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Wu H, Zhong Y, Li Y, Zhou X, Zhao T, Wan D, Zhu Y, Zhang Z, Li X, Gan X. Wnt3a Enhances Mesenchymal Stem Cell Engraftment and Differentiation in a Chronic Obstructive Pulmonary Disease Rat Model. Int J Chron Obstruct Pulmon Dis 2025; 20:69-81. [PMID: 39802038 PMCID: PMC11725259 DOI: 10.2147/copd.s486262] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2024] [Accepted: 12/28/2024] [Indexed: 01/16/2025] Open
Abstract
Background Bone marrow mesenchymal stem cell (BMSC) therapy is a novel approach for treating COPD. However, the difficulty in engraftment and easy clearance of BMSCs in vivo has hindered their clinical application. Hence, exploring effective methods to improve the engraftment and differentiation rates of BMSCs in vivo is urgent. Methods We constructed BMSCs overexpressing Wnt3a by lentivirus infection and transplanted them into a COPD rat model. The damage level of COPD rat lung tissue was assessed by pathology analysis and inflammatory cytokines analysis. The engraftment of BMSC was detected by immunofluorescence staining. Statistical analysis was performed using GraphPad Prism 7. Results We found that Wnt3a significantly enhanced the engraftment rate of BMSCs in the lungs of rats and further increased their differentiation rate into type II alveolar epithelial cells. We also assessed the expression of inflammatory factors in the lung tissues of COPD rats and discovered that Wnt3a reduced the levels of the inflammatory factors IL-6 and IL-1β while increasing the level of the anti-inflammatory factor IL-10. Our study demonstrates that Wnt3a can improve the engraftment and differentiation rates of BMSCs in the host and further alleviate COPD symptoms by regulating the secretion of inflammatory factors. Conclusion Constructing BMSCs overexpressing Wnt3a could serve as a new strategy for stem cell therapy in COPD.
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Affiliation(s)
- Huala Wu
- Department of Respiratory and Critical Care Medicine, Jiangxi Provincial Key Laboratory of Respiratory Diseases, Jiangxi Institute of Respiratory Diseases, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, Jiangxi, 330006, People’s Republic of China
| | - Yulan Zhong
- Department of Respiratory and Critical Care Medicine, Chest Hospital of Jiangxi Province, Nanchang, Jiangxi, 330006, People’s Republic of China
| | - Yangjingsi Li
- Department of Respiratory and Critical Care Medicine, Jiangxi Provincial People’s Hospital, Nanchang, Jiangxi, 330006, People’s Republic of China
| | - Xiangxiang Zhou
- Department of Respiratory and Critical Care Medicine, Chest Hospital of Jiangxi Province, Nanchang, Jiangxi, 330006, People’s Republic of China
| | - Tiantian Zhao
- Department of Respiratory and Critical Care Medicine, Jiangxi Provincial Key Laboratory of Respiratory Diseases, Jiangxi Institute of Respiratory Diseases, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, Jiangxi, 330006, People’s Republic of China
| | - Daomou Wan
- Department of Respiratory and Critical Care Medicine, Jiangxi Provincial Key Laboratory of Respiratory Diseases, Jiangxi Institute of Respiratory Diseases, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, Jiangxi, 330006, People’s Republic of China
| | - Yuanzhe Zhu
- Department of Respiratory and Critical Care Medicine, Jiangxi Provincial Key Laboratory of Respiratory Diseases, Jiangxi Institute of Respiratory Diseases, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, Jiangxi, 330006, People’s Republic of China
| | - Zhiyan Zhang
- Department of Respiratory and Critical Care Medicine, Jiangxi Provincial Key Laboratory of Respiratory Diseases, Jiangxi Institute of Respiratory Diseases, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, Jiangxi, 330006, People’s Republic of China
| | - Xiaolei Li
- Department of Respiratory and Critical Care Medicine, Jiangxi Provincial Key Laboratory of Respiratory Diseases, Jiangxi Institute of Respiratory Diseases, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, Jiangxi, 330006, People’s Republic of China
