein levels correlate with the severity of asthma, and HMGB1 may contribute to the inflammatory process of asthma.     

Importance of viral and bacterial infections in chronic obstructive pulmonary disease exacerbations

BackgroundFew studies have evaluated the contribution of both viruses and bacteria in acute exacerbation of chronic obstructive pulmonary disease (AECOPD).ObjectivesThis study estimated the burden of both types of pathogens among adults seeking care for an AECOPD during two consecutive winter seasons.Study designPatients 50 years or older who consulted within 10 days of AECOPD onset were eligible. Clinical data were collected on a standardized questionnaire, and nasopharyngeal aspirates (NPA), paired sera, and non-induced sputum were collected. Polymerase chain reaction (PRC) assays were used to identify viral, atypical and bacterial pathogens in NPA specimen.ResultsOverall, 108 patients with AECOPD were included, 88% of patients were admitted and 2 patients (2%) received intensive care. A third of patients (31%) had evidence of a viral infection, 9% with influenza A, 7% RSV and 7% with PIV-3. One patient was positive for Mycoplasma pneumoniae. Bacterial pathogens were identified in 49% of patients with available sputum, most frequently Staphylococcus aureus, Pseudomonas aeruginosa, and Haemophilus influenzae. Among virus-infected patients, 14 (58%) also had bacteria in their sputum, but co-infected patients did not present with different symptoms than patients with single infections.ConclusionsThese results suggest that influenza and RSV are frequent contributors of AECOPD, and that coinfection with bacteria does not appear to be more severe among virus-infected patients. Clinicians should be aware that AECOPD may be frequently triggered by viruses, and may consider antivirals and proper infection control measures in appropriate epidemiological setting.     

Sputum colour reported by patients is not a reliable marker of the presence of bacteria in acute exacerbations of chronic obstructive pulmonary disease

Sputum colour is regarded as a good marker of bacterial involvement in acute exacerbations of chronic obstructive pulmonary disease (COPD) and guides many physicians in deciding on antibiotic treatment. Although most doctors rely on the sputum colour that is reported by patients, it can also be assessed using a validated colour chart. In this study, reported sputum colour and assessed sputum colour were compared as markers of the presence of bacteria, bacterial load, and systemic inflammation. Data on 257 exacerbations in 216 patients hospitalized with an acute exacerbation were analysed (mean age, 72 years; mean forced expiratory volume in 1 s, 44.8% ± 17.8% (± standard deviation)). Sputum colour was reported by the patients and assessed at the laboratory with a colour chart. Subsequently, quantitative sputum cultures were performed. C-reactive protein was measured as a marker of systemic inflammation. A sputum sample was obtained in 216 exacerbations (84%), of which 177 (82%) were representative. A pathogen was identified in 155 patients (60%). Assessed sputum colour was a better marker of the presence of bacteria (OR 9.8; 95% CI 4.7–20.4; p <0.001) than reported sputum colour (OR 1.7; 95% CI 1.0–3.0; p 0.041). The sensitivity and specificity were 73% and 39% for reported sputum colour, and 90% and 52% for assessed sputum colour. Assessed sputum colour was clearly related to sputum bacterial load and C-reactive protein levels, whereas reported sputum colour was not. It is concluded that sputum colour reported by patients is an unreliable marker of the presence of bacteria in acute exacerbations of COPD. Assessed sputum colour is clearly superior and is also related to bacterial load and systemic inflammation.     

Inflammatory cell profiles and T-lymphocyte subsets in chronic obstructive pulmonary disease and severe persistent asthma

Background Severe persistent asthma (SPA) and chronic obstructive pulmonary disease (COPD) are both associated with non‐reversible airflow limitation and airway neutrophilia. Objective To compare inflammatory cell profiles and T lymphocyte subsets between SPA and COPD patients with similar severity of airflow limitation. Methods Sputum induction and lung function tests were performed in 15 COPD patients aged (mean±SD) 68±8 years, ex‐smokers, mean forced expiratory volume in 1 s (FEV₁) 45% of predicted (%pred) and 13 SPA aged 55±10 years, non‐smokers, mean FEV₁ 49%pred. All patients were on inhaled steroid treatment. Eight asthmatics exhibited irreversible airflow limitation. Differential cell count, metachromatic cell count and double immunocytochemistry for the analysis of T lymphocyte subsets were performed on sputum slides. Results COPD patients had increased sputum neutrophils in comparison with SPA (P<0.03), but similar to SPA with fixed obstruction. In COPD sputum neutrophils negatively correlated with the lung transfer factor for carbon monoxide (K_CO) (r=?0.462, P=0.04). SPA showed significantly increased eosinophils and metachromatic cells vs. COPD patients (P<0.04, P<0.007, respectively). Increased CD4/CD8 and decreased CD4‐IFN‐γ/CD4‐IL4⁺ cell ratio (P<0.001) were found in SPA vs. COPD. In SPA, CD4/CD8⁺ cell ratio correlated with sputum eosinophils (r=0.567, P=0.04). Conclusion In spite of treatment with inhaled steroids, SPA and COPD exhibit distinct sputum inflammatory cell patterns, although SPA with fixed airflow limitation and COPD patients have similar numbers of neutrophils.     

