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Kuipers ME, van Liefferinge F, van der Wal E, Rovituso M, Slats AM, Hiemstra PS, Van Doorn-Wink KC. Effect of FLASH proton therapy on primary bronchial epithelial cell organoids. Clin Transl Radiat Oncol 2025; 52:100927. [PMID: 39968050 PMCID: PMC11833640 DOI: 10.1016/j.ctro.2025.100927] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/16/2024] [Revised: 01/21/2025] [Accepted: 01/28/2025] [Indexed: 02/20/2025] Open
Abstract
Purpose The effects of conventional (CONV) and FLASH proton therapy on primary bronchial epithelial cell (PBEC) organoids from individuals with chronic obstructive pulmonary disease (COPD) were investigated. The primary objective was to compare the effect of FLASH and CONV on COPD PBEC organoids with a focus on DNA damage, organoid formation, and gene expression. Methods PBECs were obtained from six COPD donors, cultured as three-dimensional (3D) organoids and exposed to 2 and 8 Gy CONV and FLASH proton radiation at the Holland Proton Therapy Center. DNA damage was assessed by γH2AX staining. Organoid formation capacity was assessed by counting the organoids formed after reseeding irradiated cells at 24 h and 7 days. Bulk RNA sequencing (RNAseq) and qPCR analyses were performed to identify pathways and differences in the radiation response. Results γH2AX foci analysis showed a significant dose-dependent increase in DNA damage at 1 h for both CONV and FLASH treatments, without differences between the two modalities. Organoid formation assays revealed a dose-dependent decrease in organoid formation capacity at 24 h for both treatments. At 7 days, 2 Gy FLASH-treated samples showed significantly reduced organoid formation compared to 2 Gy CONV (p = 0.008). RNAseq identified CONV and FLASH-induced changes in expression of DNA-damage response and apoptosis pathway genes. A dose-dependent upregulation of MDM2, GDF15, DDB2, BAX, P21, AEN and a decrease in MKi67 expression was confirmed by qPCR analysis. Conclusion No significant differences were found in DNA damage or gene expression profiles between CONV and FLASH. The organoid formation assay showed a prolonged detrimental effect in the FLASH-treated organoids, suggesting a more complex interaction of FLASH with lung epithelial cells. The results of this study contribute to the advancement of robust in vitro human lung models for investigating the mechanisms of action of FLASH, potentially facilitating the treatment of NSCLC patients with proton FLASH therapy.
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Affiliation(s)
- Merian E. Kuipers
- Leiden University Medical Center (LUMC), Department of Pulmonology, C02-Q, Albinusdreef 2 2333 ZA Leiden, the Netherlands
| | - Floriane van Liefferinge
- Leiden University Medical Center (LUMC), Department of Pulmonology, C02-Q, Albinusdreef 2 2333 ZA Leiden, the Netherlands
| | - Ernst van der Wal
- Holland Proton Therapy Center (HollandPTC), Huismansingel 4 2629 JH Delft, the Netherlands
| | - Marta Rovituso
- Holland Proton Therapy Center (HollandPTC), Huismansingel 4 2629 JH Delft, the Netherlands
| | - Annelies M. Slats
- Leiden University Medical Center (LUMC), Department of Pulmonology, C02-Q, Albinusdreef 2 2333 ZA Leiden, the Netherlands
| | - Pieter S. Hiemstra
- Leiden University Medical Center (LUMC), Department of Pulmonology, C02-Q, Albinusdreef 2 2333 ZA Leiden, the Netherlands
| | - Krista C.J. Van Doorn-Wink
- Holland Proton Therapy Center (HollandPTC), Huismansingel 4 2629 JH Delft, the Netherlands
- Leiden University Medical Center (LUMC), Department of Radiotherapy, K01-P, Albinusdreef 2 2333 ZA Leiden, the Netherlands
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2
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Schmiedl A, Mühlfeld C. Morphological and molecular aspects of lung development. Histol Histopathol 2025; 40:411-430. [PMID: 39344418 DOI: 10.14670/hh-18-807] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/01/2024]
Abstract
Healthy breathing relies on normal morphological and functional development of the lung. This includes different prenatal and postnatal developmental stages. Depending on species and postnatal behavior as nest escapers or nest squatters, the duration of individual developmental phases and the state of differentiation of the lungs at birth differ. However, the sequence and morphology of the lung developmental stages are similar in all mammals, so knowledge gained from animal models about development-specific genetic control and regulatory mechanisms can be translated in principle to the human lung. Functional lung development comprises the maturation of the surfactant system, which is closely linked to the morphological development of the pulmonary acini. Although a number of reviews are found in the literature, a presentation that integrates the morphological and molecular regulatory mechanisms is missing. Therefore, the aim of this article was to provide an up-to-date comprehensive review of the main morphological steps and regulatory mechanisms of lung development, including clinical aspects related to developmental disorders.
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Affiliation(s)
- Andreas Schmiedl
- Institute of Functional and Applied Anatomy, Hannover Medical School, Hannover, Germany
- Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), Member of the German Center for Lung Research, Hannover, Germany
| | - Christian Mühlfeld
- Institute of Functional and Applied Anatomy, Hannover Medical School, Hannover, Germany
- Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), Member of the German Center for Lung Research, Hannover, Germany
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3
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Eiken MK, Childs CJ, Brastrom LK, Frum T, Plaster EM, Ahmed DW, Spencer RC, Shachaf O, Pfeiffer S, Levine JE, Alysandratos KD, Kotton DN, Spence JR, Loebel C. Nascent matrix deposition supports alveolar organoid formation from aggregates in synthetic hydrogels. Stem Cell Reports 2025; 20:102376. [PMID: 39672155 PMCID: PMC11784465 DOI: 10.1016/j.stemcr.2024.11.006] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2024] [Revised: 11/12/2024] [Accepted: 11/14/2024] [Indexed: 12/15/2024] Open
Abstract
Human induced pluripotent stem cell (iPSC)-derived alveolar organoids have emerged as a system to model the alveolar epithelium in homeostasis and disease. However, alveolar organoids are typically grown in Matrigel, a mouse sarcoma-derived basement membrane matrix that offers poor control over matrix properties, prompting the development of synthetic hydrogels as a Matrigel alternative. Here, we develop a two-step culture method that involves pre-aggregation of organoids in hydrogel-based microwells followed by embedding in a synthetic hydrogel that supports alveolar organoid growth, while also offering considerable control over organoid and hydrogel properties. We find that the aggregated organoids secrete their own nascent extracellular matrix (ECM) both in the microwells and upon embedding in synthetic hydrogels, which supports their growth. Thus, the synthetic hydrogels described here allow us to de-couple exogenous and nascent ECM to interrogate the role of ECM in organoid formation.
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Affiliation(s)
- Madeline K Eiken
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA
| | - Charlie J Childs
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Lindy K Brastrom
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Tristan Frum
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Eleanor M Plaster
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA
| | - Donia W Ahmed
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA
| | - Ryan C Spencer
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Orren Shachaf
- Department of Biomedical Engineering, University of Texas, Austin, TX, USA
| | - Suzanne Pfeiffer
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA
| | - Justin E Levine
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA
| | - Konstantinos-Dionysios Alysandratos
- Center for Regenerative Medicine, Boston University and Boston Medical Center, Boston, MA 02118, USA; The Pulmonary Center and Department of Medicine, Boston University Chobanian & Avedisian School of Medicine, Boston, MA 02118, USA
| | - Darrell N Kotton
- Center for Regenerative Medicine, Boston University and Boston Medical Center, Boston, MA 02118, USA; The Pulmonary Center and Department of Medicine, Boston University Chobanian & Avedisian School of Medicine, Boston, MA 02118, USA
| | - Jason R Spence
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA; Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA; Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI, USA; Program in Cell and Molecular Biology, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Claudia Loebel
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA; Department of Materials Science and Engineering, University of Michigan, Ann Arbor, MI, USA.
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4
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Eiken MK, Childs CJ, Brastrom LK, Frum T, Plaster EM, Shachaf O, Pfeiffer S, Levine JE, Alysandratos KD, Kotton DN, Spence JR, Loebel C. Nascent matrix deposition supports alveolar organoid formation from aggregates in synthetic hydrogels. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.03.19.585720. [PMID: 38562781 PMCID: PMC10983987 DOI: 10.1101/2024.03.19.585720] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 04/04/2024]
Abstract
Human induced pluripotent stem cell (iPSC) derived alveolar organoids have emerged as a system to model the alveolar epithelium in homeostasis and disease. However, alveolar organoids are typically grown in Matrigel, a mouse-sarcoma derived basement membrane matrix that offers poor control over matrix properties, prompting the development of synthetic hydrogels as a Matrigel alternative. Here, we develop a two-step culture method that involves pre-aggregation of organoids in hydrogel-based microwells followed by embedding in a synthetic hydrogel that supports alveolar organoid growth, while also offering considerable control over organoid and hydrogel properties. We find that the aggregated organoids secrete their own nascent extracellular matrix (ECM) both in the microwells and upon embedding in the synthetic hydrogels. Thus, the synthetic gels described here allow us to de-couple exogenous and nascent ECM in order to interrogate the role of ECM in organoid formation.
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Affiliation(s)
- Madeline K. Eiken
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA
| | - Charlie J. Childs
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI, USA
- Department of Computational Medicine and Bioinformatics, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Lindy K. Brastrom
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Tristan Frum
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Eleanor M. Plaster
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA
| | - Orren Shachaf
- Department of Biomedical Engineering, University of Texas, Austin, TX, USA
| | - Suzanne Pfeiffer
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA
| | - Justin E. Levine
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA
| | - Konstantinos-Dionysios Alysandratos
- Center for Regenerative Medicine, Boston University and Boston Medical Center, Boston, MA 02118, USA
- The Pulmonary Center and Department of Medicine, Boston University Chobanian & Avedisian School of Medicine, Boston, MA 02118, USA
| | - Darrell N. Kotton
- Center for Regenerative Medicine, Boston University and Boston Medical Center, Boston, MA 02118, USA
- The Pulmonary Center and Department of Medicine, Boston University Chobanian & Avedisian School of Medicine, Boston, MA 02118, USA
| | - Jason R. Spence
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI, USA
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA
- Program in Cell and Molecular Biology, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Claudia Loebel
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA
- Department of Materials Science and Engineering, University of Michigan, Ann Arbor, MI, USA
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Shrestha J, Paudel KR, Nazari H, Dharwal V, Bazaz SR, Johansen MD, Dua K, Hansbro PM, Warkiani ME. Advanced models for respiratory disease and drug studies. Med Res Rev 2023; 43:1470-1503. [PMID: 37119028 PMCID: PMC10946967 DOI: 10.1002/med.21956] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2022] [Revised: 02/02/2023] [Accepted: 03/17/2023] [Indexed: 04/30/2023]
Abstract
The global burden of respiratory diseases is enormous, with many millions of people suffering and dying prematurely every year. The global COVID-19 pandemic witnessed recently, along with increased air pollution and wildfire events, increases the urgency of identifying the most effective therapeutic measures to combat these diseases even further. Despite increasing expenditure and extensive collaborative efforts to identify and develop the most effective and safe treatments, the failure rates of drugs evaluated in human clinical trials are high. To reverse these trends and minimize the cost of drug development, ineffective drug candidates must be eliminated as early as possible by employing new, efficient, and accurate preclinical screening approaches. Animal models have been the mainstay of pulmonary research as they recapitulate the complex physiological processes, Multiorgan interplay, disease phenotypes of disease, and the pharmacokinetic behavior of drugs. Recently, the use of advanced culture technologies such as organoids and lung-on-a-chip models has gained increasing attention because of their potential to reproduce human diseased states and physiology, with clinically relevant responses to drugs and toxins. This review provides an overview of different animal models for studying respiratory diseases and evaluating drugs. We also highlight recent progress in cell culture technologies to advance integrated models and discuss current challenges and present future perspectives.
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Affiliation(s)
- Jesus Shrestha
- School of Biomedical EngineeringUniversity of Technology SydneySydneyNew South WalesAustralia
| | - Keshav Raj Paudel
- Centre for InflammationCentenary Institute and University of Technology SydneySydneyNew South WalesAustralia
| | - Hojjatollah Nazari
- School of Biomedical EngineeringUniversity of Technology SydneySydneyNew South WalesAustralia
| | - Vivek Dharwal
- Centre for InflammationCentenary Institute and University of Technology SydneySydneyNew South WalesAustralia
| | - Sajad Razavi Bazaz
- School of Biomedical EngineeringUniversity of Technology SydneySydneyNew South WalesAustralia
| | - Matt D. Johansen
- Centre for InflammationCentenary Institute and University of Technology SydneySydneyNew South WalesAustralia
| | - Kamal Dua
- Discipline of Pharmacy, Graduate School of HealthUniversity of TechnologySydneyNew South WalesAustralia
- Faculty of Health, Australian Research Centre in Complementary & Integrative MedicineUniversity of Technology SydneyUltimoNew South WalesAustralia
| | - Philip M. Hansbro
- Centre for InflammationCentenary Institute and University of Technology SydneySydneyNew South WalesAustralia
| | - Majid Ebrahimi Warkiani
- School of Biomedical EngineeringUniversity of Technology SydneySydneyNew South WalesAustralia
- Institute for Biomedical Materials and Devices, Faculty of ScienceUniversity of Technology SydneyUltimoNew South WalesAustralia
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Tiwari SK, Rana TM. Generation of 3D lung organoids from human induced pluripotent stem cells for modeling of lung development and viral infection. Heliyon 2023; 9:e19601. [PMID: 37809493 PMCID: PMC10558843 DOI: 10.1016/j.heliyon.2023.e19601] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2023] [Revised: 08/20/2023] [Accepted: 08/28/2023] [Indexed: 10/10/2023] Open
Abstract
The lack of physiologically relevant in vitro models has hampered progress in understanding human lung development and disease. Here, we describe a protocol in which human induced pluripotent stem cells (hiPSCs) undergo stepwise differentiation into definitive endoderm (>88% population) to three-dimensional (3D) lung organoids (LORGs), which contain both epithelial and mesenchymal cellular architecture and display proximal and distal airway patterning. These LORGs can maintained for more than 90 days by re-embedding in the Matrigel. We show the utility of LORGs for disease modeling and drug screening by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and treatment with antiviral drugs.
