Alveolar epithelial cells type II show a high sensitivity to cigarette smoke extract

Alveolar epithelial cells type II (AECII) play an important role in the normal pulmonary function as well as in the host defense and immune response. The human tumor cell line A549 is the most popular model of AECII.

In the presented study, the effects of cigarette smoke extract (CSE) on primary AECII and A549 as well as the epithelial adenocarcinoma cell line H1975 were investigated.

Tumor-free lung tissue from patients who underwent lobectomy due to cancer at the LungenClinic Großhansdorf was used to isolate AECII. Briefly, after crushing the lung tissues, AECII were separated by negative selection via a CD45. Subsequently, cells were seeded on collagen-coated 96-well plates at low density (4 × 10⁴ cells/well). A549 cells were seeded at 2 × 105 cells/well, H1975 at 2 × 10⁴ cells/well. All three cell types were maintained in DMEM-F12 supplemented with 10% FCS overnight. CSE was obtained using commercially available cigarettes (West light) by drawing smoke of one cigarette slowly through a water pump into a tube containing 10 mL of ddH2O (= 10% CSE). Stimulation was performed under serum-free conditions. Cells were stimulated with increasing concentrations of CSE (0.1 – 5%) for 1h. Cells were cultured for further 4h. Cell viability was measured via MTT assay.

Acute CSE exposure with 0.5% CSE induced a significant cytotoxic effect in AECII (IC50: 0.21% CSE), which was not reversible by dexamethasone (4 mg/mL) or roflumilast (15µM). In H1975, a concentration of 5% CSE caused a significant reduction of cell viability. By contrast, no cytotoxic effect was detectable in A549 cells.

Primary AECII are a model to investigate cigarette smoke induced inflammatory effects, better suited than the widely used tumor cell line A549. Moreover, CSE-induced AECII damage is not reversible by anti-inflammatory treatment.