Elevated HMGB1-related interleukin-6 is associated with dynamic responses of monocytes in patients with active pulmonary tuberculosis

There were limited studies assessing the role of HMGB1 in TB infection. In this prospective study, we aimed to assess the levels of HMGB1 in plasma or sputum from active pulmonary tuberculosis (APTB) patients positive for Mtb culture test, and to evaluate its relationship with inflammatory cytokines and innate immune cells. A total of 36 sputum Mtb culture positive APTB patients and 32 healthy volunteers (HV) were included. Differentiated THP-1 cells were treated for 6, 12 and 24 hrs with BCG at a multiplicity of infection of 10. The absolute values and percentages of white blood cells (WBC), neutrophils, lymphocytes, and monocytes were detected by an automatic blood analyzer. Levels of HMGB1, IL-6, IL-10 and TNF-α in plasma, sputum, or cell culture supernatant were measured by ELISA. The blood levels of HMGB1, IL-6, IL-10 and TNF-α, the absolute values of WBC, monocytes and neutrophils, and the percentage of monocytes were significant higher in APTB patients than those in HV groups (P<0.05). The sputum levels of HMGB1, IL-10, and TNF-α were also significantly higher in APTB patients than those in HV groups (P<0.05). Meanwhile, plasma level of HMGB1, IL-6, and IL-10 in APTB patients were positively correlated with those in sputum (P<0.05), respectively. IL-6 was positively correlated with HMGB1 both in plasma and sputum of APTB patients (P<0.05). HMGB1 and IL-6 is positively correlated with the absolute number of monocytes in APTB patients (P<0.05). BCG induced HMGB1, IL-6, IL-10 and TNF-α production effectively in PMA-treated THP-1 cells. HMGB1 may be used as an attractive biomarker for APTB diagnosis and prognosis and may reflect the inflammatory status of monocytes in patients with APTB.     

Chlamydia pneumoniae Infection in Acute Exacerbations of Chronic Obstructive Pulmonary Disease: Analysis of 250 Hospitalizations

Two hundred fifty hospitalizations were included in a serologically based prospective study to assess the role of Chlamydia pneumoniae in episodes of acute exacerbation of chronic obstructive pulmonary disease (AECOPD) and the percentage of COPD patients chronically infected with this pathogen. Chlamydia pneumoniae-specific IgG, IgA and IgM antibody titers were determined using a commercial kit with the microimmunofluorescence method. A significantly higher geometric mean titer in the COPD patients compared to the control group was found for IgG (P<0.00001) and IgA (P<0.000001). The serological criterion for chronic Chlamydia pneumoniae infection (IgG ≥128 concomitant with IgA ≥64) was positive in 73 (33.3%) COPD patients compared with 7 (7%) controls (P=0.000001). No difference was found in any serological parameter when the study population was divided by severity of COPD. When the serological profiles were compared between the first and second of 31 pairs of hospitalizations, 7 of the 62 (11.3%) hospitalizations showed evidence of acute infection with Chlamydia pneumoniae around one of the episodes of AECOPD. It is concluded that compared with the control group, the COPD patients had a significantly higher prevalence of chronic Chlamydia pneumoniae infection. In the COPD group, there was no correlation between the severity of the disease and the rate of chronic Chlamydia pneumoniae infection. In a substantial percentage of AECOPD cases, there is serological evidence of acute Chlamydia pneumoniae infection around the time of the exacerbation. The clinical and pathophysiologic implications of these findings should be clarified by further studies. Electronic Publication     

Supplementation with Astragalus polysaccharides alters Aeromonas-induced tissue-specific cellular immune response