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Affiliation(s)
- Shashi Kant Tiwari
- Division of Genetics, Department of Pediatrics, Program in Immunology, Institute for Genomic Medicine, University of California San Diego, 9500 Gilman Drive MC 0762, La Jolla, CA, 92093, USA
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Dean CH, Cheong SS. Simple Models of Lung Development. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2023; 1413:17-28. [PMID: 37195524 DOI: 10.1007/978-3-031-26625-6_2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/18/2023]
Abstract
Models are essential to further our understanding of lung development and regeneration and to facilitate identification and testing of potential treatments for lung diseases. A wide variety of rodent and human models are available that recapitulate one or more stages of lung development. This chapter describes the existing 'simple' in vitro, in silico and ex vivo models of lung development. We define which stage(s) of development each model recapitulates and highlight their pros and cons.
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Affiliation(s)
- Charlotte H Dean
- National Heart and Lung Institute, Imperial College London, London, UK.
| | - Sek-Shir Cheong
- National Heart and Lung Institute, Imperial College London, London, UK
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8
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Zamprogno P, Schulte J, Ferrari D, Rechberger K, Sengupta A, van Os L, Weber T, Zeinali S, Geiser T, Guenat OT. Lung-on-a-Chip Models of the Lung Parenchyma. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2023; 1413:191-211. [PMID: 37195532 DOI: 10.1007/978-3-031-26625-6_10] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/18/2023]
Abstract
Since the publication of the first lung-on-a-chip in 2010, research has made tremendous progress in mimicking the cellular environment of healthy and diseased alveoli. As the first lung-on-a-chip products have recently reached the market, innovative solutions to even better mimic the alveolar barrier are paving the way for the next generation lung-on-chips. The original polymeric membranes made of PDMS are being replaced by hydrogel membranes made of proteins from the lung extracellular matrix, whose chemical and physical properties exceed those of the original membranes. Other aspects of the alveolar environment are replicated, such as the size of the alveoli, their three-dimensional structure, and their arrangement. By tuning the properties of this environment, the phenotype of alveolar cells can be tuned, and the functions of the air-blood barrier can be reproduced, allowing complex biological processes to be mimicked. Lung-on-a-chip technologies also provide the possibility of obtaining biological information that was not possible with conventional in vitro systems. Pulmonary edema leaking through a damaged alveolar barrier and barrier stiffening due to excessive accumulation of extracellular matrix proteins can now be reproduced. Provided that the challenges of this young technology are overcome, there is no doubt that many application areas will benefit greatly.
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Affiliation(s)
- Pauline Zamprogno
- Organs-on-Chip Technologies Laboratory, ARTORG Center, University of Bern, Bern, Switzerland
| | - Jan Schulte
- Organs-on-Chip Technologies Laboratory, ARTORG Center, University of Bern, Bern, Switzerland
| | - Dario Ferrari
- Organs-on-Chip Technologies Laboratory, ARTORG Center, University of Bern, Bern, Switzerland
| | - Karin Rechberger
- Organs-on-Chip Technologies Laboratory, ARTORG Center, University of Bern, Bern, Switzerland
| | - Arunima Sengupta
- Organs-on-Chip Technologies Laboratory, ARTORG Center, University of Bern, Bern, Switzerland
| | - Lisette van Os
- Organs-on-Chip Technologies Laboratory, ARTORG Center, University of Bern, Bern, Switzerland
| | - Tobias Weber
- Organs-on-Chip Technologies Laboratory, ARTORG Center, University of Bern, Bern, Switzerland
| | - Soheila Zeinali
- Organs-on-Chip Technologies Laboratory, ARTORG Center, University of Bern, Bern, Switzerland
| | - Thomas Geiser
- Department of Pulmonary Medicine, University Hospital of Bern, Bern, Switzerland
| | - Olivier T Guenat
- Organs-on-Chip Technologies Laboratory, ARTORG Center, University of Bern, Bern, Switzerland.
- Department of Pulmonary Medicine, University Hospital of Bern, Bern, Switzerland.
- Department of General Thoracic Surgery, University Hospital of Bern, Bern, Switzerland.
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9
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Demchenko A, Lavrov A, Smirnikhina S. Lung organoids: current strategies for generation and transplantation. Cell Tissue Res 2022; 390:317-333. [PMID: 36178558 PMCID: PMC9522545 DOI: 10.1007/s00441-022-03686-x] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2021] [Accepted: 09/08/2022] [Indexed: 01/19/2023]
Abstract
Lung diseases occupy a leading position in human morbidity and are the third leading cause of death. Often the chronic forms of these diseases do not respond to therapy, so that lung transplantation is the only treatment option. The development of cellular and biotechnologies offers a new solution-the use of lung organoids for transplantation in such patients. Here, we review types of lung organoids, methods of their production and characterization, and experimental works on transplantation in vivo. These results show the promise of work in this direction. Despite the current problems associated with a low degree of cell engraftment, immune response, and insufficient differentiation, we are confident that organoid transplantation will find it is clinical application.
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Affiliation(s)
- Anna Demchenko
- Research Centre for Medical Genetics, Laboratory of Genome Editing, Moscow, 115522 Russia
| | - Alexander Lavrov
- Research Centre for Medical Genetics, Laboratory of Genome Editing, Moscow, 115522 Russia
| | - Svetlana Smirnikhina
- Research Centre for Medical Genetics, Laboratory of Genome Editing, Moscow, 115522 Russia
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10
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Organoid Technologies for SARS-CoV-2 Research. CURRENT STEM CELL REPORTS 2022; 8:151-163. [PMID: 36313938 PMCID: PMC9589566 DOI: 10.1007/s40778-022-00220-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 10/07/2022] [Indexed: 12/05/2022]
Abstract
Purpose of Review Organoids are an emerging technology utilizing three-dimensional (3D), multi-cellular in vitro models to represent the function and physiological responses of tissues and organs. By using physiologically relevant models, more accurate tissue responses to viral infection can be observed, and effective treatments and preventive strategies can be identified. Animals and two-dimensional (2D) cell culture models occasionally result in inaccurate disease modeling outcomes. Organoids have been developed to better represent human organ and tissue systems, and accurately model tissue function and disease responses. By using organoids to study SARS-Cov-2 infection, researchers have now evaluated the viral effects on different organs and evaluate efficacy of potential treatments. The purpose of this review is to highlight organoid technologies and their ability to model SARS-Cov-2 infection and tissue responses. Recent Findings Lung, cardiac, kidney, and small intestine organoids have been examined as potential models of SARS-CoV-2 infection. Lung organoid research has highlighted that SARS-CoV-2 shows preferential infection of club cells and have shown value for the rapid screening and evaluations of multiple anti-viral drugs. Kidney organoid research suggests human recombinant soluble ACE2 as a preventative measure during early-stage infection. Using small intestine organoids, fecal to oral transmission has been evaluated as a transmission route for the virus. Lastly in cardiac organoids drug evaluation studies have found that drugs such as bromodomain, external family inhibitors, BETi, and apabetalone may be effective treatments for SARs-CoV-2 cardiac injury. Summary Organoids are an effective tool to study the effects of viral infections and for drug screening and evaluation studies. By using organoids, more accurate disease modeling can be performed, and physiological effects of infection and treatment can be better understood.
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Hein RFC, Conchola AS, Fine AS, Xiao Z, Frum T, Brastrom LK, Akinwale MA, Childs CJ, Tsai YH, Holloway EM, Huang S, Mahoney J, Heemskerk I, Spence JR. Stable iPSC-derived NKX2-1+ lung bud tip progenitor organoids give rise to airway and alveolar cell types. Development 2022; 149:dev200693. [PMID: 36039869 PMCID: PMC9534489 DOI: 10.1242/dev.200693] [Citation(s) in RCA: 20] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2022] [Accepted: 07/28/2022] [Indexed: 12/13/2022]
Abstract
Bud tip progenitors (BTPs) in the developing lung give rise to all epithelial cell types found in the airways and alveoli. This work aimed to develop an iPSC organoid model enriched with NKX2-1+ BTP-like cells. Building on previous studies, we optimized a directed differentiation paradigm to generate spheroids with more robust NKX2-1 expression. Spheroids were expanded into organoids that possessed NKX2-1+/CPM+ BTP-like cells, which increased in number over time. Single cell RNA-sequencing analysis revealed a high degree of transcriptional similarity between induced BTPs (iBTPs) and in vivo BTPs. Using FACS, iBTPs were purified and expanded as induced bud tip progenitor organoids (iBTOs), which maintained an enriched population of bud tip progenitors. When iBTOs were directed to differentiate into airway or alveolar cell types using well-established methods, they gave rise to organoids composed of organized airway or alveolar epithelium, respectively. Collectively, iBTOs are transcriptionally and functionally similar to in vivo BTPs, providing an important model for studying human lung development and differentiation.
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Affiliation(s)
- Renee F. C. Hein
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
- Department of Computational Medicine and Bioinformatics, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Ansley S. Conchola
- Program in Cell and Molecular Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Alexis S. Fine
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Zhiwei Xiao
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Tristan Frum
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Lindy K. Brastrom
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Mayowa A. Akinwale
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Charlie J. Childs
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
- Department of Computational Medicine and Bioinformatics, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Yu-Hwai Tsai
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Emily M. Holloway
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Sha Huang
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - John Mahoney
- Therapeutics Lab, Cystic Fibrosis Foundation, Lexington, MA 02421, USA
| | - Idse Heemskerk
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
- Department of Computational Medicine and Bioinformatics, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Jason R. Spence
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
- Program in Cell and Molecular Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
- Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA
- Department of Biomedical Engineering Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA
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12
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Guan X, Huang S. Advances in the application of 3D tumor models in precision oncology and drug screening. Front Bioeng Biotechnol 2022; 10:1021966. [PMID: 36246388 PMCID: PMC9555934 DOI: 10.3389/fbioe.2022.1021966] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2022] [Accepted: 09/13/2022] [Indexed: 11/29/2022] Open
Abstract
Traditional tumor models cannot perfectly simulate the real state of tumors in vivo, resulting in the termination of many clinical trials. 3D tumor models’ technology provides new in vitro models that bridge the gap between in vitro and in vivo findings, and organoids maintain the properties of the original tissue over a long period of culture, which enables extensive research in this area. In addition, they can be used as a substitute for animal and in vitro models, and organoids can be established from patients’ normal and malignant tissues, with unique advantages in clinical drug development and in guiding individualized therapies. 3D tumor models also provide a promising platform for high-throughput research, drug and toxicity testing, disease modeling, and regenerative medicine. This report summarizes the 3D tumor model, including evidence regarding the 3D tumor cell culture model, 3D tumor slice model, and organoid culture model. In addition, it provides evidence regarding the application of 3D tumor organoid models in precision oncology and drug screening. The aim of this report is to elucidate the value of 3D tumor models in cancer research and provide a preclinical reference for the precise treatment of cancer patients.
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Affiliation(s)
- Xiaoyong Guan
- Department of Clinical Laboratory, The First Affiliated Hospital of Guangxi University of Science and Technology, Liuzhou, Guangxi, China
| | - Shigao Huang
- Department of Radiation Oncology, The First Affiliated Hospital, Air Force Medical University, Xi’an, China
- *Correspondence: Shigao Huang,
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13
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Lakhdar R, Mumby S, Abubakar-Waziri H, Porter A, Adcock IM, Chung KF. Lung toxicity of particulates and gaseous pollutants using ex-vivo airway epithelial cell culture systems. ENVIRONMENTAL POLLUTION (BARKING, ESSEX : 1987) 2022; 305:119323. [PMID: 35447256 DOI: 10.1016/j.envpol.2022.119323] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/08/2022] [Revised: 04/14/2022] [Accepted: 04/14/2022] [Indexed: 06/14/2023]
Abstract
Air pollution consists of a multi-faceted mix of gases and ambient particulate matter (PM) with diverse organic and non-organic chemical components that contribute to increasing morbidity and mortality worldwide. In particular, epidemiological and clinical studies indicate that respiratory health is adversely affected by exposure to air pollution by both causing and worsening (exacerbating) diseases such as chronic obstructive pulmonary disease (COPD), asthma, interstitial pulmonary fibrosis and lung cancer. The molecular mechanisms of air pollution-induced pulmonary toxicity have been evaluated with regards to different types of PM of various sizes and concentrations with single and multiple exposures over different time periods. These data provide a plausible interrelationship between cellular toxicity and the activation of multiple biological processes including proinflammatory responses, oxidative stress, mitochondrial oxidative damage, autophagy, apoptosis, cell genotoxicity, cellular senescence and epithelial-mesenchymal transition. However, these molecular changes have been studied predominantly in cell lines rather than in primary bronchial or nasal cells from healthy subjects or those isolated from patients with airways disease. In addition, they have been conducted under different cell culture conditions and generally in submerged culture rather than the more relevant air-liquid interface culture and with a variety of air pollutant exposure protocols. Cell types may respond differentially to pollution delivered as an aerosol rather than being bathed in media containing agglomerations of particles. As a result, the actual pathophysiological pathways activated by different PMs in primary cells from the airways of healthy and asthmatic subjects remains unclear. This review summarises the literature on the different methodologies utilised in studying the impact of submicron-sized pollutants on cells derived from the respiratory tract with an emphasis on data obtained from primary human cell. We highlight the critical underlying molecular mechanisms that may be important in driving disease processes in response to air pollution in vivo.
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Affiliation(s)
- Ramzi Lakhdar
- National Heart and Lung Institute and *Department of Materials, Imperial College London, London, SW3 6LY, United Kingdom.
| | - Sharon Mumby
- National Heart and Lung Institute and *Department of Materials, Imperial College London, London, SW3 6LY, United Kingdom.
| | - Hisham Abubakar-Waziri
- National Heart and Lung Institute and *Department of Materials, Imperial College London, London, SW3 6LY, United Kingdom.
| | - Alexandra Porter
- National Heart and Lung Institute and *Department of Materials, Imperial College London, London, SW3 6LY, United Kingdom.
| | - Ian M Adcock
- National Heart and Lung Institute and *Department of Materials, Imperial College London, London, SW3 6LY, United Kingdom.
| | - Kian Fan Chung
- National Heart and Lung Institute and *Department of Materials, Imperial College London, London, SW3 6LY, United Kingdom.