- China-Japan Friendship Jiangxi Hospital, National Regional Center for Respiratory Medicine, Nanchang, Jiangxi, 330200, People’s Republic of China
| | - Xin Gan
- Department of Respiratory and Critical Care Medicine, Jiangxi Provincial Key Laboratory of Respiratory Diseases, Jiangxi Institute of Respiratory Diseases, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, Jiangxi, 330006, People’s Republic of China
- China-Japan Friendship Jiangxi Hospital, National Regional Center for Respiratory Medicine, Nanchang, Jiangxi, 330200, People’s Republic of China
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Akita T. [Curative Treatment for COPD Based on Differentiation Induction by Synthetic Retinoid Am80 and Development of Inhalation Powder]. YAKUGAKU ZASSHI 2025; 145:1-6. [PMID: 39756917 DOI: 10.1248/yakushi.24-00139] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/07/2025]
Abstract
Chronic obstructive pulmonary disease (COPD) is characterized by chronic bronchitis and emphysema, and current drug treatments is limited to symptomatic therapy. Thus, there is an urgent need for development of new treatments to repair alveolar destruction. To regenerate the destroyed alveoli, we focused on the differentiation of alveolar epithelial progenitor cells into type I or type II alveolar epithelial cells that constitute the alveoli. Our concept of alveolar regeneration therapy is based on developing a drug delivery system (DDS) and dry powder inhalation that can efficiently deliver new alveolar regeneration drugs, which were discovered using human alveolar epithelial progenitor cells, to stem cells present on the surface of the alveoli of COPD patients, thereby inducing alveolar regeneration. This review article summarizes our data on the discovery of the synthetic retinoid Am80 as a candidate drug for alveolar regeneration, the construction of a DDS that utilizes a biological mechanism that enhances its effect on alveolar regeneration, and the formulation design of a dry powder inhalation.
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Affiliation(s)
- Tomomi Akita
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science
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Duong A, Wong A, Ramendra R, Sebben D, Moshkelgosha S, MacParland S, Liu M, Juvet S, Martinu T. A Rapid Human Lung Tissue Dissociation Protocol Maximizing Cell Yield and Minimizing Cellular Stress. Am J Respir Cell Mol Biol 2024; 71:509-518. [PMID: 38959415 DOI: 10.1165/rcmb.2023-0343ma] [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: 09/21/2023] [Accepted: 07/03/2024] [Indexed: 07/05/2024] Open
Abstract
The human lung is a complex organ that comprises diverse populations of epithelial, mesenchymal, vascular, and immune cells, which gains even greater complexity during disease states. To effectively study the lung at a single-cell level, a dissociation protocol that achieves the highest yield of viable cells of interest with minimal dissociation-associated protein or transcription changes is key. Here, we detail a rapid collagenase-based dissociation protocol (Col-Short) that provides a high-yield single-cell suspension that is suitable for a variety of downstream applications. Diseased human lung explants were obtained and dissociated through the Col-Short protocol and compared with four other dissociation protocols. Resulting single-cell suspensions were then assessed with flow cytometry, differential staining, and quantitative real-time PCR to identify major hematopoietic and nonhematopoietic cell populations, as well as their activation states. We observed that the Col-Short protocol provides the greatest number of cells per gram of lung tissue, with no reduction in viability when compared with previously described dissociation protocols. Col-Short had no observable surface protein marker cleavage as well as lower expression of protein activation markers and stress-related transcripts compared with four other protocols. The Col-Short dissociation protocol can be used as a rapid strategy to generate single cells for respiratory cell biology research.
