Treatment with anti‐OX40L or anti‐TSLP does not alter the frequency of T regulatory cells in allergic asthmatics

OX40‐OX40L interactions and thymic stromal lymphopoietin (TSLP) are important in the induction and maintenance of Th2 responses in allergic disease, whereas T regulatory cells (Treg) have been shown to suppress pro‐inflammatory Th2 responses. Both OX40L and TSLP have been implicated in the negative regulation of Treg. The effect of anti‐asthma therapies on Treg is not well known. Our aim was to assess the effects of two monoclonal antibody therapies (anti‐OX40L and anti‐TSLP) on Treg frequency using a human model of allergic asthma. We hypothesized that the anti‐inflammatory effects of these therapies would result in an increase in circulating Treg (CD4⁺CD25⁺CD127^lowFoxp3⁺ cells) frequency. We measured Treg using flow cytometry, and our results showed that neither allergen challenge nor monoclonal antibody therapy altered circulating Treg frequency. These data highlight the need for assessment of airway Treg and for a more complete understanding of Treg biology so as to develop pharmacologics/biologics that modulate Treg for asthma therapy.     

Monomeric C‐reactive protein and inflammation in age‐related macular degeneration

Age‐related macular degeneration (AMD) is a devastating disease characterized by central vision loss in elderly individuals. Previous studies have suggested a link between elevated levels of total C‐reactive protein (CRP) in the choroid, CFH genotype, and AMD status; however, the structural form of CRP present in the choroid, its relationship to CFH genotype, and its functional consequences have not been assessed. In this report, we studied genotyped human donor eyes (n = 60) and found that eyes homozygous for the high‐risk CFH (Y402H) allele had elevated monomeric CRP (mCRP) within the choriocapillaris and Bruch's membrane, compared to those with the low‐risk genotype. Treatment of choroidal endothelial cells in vitro with mCRP increased migration rate and monolayer permeability compared to treatment with pentameric CRP (pCRP) or medium alone. Organ cultures treated with mCRP exhibited dramatically altered expression of inflammatory genes as assessed by RNA sequencing, including ICAM‐1 and CA4, both of which were confirmed at the protein level. Our data indicate that mCRP is the more abundant form of CRP in human choroid, and that mCRP levels are elevated in individuals with the high‐risk CFH genotype. Moreover, pro‐inflammatory mCRP significantly affects endothelial cell phenotypes in vitro and ex vivo, suggesting a role for mCRP in choroidal vascular dysfunction in AMD. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Thymic stromal lymphopoietin is a critical mediator of IL‐13‐driven allergic inflammation

Both thymic stromal lymphopoietin (TSLP) and IL‐13 are essential cytokines for the development of allergic inflammation. However, a causal link between TSLP and IL‐13 has not yet been fully elucidated. This study aimed to investigate whether IL‐13 induces TSLP expression and whether the induction contributes to the development of allergic inflammation. We found that IL‐13 induced TSLP expression in mouse nasal tissue specimens in a Stat6‐dependent manner. In addition, intranasal challenge of mice with IL‐13 induced TSLP expression in the nasal epithelium. Importantly, intranasal IL‐13 challenge induced eosinophilia and goblet cell hyperplasia in the nasal mucosa in mice, which was inhibited by the blockade of TSLP activity with anti‐TSLP Ab. These findings suggest that TSLP is an important mediator of IL‐13‐driven allergic inflammation in the nasal mucosa. Taken together with the recent findings that IL‐13 is a critical downstream element for TSLP‐driven allergic inflammation, TSLP may function both upstream and downstream of IL‐13, thus providing an additional rationale as to why TSLP plays such a central role in the development of allergic inflammation.     

The use of exhaled nitric oxide concentration to identify eosinophilic airway inflammation: an observational study in adults with asthma

BACKGROUND: Assessment of eosinophilic airway inflammation may be helpful in the management of asthma. Nitric oxide (NO) has potential advantages as a tool to monitor airway inflammation although little is known about the relationship between NO and eosinophilic airway inflammation and the factors which influence it. METHODS: We set out to define the relationship between exhaled NO and the sputum eosinophil count, identify the exhaled NO concentration that best identified a sputum eosinophil count >3% and investigate the impact of several potential confounding factors in 566 consecutive patients with varying severity of asthma. Finally we examined the ability of exhaled NO concentrations measured at differing exhalation flows to identify the presence of a sputum eosinophilia. RESULTS: We found a significant positive relationship between exhaled NO and sputum eosinophil count (R(2)=0.26, P<0.001) which was best described using a non-linear model. There were no clinically important confounding factors to this model. In non-smokers an exhaled NO concentration of >8.3 p.p.b. at 250 mL/s gave 71% sensitivity and 72% specificity for identifying a sputum eosinophil count of >3%. CONCLUSIONS: This value of exhaled NO would seem to be the best for identifying significant eosinophilic airway inflammation. It is applicable to a wide range of non-smoking patients with asthma; exhalation flow does not alter the ability of exhaled NO concentration to detect a sputum eosinophilia.     