Members of the genus Aeromonas inhabit various aquatic environments and are responsible for a number of intestinal and extra-intestinal infections in humans as well as other animals. Astragalus species are used in Chinese traditional medicine as antiperspirant, antihypertensive, diuretic and tonic treatments and have been used for treatment of patients with leukemia and uterine cancers. The present study was aimed to investigate immunomodulatory effect of Astragalus polysaccharides (APS) treatment on Aeromonas hydrophila-infected mice. The present data showed that APS-treatment ameliorated neutrophils phagocytic activity and reactive oxygen species (ROS) production in intestinal tissues of infected mice. Moreover, APS treatment induced a highly significantly (P < 0.001) increase in the number of CD4⁺ T cells in the intestinal tissues and thymus, however, number of CD4⁺ T cells in the spleens of infected mice not significantly changed with APS treatment. On the other hand, APS-treatment caused a very highly significant (P < 0.001) decrease in the number of CD8⁺ T cells in the spleens and thymus of infected mice. In conclusion, the present data suggested that APS treatment reduced ROS production, downmodulated neutrophils activity, and increased CD4⁺/CD8⁺ T cells ratio in A. hydrophila-infected mice.     

Assessment of the efficacy of polyclonal intravenous immunoglobulin G (IVIG) against the infectivity of clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) in vitro and in vivo

The response to treatment of severe methicillin-resistant Staphylococcus aureus (MRSA) infections with the traditional antibiotics is sometimes unsatisfactory and multiple antibiotic resistance is common. Adjuvant therapy such as intravenous immunoglobulin G (IVIG) could possibly be helpful in the treatment of such infections. The effect of IVIG on the capacity of human neutrophils to phagocytose and kill MRSA was investigated in vitro using the MTT assay and measuring the production of reactive oxygen species (ROS) and nitric oxide (NO). The efficiency of IVIG in neutralizing α-hemolysin and coagulase of MRSA was also assessed. The capability of IVIG in the treatment and prevention of MRSA infections was also evaluated in a murine peritonitis model. IVIG significantly enhanced (p?<?0.01) the killing of MRSA by neutrophils at all concentrations tested (0.1–5 mg/ml) by 30–80 % of control values. It significantly (p?<?0.01) increased the level of NO production in a dose-dependent manner, giving up to 60 μM at 5 mg/ml. The ROS level significantly increased (p?<?0.01) in the presence of IVIG. In addition, IVIG significantly reduced the hemolytic activity of MRSA 10-fold and its coagulation capabilities by 50 %. When tested in vivo, groups receiving IVIG via tail vein infusion showed no significant improvement in their survival. Only when delivered to the same site of infection did IVIG show an improvement in the survival of mice (n?=?80). These results could pave the way for a better understanding of the mechanism of action of IVIG and suggest its clinical potential as an adjuvant preventive and therapeutic agent against life-threatening infections caused by MRSA and other bacteria.     

Pseudomonas aeruginosa and risk of death and exacerbations in patients with chronic obstructive pulmonary disease: an observational cohort study of 22 053 patients

ObjectivesThe role of Pseudomonas aeruginosa in the long-term prognosis of chronic obstructive pulmonary disease (COPD) is unknown. The purpose of this study was to determine whether P. aeruginosa is associated with increased risk of exacerbations or death in patients with COPD.MethodsThis is a multiregional epidemiological study based on complete data on COPD outpatients between 1 January 2010 and 31 October 2017 and corresponding microbiology and national register data. Time-dependent Cox proportional hazards models and propensity matching was used to estimate hospitalization-demanding exacerbations and death after 2 years, separately and in combination.ResultsA total of 22 053 COPD outpatients were followed for a median of 1082 days (interquartile-range: 427–1862). P. aeruginosa was present in 905 (4.1%) patients. During 730 days of follow-up, P. aeruginosa strongly and independently predicted an increased risk of hospitalization for exacerbation or all-cause death (HR 2.8, 95%CI 2.2–3.6; p <0.0001) and all-cause death (HR 2.7, 95%CI 2.3–3.4; p <0.0001) in analyses adjusted for known and suspected confounders. The signal remained unchanged in unadjusted analyses as well as propensity-matched subgroup analyses. Among patients ‘ever colonized’ with P. aeruginosa, the incidence of hospital-demanding exacerbations doubled after the time of the first colonization.ConclusionsCOPD patients in whom P. aeruginosa can be cultured from the airways had a markedly increased risk of exacerbations and death. It is still not clear whether this risk can be reduced by offering patients targeted antipseudomonal antibiotics. A randomized trial is currently recruiting patients to clarify this (ClinicalTrials.gov: NCT03262142).     