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Affiliation(s)
- Allen Duong
- Toronto Lung Transplant Program
- Toronto General Hospital Research Institute, and
- Institute of Medical Science
| | - Aaron Wong
- Toronto Lung Transplant Program
- Toronto General Hospital Research Institute, and
- Institute of Medical Science
| | - Rayoun Ramendra
- Toronto Lung Transplant Program
- Toronto General Hospital Research Institute, and
| | - David Sebben
- Toronto Lung Transplant Program
- Toronto General Hospital Research Institute, and
| | - Sajad Moshkelgosha
- Toronto Lung Transplant Program
- Toronto General Hospital Research Institute, and
| | - Sonya MacParland
- Toronto General Hospital Research Institute, and
- Ajmera Transplant Centre, University Health Network, Toronto, Ontario, Canada; and
- Institute of Medical Science
- Department of Laboratory Medicine and Pathobiology, and
| | - Mingyao Liu
- Toronto Lung Transplant Program
- Toronto General Hospital Research Institute, and
- Ajmera Transplant Centre, University Health Network, Toronto, Ontario, Canada; and
- Institute of Medical Science
| | - Stephen Juvet
- Toronto Lung Transplant Program
- Toronto General Hospital Research Institute, and
- Ajmera Transplant Centre, University Health Network, Toronto, Ontario, Canada; and
- Institute of Medical Science
- Division of Respirology, Department of Medicine, University of Toronto, Toronto, Ontario, Canada
| | - Tereza Martinu
- Toronto Lung Transplant Program
- Toronto General Hospital Research Institute, and
- Ajmera Transplant Centre, University Health Network, Toronto, Ontario, Canada; and
- Institute of Medical Science
- Division of Respirology, Department of Medicine, University of Toronto, Toronto, Ontario, Canada
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4
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Duong A, Juvet S, Martinu T. Reply to Fujino et al.: Human Lung Cell Separation Strategies for Translational Research. Am J Respir Cell Mol Biol 2024; 71:622-623. [PMID: 39226579 PMCID: PMC11568469 DOI: 10.1165/rcmb.2024-0369le] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2024] [Accepted: 08/28/2024] [Indexed: 09/05/2024] Open
Affiliation(s)
- Allen Duong
- Toronto Lung Transplant Program, Ajmera Transplant Centre,University Health NetworkToronto, Ontario, Canada
- University of TorontoToronto, Ontario, Canada
| | - Stephen Juvet
- Toronto Lung Transplant Program, Ajmera Transplant Centre,University Health NetworkToronto, Ontario, Canada
- University of TorontoToronto, Ontario, Canada
| | - Tereza Martinu
- Toronto Lung Transplant Program, Ajmera Transplant Centre,University Health NetworkToronto, Ontario, Canada
- University of TorontoToronto, Ontario, Canada
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Li D, Zhang X, Song Z, Zhao S, Huang Y, Qian W, Cai X. Advances in common in vitro cellular models of pulmonary fibrosis. Immunol Cell Biol 2024; 102:557-569. [PMID: 38714318 DOI: 10.1111/imcb.12756] [Citation(s) in RCA: 2] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2023] [Revised: 12/24/2023] [Accepted: 04/09/2024] [Indexed: 05/09/2024]
Abstract
The development of in vitro models is essential for a comprehensive understanding and investigation of pulmonary fibrosis (PF) at both cellular and molecular levels. This study presents a literature review and an analysis of various cellular models used in scientific studies, specifically focusing on their applications in elucidating the pathogenesis of PF. Our study highlights the importance of taking a comprehensive approach to studing PF, emphasizing the necessity of considering multiple cell types and organs and integrating diverse analytical perspectives. Notably, primary cells demonstrate remarkable cell growth characteristics and gene expression profiles; however, their limited availability, maintenance challenges, inability for continuous propagation and susceptibility to phenotypic changes over time significantly limit their utility in scientific investigation. By contrast, immortalized cell lines are easily accessible, cultured and continuously propagated, although they may have some phenotypic differences from primary cells. Furthermore, in vitro coculture models offer a more practical and precise method to explore complex interactions among cells, tissues and organs. Consequently, when developing models of PF, researchers should thoroughly assess the advantages, limitations and relevant mechanisms of different cell models to ensure their selection is consistent with the research objectives.
