The Role of Alveolar Epithelial Cells in Initiating and Shaping Pulmonary Immune Responses: Communication between Innate and Adaptive Immune Systems

Abstract

Macrophages and dendritic cells have been recognized as key players in the defense against mycobacterial infection. However, more recently, other cells in the lungs such as alveolar epithelial cells (AEC) have been found to play important roles in the defense and pathogenesis of infection. In the present study we first compared AEC with pulmonary macrophages (PuM) isolated from mice in their ability to internalize and control Bacillus Calmette-Guérin (BCG) growth and their capacity as APCs. AEC were able to internalize and control bacterial growth as well as present antigen to primed T cells. Secondly, we compared both cell types in their capacity to secrete cytokines and chemokines upon stimulation with various molecules including mycobacterial products. Activated PuM and AEC displayed different patterns of secretion. Finally, we analyzed the profile of response of AEC to diverse stimuli. AEC responded to both microbial and internal stimuli exemplified by TLR ligands and IFNs, respectively. The response included synthesis by AEC of several factors, known to have various effects in other cells. Interestingly, TNF could stimulate the production of CCL2/MCP-1. Since MCP-1 plays a role in the recruitment of monocytes and macrophages to sites of infection and macrophages are the main producers of TNF, we speculate that both cell types can stimulate each other. Also, another cell-cell interaction was suggested when IFNs (produced mainly by lymphocytes) were able to induce expression of chemokines (IP-10 and RANTES) by AEC involved in the recruitment of circulating lymphocytes to areas of injury, inflammation, or viral infection. In the current paper we confirm previous data on the capacity of AEC regarding internalization of mycobacteria and their role as APC, and extend the knowledge of AEC as a multifunctional cell type by assessing the secretion of a broad array of factors in response to several different types of stimuli.

Alveolar Epithelial Cells Are Critical in Protection of the Respiratory Tract by Secretion of Factors Able To Modulate the Activity of Pulmonary Macrophages and Directly Control Bacterial Growth

“…Furthermore, Thorley et al (11) showed that other factors, such as IL-8 and GRO-␣, also are important, as neutralizing these chemokines reduced monocyte migration toward culture medium from primary human type II AEC by 39% and 51%, respectively (11). We have previously published that in our system, AEC secrete high levels of MCP-1 and the murine IL-8 homologs KC and MIP-2 upon stimulation with LPS (20). It is thus likely that these factors play a major role in inducing the macrophage migration in our system.…”

“…Upon activation, AEC secrete a number of chemokines, including keratinocyte cytokine (KC), monocyte chemoattractant protein (MCP-1), and MIP-2␣ (20). To determine whether AEC promote migration of pulmonary cells and whether a specific cell type is preferentially influenced by AEC-derived factors, we set up a Transwell migration assay, using total lung cells as responder cells, and let these migrate toward supernatants from either AEC sup or AEC LPS .…”

“…The use of this construct is convenient, since bacteria can be quantified immediately by luminescence, while the classical evaluation of BCG growth in agar plates takes between 2 to 3 weeks. Bacterial contents are expressed as relative luminescence units (RLU), which correlate with the number of CFU (20,21). GFP-BCG was grown in Middlebrook 7H9 broth (Difco, Sparks, MD) supplemented with albumin-dextrose-catalase (ADC), 0.5% glycerol, 0.05% Tween 80 (vol/vol), and 50 g/ml hygromycin for 10 to 15 days.…”

“…Total pulmonary cells were prepared using Corti's protocol (22) with previously described modifications (20,21). In short, CD45 ϩ cells were obtained from total lung cells using magneticactivated cell sorting (MACS; Miltenyi Biotec, Bergisch Gladbach, Germany) and subsequently cultured for 48 h in RPMI (Gibco-Invitrogen, Paisley, United Kingdom) supplemented with 10% fetal calf serum (FCS), 2 mM L-glutamine, 100 U/ml penicillin, 100 g/ml streptomycin, 0.02 M HEPES, and 0.05 M 2-mercaptoethanol (Sigma) at 37°C and 5% CO 2 .…”

Inhibition of acute lethal pulmonary inflammation by the IDO–AhR pathway

“…Purified CD4 + T cells (1.5 × 10 6 cells per milliliter) were treated with soluble anti-CD3 (3 μg/mL) or recombinant human IL-2 (20 ng/mL; R&D Systems) with or without L-Kyn (300 μM) in a 96-well plate (Corning Costar) at 37°C for 72 h. During the final 15∼17 h, the cells were pulsed with [ Isolation of Alveolar Epithelial Cells. Primary alveolar epithelial cells were isolated as previously described (48). In brief, dispase (Gibco-Invitrogen) was instilled into lung tissues of C57BL/6 mice via the trachea.…”