Proliferation and production of interferon-gamma (IFN-γ) and IL-4 in response to Staphylococcus aureus and Staphylococcal superantigen in childhood atopic dermatitis

We have examined the cell-mediated immunity (CMI) to Staphylococcus aureus(S. aureus) and Staphylococcal enterotoxin B (SEB) in peripheral blood mononuclear cells (PBMC) from children with atopic dermatitis (AD) and from non-atopic child controls by measurement of proliferative responses and production of the cytokines IFN-γ and IL-4. PBMC from children with AD showed significantly higher proliferative responses to both S. aureus (P <0.01) and SEB (P < 0.05). Despite this enhanced proliferation, production of IFN-γ in response to S. aureus (P <0.001) and SEB (P < 0.01) from these PBMC was significantly diminished. In contrast, PBMC from children with AD were significantly more likely to produce IL-4 in response to S. aureus (P < 0.01). These findings demonstrate in vitro heightened CMI to S. aureus in children with AD, and implicate S. aureus as a potent inflammatory stimulant. Impaired IFN-γ production to S. aureusin vivo may result in failure to eradicate S. aureus from skin. The organism's persistence on skin would contribute to inflammation by causing continued T cell activation and release of pro-inflammatory mediators.     

Increased CTLA‐4+ T cells may contribute to impaired T helper type 1 immune responses in patients with chronic obstructive pulmonary disease

Impaired T helper type 1 (Th1) function is implicated in the susceptibility of patients with chronic obstructive pulmonary disease (COPD) to respiratory infections, which are common causes of acute exacerbations of COPD (AECOPD). To understand the underlying mechanisms, we assessed regulatory T (Treg) cells and the expression of an inhibitory T‐cell receptor, cytotoxic T‐lymphocyte‐associated antigen 4 (CTLA‐4). Cryopreserved peripheral blood mononuclear cells (PBMC) from patients with AECOPD (n = 17), patients with stable COPD (sCOPD; n = 24) and age‐matched healthy non‐smoking controls (n = 26) were cultured for 24 hr with brefeldin‐A or monensin to detect intracellular or surface CTLA‐4 (respectively) by flow cytometry. T cells in PBMC from AECOPD (n = 9), sCOPD (n = 14) and controls (n = 12) were stimulated with anti‐CD3 with and without anti‐CTLA‐4 blocking antibodies and cytokines were quantified by ELISA. Frequencies of circulating T cells expressing intracellular CTLA‐4 were higher in sCOPD (P = 0·01), whereas patients with AECOPD had more T cells expressing surface CTLA‐4 than healthy controls (P = 0·03). Increased frequencies of surface CTLA‐4⁺ CD4⁺ T cells and CTLA‐4⁺ Treg cells paralleled increases in plasma soluble tumour necrosis factor receptor‐1 levels (r = 0·32, P = 0·01 and r = 0·29, P = 0·02, respectively) in all subjects. Interferon‐γ responses to anti‐CD3 stimulation were inversely proportional to frequencies of CD4⁺ T cells expressing intracellular CTLA‐4 (r = −0·43, P = 0·01). Moreover, CTLA‐4 blockade increased the induction of interferon‐γ, tumour necrosis factor‐α and interleukin‐6 in PBMC stimulated with anti‐CD3. Overall, chronic inflammation may expand sub‐populations of T cells expressing CTLA‐4 in COPD patients and therefore impair T‐cell function. CTLA‐4 blockade may restore Th1 function in patients with COPD and so aid the clearance of bacterial pathogens responsible for AECOPD.     