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Affiliation(s)
- Die Li
- First Clinical Medical College, Shandong University of Traditional Chinese Medicine, Jinan, Shandong, China
| | - Xinyue Zhang
- Department of Lung Disease, Affiliated Hospital of Shandong University of Traditional Chinese Medicine, Jinan, Shandong, China
| | - Ziqiong Song
- Shandong First Medical University & Shandong Academy of Medical Sciences, Shandong Academy of Occupational Health and Occupational Medicine, Jinan, Shandong, China
| | - Shan Zhao
- First Clinical Medical College, Shandong University of Traditional Chinese Medicine, Jinan, Shandong, China
| | - Yuan Huang
- First Clinical Medical College, Shandong University of Traditional Chinese Medicine, Jinan, Shandong, China
| | - Weibin Qian
- Department of Lung Disease, Affiliated Hospital of Shandong University of Traditional Chinese Medicine, Jinan, Shandong, China
| | - Xinrui Cai
- Shandong Academy of Occupational Health and Occupational Medicine, Shandong First Medical University & Shandong Academy of Medical Sciences, Jinan, Shandong, China
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Kristensen C, Larsen LE, Trebbien R, Jensen HE. The avian influenza A virus receptor SA-α2,3-Gal is expressed in the porcine nasal mucosa sustaining the pig as a mixing vessel for new influenza viruses. Virus Res 2024; 340:199304. [PMID: 38142890 PMCID: PMC10793167 DOI: 10.1016/j.virusres.2023.199304] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2023] [Revised: 12/15/2023] [Accepted: 12/17/2023] [Indexed: 12/26/2023]
Abstract
Influenza A viruses (IAVs) originate from wild birds but have on several occasions jumped host barriers and are now also circulating in humans and mammals. The IAV host receptors (glycans with galactose linked to a sialic acid (SA) in an α2,3 or α2,6 linkage) are crucial host factors restricting inter-species transmission. In general, avian-origin IAVs show a preference for SA-α2,3 (avian receptor), whereas IAVs isolated from humans and pigs prefer SA-α2,6 (human receptor). N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc) are the two major SAs. Neu5Ac is expressed in all species, whereas Neu5Gc is only expressed in a limited number of domestic species such as pigs and horses, but not in humans. Despite that previous studies have shown that the IAV host receptor distribution appears to be similar in pigs and humans, none of these studies have investigated the expression of Neu5Gc-α2,6 in situ in porcine tissues. Thus, the aim of this study was to elucidate the distribution of IAV host receptors expressed in the porcine respiratory tract and relate the expression to the viral tropism of diverse host-adapted IAVs. The IAV receptor (SA-α2,3 and SA-α2,6) distribution and the presence of specifically Neu5Gc-α2,6 in the porcine nasal, tracheal, and lung tissues was investigated by lectin histochemistry. Furthermore, IAV immunohistochemistry was performed on tissues from pigs experimentally infected with IAVs, either adapted to pigs or humans, to investigate the significance of the IAV host receptors and the tropism of the diverse host-adapted IAVs. We document for the first time the expression of the avian receptor on the surface of the porcine nasal mucosa and an equal expression of Neu5Ac-α2,6 and Neu5Gc-α2,6 on the surface of the tracheal epithelium and alveoli. In all IAV-infected pigs, we found a low amount of IAV-positive cells in the trachea despite a high expression of the human receptor. Cumulatively, these findings suggest that optimal IAV replication involves a complex interplay between the viruses and their host receptors and that there might be other less clearly defined host factors that determine the site of replication.
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Affiliation(s)
- Charlotte Kristensen
- Department of Veterinary and Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark.
| | - Lars E Larsen
- Department of Veterinary and Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark
| | - Ramona Trebbien
- Department of Virus and Microbiological Special Diagnostics, Statens Serum Institut, Copenhagen S, Denmark
| | - Henrik E Jensen
- Department of Veterinary and Animal Sciences, University of Copenhagen, Frederiksberg C, Denmark
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Li Z, Jin T, Yang R, Guo J, Niu Z, Gao H, Song X, Zhang Q, Ning Z, Ren L, Wang Y, Fan X, Liang H, Li T, He W. Long non-coding RNA PFI inhibits apoptosis of alveolar epithelial cells to alleviate lung injury via miR-328-3p/Creb1 axis. Exp Cell Res 2023:113685. [PMID: 37330182 DOI: 10.1016/j.yexcr.2023.113685] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2023] [Revised: 06/05/2023] [Accepted: 06/06/2023] [Indexed: 06/19/2023]
Abstract
Acute lung injury (ALI), a common clinical type of critical illness, is an acute hypoxic respiratory insufficiency caused by the damage of alveolar epithelial cells and capillary endothelial cells. In a previous study, we reported a novel lncRNA, lncRNA PFI, which could protect against pulmonary fibrosis in pulmonary fibroblasts. The present study demonstrated that lncRNA PFI was downregulated in alveolar epithelial cell of mice injury lung tissues, and further investigated the role of lncRNA PFI in regulating inflammation-induced alveolar epithelial cell apoptosis. Overexpression of lncRNA PFI could partially abrogated bleomycin induced type II AECs injured. Subsequently, bioinformatic prediction revealed that lncRNA PFI might directly bind to miR-328-3p, and further AGO-2 RNA binding protein immunoprecipitation (RIP) assay confirmed their binding relationship. Furthermore, miR-328-3p promoted apoptosis in MLE-12 cells by limiting the activation of the Creb1, a protein correlated with cell apoptosis, whereas AMO-328-3p ablated the pro-apoptosis effect of silencing lncRNA PFI in MLE-12 cells. While miR-328-3p could also ablate the function of lncRNA PFI in bleomycin treated human lung epithelial cells. Enhanced expression of lncRNA PFI reversed the LPS-induced lung injury in mice. Overall, these data reveal that lncRNA PFI mitigated acute lung injury through miR-328-3p/Creb1 pathway in alveolar epithelial cells.
