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RESEARCH ARTICLE

Fabrication and Characterization of 3D Bioprinted

Triple-layered Human Alveolar Lung Models

Wei Long Ng 1,2† , Teck Choon Ayi , Yi-Chun Liu , Swee Leong Sing , Wai Yee Yeong *,
3
1
1,2
3†
Boon-Huan Tan 3,4
1 Singapore Centre for 3D Printing, School of Mechanical and Aerospace Engineering, Nanyang Technological University,
50 Nanyang Avenue, 639798, Singapore
2 HP-NTU Digital Manufacturing Corporate Lab, 65 Nanyang Avenue, 637460, Singapore
3 Defence Medical and Environmental Research Institute, DSO National Laboratories, 27 Medical Drive, 117510, Singapore
4 Lee Kong Chian School of Medicine, Novena Campus, Nanyang Technological University, 11 Mandalay Road, 308232,
Singapore
† These authors contributed equally to this work.

Abstract: The global prevalence of respiratory diseases caused by infectious pathogens has resulted in an increased demand
for realistic in-vitro alveolar lung models to serve as suitable disease models. This demand has resulted in the fabrication of
numerous two-dimensional (2D) and three-dimensional (3D) in-vitro alveolar lung models. The ability to fabricate these 3D
in-vitro alveolar lung models in an automated manner with high repeatability and reliability is important for potential scalable
production. In this study, we reported the fabrication of human triple-layered alveolar lung models comprising of human
lung epithelial cells, human endothelial cells, and human lung fibroblasts using the drop-on-demand (DOD) 3D bioprinting
technique. The polyvinylpyrrolidone-based bio-inks and the use of a 300 µm nozzle diameter improved the repeatability of the
bioprinting process by achieving consistent cell output over time using different human alveolar lung cells. The 3D bioprinted
human triple-layered alveolar lung models were able to maintain cell viability with relative similar proliferation profile over
time as compared to non-printed cells. This DOD 3D bioprinting platform offers an attractive tool for highly repeatable and
scalable fabrication of 3D in-vitro human alveolar lung models.
Keywords: 3D bioprinting; 3D printing; Biofabrication; Lung bioprinting; In-vitro human tissue models; Drop-on-demand

*Correspondence to: Wai Yee Yeong, Singapore Centre for 3D Printing, School of Mechanical and Aerospace Engineering, Nanyang
Technological University, 50 Nanyang Avenue, Singapore 639798; wyyeong@ntu.edu.sg
Received: December 22, 2020; Accepted: January 28, 2021; Published Online: April 9, 2021

Citation: Ng WL, Ayi TC, Liu YC, et al., 2021, Fabrication and Characterization of 3D Bioprinted Triple-layered Human
Alveolar Lung Models. Int J Bioprint, 7(2):332. http://doi.org/10.18063/ijb.v7i2.332

1. Introduction advanced Biosystems by facilitating critical cell-cell and
cell-matrix interactions found within the native tissue and
With the increase in respiratory diseases over the last two its microenvironment . The main drawback of these 3D
[7]
decades [1-4] , it is critical to elucidate how these pathogens tissue constructs is that the conventional manual production
interact and penetrate through the pulmonary epithelial of in-vitro tissue models is often laborious and unrepeatable.
tissue barrier and evaluate the potential severity of these 3D bioprinting has emerged as one of the leading
respiratory diseases. As such, numerous in-vitro alveolar manufacturing platforms for automated fabrication of highly
lung models ranging from simple, mono-cultured two- complex 3D tissues and/or organs in a scalable manner [8-11] .
dimensional (2D) models to more sophisticated three- The envisioned long-term goal of 3D bioprinting is to
[5]
dimensional (3D) constructs have been developed to fabricate highly-functional 3D tissue-engineered constructs
[6]
closely emulate the human pulmonary epithelial tissue in a layer-by-layer fabrication approach [12,13] . This would
barrier. The 3D tissue constructs are considered more promote important cell-extracellular matrix (ECM) and cell-
© 2021 Ng, et al. This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International License
(http://creativecommons.org/licenses/by-nc/4.0/), permitting all non-commercial use, distribution, and reproduction in any medium, provided the original
work is properly cited.
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