Kerstin A. Heyl

Chronic obstructive pulmonary disease (COPD) is complicated by infectious exacerbations with acute worsening of respiratory symptoms. Coinfections of bacterial and viral pathogens are associated with more severe exacerbations. Moraxella catarrhalis is one of the most frequent lower respiratory tract pathogens detected in COPD. We therefore studied the impact of M. catarrhalis on the antiviral innate immune response that is mediated via TLR3 and p53. Molecular interactions between M. catarrhalis and normal human bronchial epithelial (NHBE) cells as well as Beas‐2B cells were studied using flow cytometry, quantitative PCR analysis, chromatin immunoprecipitation, RNA interference, and ELISA. M. catarrhalis induces a significant down‐regulation of TLR3 in human bronchial epithelial cells. In M. catarrhalis ‐infected cells, expression of p53 was decreased. We detected a reduced binding of p53 to the tlr3 promoter, resulting in reduced TLR3 gene transcription. M. catarrhalis diminished the TLR3‐dependent secretion of IFN‐β, IFN‐λ, and chemokine (C‐X‐C motif) ligand 8. In addition in M. catarrhalis infected cells , expression of rhinovirus type 1A RNA was increased compared with uninfected cells. M. catarrhalis reduces antiviral defense functions of bronchial epithelial cells, which may increase susceptibility to viral infections.—Heinrich, A., Haarmann, H., Zahradnik, S., Frenzel, K., Schreiber, F., Klassert, T. E., Heyl, K. A., Endres, A.‐S., Schmidtke, M., Hofmann, J., Slevogt, H. Moraxella catarrhalis decreases antiviral innate immune responses by down‐regulation of TLR3 via inhibition of p53 in human bronchial epithelial cells. FASEB J. 30, 2426–2434 (2016). www.fasebj.org

The C-type lectin receptor Dectin-1 is expressed mainly on myeloid cells mediating the immune response targeting respiratory pathogens such as Aspergillus fumigatus and Mycobacterium tuberculosis . The pulmonary epithelium serves as an important interface for interactions between these pathogens and the respiratory tract. Therefore, we analyzed the expression pattern of Dectin-1 in the human lung. Immunohistochemically stained human lung sections from 17 out of 19 individuals were positive for Dectin-1, which was expressed mainly apically on bronchial and alveolar epithelium. Our results showed no correlation with chronic obstructive pulmonary disease (COPD) or the smoking habits of the patients. Nontypeable Haemophilus influenzae (NTHI), an important bacterial pathogen of the respiratory tract with significant importance in COPD, has also been proposed to be recognized by Dectin-1, suggesting a possible impact on the NTHI-dependent immune response in human airways. Therefore, the involvement of Dectin-1 in NTHI-triggered cytokine responses was investigated in primary normal human bronchial epithelial (NHBE) cells and in the A549 cell line stably transfected with Dectin-1. The presence of Dectin-1 significantly increased cytokine release in response to NTHI in NHBE and A549 cells. In addition, phosphorylation of the Dectin-1 hem-immunoreceptor tyrosine-based activation motif (hemITAM) was essential for the Dectin-1-triggered response to NTHI in A549 cells. In conclusion, in human airways, epithelium-expressed Dectin-1 may play a significant role in generating an NTHI-mediated, proinflammatory immune response. IMPORTANCE In this study, we demonstrated, for the first time, the expression of Dectin-1 on human lung tissues and, in particular, pulmonary epithelium by making use of immunohistochemical staining. The epithelial lining of the human airways is an important interface for host-pathogen interactions. Therefore, our data suggest that epithelium-expressed Dectin-1 is of considerable importance for the interaction of the human airways with pathogens detected by this receptor, such as A. fumigatus and M. tuberculosis . Moreover, we further demonstrated that, in pulmonary epithelial cells, Dectin-1 enhances the proinflammatory immune response to NTHI. In COPD patients, NTHI is a major cause of respiratory tract infections and is associated with proinflammatory immune responses in the lower airways. Therefore, our data suggest that the functional interaction of Dectin-1 with NTHI in human airways may have an important impact on the pathogenesis of COPD.