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Affiliation(s)
- Zhixin Li
- Department of Thoracic Surgery, Shanghai Pulmonary Hospital, Tongji University, Shanghai, 200433, China
| | - Tongzhu Jin
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China
| | - Ruoxuan Yang
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China
| | - Jiayu Guo
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China
| | - Zhihui Niu
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China
| | - Huiying Gao
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China
| | - Xiaoying Song
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China
| | - Qing Zhang
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China
| | - Zhiwei Ning
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China
| | - Lingxue Ren
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China
| | - Yan Wang
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China
| | - Xingxing Fan
- State Key Laboratory of Quality Research in Chinese Medicine/Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Macau (SAR), China
| | - Haihai Liang
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Research Unit of Noninfectious Chronic Diseases in Frigid Zone (2019RU070), Chinese Academy of Medical Sciences, Harbin, Heilongjiang, 150081, PR China
| | - Tianyu Li
- Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China; Northern Translational Medicine Research and Cooperation Center, Heilongjiang Academy of Medical Sciences, Harbin Medical University, Harbin, Heilongjiang, 150081, PR China.
| | - Wenxin He
- Department of Thoracic Surgery, Shanghai Pulmonary Hospital, Tongji University, Shanghai, 200433, China.
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Akita T, Oda K, Narukawa S, Morita Y, Tange K, Nakai Y, Yamashita C. Intracellular Drug Delivery Process of Am80-Encapsulated Lipid Nanoparticles Aiming for Alveolar Regeneration. Pharmaceuticals (Basel) 2023; 16:838. [PMID: 37375785 DOI: 10.3390/ph16060838] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2023] [Revised: 05/29/2023] [Accepted: 05/31/2023] [Indexed: 06/29/2023] Open
Abstract
Chronic obstructive pulmonary disease (COPD) results in obstructive ventilatory impairment caused by emphysema, and current treatment is limited to symptomatic therapy or lung transplantation. Therefore, the development of new treatments to repair alveolar destruction is especially urgent. Our previous study revealed that 1.0 mg/kg of synthetic retinoid Am80 had a repair effect on collapsed alveoli in a mouse model of elastase-induced emphysema. From these results, however, the clinical dose calculated in accordance with FDA guidance is estimated to be 5.0 mg/60 kg, and it is desirable to further reduce the dose to allow the formulation of a powder inhaler for clinical application. To efficiently deliver Am80 to the retinoic acid receptor in the cell nucleus, which is the site of action, we focused on SS-cleavable proton-activated lipid-like material O-Phentyl-P4C2COATSOME®SS-OP, hereinafter referred to as "SS-OP"). In this study, we investigated the cellular uptake and intracellular drug delivery process of Am80-encapsulated SS-OP nanoparticles to elucidate the mechanism of Am80 by nanoparticulation. Am80-encapsulated SS-OP nanoparticles were taken up into the cells via ApoE, and then Am80 was efficiently delivered into the nucleus via RARα. These results indicated the usefulness of SS-OP nanoparticles as drug delivery system carriers of Am80 for COPD treatment.