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14
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Bosáková V, De Zuani M, Sládková L, Garlíková Z, Jose SS, Zelante T, Hortová Kohoutková M, Frič J. Lung Organoids—The Ultimate Tool to Dissect Pulmonary Diseases? Front Cell Dev Biol 2022; 10:899368. [PMID: 35912110 PMCID: PMC9326165 DOI: 10.3389/fcell.2022.899368] [Citation(s) in RCA: 23] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2022] [Accepted: 06/24/2022] [Indexed: 11/15/2022] Open
Abstract
Organoids are complex multicellular three-dimensional (3D) in vitro models that are designed to allow accurate studies of the molecular processes and pathologies of human organs. Organoids can be derived from a variety of cell types, such as human primary progenitor cells, pluripotent stem cells, or tumor-derived cells and can be co-cultured with immune or microbial cells to further mimic the tissue niche. Here, we focus on the development of 3D lung organoids and their use as disease models and drug screening tools. We introduce the various experimental approaches used to model complex human diseases and analyze their advantages and disadvantages. We also discuss validation of the organoids and their physiological relevance to the study of lung diseases. Furthermore, we summarize the current use of lung organoids as models of host-pathogen interactions and human lung diseases such as cystic fibrosis, chronic obstructive pulmonary disease, or SARS-CoV-2 infection. Moreover, we discuss the use of lung organoids derived from tumor cells as lung cancer models and their application in personalized cancer medicine research. Finally, we outline the future of research in the field of human induced pluripotent stem cell-derived organoids.
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Affiliation(s)
- Veronika Bosáková
- International Clinical Research Center, St. Anne’s University Hospital Brno, Brno, Czechia
- Department of Biology, Faculty of Medicine, Masaryk University, Brno, Czechia
| | - Marco De Zuani
- International Clinical Research Center, St. Anne’s University Hospital Brno, Brno, Czechia
| | - Lucie Sládková
- Institute of Hematology and Blood Transfusion, Prague, Czechia
- Department of Cell Biology, Faculty of Science, Charles University, Prague, Czechia
| | - Zuzana Garlíková
- International Clinical Research Center, St. Anne’s University Hospital Brno, Brno, Czechia
| | - Shyam Sushama Jose
- International Clinical Research Center, St. Anne’s University Hospital Brno, Brno, Czechia
| | - Teresa Zelante
- Department of Medicine and Surgery, University of Perugia, Perugia, Italy
| | | | - Jan Frič
- International Clinical Research Center, St. Anne’s University Hospital Brno, Brno, Czechia
- Institute of Hematology and Blood Transfusion, Prague, Czechia
- *Correspondence: Jan Frič,
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15
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Hein RFC, Wu JH, Holloway EM, Frum T, Conchola AS, Tsai YH, Wu A, Fine AS, Miller AJ, Szenker-Ravi E, Yan KS, Kuo CJ, Glass I, Reversade B, Spence JR. R-SPONDIN2 + mesenchymal cells form the bud tip progenitor niche during human lung development. Dev Cell 2022; 57:1598-1614.e8. [PMID: 35679862 PMCID: PMC9283295 DOI: 10.1016/j.devcel.2022.05.010] [Citation(s) in RCA: 16] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2021] [Revised: 04/18/2022] [Accepted: 05/16/2022] [Indexed: 01/23/2023]
Abstract
The human respiratory epithelium is derived from a progenitor cell in the distal buds of the developing lung. These "bud tip progenitors" are regulated by reciprocal signaling with surrounding mesenchyme; however, mesenchymal heterogeneity and function in the developing human lung are poorly understood. We interrogated single-cell RNA sequencing data from multiple human lung specimens and identified a mesenchymal cell population present during development that is highly enriched for expression of the WNT agonist RSPO2, and we found that the adjacent bud tip progenitors are enriched for the RSPO2 receptor LGR5. Functional experiments using organoid models, explant cultures, and FACS-isolated RSPO2+ mesenchyme show that RSPO2 is a critical niche cue that potentiates WNT signaling in bud tip progenitors to support their maintenance and multipotency.
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Affiliation(s)
- Renee F C Hein
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Joshua H Wu
- Department of Internal Medicine, Gastroenterology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Emily M Holloway
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Tristan Frum
- Department of Internal Medicine, Gastroenterology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Ansley S Conchola
- Program in Cell and Molecular Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Yu-Hwai Tsai
- Department of Internal Medicine, Gastroenterology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Angeline Wu
- Department of Internal Medicine, Gastroenterology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Alexis S Fine
- Department of Internal Medicine, Gastroenterology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Alyssa J Miller
- Program in Cell and Molecular Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA
| | - Emmanuelle Szenker-Ravi
- Laboratory of Human Genetics & Therapeutics, Genome Institute of Singapore, A(∗)STAR, Singapore 138648, Singapore
| | - Kelley S Yan
- Columbia Center for Human Development, Columbia Stem Cell Initiative, Departments of Medicine and Genetics and Development, Columbia University Irving Medical Center, New York, NY 10032, USA
| | - Calvin J Kuo
- Department of Medicine, Division of Hematology, Stanford University School of Medicine, Stanford, CA 94305, USA
| | - Ian Glass
- Department of Pediatrics, Genetic Medicine, University of Washington, Seattle, WA 98195, USA
| | - Bruno Reversade
- Laboratory of Human Genetics & Therapeutics, Genome Institute of Singapore, A(∗)STAR, Singapore 138648, Singapore; Laboratory of Human Genetics & Therapeutics, Institute of Molecular and Cell Biology (IMCB), A∗STAR, Singapore; Medical Genetics Department, Koç University School of Medicine (KUSOM), Istanbul, Turkey
| | - Jason R Spence
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA; Department of Internal Medicine, Gastroenterology, University of Michigan Medical School, Ann Arbor, MI 48109, USA; Program in Cell and Molecular Biology, University of Michigan Medical School, Ann Arbor, MI 48109, USA; Department of Biomedical Engineering, University of Michigan College of Engineering, Ann Arbor, MI 48109, USA.
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16
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Campion S, Inselman A, Hayes B, Casiraghi C, Joseph D, Facchinetti F, Salomone F, Schmitt G, Hui J, Davis-Bruno K, Van Malderen K, Morford L, De Schaepdrijver L, Wiesner L, Kourula S, Seo S, Laffan S, Urmaliya V, Chen C. The benefits, limitations and opportunities of preclinical models for neonatal drug development. Dis Model Mech 2022; 15:dmm049065. [PMID: 35466995 PMCID: PMC9066504 DOI: 10.1242/dmm.049065] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022] Open
Abstract
Increased research to improve preclinical models to inform the development of therapeutics for neonatal diseases is an area of great need. This article reviews five common neonatal diseases - bronchopulmonary dysplasia, retinopathy of prematurity, necrotizing enterocolitis, perinatal hypoxic-ischemic encephalopathy and neonatal sepsis - and the available in vivo, in vitro and in silico preclinical models for studying these diseases. Better understanding of the strengths and weaknesses of specialized neonatal disease models will help to improve their utility, may add to the understanding of the mode of action and efficacy of a therapeutic, and/or may improve the understanding of the disease pathology to aid in identification of new therapeutic targets. Although the diseases covered in this article are diverse and require specific approaches, several high-level, overarching key lessons can be learned by evaluating the strengths, weaknesses and gaps in the available models. This Review is intended to help guide current and future researchers toward successful development of therapeutics in these areas of high unmet medical need.
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Affiliation(s)
- Sarah Campion
- Pfizer Worldwide Research, Development, and Medical, Groton, CT 06340, USA
| | - Amy Inselman
- U.S. Food and Drug Administration, National Center for Toxicological Research, Division of Systems Biology, Jefferson, AR 72079, USA
| | - Belinda Hayes
- U.S. Food and Drug Administration, Center for Drug Evaluation and Research, Office of New Drugs, Silver Spring, MD 20993, USA
| | - Costanza Casiraghi
- Department of Experimental Pharmacology and Translational Science, Chiesi Farmaceutici S.p.A., 43122 Parma, Italy
| | - David Joseph
- U.S. Food and Drug Administration, Center for Drug Evaluation and Research, Office of New Drugs, Silver Spring, MD 20993, USA
| | - Fabrizio Facchinetti
- Department of Experimental Pharmacology and Translational Science, Chiesi Farmaceutici S.p.A., 43122 Parma, Italy
| | - Fabrizio Salomone
- Department of Experimental Pharmacology and Translational Science, Chiesi Farmaceutici S.p.A., 43122 Parma, Italy
| | - Georg Schmitt
- Pharma Research and Early Development, Roche Innovation Center Basel, Pharmaceutical Sciences, F. Hoffmann-La Roche, 4070 Basel, Switzerland
| | - Julia Hui
- Bristol Myers Squibb, Nonclinical Research and Development, Summit, NJ 07901, USA
| | - Karen Davis-Bruno
- U.S. Food and Drug Administration, Center for Drug Evaluation and Research, Office of New Drugs, Silver Spring, MD 20993, USA
| | - Karen Van Malderen
- Federal Agency for Medicines and Health Products (FAMHP), Department DG PRE authorization, 1210 Brussels, Belgium
| | - LaRonda Morford
- Eli Lilly, Global Regulatory Affairs, Indianapolis, IN 46285, USA
| | | | - Lutz Wiesner
- Federal Institute for Drugs and Medical Devices, Clinical Trials, 53175 Bonn, Germany
| | - Stephanie Kourula
- Janssen R&D, Drug Metabolism & Pharmacokinetics, 2340 Beerse, Belgium
| | - Suna Seo
- U.S. Food and Drug Administration, Center for Drug Evaluation and Research, Office of New Drugs, Silver Spring, MD 20993, USA
| | - Susan Laffan
- GlaxoSmithKline, Non-Clinical Safety, Collegeville, PA 19406, USA
| | | | - Connie Chen
- Health and Environmental Sciences Institute, Washington, DC 20005, USA
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17
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Mahapatra C, Lee R, Paul MK. Emerging role and promise of nanomaterials in organoid research. Drug Discov Today 2022; 27:890-899. [PMID: 34774765 DOI: 10.1016/j.drudis.2021.11.007] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2021] [Revised: 09/27/2021] [Accepted: 11/04/2021] [Indexed: 12/30/2022]
Abstract
Organoids are 3D stem cell-derived self-organization of cells. Organoid bioengineering helps recreate and tailor their architecture in vitro to generate mini organ-like properties, providing the opportunity to study fundamental cell behavior in heterogeneous populations and as a tool to model various diseases. Nanomaterials (NMs) are becoming indispensable in regenerative medicine and in developing treatment modalities for various diseases. Therefore, organoid-NM interactions are set to gain traction for the development of advanced diagnostics and therapeutics. Here, we discuss the interactions of NMs with distinctive organoid types, organoid matrices, trafficking and cargo delivery, organs-on-a-chip, bioprinting, downstream therapeutic implications, and future approaches.
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Affiliation(s)
- Chinmaya Mahapatra
- Department of Biotechnology, National Institute of Technology Raipur, Raipur, Chhattisgarh 492010, India
| | - Ruda Lee
- International Research Organization for Advanced Science and Technology (IROAST), Kumamoto University, Kumamoto 860-8555, Japan
| | - Manash K Paul
- Department of Pulmonary and Critical Care Medicine, David Geffen School of Medicine, University of California Los Angeles (UCLA), Los Angeles, CA 90095, USA.
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18
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Unravelling the molecular mechanisms underlying chronic respiratory diseases for the development of novel therapeutics via in vitro experimental models. Eur J Pharmacol 2022; 919:174821. [DOI: 10.1016/j.ejphar.2022.174821] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/20/2021] [Revised: 02/01/2022] [Accepted: 02/09/2022] [Indexed: 12/11/2022]
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19
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Three-Dimensional Airway Spheroids and Organoids for Cystic Fibrosis Research. JOURNAL OF RESPIRATION 2021. [DOI: 10.3390/jor1040022] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Cystic fibrosis (CF) is an autosomal recessive multi-organ disease caused by mutations in the CF Transmembrane Conductance Regulator (CFTR) gene, with morbidity and mortality primacy related to the lung disease. The CFTR protein, a chloride/bicarbonate channel, is expressed at the apical side of airway epithelial cells and is mainly involved in appropriate ion and fluid transport across the epithelium. Although many animal and cellular models have been developed to study the pathophysiological consequences of the lack/dysfunction of CFTR, only the three-dimensional (3D) structures termed “spheroids” and “organoids” can enable the reconstruction of airway mucosa to model organ development, disease pathophysiology, and drug screening. Airway spheroids and organoids can be derived from different sources, including adult lungs and induced pluripotent stem cells (iPSCs), each with its advantages and limits. Here, we review the major features of airway spheroids and organoids, anticipating that their potential in the CF field has not been fully shown. Further work is mandatory to understand whether they can accomplish better outcomes than other culture conditions of airway epithelial cells for CF personalized therapies and tissue engineering aims.
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20
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Viana F, O'Kane CM, Schroeder GN. Precision-cut lung slices: A powerful ex vivo model to investigate respiratory infectious diseases. Mol Microbiol 2021; 117:578-588. [PMID: 34570407 PMCID: PMC9298270 DOI: 10.1111/mmi.14817] [Citation(s) in RCA: 43] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2021] [Accepted: 09/23/2021] [Indexed: 11/30/2022]
Abstract
Respiratory infections are a leading cause of mortality worldwide. Most of the research on the underlying disease mechanisms is based on cell culture, organoid, or surrogate animal models. Although these provide important insights, they have limitations. Cell culture models fail to recapitulate cellular interactions in the lung and animal models often do not permit high‐throughput analysis of drugs or pathogen isolates; hence, there is a need for improved, scalable models. Precision‐cut lung slices (PCLS), small, uniform tissue slices generated from animal or human lungs are increasingly recognized and employed as an ex vivo organotypic model. PCLS retain remarkable cellular complexity and the architecture of the lung, providing a platform to investigate respiratory pathogens in a near‐native environment. Here, we review the generation and features of PCLS, their use to investigate the pathogenesis of viral and bacterial pathogens, and highlight their potential to advance respiratory infection research in the future.