High serum levels of tumour necrosis factor‐α and interleukin‐8 in severe asthma: markers of systemic inflammation?

BACKGROUND: Severe asthma is characterized by elevated levels of pro-inflammatory cytokines and neutrophilic inflammation in the airways. Blood cytokines, markers of 'systemic' inflammation, may be a feature of amplified inflammation in severe asthma. OBJECTIVE: To detect differences in IL-8, TNF-alpha, IL-16 and IL-13 levels in the serum(s) of stable severe and mild-moderate asthmatics related to blood leucocytes proportion, airway calibre and exhaled nitric oxide (NO) levels. METHODS: We assessed cytokine serum levels by ELISA and blood leucocyte counts by an alkaline peroxidase method in 20 healthy controls, 22 mild-moderate [forced expiratory volume in 1 s (FEV1)(%pred): 89+/-3] and 14 severe asthmatics [FEV1(%pred): 49+/-2]. RESULTS: IL-8 and TNF-alpha levels were higher in severe asthmatics than in mild-moderate asthmatics or in controls (P<0.05). No differences in IL-16 and IL-13 levels were detected. Severe asthmatics showed higher circulating neutrophil and eosinophil number than controls (P<0.05). In severe asthmatics, exhaled NO levels were superior than in controls (P<0.05), but inferior than in mild-moderate asthmatics (P<0.05). We found positive correlation between TNF-alpha levels and exhaled NO (r=0.67; P=0.01) or circulating neutrophil counts (r=0.57; P=0.03) in severe asthmatics. CONCLUSION: sTNF-alpha and sIL-8 are markers of 'systemic' inflammation in severe asthmatics, in conjunction with augmented circulating neutrophils, suggesting the involvement of neutrophil-derived cytokine pattern in severe asthma.     

Allergen inhalation decreases adenosine receptor expression in sputum and blood of asthma patients

Background: Adenosine is a signalling nucleoside that has been proposed to contribute to the pathogenesis of asthma. Adenosine is produced in inflammatory environments and acts via adenosine receptors (A₁R, A_2AR, A_2BR, and A₃R) expressed by a wide variety of cells, resulting in pro‐ and anti‐inflammatory effects. Objective: To compare AR expression in asthma patients and healthy subjects, and to assess the effect of allergen challenge on AR expression of inflammatory cells and on cytokines in peripheral blood and sputum in asthma. Methods: Asthma patients underwent an allergen challenge, and blood and induced sputum samples were taken before and 24 h after allergen challenge to study inflammatory cells numbers, AR expression and cytokine production. Blood and sputum were investigated at one time point in healthy subjects. AR expression was measured by flow cytometry (blood) or on cytospins using immunocytochemistry (sputum). Cytokines (luminex, ELISA) and adenosine (HPLC) were measured in sputum supernatant. Results: The percentage of A_2BR expressing neutrophils in sputum was lower in asthma patients than in healthy subjects (P = 0.016). Allergen challenge decreased A₁R and A_2AR expression on neutrophils and A₁R expression on T cells in peripheral blood (all P < 0.05). Allergen challenge increased IL‐8 levels and eosinophil numbers (P < 0.05), whereas it decreased thymic stromal lymphopoietin levels and the percentage of A₁R expressing macrophages in induced sputum (P < 0.05). Conclusions: Allergen challenge has a down‐regulatory effect on AR expression in asthma, suggesting a contribution of adenosine‐related effector mechanisms in the pathophysiology.     

Nitric oxide attenuates vascular endothelial cadherin‐mediated vascular integrity in human chronic inflammation

In this study, we examined the role of nitric oxide (NO) in controlling vascular integrity mediated by vascular endothelial (VE)‐cadherin in chronic inflammation. Periapical granulomas were analysed for the expression of inducible NO synthase (iNOS) and VE‐cadherin, and more iNOS expression than VE‐cadherin was shown. Human umbilical vein endothelial cells (HUVECs) were stimulated with proinflammatory cytokines and lipopolysaccharide extracted from Porphyromonas gingivalis and it induced iNOS expression, whereas it reduced VE‐cadherin expression, compared with negative controls. On the other hand, pre‐incubation with 1400W, an iNOS‐specific inhibitor, markedly reduced iNOS expression in stimulated HUVECs and restored VE‐cadherin expression to its control level, suggesting that vascular integrity was modulated in conjunction with the reduction of NO. Immunocytochemistry confirmed the functional role of NO in cultured HUVEC monolayers with or without 1400W. These data are consistent with a hypothesis suggesting that NO could attenuate VE‐cadherin‐mediated vascular integrity in human chronic inflammation.