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Affiliation(s)
- Tomomi Akita
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510, Japan
| | - Kazuaki Oda
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510, Japan
| | - Satoru Narukawa
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510, Japan
| | - Yuki Morita
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510, Japan
| | - Kota Tange
- Life Science Research Laboratory, NOF CORPORATION, 3-3 Chidori-cho, Kawasaki-ku, Kawasaki City 210-0865, Japan
| | - Yuta Nakai
- Life Science Research Laboratory, NOF CORPORATION, 3-3 Chidori-cho, Kawasaki-ku, Kawasaki City 210-0865, Japan
| | - Chikamasa Yamashita
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510, Japan
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Mižíková I, Thébaud B. Perinatal origins of bronchopulmonary dysplasia-deciphering normal and impaired lung development cell by cell. Mol Cell Pediatr 2023; 10:4. [PMID: 37072570 PMCID: PMC10113423 DOI: 10.1186/s40348-023-00158-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2022] [Accepted: 03/26/2023] [Indexed: 04/20/2023] Open
Abstract
Bronchopulmonary dysplasia (BPD) is a multifactorial disease occurring as a consequence of premature birth, as well as antenatal and postnatal injury to the developing lung. BPD morbidity and severity depend on a complex interplay between prenatal and postnatal inflammation, mechanical ventilation, and oxygen therapy as well as associated prematurity-related complications. These initial hits result in ill-explored aberrant immune and reparative response, activation of pro-fibrotic and anti-angiogenic factors, which further perpetuate the injury. Histologically, the disease presents primarily by impaired lung development and an arrest in lung microvascular maturation. Consequently, BPD leads to respiratory complications beyond the neonatal period and may result in premature aging of the lung. While the numerous prenatal and postnatal stimuli contributing to BPD pathogenesis are relatively well known, the specific cell populations driving the injury, as well as underlying mechanisms are still not well understood. Recently, an effort to gain a more detailed insight into the cellular composition of the developing lung and its progenitor populations has unfold. Here, we provide an overview of the current knowledge regarding perinatal origin of BPD and discuss underlying mechanisms, as well as novel approaches to study the perturbed lung development.
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Affiliation(s)
- I Mižíková
- Experimental Pulmonology, Department of Pediatrics and Adolescent Medicine, Faculty of Medicine and University Hospital Cologne, University of Cologne, Cologne, Germany.
| | - B Thébaud
- Sinclair Centre for Regenerative Medicine, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Department of Cellular and Molecular Medicine, University of Ottawa, Ottawa, ON, Canada
- Department of Pediatrics, Children's Hospital of Eastern Ontario (CHEO), CHEO Research Institute, University of Ottawa, Ottawa, ON, Canada
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10
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The Cultivation Modality and Barrier Maturity Modulate the Toxicity of Industrial Zinc Oxide and Titanium Dioxide Nanoparticles on Nasal, Buccal, Bronchial, and Alveolar Mucosa Cell-Derived Barrier Models. Int J Mol Sci 2023; 24:ijms24065634. [PMID: 36982705 PMCID: PMC10056597 DOI: 10.3390/ijms24065634] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2023] [Revised: 03/07/2023] [Accepted: 03/12/2023] [Indexed: 03/18/2023] Open
Abstract
As common industrial by-products, airborne engineered nanomaterials are considered important environmental toxins to monitor due to their potential health risks to humans and animals. The main uptake routes of airborne nanoparticles are nasal and/or oral inhalation, which are known to enable the transfer of nanomaterials into the bloodstream resulting in the rapid distribution throughout the human body. Consequently, mucosal barriers present in the nose, buccal, and lung have been identified and intensively studied as the key tissue barrier to nanoparticle translocation. Despite decades of research, surprisingly little is known about the differences among various mucosa tissue types to tolerate nanoparticle exposures. One limitation in comparing nanotoxicological data sets can be linked to a lack of harmonization and standardization of cell-based assays, where (a) different cultivation conditions such as an