Modelling the support function of vascularized lung endothelial cells for alveolar progenitor function using an alveolus-on-chip

Abstract

Lung microvascular endothelial cells are hypothesized to contribute to alveolar repair and regeneration. We aim to develop an open-top alveolus-on-chip system with vascularized lung endothelial cells and pericytes, cultured in networks, and study interactions with alveolar epithelial cells. Primary endothelial cells (CD31+), pericytes (NG2+) and alveolar epithelial type-2 cells (AEC2; HTII-280+) were isolated from resected peripheral human lung tissue and expanded in vitro. Cells were seeded in a fibrin-collagen hydrogel inside an open-top alveolus-on-chip (2 mm diameter, 500 µm high culture chamber) to from vascular networks. The open-top design enables controlled co-culture of vascular networks with monolayers of AEC2. The expanded endothelial cells were analyzed by flow cytometry and shown to be CD31+ and CD140b-, whereas pericytes were NG2+ and CD140b+. The endothelial cells (CD31+ CD144+) formed vascular networks by themselves, as well as in co-cultures with pericytes (NG2+) and in co-cultures with MRC-5 fibroblasts (α-SMA+) at day 5. Endothelial cells alone were able to form networks with typical vessel diameters of 5 to 10 μm compared to 1 to 3 μm in networks formed with pericytes. In presence of pericytes, a more continuous endothelial network was observed when compared to co-cultures with fibroblasts, while the vessel diameters were comparable in both conditions. Monolayers of AEC2 (surfactant protein C+) were successfully cultured at day 5 on top of the vascular networks until day 12. This open-top alveolus-on-a-chip model enables controlled studies of the role of vascular networks in alveolar repair, e.g. upon exposure to cigarette smoke.

Authors 9

1. Abilash Ravi Aachen PulmoScience Lab Dept of Pulmonology, Leiden University Medical Center

   Leiden University Medical Center

   Affiliation as printed

   PulmoScience Lab, Department of Pulmonology, Leiden University Medical Center, Leiden, Netherlands

2. Tarek Gensheimer

   University of Twente

   Affiliation as printed

   Department of Applied Stem Cell Technologies, University of Twente, Enschede, Netherlands

3. Xinhui Wu

   University of Groningen

   Affiliation as printed

   Department of Molecular Pharmacology, Faculty of Science and Engineering, Groningen, Netherlands

4. Jill R. Johnson

   Aston University

   Affiliation as printed

   School of Biosciences, College of Health & Life Sciences, Aston University, Birmingham, United Kingdom

5. Martin C. Harmsen

   University of Groningen

   Affiliation as printed

   Department of Molecular Pharmacology, Faculty of Science and Engineering, Groningen, Netherlands

6. Reinoud Gosens

   University of Groningen

   Affiliation as printed

   Department of Molecular Pharmacology, Faculty of Science and Engineering, Groningen, Netherlands

7. Pieter Sicco Hiemstra Aachen PulmoScience Lab Dept of Pulmonology, Leiden University Medical Center

   Leiden University Medical Center

   Affiliation as printed

   PulmoScience Lab, Department of Pulmonology, Leiden University Medical Center, Leiden, Netherlands

8. Andries D. van der Meer

   University of Twente

   Affiliation as printed

   Department of Applied Stem Cell Technologies, University of Twente, Enschede, Netherlands

9. Anne M. van der Does Aachen PulmoScience Lab Dept of Pulmonology, Leiden University Medical Center

   Leiden University Medical Center

   Affiliation as printed

   PulmoScience Lab, Department of Pulmonology, Leiden University Medical Center, Leiden, Netherlands