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Affiliation(s)
- Flávia Viana
- Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry and Biomedical Sciences, Queen's University Belfast, Belfast, UK
| | - Cecilia M O'Kane
- Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry and Biomedical Sciences, Queen's University Belfast, Belfast, UK
| | - Gunnar N Schroeder
- Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry and Biomedical Sciences, Queen's University Belfast, Belfast, UK
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21
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Prasad M, Kumar R, Buragohain L, Kumari A, Ghosh M. Organoid Technology: A Reliable Developmental Biology Tool for Organ-Specific Nanotoxicity Evaluation. Front Cell Dev Biol 2021; 9:696668. [PMID: 34631696 PMCID: PMC8495170 DOI: 10.3389/fcell.2021.696668] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/17/2021] [Accepted: 08/13/2021] [Indexed: 12/14/2022] Open
Abstract
Engineered nanomaterials are bestowed with certain inherent physicochemical properties unlike their parent materials, rendering them suitable for the multifaceted needs of state-of-the-art biomedical, and pharmaceutical applications. The log-phase development of nano-science along with improved "bench to beside" conversion carries an enhanced probability of human exposure with numerous nanoparticles. Thus, toxicity assessment of these novel nanoscale materials holds a key to ensuring the safety aspects or else the global biome will certainly face a debacle. The toxicity may span from health hazards due to direct exposure to indirect means through food chain contamination or environmental pollution, even causing genotoxicity. Multiple ways of nanotoxicity evaluation include several in vitro and in vivo methods, with in vitro methods occupying the bulk of the "experimental space." The underlying reason may be multiple, but ethical constraints in in vivo animal experiments are a significant one. Two-dimensional (2D) monoculture is undoubtedly the most exploited in vitro method providing advantages in terms of cost-effectiveness, high throughput, and reproducibility. However, it often fails to mimic a tissue or organ which possesses a defined three-dimensional structure (3D) along with intercellular communication machinery. Instead, microtissues such as spheroids or organoids having a precise 3D architecture and proximate in vivo tissue-like behavior can provide a more realistic evaluation than 2D monocultures. Recent developments in microfluidics and bioreactor-based organoid synthesis have eased the difficulties to prosper nano-toxicological analysis in organoid models surpassing the obstacle of ethical issues. The present review will enlighten applications of organoids in nanotoxicological evaluation, their advantages, and prospects toward securing commonplace nano-interventions.
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Affiliation(s)
- Minakshi Prasad
- Department of Animal Biotechnology, Lala Lajpat Rai University of Veterinary and Animal Sciences, Hisar, India
| | - Rajesh Kumar
- Department of Veterinary Physiology and Biochemistry, Lala Lajpat Rai University of Veterinary and Animal Sciences, Hisar, India
| | - Lukumoni Buragohain
- Department of Animal Biotechnology, College of Veterinary Science, Assam Agricultural University, Guwahati, India
| | | | - Mayukh Ghosh
- Department of Veterinary Physiology and Biochemistry, RGSC, Banaras Hindu University, Varanasi, India
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22
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Yaqub N, Wayne G, Birchall M, Song W. Recent advances in human respiratory epithelium models for drug discovery. Biotechnol Adv 2021; 54:107832. [PMID: 34481894 DOI: 10.1016/j.biotechadv.2021.107832] [Citation(s) in RCA: 30] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2021] [Revised: 07/08/2021] [Accepted: 08/30/2021] [Indexed: 12/12/2022]
Abstract
The respiratory epithelium is intimately associated with the pathophysiologies of highly infectious viral contagions and chronic illnesses such as chronic obstructive pulmonary disorder, presently the third leading cause of death worldwide with a projected economic burden of £1.7 trillion by 2030. Preclinical studies of respiratory physiology have almost exclusively utilised non-humanised animal models, alongside reductionistic cell line-based models, and primary epithelial cell models cultured at an air-liquid interface (ALI). Despite their utility, these model systems have been limited by their poor correlation to the human condition. This has undermined the ability to identify novel therapeutics, evidenced by a 15% chance of success for medicinal respiratory compounds entering clinical trials in 2018. Consequently, preclinical studies require new translational efficacy models to address the problem of respiratory drug attrition. This review describes the utility of the current in vivo (rodent), ex vivo (isolated perfused lungs and precision cut lung slices), two-dimensional in vitro cell-line (A549, BEAS-2B, Calu-3) and three-dimensional in vitro ALI (gold-standard and co-culture) and organoid respiratory epithelium models. The limitations to the application of these model systems in drug discovery research are discussed, in addition to perspectives of the future innovations required to facilitate the next generation of human-relevant respiratory models.
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Affiliation(s)
- Naheem Yaqub
- UCL Centre for Biomaterials in Surgical Reconstruction and Regeneration, Department of Surgical Biotechnology, Division of Surgery & Interventional Science, University College London, London NW3 2PF, UK
| | - Gareth Wayne
- Novel Human Genetics, GlaxoSmithKline, Stevenage SG1 2NY, UK
| | - Martin Birchall
- The Ear Institute, Faculty of Brain Sciences, University College London, London WC1X 8EE, UK.
| | - Wenhui Song
- UCL Centre for Biomaterials in Surgical Reconstruction and Regeneration, Department of Surgical Biotechnology, Division of Surgery & Interventional Science, University College London, London NW3 2PF, UK.
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23
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Tisdale-Macioce N, Green J, Perl AKT, Ashbaugh A, Wiederhold NP, Patterson TF, Cushion MT. The Promise of Lung Organoids for Growth and Investigation of Pneumocystis Species. FRONTIERS IN FUNGAL BIOLOGY 2021; 2:740845. [PMID: 37744131 PMCID: PMC10512221 DOI: 10.3389/ffunb.2021.740845] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 07/13/2021] [Accepted: 08/06/2021] [Indexed: 09/26/2023]
Abstract
Pneumocystis species (spp.) are host-obligate fungal parasites that colonize and propagate almost exclusively in the alveolar lumen within the lungs of mammals where they can cause a lethal pneumonia. The emergence of this pneumonia in non-HIV infected persons caused by Pneumocystis jirovecii (PjP), illustrates the continued importance of and the need to understand its associated pathologies and to develop new therapies and preventative strategies. In the proposed life cycle, Pneumocystis spp. attach to alveolar type 1 epithelial cells (AEC1) and prevent gas exchange. This process among other mechanisms of Pneumocystis spp. pathogenesis is challenging to observe in real time due to the absence of a continuous ex vivo or in vitro culture system. The study presented here provides a proof-of-concept for the development of murine lung organoids that mimic the lung alveolar sacs expressing alveolar epithelial type 1 cells (AEC1) and alveolar type 2 epithelial cells (AEC2). Use of these 3-dimensional organoids should facilitate studies of a multitude of unanswered questions and serve as an improved means to screen new anti- PjP agents.
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Affiliation(s)
- Nikeya Tisdale-Macioce
- Department of Internal Medicine, University of Cincinnati College of Medicine, Cincinnati, OH, United States
- Medical Research Service, Cincinnati Veterans Affairs Medical Center, Cincinnati, OH, United States
| | - Jenna Green
- Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, OH, United States
| | - Anne-Karina T. Perl
- Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, OH, United States
- Perinatal and Pulmonary Biology, Cincinnati Children's Hospital Medical Center, The Perinatal Institute and Section of Neonatology, Cincinnati, OH, United States
| | - Alan Ashbaugh
- Department of Internal Medicine, University of Cincinnati College of Medicine, Cincinnati, OH, United States
- Medical Research Service, Cincinnati Veterans Affairs Medical Center, Cincinnati, OH, United States
| | - Nathan P. Wiederhold
- Department of Pathology, The University of Texas Health Science Center, San Antonio, TX, United States
| | - Thomas F. Patterson
- Department of Medicine, The University of Texas Health Science Center, San Antonio, TX, United States
- Section of Infectious Diseases, South Texas Veterans Health Care System, San Antonio, TX, United States
| | - Melanie T. Cushion
- Department of Internal Medicine, University of Cincinnati College of Medicine, Cincinnati, OH, United States
- Medical Research Service, Cincinnati Veterans Affairs Medical Center, Cincinnati, OH, United States
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Tindle C, Fuller M, Fonseca A, Taheri S, Ibeawuchi SR, Beutler N, Katkar GD, Claire A, Castillo V, Hernandez M, Russo H, Duran J, Crotty Alexander LE, Tipps A, Lin G, Thistlethwaite PA, Chattopadhyay R, Rogers TF, Sahoo D, Ghosh P, Das S. Adult stem cell-derived complete lung organoid models emulate lung disease in COVID-19. eLife 2021; 10:e66417. [PMID: 34463615 PMCID: PMC8463074 DOI: 10.7554/elife.66417] [Citation(s) in RCA: 79] [Impact Index Per Article: 19.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/10/2021] [Accepted: 08/11/2021] [Indexed: 12/13/2022] Open
Abstract
Background SARS-CoV-2, the virus responsible for COVID-19, causes widespread damage in the lungs in the setting of an overzealous immune response whose origin remains unclear. Methods We present a scalable, propagable, personalized, cost-effective adult stem cell-derived human lung organoid model that is complete with both proximal and distal airway epithelia. Monolayers derived from adult lung organoids (ALOs), primary airway cells, or hiPSC-derived alveolar type II (AT2) pneumocytes were infected with SARS-CoV-2 to create in vitro lung models of COVID-19. Results Infected ALO monolayers best recapitulated the transcriptomic signatures in diverse cohorts of COVID-19 patient-derived respiratory samples. The airway (proximal) cells were critical for sustained viral infection, whereas distal alveolar differentiation (AT2→AT1) was critical for mounting the overzealous host immune response in fatal disease; ALO monolayers with well-mixed proximodistal airway components recapitulated both. Conclusions Findings validate a human lung model of COVID-19, which can be immediately utilized to investigate COVID-19 pathogenesis and vet new therapies and vaccines. Funding This work was supported by the National Institutes for Health (NIH) grants 1R01DK107585-01A1, 3R01DK107585-05S1 (to SD); R01-AI141630, CA100768 and CA160911 (to PG) and R01-AI 155696 (to PG, DS and SD); R00-CA151673 and R01-GM138385 (to DS), R01- HL32225 (to PT), UCOP-R00RG2642 (to SD and PG), UCOP-R01RG3780 (to P.G. and D.S) and a pilot award from the Sanford Stem Cell Clinical Center at UC San Diego Health (P.G, S.D, D.S). GDK was supported through The American Association of Immunologists Intersect Fellowship Program for Computational Scientists and Immunologists. L.C.A's salary was supported in part by the VA San Diego Healthcare System. This manuscript includes data generated at the UC San Diego Institute of Genomic Medicine (IGC) using an Illumina NovaSeq 6000 that was purchased with funding from a National Institutes of Health SIG grant (#S10 OD026929).
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Affiliation(s)
- Courtney Tindle
- Department of Cellular and Molecular Medicine, University of California San DiegoSan DiegoUnited States
- HUMANOID CoRE, University of California San DiegoSan DiegoUnited States
| | - MacKenzie Fuller
- Department of Cellular and Molecular Medicine, University of California San DiegoSan DiegoUnited States
- HUMANOID CoRE, University of California San DiegoSan DiegoUnited States
| | - Ayden Fonseca
- Department of Cellular and Molecular Medicine, University of California San DiegoSan DiegoUnited States
- HUMANOID CoRE, University of California San DiegoSan DiegoUnited States
| | - Sahar Taheri
- Department of Computer Science and Engineering, Jacobs School of Engineering, University of California San DiegoSan DiegoUnited States
| | | | - Nathan Beutler
- Department of Immunology and Microbiology, The Scripps Research InstituteLa JollaUnited States
| | - Gajanan Dattatray Katkar
- Department of Cellular and Molecular Medicine, University of California San DiegoSan DiegoUnited States
| | - Amanraj Claire
- Department of Cellular and Molecular Medicine, University of California San DiegoSan DiegoUnited States
- HUMANOID CoRE, University of California San DiegoSan DiegoUnited States
| | - Vanessa Castillo
- Department of Cellular and Molecular Medicine, University of California San DiegoSan DiegoUnited States
| | - Moises Hernandez
- Division of Cardiothoracic Surgery, University of California San DiegoSan DiegoUnited States
| | - Hana Russo
- Department of Pathology, University of California San DiegoSan DiegoUnited States
| | - Jason Duran
- Division of Cardiology, Department of Internal Medicine, UC San Diego Medical CenterSan DiegoUnited States
| | - Laura E Crotty Alexander
- Pulmonary Critical Care Section, Veterans Affairs (VA) San Diego Healthcare SystemLa JollaUnited States
- Division of Pulmonary and Critical Care, Department of Medicine, University of California, San DiegoLa Jolla, CAUnited States
| | - Ann Tipps
- Department of Pathology, University of California San DiegoSan DiegoUnited States
| | - Grace Lin
- Department of Pathology, University of California San DiegoSan DiegoUnited States
| | | | - Ranajoy Chattopadhyay
- Department of Cellular and Molecular Medicine, University of California San DiegoSan DiegoUnited States
- HUMANOID CoRE, University of California San DiegoSan DiegoUnited States
- Cell Applications Inc.La Jolla, CAUnited States
| | - Thomas F Rogers
- Department of Immunology and Microbiology, The Scripps Research InstituteLa JollaUnited States
- Division of Infectious Diseases, Department of Medicine, University of California, San DiegoLa JollaUnited States
- Department of Immunology and Microbiology, The Scripps Research InstituteLa JollaUnited States
| | - Debashis Sahoo
- Department of Computer Science and Engineering, Jacobs School of Engineering, University of California San DiegoSan DiegoUnited States
- Department of Pediatrics, University of California, San DiegoLa Jolla, CAUnited States
| | - Pradipta Ghosh
- Department of Cellular and Molecular Medicine, University of California San DiegoSan DiegoUnited States
- HUMANOID CoRE, University of California San DiegoSan DiegoUnited States
- Department of Medicine, University of California, San DiegoLa Jolla, CAUnited States
| | - Soumita Das
- HUMANOID CoRE, University of California San DiegoSan DiegoUnited States
- Department of Pathology, University of California San DiegoSan DiegoUnited States
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Lv T, Meng F, Yu M, Huang H, Lin X, Zhao B. Defense of COVID-19 by Human Organoids. PHENOMICS (CHAM, SWITZERLAND) 2021; 1:113-128. [PMID: 35233559 PMCID: PMC8277987 DOI: 10.1007/s43657-021-00015-0] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/27/2021] [Revised: 05/08/2021] [Accepted: 05/11/2021] [Indexed: 01/08/2023]
Abstract
Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), has created an immense menace to public health worldwide, exerting huge effects on global economic and political conditions. Understanding the biology and pathogenesis mechanisms of this novel virus, in large parts, relies on optimal physiological models that allow replication and propagation of SARS-CoV-2. Human organoids, derived from stem cells, are three-dimensional cell cultures that recapitulate the cellular organization, transcriptional and epigenetic signatures of their counterpart organs. Recent studies have indicated their great values as experimental virology platforms, making human organoid an ideal tool for investigating host-pathogen interactions. Here, we summarize research developments for SARS-CoV-2 infection of various human organoids involved in multiple systems, including lung, liver, brain, intestine, kidney and blood vessel organoids. These studies help us reveal the pathogenesis mechanism of COVID-19, and facilitate the development of effective vaccines and drugs as well as other therapeutic regimes.
