Increased CTLA-4 + T-cells may contribute to impaired Th 1 immune responses in patients with chronic obstructive pulmonary disease ( COPD )

Impaired T-helper (Th)-1 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-cells (Treg) and the expression of an inhibitory T-cell receptor, cytotoxic T-lymphocyte-associated antigen (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 24hr 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), whilst AECOPD patients had more Tcells expressing surface CTLA-4 than healthy controls (p=0.03). Increased frequencies of surface CTLA-4 CD4 T-cells and CTLA-4Treg paralleled increases in plasma sTNFR1 levels (r=0.32, p=0.01 and r=0.29, p=0.02 respectively) in all subjects. IFNγ 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 IFNγ, TNFα and IL-6 in PBMC stimulated with anti-CD3. Overall, chronic inflammation may expand sub-populations of T-cells expressing CTLA-4 in COPD patients A cc ep te d A rt ic le This article is protected by copyright. All rights reserved. and therefore impair T-cell function. CTLA-4 blockade may restore Th1 function in COPD patients and thus aid the clearance of bacterial pathogens responsible for AECOPD. INTRODUCTION Chronic obstructive pulmonary disease (COPD) is one of the top five causes of morbidity and mortality worldwide. Smoking is the most common cause of COPD, but genetic factors and environmental pollutions are also implicated (1). The disease is characterized by progressive limitation of airflow, with abnormal inflammatory responses to inhaled particles or gases (2). The chronic inflammation of the airways involves infiltration and activation of neutrophils, macrophages and T-cells (3-5). Subsequent systemic inflammation contributes to comorbidities (e.g. cardiovascular disease and cachexia) (6). Most patients are relatively stable with treatment (sCOPD), but some experience acute exacerbations of COPD (AECOPD), which increase mortality and morbidity (7). AECOPD is defined as “an event in the natural course of the disease characterized by a change in the patient’s baseline dyspnea, cough and/or sputum, and beyond normal day-to-day variations, that is acute in onset and may warrant a change in regular medication in a patient with underlying COPD’’ (2). Respiratory bacterial infections account for up to 50% of AECOPD events. Non-typeable Haemophilus influenzae (NTHI), Streptococcus pneumoniae and Moraxella catarrhalis are the major bacterial pathogens isolated from patients with AECOPD (8). As NTHI oral vaccines do not reduce the frequency and severity of AECOPD (9), the capacity to mount a protective anti-bacterial immune response may be limited in patients with COPD. A cc ep te d A rt ic le This article is protected by copyright. All rights reserved. Despite its inflammatory aetiology, COPD is considered as an immune-deficient state as the abundant activated T-cells in the airways of COPD patients do not eradicate bacterial infections. Indeed, T-helper (Th)-1 immune responses [e.g. production of interferon (IFN)γ and phagocytosis] are impaired in COPD patients (10-13). Exogenous IFNγ can enhance killing of NTHI by monocytes from patients with bronchiectasis (14), confirming the necessity for appropriate Th1 responses for clearance of bacterial infections. Here we address the regulators of T-cell responses in COPD patients and search for means to improve host production of IFNγ. An important regulator of T-cell function is cytotoxic T-lymphocyte antigen-4 (CTLA-4), which blocks the CD28-mediated activation of T-cells (15). CTLA-4 is constitutively expressed by regulatory T-cells (Tregs) and activated effector T-cells. It inhibits the proliferation of T-cells and production of pro-inflammatory cytokines, and so prevents continuous T-cell activation (15, 16). Polymorphisms in the gene encoding CTLA-4 (e.g. rs231775 and rs5742909) and increased serum levels of the soluble form of CTLA-4 have been associated with dysregulated T-cell responses and increased susceptibility to COPD and chronic bronchitis (17-20). Furthermore, increased levels of soluble CTLA-4 in the serum of COPD patients paralleled decreased lung function and increased C-reactive protein (CRP) (21). Frequencies of circulating CTLA-4 T-cells and CTLA-4 Tregs were also higher in COPD patients than controls (22, 23). Kalathil et al. reported that CTLA-4 blockade in vitro increased the proliferation of CD4 and CD8 T-cells and production of IFNγ by peripheral blood mononuclear cells (PBMC) from three COPD patients (24).