Toxicology Letters

Andreas O. Stucki, Giulia Raggi, Serena Sigrist, Pauline Zamprogno, Nicole Schneider-Daum, Claus Michael-Lehr, Hanno Huwer, Janick D. Stucki, Nina Hobi & Olivier T. Guenat


Introduction: The pulmonary alveolar barrier is one of the largest entry ports for xenobiotics into the body and consists of a thin barrier of epithelial and endothelial cells. Yet, establishing a biologically relevant model of the pulmonary alveolar-capillary barrier is challenging, due to poor availability of human primary alveolar epithelial cells. In addition, the effect of mechanical stretch on the cells due to breathing is thought to be important in regulating lung functions but has so far not thoroughly been considered due to the lack of appropriate tools. 

Aims: The goal of our work is to establish a stable pulmonary alveolar-capillary barrier and understand the interplay between lung alveolar epithelial and endothelial cells in an in vivo-like environment. 

Methods: Primary human pulmonary alveolar epithelial cells (pHPAEC) from patients undergoing lung resections were cultured on inserts (0.4 µm pore size) or lung-on-a-chips (3.5 µm pore size) with and without four different endothelial cell types. Medium was exchanged every other day, following which trans-epithelial electrical resistance (TEER) was measured for up to 14 days. 

Results: The TEER of pHPAEC monoculture rises above 1000 cm2 until day 4-6 and then begins to drop reaching values below 500 cm2 at days 8 to 14. Co-culturing pHPAEC with human VeraVec endothelial cells increased TEER values significantly and lead to a more stable culture (values above 4000 cm2 at day 12). Culturing pHPAEC together with primary human pulmonary microvascular (HPMEC) or umbilical vein (HUVEC) endothelial cells, or with immortalized HPMEC ST1.6R resulted in slightly lower TEER values compared to VeraVec cultures but significantly higher compared to monoculture of pHPAEC.  

Conclusion: The integrity of the alveolar-capillary barrier is a key parameter that prevents any unwanted leakage from either side of the barrier. TEER is often used to assess the quality of the barrier integrity. Here, we show that a co-culture of primary human alveolar epithelial cells together with endothelial cells does not only significantly increase the barrier integrity but is also a factor of a long-term stability.