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Affiliation(s)
- Ting Lv
- State Key Laboratory of Genetic Engineering, School of Life Sciences, Zhongshan Hospital, Fudan University, Shanghai, 200438 China
- Institute of Antibiotics, Huashan Hospital, Fudan University, Shanghai, 200040 China
| | - Fanlu Meng
- Shandong Key Laboratory of Biophysics, Institute of Biophysics, Dezhou University, Dezhou, 253023 China
| | - Meng Yu
- State Key Laboratory of Genetic Engineering, School of Life Sciences, Zhongshan Hospital, Fudan University, Shanghai, 200438 China
| | - Haihui Huang
- Institute of Antibiotics, Huashan Hospital, Fudan University, Shanghai, 200040 China
| | - Xinhua Lin
- State Key Laboratory of Genetic Engineering, School of Life Sciences, Zhongshan Hospital, Fudan University, Shanghai, 200438 China
| | - Bing Zhao
- State Key Laboratory of Genetic Engineering, School of Life Sciences, Zhongshan Hospital, Fudan University, Shanghai, 200438 China
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Harman RM, Theoret CL, Van de Walle GR. The Horse as a Model for the Study of Cutaneous Wound Healing. Adv Wound Care (New Rochelle) 2021; 10:381-399. [PMID: 34042536 DOI: 10.1089/wound.2018.0883] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
Significance: Cutaneous wounds are a major problem in both human and equine medicine. The economic cost of treating skin wounds and related complications in humans and horses is high, and in both species, particular types of chronic wounds do not respond well to current therapies, leading to suffering and morbidity. Recent Advances: Conventional methods for the treatment of cutaneous wounds are generic and have not changed significantly in decades. However, as more is learned about the mechanisms involved in normal skin wound healing, and how failure of these processes leads to chronic nonhealing wounds, novel therapies targeting the specific pathologies of hard-to-heal wounds are being developed and evaluated. Critical Issues: Physiologically relevant animal models are needed to (1) study the mechanisms involved in normal and impaired skin wound healing and (2) test newly developed therapies. Future Directions: Similarities in normal wound healing in humans and horses, and the natural development of distinct types of hard-to-heal chronic wounds in both species, make the horse a physiologically relevant model for the study of mechanisms involved in wound repair. Horses are also well-suited models to test novel therapies. In addition, studies in horses have the potential to benefit veterinary, as well as human medicine.
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Affiliation(s)
- Rebecca M. Harman
- Baker Institute for Animal Health, College of Veterinary Medicine, Cornell University, Ithaca, New York
| | | | - Gerlinde R. Van de Walle
- Baker Institute for Animal Health, College of Veterinary Medicine, Cornell University, Ithaca, New York
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27
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Bukowy-Bieryłło Z. Long-term differentiating primary human airway epithelial cell cultures: how far are we? Cell Commun Signal 2021; 19:63. [PMID: 34044844 PMCID: PMC8159066 DOI: 10.1186/s12964-021-00740-z] [Citation(s) in RCA: 38] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/02/2021] [Accepted: 04/16/2021] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND Human airway epithelial (HAE) cellular models are widely used in applicative studies of the airway physiology and disease. In vitro expanded and differentiated primary HAE cells collected from patients seem to be an accurate model of human airway, offering a quicker and cheaper alternative to the induced pluripotent stem cell (iPSCs) models. However, the biggest drawback of primary HAE models is their limited proliferative lifespan in culture. Much work has been devoted to understand the factors, which govern the HAE cell proliferation and differentiation, both in vivo and in vitro. Here, I have summarized recent achievements in primary HAE culture, with the special emphasis on the models of conditionally reprogrammed cells (CRC), which allow longer in vitro proliferation and differentiation of HAE cells. The review compares the CRC HAE technique variants (feeder culture or HAE mono-culture), based on recently published studies exploiting this model. The advantages and limitations of each CRC HAE model variant are summarized, along with the description of other factors affecting the CRC HAE culture success (tissue type, sampling method, sample quality). CONCLUSIONS CRC HAE cultures are a useful technique in respiratory research, which in many cases exceeds the iPSCs and organoid culture methods. Until the current limitations of the iPSCs and organoid culture methods will be alleviated, the primary CRC HAE cultures might be a useful model in respiratory research. Airway epithelium (AE) is a type of tissue, which lines the whole length of human airways, from the nose to the bronchi. Improper functioning of AE causes several human airway disorders, such as asthma, chronic obstructive pulmonary disease (COPD) or cystic fibrosis (CF). Much work has been devoted to finding the best scientific model of human AE, in order to learn about its functioning in health and disease. Among the popular AE models are the primary in vitro cultured AE cells collected from human donors. Unfortunately, such human AE (HAE) cells do not easily divide (expand) in vitro; this poses a large logistic and ethical problem for the researchers. Here, I summarize recent achievements in the methods for in vitro culture of human AE cells, with special emphasis on the conditionally reprogrammed cell (CRC) models, which allow longer and more effective expansion of primary human AE cells in vitro. The review describes how the specific chemicals used in the CRC models work to allow the increased HAE divisions and compares the effects of the different so-far developed variants of the CRC HAE culture. The review also pinpoints the areas which need to be refined, in order to maximize the usefulness of the CRC AE cultures from human donors in research on human airway disorders. Video abstract.
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Tindle C, Fuller M, Fonseca A, Taheri S, Ibeawuchi SR, Beutler N, Katkar G, Claire A, Castillo V, Hernandez M, Russo H, Duran J, Crotty Alexander LE, Tipps A, Lin G, Thistlethwaite PA, Chattopadhyay R, Rogers TF, Sahoo D, Ghosh P, Das S. Adult Stem Cell-derived Complete Lung Organoid Models Emulate Lung Disease in COVID-19. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2021:2020.10.17.344002. [PMID: 33106807 PMCID: PMC7587781 DOI: 10.1101/2020.10.17.344002] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 04/12/2023]
Abstract
SARS-CoV-2, the virus responsible for COVID-19, causes widespread damage in the lungs in the setting of an overzealous immune response whose origin remains unclear. We present a scalable, propagable, personalized, cost-effective adult stem cell-derived human lung organoid model that is complete with both proximal and distal airway epithelia. Monolayers derived from adult lung organoids (ALOs), primary airway cells, or hiPSC-derived alveolar type-II (AT2) pneumocytes were infected with SARS-CoV-2 to create in vitro lung models of COVID-19. Infected ALO-monolayers best recapitulated the transcriptomic signatures in diverse cohorts of COVID-19 patient-derived respiratory samples. The airway (proximal) cells were critical for sustained viral infection whereas distal alveolar differentiation (AT2→AT1) was critical for mounting the overzealous host immune response in fatal disease; ALO monolayers with well-mixed proximodistal airway components recapitulated both. Findings validate a human lung model of COVID-19 which can be immediately utilized to investigate COVID-19 pathogenesis, and vet new therapies and vaccines.
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29
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Lu T, Cao Y, Zhao P, Shen S, Xi Y. Organoid: a powerful tool to study lung regeneration and disease. CELL REGENERATION (LONDON, ENGLAND) 2021; 10:21. [PMID: 33900491 PMCID: PMC8074347 DOI: 10.1186/s13619-021-00082-8] [Citation(s) in RCA: 30] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 12/31/2020] [Accepted: 04/02/2021] [Indexed: 02/07/2023]
Abstract
Organoids are three-dimensional self-organizing structures formed by adult tissue stem cells or pluripotent stem cells. They recapitulate cell-cell, cell-niche interactions in tissue development, homeostasis, regeneration and disease, and provide an in vitro model for drug screening. This review summarizes the recent advances of organoid cultures derived from adult lung stem cells and human pluripotent stem cells, especially focusing on the organoids of the distal airway stem/progenitor cells. We also discuss the applications of organoids in studying lung regeneration and pulmonary diseases, including pulmonary fibrosis, airway diseases and Coronavirus disease 2019 (COVID-19).
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Affiliation(s)
- Tiantian Lu
- School of Life Science and Technology, ShanghaiTech University, Shanghai, China
| | - Yiyuan Cao
- School of Life Science and Technology, ShanghaiTech University, Shanghai, China
| | - Peng Zhao
- School of Life Science and Technology, ShanghaiTech University, Shanghai, China
| | - Shengxi Shen
- School of Life Science and Technology, ShanghaiTech University, Shanghai, China
| | - Ying Xi
- School of Life Science and Technology, ShanghaiTech University, Shanghai, China.
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30
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Wanczyk H, Jensen T, Weiss DJ, Finck C. Advanced single-cell technologies to guide the development of bioengineered lungs. Am J Physiol Lung Cell Mol Physiol 2021; 320:L1101-L1117. [PMID: 33851545 DOI: 10.1152/ajplung.00089.2021] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
Lung transplantation remains the only viable option for individuals suffering from end-stage lung failure. However, a number of current limitations exist including a continuing shortage of suitable donor lungs and immune rejection following transplantation. To address these concerns, engineering a decellularized biocompatible lung scaffold from cadavers reseeded with autologous lung cells to promote tissue regeneration is being explored. Proof-of-concept transplantation of these bioengineered lungs into animal models has been accomplished. However, these lungs were incompletely recellularized with resulting epithelial and endothelial leakage and insufficient basement membrane integrity. Failure to repopulate lung scaffolds with all of the distinct cell populations necessary for proper function remains a significant hurdle for the progression of current engineering approaches and precludes clinical translation. Advancements in 3D bioprinting, lung organoid models, and microfluidic device and bioreactor development have enhanced our knowledge of pulmonary lung development, as well as important cell-cell and cell-matrix interactions, all of which will help in the path to a bioengineered transplantable lung. However, a significant gap in knowledge of the spatiotemporal interactions between cell populations as well as relative quantities and localization within each compartment of the lung necessary for its proper growth and function remains. This review will provide an update on cells currently used for reseeding decellularized scaffolds with outcomes of recent lung engineering attempts. Focus will then be on how data obtained from advanced single-cell analyses, coupled with multiomics approaches and high-resolution 3D imaging, can guide current lung bioengineering efforts for the development of fully functional, transplantable lungs.
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Affiliation(s)
- Heather Wanczyk
- Department of Pediatrics, University of Connecticut Health Center, Farmington, Connecticut
| | - Todd Jensen
- Department of Pediatrics, University of Connecticut Health Center, Farmington, Connecticut
| | - Daniel J Weiss
- Department of Medicine, University of Vermont, Burlington, Vermont
| | - Christine Finck
- Department of Pediatrics, University of Connecticut Health Center, Farmington, Connecticut.,Department of Surgery, Connecticut Children's Medical Center, Hartford, Connecticut
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31
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Lamers MM, van der Vaart J, Knoops K, Riesebosch S, Breugem TI, Mykytyn AZ, Beumer J, Schipper D, Bezstarosti K, Koopman CD, Groen N, Ravelli RBG, Duimel HQ, Demmers JAA, Verjans GMGM, Koopmans MPG, Muraro MJ, Peters PJ, Clevers H, Haagmans BL. An organoid-derived bronchioalveolar model for SARS-CoV-2 infection of human alveolar type II-like cells. EMBO J 2021; 40:e105912. [PMID: 33283287 PMCID: PMC7883112 DOI: 10.15252/embj.2020105912] [Citation(s) in RCA: 148] [Impact Index Per Article: 37.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2020] [Revised: 12/03/2020] [Accepted: 12/04/2020] [Indexed: 12/15/2022] Open
Abstract
Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) causes coronavirus disease 2019 (COVID‐19), which may result in acute respiratory distress syndrome (ARDS), multiorgan failure, and death. The alveolar epithelium is a major target of the virus, but representative models to study virus host interactions in more detail are currently lacking. Here, we describe a human 2D air–liquid interface culture system which was characterized by confocal and electron microscopy and single‐cell mRNA expression analysis. In this model, alveolar cells, but also basal cells and rare neuroendocrine cells, are grown from 3D self‐renewing fetal lung bud tip organoids. These cultures were readily infected by SARS‐CoV‐2 with mainly surfactant protein C‐positive alveolar type II‐like cells being targeted. Consequently, significant viral titers were detected and mRNA expression analysis revealed induction of type I/III interferon response program. Treatment of these cultures with a low dose of interferon lambda 1 reduced viral replication. Hence, these cultures represent an experimental model for SARS‐CoV‐2 infection and can be applied for drug screens.