Beyond its well-documented role in reproduction, embryogenesis and maintenance of body tissues, vitamin A has attracted considerable attention due to its immunomodulatory effects on both the innate and the adaptive immune responses. In infectious diseases, vitamin A has been shown to have a host-protective effect in infections of bacterial, viral or protozoan origin. Nevertheless, its impact in fungal infections remains unknown. Meanwhile, the frequency of invasive mycoses keeps on growing, with Candida albicans being the major opportunistic fungal pathogen and associated with high mortality. In the present work, we explored the impact of all-trans retinoic acid (atRA), the most active metabolite of vitamin A, on the innate immune response against C. albicans in human monocytes. Our results show a strong immunomodulatory role for atRA, leading to a significant down-regulation of the fungi-induced expression and secretion of the pro-inflammatory cytokines TNFα, IL6 and IL12. Moreover, atRA significantly suppressed the expression of Dectin-1, a major fungal pattern recognition receptor, as well as the Dectin-1-dependent cytokine production. Both RAR-dependent and RAR-independent mechanisms seem to play a role in the atRA-mediated immunomodulation. Our findings open a new direction to elucidate the role of vitamin A on the immune function during fungal infections.

Lower respiratory tract bacterial infections are characterized by neutrophilic inflammation in the airways. The carcinoembryonic antigen-related cell adhesion molecule (CEACAM) 8 is expressed in and released by human granulocytes. Our study demonstrates that human granulocytes release CEACAM8 in response to bacterial DNA in a TLR9-dependent manner. Individuals with a high percentage of bronchial lavage fluid (BALF) granulocytes were more likely to have detectable levels of released CEACAM8 in the BALF than those with a normal granulocyte count. Soluble, recombinant CEACAM8-Fc binds to CEACAM1 expressed on human airway epithelium. Application of CEACAM8-Fc to CEACAM1-positive human pulmonary epithelial cells resulted in reduced TLR2-dependent inflammatory responses. These inhibitory effects were accompanied by tyrosine phosphorylation of the immunoreceptor tyrosine-based inhibitory motif (ITIM) of CEACAM1 and by recruitment of the phosphatase SHP-1, which could negatively regulate Toll-like receptor 2-dependent activation of the phosphatidylinositol 3-OH kinase-Akt kinase pathway. Our results suggest a new mechanism by which granulocytes reduce pro-inflammatory immune responses in human airways via secretion of CEACAM8 in neutrophil-driven bacterial infections.

Background: The carcinoembryonic antigen (CEA)-related cell adhesion molecules CEACAM1 (BGP, CD66a), CEACAM5 (CEA, CD66e) and CEACAM6 (NCA, CD66c) are expressed in human lung. They play a role in innate and adaptive immunity and are targets for various bacterial and viral adhesins. Two pathogens that colonize the normally sterile lower respiratory tract in patients with chronic obstructive pulmonary disease (COPD) are non-typableHaemophilus influenzae(NTHI) andMoraxella catarrhalis. Both pathogens bind to CEACAMs and elicit a variety of cellular reactions, including bacterial internalization, cell adhesion and apoptosis.Methods: To analyze the (co-) expression of CEACAM1, CEACAM5 and CEACAM6 in different lung tissues with respect to COPD, smoking status and granulocyte infiltration, immunohistochemically stained paraffin sections of 19 donors were studied. To address short-term effects of cigarette smoke and acute inflammation, transcriptional regulation of CEACAM5, CEACAM6 and different CEACAM1 isoforms by cigarette smoke extract, interferons, Toll-like receptor agonists, and bacteria was tested in normal human bronchial epithelial (NHBE) cells by quantitative PCR. Corresponding CEACAM protein levels were determined by flow cytometry.Results: Immunohistochemical analysis of lung sections showed the most frequent and intense staining for CEACAM1, CEACAM5 and CEACAM6 in bronchial and alveolar epithelium, but revealed no significant differences in connection with COPD, smoking status and granulocyte infiltration. In NHBE cells, mRNA expression of CEACAM1 isoforms CEACAM1-4L, CEACAM1-4S, CEACAM1-3L and CEACAM1-3S were up-regulated by interferons alpha, beta and gamma, as well as the TLR3 agonist polyinosinic:polycytidylic acid (poly I:C). Interferon-gamma also increased CEACAM5 expression. These results were confirmed on protein level by FACS analysis. Importantly, also NTHI andM. catarrhalisincreased CEACAM1 mRNA levels. This effect was independent of the ability to bind to CEACAM1. The expression of CEACAM6 was not affected by any treatment or bacterial infection.Conclusions: While we did not find a direct correlation between CEACAM1 expression and COPD, the COPD-associated bacteria NTHi andM. catarrhaliswere able to increase the expression of their own receptor on host cells. Further, the data suggest a role for CEACAM1 and CEACAM5 in the phenomenon of increased host susceptibility to bacterial infection upon viral challenge in the human respiratory tract.