Febrile infection changes the expression of IgG Fc receptors and complement receptors in human neutrophils in vivo

We have examined the expression of the IgG Fc receptors (FcRI, FcRII, FcRIII) and complement receptors (CR1, CR3) in neutrophils from 42 patients with febrile bacterial infection, 20 patients with febrile viral infection and 69 non-febrile healthy individuals. Using receptor-specific MoAbs and immunofluorescence flow cytometry the relative fluorescence intensity (as a measure of receptor number) and the proportion of receptor-positive cells were determined in peripheral blood neutrophils exposed to minimal processing, consisting only of erythrolysis. Both the percentage of FcRI-positive neutrophils and FcRI number per neutrophil were significantly (P<0.001 and P<0.0001) increased in bacterial infected patients compared with controls, whereas in viral infected patients only the FcRI percentage was markedly elevated (P<0.05). In addition, both FcRII and CR1 levels were significantly higher in the bacterial infection group than in the viral infection and control groups (bacterial versus control P<0.001, bacterial versus viral P<0.0001). No changes in expression of FcRIII or CR3 were found in the patient groups. The kinetic analysis of receptor expression in bacterial infection patients revealed a shift in the percentage of FcRI-bearing neutrophils towards normal values already on day 2 after the first analysis. On the other hand, the levels of FcRI, FcRII and CR1 remained clearly elevated in these patients during 1 week's follow-up period. We conclude that febrile infection may cause systemic activation of the entire pool of circulating neutrophils, resulting in alterations in cell surface receptor expression, some of which are characteristic of the nature of the infectious agent.     

Cytokines and Inflammatory Mediators Do Not Indicate Acute Infection in Cystic Fibrosis

Various treatment regimens and difficulties with research design are encountered with cystic fibrosis (CF) because no standard diagnostic criteria exist for defining acute respiratory exacerbations. This study evaluated the role of serial monitoring of concentrations of selected cytokines and inflammatory mediators in serum and sputum as predictors of respiratory exacerbation, as useful outcome measures for CF, and to guide therapy. Interleukin-8 (IL-8), tumor necrosis factor alpha (TNF-α), neutrophil elastase-α-1-protease inhibitor complex (NE complex), protein, and α-1-protease inhibitor (α-1-PI) were measured in serum and sputum collected from CF patients during respiratory exacerbations and periods of well-being. Levels of NE complex, protein, and α-1-PI in sputum rose during respiratory exacerbations and fell after institution of antibiotic therapy (P = 0.078, 0.001, and 0.002, respectively). Mean (± standard error of the mean) levels of IL-8 and TNF-α were extremely high in sputum (13,780 ± 916 and 249.4 ± 23.5 ng/liter, respectively) but did not change significantly with clinical deterioration of the patient (P > 0.23). IL-8 and TNF-α were generally undetectable in serum, and therefore these measures were unhelpful. Drop in forced expiratory volume in 1 s was the only clinical or laboratory parameter that was close to being a determinant of respiratory exacerbation (P = 0.055). This study provides evidence of intense immunological activity occurring continually within the lungs of adult CF patients. Measurement of cytokines and inflammatory mediators in CF sputum is not helpful for identifying acute respiratory exacerbations.     

Assembly of NADPH Oxidase in Human Neutrophils Is Modulated by the Opacity-Associated Protein Expression State of Neisseria gonorrhoeae

Neisseria gonorrhoeae (the gonococcus, Gc) triggers a potent inflammatory response and recruitment of neutrophils to the site of infection. Gc survives exposure to neutrophils despite these cells'' antimicrobial products, such as reactive oxygen species (ROS). ROS production in neutrophils is initiated by NADPH oxidase, which converts oxygen into superoxide. The subunits of NADPH oxidase are spatially separated between granules (gp91^phox/p22^phox) and the cytoplasm (p47^phox, p67^phox, and p40^phox). Activation of neutrophils promotes the coassembly of NADPH oxidase subunits at phagosome and/or plasma membranes. While Gc-expressing opacity-associated (Opa) proteins can induce neutrophils to produce ROS, Opa-negative (Opa⁻) Gc does not stimulate neutrophil ROS production. Using constitutively Opa⁻ and OpaD-positive (OpaD⁺) Gc bacteria in strain FA1090, we now show that the difference in ROS production levels in primary human neutrophils between these backgrounds can be attributed to differential assembly of NADPH oxidase. Neutrophils infected with Opa⁻ Gc showed limited translocation of NADPH oxidase cytoplasmic subunits to cellular membranes, including the bacterial phagosome. In contrast, these subunits rapidly translocated to neutrophil membranes following infection with OpaD⁺ Gc. gp91^phox and p22^phox were recruited to Gc phagosomes regardless of bacterial Opa expression. These results suggest that Opa⁻ Gc interferes with the recruitment of neutrophil NADPH oxidase cytoplasmic subunits to membranes, in particular, the p47^phox “organizing” subunit, to prevent assembly of the holoenzyme, resulting in an absence of the oxidative burst.