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Affiliation(s)
- Mart M Lamers
- Viroscience Department, Erasmus University Medical Center, Rotterdam, The Netherlands
| | - Jelte van der Vaart
- Oncode Institute, Hubrecht Institute, Royal Netherlands Academy of Arts and Sciences and University Medical Center, Utrecht, The Netherlands
| | - Kèvin Knoops
- The Maastricht Multimodal Molecular Imaging Institute, Maastricht University, Maastricht, The Netherlands
| | - Samra Riesebosch
- Viroscience Department, Erasmus University Medical Center, Rotterdam, The Netherlands
| | - Tim I Breugem
- Viroscience Department, Erasmus University Medical Center, Rotterdam, The Netherlands
| | - Anna Z Mykytyn
- Viroscience Department, Erasmus University Medical Center, Rotterdam, The Netherlands
| | - Joep Beumer
- Oncode Institute, Hubrecht Institute, Royal Netherlands Academy of Arts and Sciences and University Medical Center, Utrecht, The Netherlands
| | - Debby Schipper
- Viroscience Department, Erasmus University Medical Center, Rotterdam, The Netherlands
| | - Karel Bezstarosti
- Proteomics Center, Erasmus University Medical Center, Rotterdam, The Netherlands
| | | | | | - Raimond B G Ravelli
- The Maastricht Multimodal Molecular Imaging Institute, Maastricht University, Maastricht, The Netherlands
| | - Hans Q Duimel
- The Maastricht Multimodal Molecular Imaging Institute, Maastricht University, Maastricht, The Netherlands
| | - Jeroen A A Demmers
- Proteomics Center, Erasmus University Medical Center, Rotterdam, The Netherlands
| | - Georges M G M Verjans
- Viroscience Department, Erasmus University Medical Center, Rotterdam, The Netherlands
| | - Marion P G Koopmans
- Viroscience Department, Erasmus University Medical Center, Rotterdam, The Netherlands
| | | | - Peter J Peters
- The Maastricht Multimodal Molecular Imaging Institute, Maastricht University, Maastricht, The Netherlands
| | - Hans Clevers
- Oncode Institute, Hubrecht Institute, Royal Netherlands Academy of Arts and Sciences and University Medical Center, Utrecht, The Netherlands
| | - Bart L Haagmans
- Viroscience Department, Erasmus University Medical Center, Rotterdam, The Netherlands
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Liu NN, Ma Q, Ge Y, Yi CX, Wei LQ, Tan JC, Chu Q, Li JQ, Zhang P, Wang H. Microbiome dysbiosis in lung cancer: from composition to therapy. NPJ Precis Oncol 2020; 4:33. [PMID: 33303906 PMCID: PMC7730185 DOI: 10.1038/s41698-020-00138-z] [Citation(s) in RCA: 74] [Impact Index Per Article: 14.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2020] [Accepted: 10/02/2020] [Indexed: 02/07/2023] Open
Abstract
The correlations between microbiota dysbiosis and cancer have gained extensive attention and been widely explored. As a leading cancer diagnosis worldwide, lung cancer poses a great threat to human health. The healthy human lungs are consistently exposed to external environment and harbor a specific pattern of microbiota, sharing many key pathological and physiological characteristics with the intestinal tract. Although previous findings uncovered the critical roles of microbiota in tumorigenesis and response to anticancer therapy, most of them were focused on the intestinal microbiota rather than lung microbiota. Notably, the considerable functions of microbiota in maintaining lung homeostasis should not be neglected as the microbiome dysbiosis may promote tumor development and progression through production of cytokines and toxins and multiple other pathways. Despite the fact that increasing studies have revealed the effect of microbiome on the induction of lung cancer and different disease status, the underlying mechanisms and potential therapeutic strategies remained unclear. Herein, we summarized the recent progresses about microbiome in lung cancer and further discussed the role of microbial communities in promoting lung cancer progression and the current status of therapeutic approaches targeting microbiome to alleviate and even cure lung cancer.
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Affiliation(s)
- Ning-Ning Liu
- State Key Laboratory of Oncogenes and Related Genes, Center for Single-Cell Omics, School of Public Health, Shanghai Jiao Tong University School of Medicine, 200025, Shanghai, China
| | - Qiang Ma
- Department of Thoracic Surgery, Shanghai Pulmonary Hospital Tongji University, Shanghai, China
| | - Yang Ge
- State Key Laboratory of Oncogenes and Related Genes, Center for Single-Cell Omics, School of Public Health, Shanghai Jiao Tong University School of Medicine, 200025, Shanghai, China
| | - Cheng-Xiang Yi
- Department of Thoracic Surgery, Shanghai Pulmonary Hospital Tongji University, Shanghai, China
| | - Lu-Qi Wei
- State Key Laboratory of Oncogenes and Related Genes, Center for Single-Cell Omics, School of Public Health, Shanghai Jiao Tong University School of Medicine, 200025, Shanghai, China
| | - Jing-Cong Tan
- State Key Laboratory of Oncogenes and Related Genes, Center for Single-Cell Omics, School of Public Health, Shanghai Jiao Tong University School of Medicine, 200025, Shanghai, China
| | - Qiao Chu
- State Key Laboratory of Oncogenes and Related Genes, Center for Single-Cell Omics, School of Public Health, Shanghai Jiao Tong University School of Medicine, 200025, Shanghai, China
| | - Jing-Quan Li
- State Key Laboratory of Oncogenes and Related Genes, Center for Single-Cell Omics, School of Public Health, Shanghai Jiao Tong University School of Medicine, 200025, Shanghai, China
| | - Peng Zhang
- Department of Thoracic Surgery, Shanghai Pulmonary Hospital Tongji University, Shanghai, China.
| | - Hui Wang
- State Key Laboratory of Oncogenes and Related Genes, Center for Single-Cell Omics, School of Public Health, Shanghai Jiao Tong University School of Medicine, 200025, Shanghai, China.
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Juul NH, Stockman CA, Desai TJ. Niche Cells and Signals that Regulate Lung Alveolar Stem Cells In Vivo. Cold Spring Harb Perspect Biol 2020; 12:a035717. [PMID: 32179507 PMCID: PMC7706567 DOI: 10.1101/cshperspect.a035717] [Citation(s) in RCA: 21] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023]
Abstract
The distal lung is a honeycomb-like collection of delicate gas exchange sacs called alveoli lined by two interspersed epithelial cell types: the cuboidal, surfactant-producing alveolar type II (AT2) and the flat, gas-exchanging alveolar type I (AT1) cell. During aging, a subset of AT2 cells expressing the canonical Wnt target gene, Axin2, function as stem cells, renewing themselves while generating new AT1 and AT2 cells. Wnt activity endows AT2 cells with proliferative competency, enabling them to respond to activating cues, and simultaneously blocks AT2 to AT1 cell transdifferentiation. Acute alveolar injury rapidly expands the AT2 stem cell pool by transiently inducing Wnt signaling activity in "bulk" AT2 cells, facilitating rapid epithelial repair. AT2 cell "stemness" is thus tightly regulated by access to Wnts, supplied by a specialized single-cell fibroblast niche during maintenance and by AT2 cells themselves during injury repair. Two non-AT2 "reserve" cell populations residing in the distal airways also contribute to alveolar repair, but only after widespread epithelial injury, when they rapidly proliferate, migrate, and differentiate into airway and alveolar lineages. Here, we review alveolar renewal and repair with a focus on the niches, rather than the stem cells, highlighting what is known about the cellular and molecular mechanisms by which they control stem cell activity in vivo.
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Affiliation(s)
- Nicholas H Juul
- Department of Medicine, Division of Pulmonary, Allergy & Critical Care
- Institute for Stem Cell Biology & Regenerative Medicine, Stanford University School of Medicine, Stanford, California 94305, USA
| | - Courtney A Stockman
- Institute for Stem Cell Biology & Regenerative Medicine, Stanford University School of Medicine, Stanford, California 94305, USA
| | - Tushar J Desai
- Department of Medicine, Division of Pulmonary, Allergy & Critical Care
- Institute for Stem Cell Biology & Regenerative Medicine, Stanford University School of Medicine, Stanford, California 94305, USA
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Salahudeen AA, Choi SS, Rustagi A, Zhu J, van Unen V, de la O SM, Flynn RA, Margalef-Català M, Santos AJM, Ju J, Batish A, Usui T, Zheng GXY, Edwards CE, Wagar LE, Luca V, Anchang B, Nagendran M, Nguyen K, Hart DJ, Terry JM, Belgrader P, Ziraldo SB, Mikkelsen TS, Harbury PB, Glenn JS, Garcia KC, Davis MM, Baric RS, Sabatti C, Amieva MR, Blish CA, Desai TJ, Kuo CJ. Progenitor identification and SARS-CoV-2 infection in human distal lung organoids. Nature 2020; 588:670-675. [PMID: 33238290 PMCID: PMC8003326 DOI: 10.1038/s41586-020-3014-1] [Citation(s) in RCA: 253] [Impact Index Per Article: 50.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2017] [Accepted: 11/18/2020] [Indexed: 12/17/2022]
Abstract
The distal lung contains terminal bronchioles and alveoli that facilitate gas exchange. Three-dimensional in vitro human distal lung culture systems would strongly facilitate the investigation of pathologies such as interstitial lung disease, cancer and coronavirus disease 2019 (COVID-19) pneumonia caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here we describe the development of a long-term feeder-free, chemically defined culture system for distal lung progenitors as organoids derived from single adult human alveolar epithelial type II (AT2) or KRT5+ basal cells. AT2 organoids were able to differentiate into AT1 cells, and basal cell organoids developed lumens lined with differentiated club and ciliated cells. Single-cell analysis of KRT5+ cells in basal organoids revealed a distinct population of ITGA6+ITGB4+ mitotic cells, whose offspring further segregated into a TNFRSF12Ahi subfraction that comprised about ten per cent of KRT5+ basal cells. This subpopulation formed clusters within terminal bronchioles and exhibited enriched clonogenic organoid growth activity. We created distal lung organoids with apical-out polarity to present ACE2 on the exposed external surface, facilitating infection of AT2 and basal cultures with SARS-CoV-2 and identifying club cells as a target population. This long-term, feeder-free culture of human distal lung organoids, coupled with single-cell analysis, identifies functional heterogeneity among basal cells and establishes a facile in vitro organoid model of human distal lung infections, including COVID-19-associated pneumonia.
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Affiliation(s)
- Ameen A Salahudeen
- Division of Hematology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
- Division of Hematology and Oncology, Department of Medicine, University of Illinois at Chicago College of Medicine, Chicago, IL, USA
| | - Shannon S Choi
- Division of Hematology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Arjun Rustagi
- Division of Infectious Disease and Geographic Medicine, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Junjie Zhu
- Stanford University School of Engineering, Department of Electrical Engineering, Stanford, CA, USA
| | - Vincent van Unen
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA
- Stanford Institute of Immunity, Transplantation and Infection, Stanford University School of Medicine, Stanford, CA, USA
| | - Sean M de la O
- Division of Hematology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Ryan A Flynn
- Stanford ChEM-H, Stanford University, Stanford, CA, USA
- Department of Chemistry, Stanford University, Stanford, CA, USA
| | - Mar Margalef-Català
- Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, USA
| | - António J M Santos
- Division of Hematology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Jihang Ju
- Division of Hematology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Arpit Batish
- Division of Hematology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Tatsuya Usui
- Division of Hematology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | | | - Caitlin E Edwards
- Department of Epidemiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
| | - Lisa E Wagar
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA
- Stanford Institute of Immunity, Transplantation and Infection, Stanford University School of Medicine, Stanford, CA, USA
| | - Vincent Luca
- Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA, USA
| | - Benedict Anchang
- Division of Biomedical Data Science, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Monica Nagendran
- Division of Pulmonary, Allergy and Critical Care, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Khanh Nguyen
- Division of Gastroenterology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Daniel J Hart
- Division of Hematology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | | | | | | | | | - Pehr B Harbury
- Department of Biochemistry, Stanford University School of Medicine, Stanford, CA, USA
| | - Jeffrey S Glenn
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA
- Division of Gastroenterology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - K Christopher Garcia
- Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA, USA
- Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA, USA
| | - Mark M Davis
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA
- Stanford Institute of Immunity, Transplantation and Infection, Stanford University School of Medicine, Stanford, CA, USA
- Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA, USA
| | - Ralph S Baric
- Department of Epidemiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
- Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
| | - Chiara Sabatti
- Division of Biomedical Data Science, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
| | - Manuel R Amieva
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA
- Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, USA
| | - Catherine A Blish
- Division of Infectious Disease and Geographic Medicine, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA.
- Chan Zuckerberg Biohub, San Francisco, CA, USA.
| | - Tushar J Desai
- Division of Pulmonary, Allergy and Critical Care, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA.
| | - Calvin J Kuo
- Division of Hematology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA.
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Salahudeen AA, Choi SS, Rustagi A, Zhu J, de la O SM, Flynn RA, Margalef-Català M, Santos AJM, Ju J, Batish A, van Unen V, Usui T, Zheng GXY, Edwards CE, Wagar LE, Luca V, Anchang B, Nagendran M, Nguyen K, Hart DJ, Terry JM, Belgrader P, Ziraldo SB, Mikkelsen TS, Harbury PB, Glenn JS, Garcia KC, Davis MM, Baric RS, Sabatti C, Amieva MR, Blish CA, Desai TJ, Kuo CJ. Progenitor identification and SARS-CoV-2 infection in long-term human distal lung organoid cultures. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2020:2020.07.27.212076. [PMID: 32743583 PMCID: PMC7386503 DOI: 10.1101/2020.07.27.212076] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/26/2022]
Abstract
The distal lung contains terminal bronchioles and alveoli that facilitate gas exchange and is affected by disorders including interstitial lung disease, cancer, and SARS-CoV-2-associated COVID-19 pneumonia. Investigations of these localized pathologies have been hindered by a lack of 3D in vitro human distal lung culture systems. Further, human distal lung stem cell identification has been impaired by quiescence, anatomic divergence from mouse and lack of lineage tracing and clonogenic culture. Here, we developed robust feeder-free, chemically-defined culture of distal human lung progenitors as organoids derived clonally from single adult human alveolar epithelial type II (AT2) or KRT5 + basal cells. AT2 organoids exhibited AT1 transdifferentiation potential, while basal cell organoids progressively developed lumens lined by differentiated club and ciliated cells. Organoids consisting solely of club cells were not observed. Upon single cell RNA-sequencing (scRNA-seq), alveolar organoids were composed of proliferative AT2 cells; however, basal organoid KRT5 + cells contained a distinct ITGA6 + ITGB4 + mitotic population whose proliferation segregated to a TNFRSF12A hi subfraction. Clonogenic organoid growth was markedly enriched within the TNFRSF12A hi subset of FACS-purified ITGA6 + ITGB4 + basal cells from human lung or derivative organoids. In vivo, TNFRSF12A + cells comprised ~10% of KRT5 + basal cells and resided in clusters within terminal bronchioles. To model COVID-19 distal lung disease, we everted the polarity of basal and alveolar organoids to rapidly relocate differentiated club and ciliated cells from the organoid lumen to the exterior surface, thus displaying the SARS-CoV-2 receptor ACE2 on the outwardly-facing apical aspect. Accordingly, basal and AT2 apical-out organoids were infected by SARS-CoV-2, identifying club cells as a novel target population. This long-term, feeder-free organoid culture of human distal lung alveolar and basal stem cells, coupled with single cell analysis, identifies unsuspected basal cell functional heterogeneity and exemplifies progenitor identification within a slowly proliferating human tissue. Further, our studies establish a facile in vitro organoid model for human distal lung infectious diseases including COVID-19-associated pneumonia.
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Miller PG, Chen CY, Wang YI, Gao E, Shuler ML. Multiorgan microfluidic platform with breathable lung chamber for inhalation or intravenous drug screening and development. Biotechnol Bioeng 2019; 117:486-497. [PMID: 31608985 DOI: 10.1002/bit.27188] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/22/2019] [Revised: 10/02/2019] [Accepted: 10/06/2019] [Indexed: 12/13/2022]
Abstract
Efficient and economical delivery of pharmaceuticals to patients is critical for effective therapy. Here we describe a multiorgan (lung, liver, and breast cancer) microphysiological system ("Body-on-a-Chip") designed to mimic both inhalation therapy and/or intravenous therapy using curcumin as a model drug. This system is "pumpless" and self-contained using a rocker platform for fluid (blood surrogate) bidirectional recirculation. Our lung chamber is constructed to maintain an air-liquid interface and contained a "breathable" component that was designed to mimic breathing by simulating gas exchange, contraction and expansion of the "lung" using a reciprocating pump. Three cell lines were used: A549 for the lung, HepG2 C3A for the liver, and MDA MB231 for breast cancer. All cell lines were maintained with high viability (>85%) in the device for at least 48 hr. Curcumin is used to treat breast cancer and this allowed us to compare inhalation delivery versus intravenous delivery of the drug in terms of effectiveness and potentially toxicity. Inhalation therapy could be potentially applied at home by the patient while intravenous therapy would need to be applied in a clinical setting. Inhalation therapy would be more economical and allow more frequent dosing with a potentially lower level of drug. For 24 hr exposure to 2.5 and 25 µM curcumin in the flow device the effect on lung and liver viability was small to insignificant, while there was a significant decrease in viability of the breast cancer (to 69% at 2.5 µM and 51% at 25 µM). Intravenous delivery also selectively decreased breast cancer viability (to 88% at 2.5 µM and 79% at 25 µM) but was less effective than inhalation therapy. The response in the static device controls was significantly reduced from that with recirculation demonstrating the effect of flow. These results demonstrate for the first time the feasibility of constructing a multiorgan microphysiological system with recirculating flow that incorporates a "breathable" lung module that maintains an air-liquid interface.
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Affiliation(s)
- Paula G Miller
- Department of Biomedical Engineering, Nancy E. and Peter C. Meinig School of Biomedical Engineering, Cornell University, Ithaca, New York.,Department of Chemical and Biomolecular Engineering, Robert Frederick Smith School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York
| | | | - Ying I Wang
- Department of Biomedical Engineering, Nancy E. and Peter C. Meinig School of Biomedical Engineering, Cornell University, Ithaca, New York
| | | | - Michael L Shuler
- Department of Biomedical Engineering, Nancy E. and Peter C. Meinig School of Biomedical Engineering, Cornell University, Ithaca, New York.,Department of Chemical and Biomolecular Engineering, Robert Frederick Smith School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York
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Abstract
As the prevalence and impact of lung diseases continue to increase worldwide, new therapeutic strategies are desperately needed. Advances in lung-regenerative medicine, a broad field encompassing stem cells, cell-based therapies, and a range of bioengineering approaches, offer new insights into and new techniques for studying lung physiology and pathophysiology. This provides a platform for the development of novel therapeutic approaches. Applicability to chronic obstructive pulmonary disease of recent advances and applications in cell-based therapies, predominantly those with mesenchymal stromal cell-based approaches, and bioengineering approaches for lung diseases are reviewed.
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Furuya K, Zheng YW, Sako D, Iwasaki K, Zheng DX, Ge JY, Liu LP, Furuta T, Akimoto K, Yagi H, Hamada H, Isoda H, Oda T, Ohkohchi N. Enhanced hepatic differentiation in the subpopulation of human amniotic stem cells under 3D multicellular microenvironment. World J Stem Cells 2019; 11:705-721. [PMID: 31616545 PMCID: PMC6789189 DOI: 10.4252/wjsc.v11.i9.705] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/26/2019] [Revised: 08/06/2019] [Accepted: 08/27/2019] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND To solve the problem of liver transplantation donor insufficiency, an alternative cell transplantation therapy was investigated. We focused on amniotic epithelial cells (AECs) as a cell source because, unlike induced pluripotent stem cells, they are cost-effective and non-tumorigenic. The utilization of AECs in regenerative medicine, however, is in its infancy. A general profile for AECs has not been comprehensively analyzed. Moreover, no hepatic differentiation protocol for AECs has yet been established. To this end, we independently compiled human AEC libraries, purified amniotic stem cells (ASCs), and co-cultured them with mesenchymal stem cells (MSCs) and human umbilical vein endothelial cell (HUVECs) in a 3D system which induces functional hepatic organoids. AIM To characterize AECs and generate functional hepatic organoids from ASCs and other somatic stem cells. METHODS AECs, MSCs, and HUVECs were isolated from the placentae and umbilical cords of cesarean section patients. Amnion and primary AEC stemness characteristics and heterogeneity were analyzed by immunocytochemistry, Alkaline phosphatase (AP) staining, and flow cytometry. An adherent AEC subpopulation was selected and evaluated for ASC purification quality by a colony formation assay. AEC transcriptomes were compared with those for other hepatocytes cell sources by bioinformatics. The 2D and 3D culture were compared by relative gene expression using several differentiation protocols. ASCs, MSCs, and HUVECs were combined in a 3D co-culture system to generate hepatic organoids whose structure was compared with a 3D AEC sphere and whose function was elucidated by immunofluorescence imaging, periodic acid Schiff, and an indocyanine green (ICG) test. RESULTS AECs have certain stemness markers such as EPCAM, SSEA4, and E-cadherin. One AEC subpopulation was also either positive for AP staining or expressed the TRA-1-60 and TRA-1-81 stemness markers. Moreover, it could form colonies and its frequency was enhanced ten-fold in the adherent subpopulation after selective primary passage. Bioinformatics analysis of ribose nucleic acid sequencing revealed that the total AEC gene expression was distant from those of pluripotent stem cells and hepatocytes but some gene expression overlapped among these cells. TJP1, associated with epidermal growth factor receptor, and MET, associated with hepatocyte growth factor receptor, were upregulated and may be important for hepatic differentiation. In conventional flat culture, the cells turned unviable and did not readily differentiate into hepatocytes. In 3D culture, however, hepatic gene expression of the AEC sphere was elevated even under a two-step differentiation protocol. Furthermore, the organoids derived from the MSC and HUVEC co-culture showed 3D structure with polarity, hepatic-like glycogen storage, and ICG absorption/elimination. CONCLUSION Human amniotic epithelial cells are heterogeneous and certain subpopulations have high stemness. Under a 3D co-culture system, functional hepatic organoids were generated in a multicellular microenvironment.
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Affiliation(s)
- Kinji Furuya
- Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
| | - Yun-Wen Zheng
- Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
- Institute of Regenerative Medicine and Affiliated Hospital, Jiangsu University, Zhenjiang 212001, Jiangsu Province, China
- Department of Regenerative Medicine, School of Medicine, Yokohama City University, Yokohama 236-0004, Japan.
| | - Daisuke Sako
- Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
- Department of Medicinal and Life Sciences, Faculty of Pharmaceutical Sciences, Tokyo University of Science, Noda 278-8510, Japan
| | - Kenichi Iwasaki
- Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
| | - Dong-Xu Zheng
- Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
| | - Jian-Yun Ge
- Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
| | - Li-Ping Liu
- Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
- Institute of Regenerative Medicine and Affiliated Hospital, Jiangsu University, Zhenjiang 212001, Jiangsu Province, China
| | - Tomoaki Furuta
- Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
| | - Kazunori Akimoto
- Department of Medicinal and Life Sciences, Faculty of Pharmaceutical Sciences, Tokyo University of Science, Noda 278-8510, Japan
| | - Hiroya Yagi
- Department of Obstetrics and Gynecology, Faculty of Medicine, University of Tsukuba, Tsukuba 305-8575, Japan
| | - Hiromi Hamada
- Department of Obstetrics and Gynecology, Faculty of Medicine, University of Tsukuba, Tsukuba 305-8575, Japan
| | - Hiroko Isoda
- Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba 305-8572, Japan
| | - Tatsuya Oda
- Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
| | - Nobuhiro Ohkohchi
- Department of Gastrointestinal and Hepato-Biliary-Pancreatic Surgery, Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
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Hiemstra PS, Tetley TD, Janes SM. Airway and alveolar epithelial cells in culture. Eur Respir J 2019; 54:13993003.00742-2019. [DOI: 10.1183/13993003.00742-2019] [Citation(s) in RCA: 37] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2019] [Accepted: 08/13/2019] [Indexed: 12/29/2022]
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Kim M, Mun H, Sung CO, Cho EJ, Jeon HJ, Chun SM, Jung DJ, Shin TH, Jeong GS, Kim DK, Choi EK, Jeong SY, Taylor AM, Jain S, Meyerson M, Jang SJ. Patient-derived lung cancer organoids as in vitro cancer models for therapeutic screening. Nat Commun 2019; 10:3991. [PMID: 31488816 PMCID: PMC6728380 DOI: 10.1038/s41467-019-11867-6] [Citation(s) in RCA: 487] [Impact Index Per Article: 81.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2018] [Accepted: 08/06/2019] [Indexed: 12/16/2022] Open
Abstract
Lung cancer shows substantial genetic and phenotypic heterogeneity across individuals, driving a need for personalised medicine. Here, we report lung cancer organoids and normal bronchial organoids established from patient tissues comprising five histological subtypes of lung cancer and non-neoplastic bronchial mucosa as in vitro models representing individual patient. The lung cancer organoids recapitulate the tissue architecture of the primary lung tumours and maintain the genomic alterations of the original tumours during long-term expansion in vitro. The normal bronchial organoids maintain cellular components of normal bronchial mucosa. Lung cancer organoids respond to drugs based on their genomic alterations: a BRCA2-mutant organoid to olaparib, an EGFR-mutant organoid to erlotinib, and an EGFR-mutant/MET-amplified organoid to crizotinib. Considering the short length of time from organoid establishment to drug testing, our newly developed model may prove useful for predicting patient-specific drug responses through in vitro patient-specific drug trials.
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Affiliation(s)
- Minsuh Kim
- Asan Center for Cancer Genome Discovery, Asan Institute for Life Sciences, Seoul, South Korea
| | - Hyemin Mun
- Asan Center for Cancer Genome Discovery, Asan Institute for Life Sciences, Seoul, South Korea
| | - Chang Oak Sung
- Asan Center for Cancer Genome Discovery, Asan Institute for Life Sciences, Seoul, South Korea
- Department of Pathology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, South Korea
| | - Eun Jeong Cho
- Asan Center for Cancer Genome Discovery, Asan Institute for Life Sciences, Seoul, South Korea
| | - Hye-Joon Jeon
- Asan Center for Cancer Genome Discovery, Asan Institute for Life Sciences, Seoul, South Korea
| | - Sung-Min Chun
- Asan Center for Cancer Genome Discovery, Asan Institute for Life Sciences, Seoul, South Korea
- Department of Pathology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, South Korea
| | - Da Jung Jung
- Biomedical Engineering Research Center, Asan Institute for Life Sciences, Seoul, South Korea
| | - Tae Hoon Shin
- Biomedical Engineering Research Center, Asan Institute for Life Sciences, Seoul, South Korea
| | - Gi Seok Jeong
- Biomedical Engineering Research Center, Asan Institute for Life Sciences, Seoul, South Korea
| | - Dong Kwan Kim
- Department of Thoracic Surgery, Asan Medical Center, University of Ulsan College of Medicine, Seoul, South Korea
| | - Eun Kyung Choi
- Department of Radiation Oncology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, South Korea
| | - Seong-Yun Jeong
- Department of Radiation Oncology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, South Korea
| | - Alison M Taylor
- Department of Medical Oncology and Center for Cancer-Genome Discovery, Dana-Farber Cancer Institute and Department of Pathology, Harvard Medical School, Boston, MA, USA
| | - Sejal Jain
- Department of Medical Oncology and Center for Cancer-Genome Discovery, Dana-Farber Cancer Institute and Department of Pathology, Harvard Medical School, Boston, MA, USA
| | - Matthew Meyerson
- Department of Medical Oncology and Center for Cancer-Genome Discovery, Dana-Farber Cancer Institute and Department of Pathology, Harvard Medical School, Boston, MA, USA
| | - Se Jin Jang
- Asan Center for Cancer Genome Discovery, Asan Institute for Life Sciences, Seoul, South Korea.
- Department of Pathology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, South Korea.
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Miller AJ, Dye BR, Ferrer-Torres D, Hill DR, Overeem AW, Shea LD, Spence JR. Generation of lung organoids from human pluripotent stem cells in vitro. Nat Protoc 2019; 14:518-540. [PMID: 30664680 DOI: 10.1038/s41596-018-0104-8] [Citation(s) in RCA: 273] [Impact Index Per Article: 45.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
The lung epithelium is derived from the endodermal germ layer, which undergoes a complex series of endoderm-mesoderm-mediated signaling events to generate the final arborized network of conducting airways (bronchi, bronchioles) and gas-exchanging units (alveoli). These stages include endoderm induction, anterior-posterior and dorsal-ventral patterning, lung specification, lung budding, branching morphogenesis, and, finally, maturation. Here we describe a protocol that recapitulates several of these milestones in order to differentiate human pluripotent stem cells (hPSCs) into ventral-anterior foregut spheroids and further into two distinct types of organoids: human lung organoids and bud tip progenitor organoids. The resulting human lung organoids possess cell types and structures that resemble the bronchi/bronchioles of the developing human airway surrounded by lung mesenchyme and cells expressing alveolar-cell markers. The bud tip progenitor organoids possess a population of highly proliferative multipotent cells with in vitro multilineage differentiation potential and in vivo engraftment potential. Human lung organoids can be generated from hPSCs in 50-85 d, and bud tip progenitor organoids can be generated in 22 d. The two hPSC-derived models presented here have been benchmarked with human fetal tissue and found to be representative of human fetal-like tissue. The bud tip progenitor organoids are thus ideal for exploring epithelial fate decisions, while the human lung organoids can be used to model epithelial-mesenchymal cross-talk during human lung development. In addition to their applications in developmental biology, human lung organoids and bud tip progenitor organoids may be implemented in regenerative medicine, tissue engineering, and pharmaceutical safety and efficacy testing.
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Affiliation(s)
- Alyssa J Miller
- Program in Cell and Molecular Biology, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Briana R Dye
- Department of Biomedical Engineering, University of Michigan College of Engineering, Ann Arbor, MI, USA
| | - Daysha Ferrer-Torres
- Division of Gastroenterology, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA
| | - David R Hill
- Division of Gastroenterology, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA
| | - Arend W Overeem
- Department of Cell Biology, University of Groningen, University Medical Center Groningen, Groningen, the Netherlands
| | - Lonnie D Shea
- Department of Biomedical Engineering, University of Michigan College of Engineering, Ann Arbor, MI, USA
| | - Jason R Spence
- Program in Cell and Molecular Biology, University of Michigan Medical School, Ann Arbor, MI, USA. .,Department of Biomedical Engineering, University of Michigan College of Engineering, Ann Arbor, MI, USA. .,Division of Gastroenterology, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, USA. .,Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI, USA. .,Center for Organogenesis, University of Michigan Medical School, Ann Arbor, MI, USA.
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42
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Hosseini ZF, Nelson DA, Moskwa N, Larsen M. Generating Embryonic Salivary Gland Organoids. ACTA ACUST UNITED AC 2018; 83:e76. [PMID: 30394683 DOI: 10.1002/cpcb.76] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022]
Abstract
Organoids are important research tools for studying organ morphogenesis and differentiation because they recapitulate ex vivo the native 3D organization of cells that is essential for proper cell and organ function. The composition of organoids can be manipulated to incorporate specific cell types to facilitate molecular interrogation of cell-cell interactions during organoid formation. A method for generating organoids derived from both embryonic salivary gland epithelial progenitor cells and mesenchymal support cells is described. Methods for isolating enriched populations of the epithelial cells as clusters and the mesenchyme cells as single cells from mouse embryonic submandibular salivary glands are also provided. Separating the epithelial and mesenchymal cell populations allows for independent molecular manipulation of each cell type. In addition, methods for lentiviral transduction of the mesenchyme cells and quantitative image analysis of organoids are provided. The methods described here are useful for exploring mechanisms driving organ formation. © 2018 by John Wiley & Sons, Inc.
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Affiliation(s)
- Zeinab F Hosseini
- Department of Biological Sciences, University at Albany, State University of New York, Albany, New York.,Graduate Program in Molecular, Cellular, Developmental and Neural Biology, University at Albany, State University of New York, Albany, New York
| | - Deirdre A Nelson
- Department of Biological Sciences, University at Albany, State University of New York, Albany, New York
| | - Nicholas Moskwa
- Department of Biological Sciences, University at Albany, State University of New York, Albany, New York.,Graduate Program in Molecular, Cellular, Developmental and Neural Biology, University at Albany, State University of New York, Albany, New York
| | - Melinda Larsen
- Department of Biological Sciences, University at Albany, State University of New York, Albany, New York
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43
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Abstract
Bronchopulmonary dysplasia (BPD) continues to be one of the most common complications of preterm birth and is characterized histopathologically by impaired lung alveolarization. Extremely preterm born infants remain at high risk for the development of BPD, highlighting a pressing need for continued efforts to understand the pathomechanisms at play in affected infants. This brief review summarizes recent progress in our understanding of the how the development of the newborn lung is stunted, highlighting recent reports on roles for growth factor signaling, oxidative stress, inflammation, the extracellular matrix and proteolysis, non-coding RNA, and fibroblast and epithelial cell plasticity. Additionally, some concerns about modeling BPD in experimental animals are reviewed, as are new developments in the in vitro modeling of pathophysiological processes relevant to impaired lung alveolarization in BPD.
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Affiliation(s)
- Rory E Morty
- Department of Lung Development and Remodelling, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany; Department of Internal Medicine (Pulmonology), University of Giessen and Marburg Lung Center (UGMLC), Member of the German Center for Lung Research (DZL), Giessen, Germany.
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Modeling Host-Pathogen Interactions in the Context of the Microenvironment: Three-Dimensional Cell Culture Comes of Age. Infect Immun 2018; 86:IAI.00282-18. [PMID: 30181350 DOI: 10.1128/iai.00282-18] [Citation(s) in RCA: 104] [Impact Index Per Article: 14.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
Tissues and organs provide the structural and biochemical landscapes upon which microbial pathogens and commensals function to regulate health and disease. While flat two-dimensional (2-D) monolayers composed of a single cell type have provided important insight into understanding host-pathogen interactions and infectious disease mechanisms, these reductionist models lack many essential features present in the native host microenvironment that are known to regulate infection, including three-dimensional (3-D) architecture, multicellular complexity, commensal microbiota, gas exchange and nutrient gradients, and physiologically relevant biomechanical forces (e.g., fluid shear, stretch, compression). A major challenge in tissue engineering for infectious disease research is recreating this dynamic 3-D microenvironment (biological, chemical, and physical/mechanical) to more accurately model the initiation and progression of host-pathogen interactions in the laboratory. Here we review selected 3-D models of human intestinal mucosa, which represent a major portal of entry for infectious pathogens and an important niche for commensal microbiota. We highlight seminal studies that have used these models to interrogate host-pathogen interactions and infectious disease mechanisms, and we present this literature in the appropriate historical context. Models discussed include 3-D organotypic cultures engineered in the rotating wall vessel (RWV) bioreactor, extracellular matrix (ECM)-embedded/organoid models, and organ-on-a-chip (OAC) models. Collectively, these technologies provide a more physiologically relevant and predictive framework for investigating infectious disease mechanisms and antimicrobial therapies at the intersection of the host, microbe, and their local microenvironments.
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Ferguson KT, McQuattie-Pimentel AC, Malsin ES, Sporn PHS. Dynamics of Influenza-induced Lung-Resident Memory T Cells, Anatomically and Functionally Distinct Lung Mesenchymal Populations, and Dampening of Acute Lung Injury by Neutrophil Transfer of Micro-RNA-223 to Lung Epithelial Cells. Am J Respir Cell Mol Biol 2018; 59:397-399. [PMID: 29641210 PMCID: PMC6189642 DOI: 10.1165/rcmb.2018-0047ro] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2018] [Accepted: 04/09/2018] [Indexed: 12/15/2022] Open
Affiliation(s)
- Keith T. Ferguson
- Division of Pulmonary and Critical Care Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois; and
| | - Alexandra C. McQuattie-Pimentel
- Division of Pulmonary and Critical Care Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois; and
| | - Elizabeth S. Malsin
- Division of Pulmonary and Critical Care Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois; and
| | - Peter H. S. Sporn
- Division of Pulmonary and Critical Care Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois; and
- Medical and Research Services, Jesse Brown Veterans Affairs Medical Center, Chicago, Illinois
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Nerger BA, Nelson CM. 3D culture models for studying branching morphogenesis in the mammary gland and mammalian lung. Biomaterials 2018; 198:135-145. [PMID: 30174198 DOI: 10.1016/j.biomaterials.2018.08.043] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2018] [Revised: 06/20/2018] [Accepted: 08/20/2018] [Indexed: 12/13/2022]
Abstract
The intricate architecture of branched tissues and organs has fascinated scientists and engineers for centuries. Yet-despite their ubiquity-the biophysical and biochemical mechanisms by which tissues and organs undergo branching morphogenesis remain unclear. With the advent of three-dimensional (3D) culture models, an increasingly powerful and diverse set of tools are available for investigating the development and remodeling of branched tissues and organs. In this review, we discuss the application of 3D culture models for studying branching morphogenesis of the mammary gland and the mammalian lung in the context of normal development and disease. While current 3D culture models lack the cellular and molecular complexity observed in vivo, we emphasize how these models can be used to answer targeted questions about branching morphogenesis. We highlight the specific advantages and limitations of using 3D culture models to study the dynamics and mechanisms of branching in the mammary gland and mammalian lung. Finally, we discuss potential directions for future research and propose strategies for engineering the next generation of 3D culture models for studying tissue morphogenesis.
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Affiliation(s)
- Bryan A Nerger
- Department of Chemical & Biological Engineering, Princeton, NJ, 08544, USA
| | - Celeste M Nelson
- Department of Chemical & Biological Engineering, Princeton, NJ, 08544, USA; Department of Molecular Biology, Princeton University, Princeton, NJ, 08544, USA.
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47
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Cell culture keeps pace with influenza virus. THE LANCET RESPIRATORY MEDICINE 2018; 6:805-806. [PMID: 30001993 DOI: 10.1016/s2213-2600(18)30245-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/06/2018] [Accepted: 06/07/2018] [Indexed: 11/21/2022]
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48
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Bhowmick R, Derakhshan T, Liang Y, Ritchey J, Liu L, Gappa-Fahlenkamp H. A Three-Dimensional Human Tissue-Engineered Lung Model to Study Influenza A Infection. Tissue Eng Part A 2018; 24:1468-1480. [PMID: 29732955 DOI: 10.1089/ten.tea.2017.0449] [Citation(s) in RCA: 39] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Abstract
Influenza A virus (IAV) claims ∼250,000-500,000 lives annually worldwide. Currently, there are a few in vitro models available to study IAV immunopathology. Monolayer cultures of cell lines and primary lung cells (two-dimensional [2D] cell culture) is the most commonly used tool, however, this system does not have the in vivo-like structure of the lung and immune responses to IAV as it lacks the three-dimensional (3D) tissue structure. To recapitulate the lung physiology in vitro, a system that contains multiple cell types within a 3D environment that allows cell movement and interaction would provide a critical tool. In this study, as a first step in designing a 3D-Human Tissue-Engineered Lung Model (3D-HTLM), we describe the 3D culture of primary human small airway epithelial cells (HSAEpCs) and determined the immunophenotype of this system in response to IAV infections. We constructed a 3D chitosan-collagen scaffold and cultured HSAEpCs on these scaffolds at air-liquid interface (ALI). These 3D cultures were compared with 2D-cultured HSAEpCs for viability, morphology, marker protein expression, and cell differentiation. Results showed that the 3D-cultured HSAEpCs at ALI yielded maximum viable cells and morphologically resembled the in vivo lower airway epithelium. There were also significant increases in aquaporin-5 and cytokeratin-14 expression for HSAEpCs cultured in 3D compared to 2D. The 3D culture system was used to study the infection of HSAEpCs with two major IAV strains, H1N1 and H3N2. The HSAEpCs showed distinct changes in marker protein expression, both at mRNA and protein levels, and the release of proinflammatory cytokines. This study is the first step in the development of the 3D-HTLM, which will have wide applicability in studying pulmonary pathophysiology and therapeutics development.
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Affiliation(s)
- Rudra Bhowmick
- 1 School of Chemical Engineering, Oklahoma State University , Stillwater, Oklahoma
| | - Tahereh Derakhshan
- 1 School of Chemical Engineering, Oklahoma State University , Stillwater, Oklahoma
| | - Yurong Liang
- 2 Department of Physiological Sciences, Center for Veterinary Health Sciences, Oklahoma State University , Stillwater, Oklahoma
| | - Jerry Ritchey
- 3 Department of Veterinary Pathobiology, Center for Veterinary Health Sciences, Oklahoma State University , Stillwater, Oklahoma
| | - Lin Liu
- 2 Department of Physiological Sciences, Center for Veterinary Health Sciences, Oklahoma State University , Stillwater, Oklahoma
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Faber SC, McCullough SD. Through the Looking Glass: In Vitro Models for Inhalation Toxicology and Interindividual Variability in the Airway. ACTA ACUST UNITED AC 2018; 4:115-128. [PMID: 31380467 DOI: 10.1089/aivt.2018.0002] [Citation(s) in RCA: 36] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
With 7 million deaths reported annually from air pollution alone, it is evident that adverse effects of inhaled toxicant exposures remain a major public health concern in the 21st century. Assessment and characterization of the impacts of air pollutants on human health stems from epidemiological and clinical studies, which have linked both outdoor and indoor air contaminant exposure to adverse pulmonary and cardiovascular health outcomes. Studies in animal models support epidemiological findings and have been critical in identifying systemic effects of environmental chemicals on cognitive abilities, liver disease, and metabolic dysfunction following inhalation exposure. Likewise, traditional monoculture systems have aided in identifying biomarkers of susceptibility to inhaled toxicants and served as a screening platform for safety assessment of pulmonary toxicants. Despite their contributions, in vivo and classic in vitro models have not been able to accurately represent the heterogeneity of the human population and account for interindividual variability in response to inhaled toxicants and susceptibility to the adverse health effects. Development of new technologies that can investigate genetic predisposition, are cost and time efficient, and are ethically sound, will enhance elucidation of mechanisms of inhalation toxicity, and aid in the development of novel pharmaceuticals and/or safety evaluation. This review will describe the classic and novel cell-based inhalation toxicity models and how these emerging technologies can be incorporated into regulatory or nonregulatory testing to address interindividual variability and improve overall human health.
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Affiliation(s)
- Samantha C Faber
- Curriculum in Toxicology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina
| | - Shaun D McCullough
- National Health and Environmental Effects Research Laboratory, US Environmental Protection Agency, Research Triangle Park, North Carolina
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50
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Niemeyer BF, Zhao P, Tuder RM, Benam KH. Advanced Microengineered Lung Models for Translational Drug Discovery. SLAS DISCOVERY 2018; 23:777-789. [PMID: 29447055 DOI: 10.1177/2472555218760217] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Lung diseases impose a significant socioeconomic burden and are a leading cause of morbidity and mortality worldwide. Moreover, respiratory medicine, unlike several other therapeutic areas, faces a disappointingly low number of new approved therapies. This is partly due to lack of reliable in vitro or in vivo models that can reproduce organ-level complexity and pathophysiological responses of human lung. Here, we examine new opportunities in application of recently emerged organ-on-chip technology to model human lung alveolus and small airway in preclinical drug development and biomarker discovery. We also discuss challenges that need to be addressed in coming years to further enhance the physiological and clinical relevance of these microsystems, enable their increased accessibility, and support their leap into personalized medicine.
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Affiliation(s)
- Brian F Niemeyer
- 1 Division of Pulmonary Sciences and Critical Care Medicine, Department of Medicine, University of Colorado, Anschutz Medical Campus, Aurora, CO, USA
| | - Peng Zhao
- 1 Division of Pulmonary Sciences and Critical Care Medicine, Department of Medicine, University of Colorado, Anschutz Medical Campus, Aurora, CO, USA
| | - Rubin M Tuder
- 1 Division of Pulmonary Sciences and Critical Care Medicine, Department of Medicine, University of Colorado, Anschutz Medical Campus, Aurora, CO, USA
| | - Kambez H Benam
- 1 Division of Pulmonary Sciences and Critical Care Medicine, Department of Medicine, University of Colorado, Anschutz Medical Campus, Aurora, CO, USA.,2 Department of Bioengineering, University of Colorado Denver, Aurora, CO, USA
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