air-liquid interface or submerged cultures, (b) varying barrier maturity, and (c) diverse media substitutes have been used. The current comparative nanotoxicological study, therefore, aims at analyzing the toxic effects of nanomaterials on four human mucosa barrier models including nasal (RPMI2650), buccal (TR146), alveolar (A549), and bronchial (Calu-3) mucosal cell lines to better understand the modulating effects of tissue maturity, cultivation conditions, and tissue type using standard transwell cultivations at liquid-liquid and air-liquid interfaces. Overall, cell size, confluency, tight junction localization, and cell viability as well as barrier formation using 50% and 100% confluency was monitored using trans-epithelial-electrical resistance (TEER) measurements and resazurin-based Presto Blue assays of immature (e.g., 5 days) and mature (e.g., 22 days) cultures in the presence and absence of corticosteroids such as hydrocortisone. Results of our study show that cellular viability in response to increasing nanoparticle exposure scenarios is highly compound and cell-type specific (TR146 6 ± 0.7% at 2 mM ZnO (ZnO) vs. ~90% at 2 mM TiO2 (TiO2) for 24 h; Calu3 93.9 ± 4.21% at 2 mM ZnO vs. ~100% at 2 mM TiO2). Nanoparticle-induced cytotoxic effects under air-liquid cultivation conditions declined in RPMI2650, A549, TR146, and Calu-3 cells (~0.7 to ~0.2-fold), with increasing 50 to 100% barrier maturity under the influence of ZnO (2 mM). Cell viability in early and late mucosa barriers where hardly influenced by TiO2 as well as most cell types did not fall below 77% viability when added to Individual ALI cultures. Fully maturated bronchial mucosal cell barrier models cultivated under ALI conditions showed less tolerance to acute ZnO nanoparticle exposures (~50% remaining viability at 2 mM ZnO for 24 h) than the similarly treated but more robust nasal (~74%), buccal (~73%), and alveolar (~82%) cell-based models.
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11
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Akita T, Morita Y, Kawai T, Oda K, Tange K, Nakai Y, Yamashita C. Am80-Encapsulated Lipid Nanoparticles, Developed with the Aim of Achieving Alveolar Regeneration, Have an Improvement Effect on Pulmonary Emphysema. Pharmaceutics 2022; 15:pharmaceutics15010037. [PMID: 36678666 PMCID: PMC9860907 DOI: 10.3390/pharmaceutics15010037] [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: 11/04/2022] [Revised: 12/10/2022] [Accepted: 12/15/2022] [Indexed: 12/25/2022] Open
Abstract
Chronic obstructive pulmonary disease (COPD) is characterized by chronic bronchitis and emphysema, and current drug treatments target its symptoms. Thus, the development of a therapeutic drug to repair alveolar destruction is urgently needed. Our previous research revealed that the synthetic retinoic acid Am80 (1.0 mg/kg) showed a repairing effect on collapsed alveoli in a mouse model of elastase-induced emphysema. However, a further reduction in the dose is desirable to facilitate the development of a powder inhalation formulation for clinical application. We, therefore, focused on SS-OP to deliver Am80 efficiently. As a result, 0.01 mg/kg of Am80-encapsulated SS-OP nanoparticles repaired collapsed alveoli and improved the respiratory function in the mouse model of elastase induced emphysema. The results suggested that, with the use of SS-OP, the Am80 dose could be reduced. This could contribute to the development of a powder inhalation system as a curative medicine for COPD.
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Affiliation(s)
- Tomomi Akita
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510, Japan
| | - Yuki Morita
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510, Japan
| | - Takehiro Kawai
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510, Japan
| | - Kazuaki Oda
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510, Japan
| | - Kota Tange
- DDS Research Laboratory, NOF CORPORATION, 3-3 Chidori-cho, Kawasaki-ku, Kawasaki City 210-0865, Japan
| | - Yuta Nakai
- DDS Research Laboratory, NOF CORPORATION, 3-3 Chidori-cho, Kawasaki-ku, Kawasaki City 210-0865, Japan
| | - Chikamasa Yamashita
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510, Japan
- Correspondence: ; Tel.: +81-4-7124-1501
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12
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Petrosyan A, Montali F, Peloso A, Citro A, Byers LN, La Pointe C, Suleiman M, Marchetti A, Mcneill EP, Speer AL, Ng WH, Ren X, Bussolati B, Perin L, Di Nardo P, Cardinale V, Duisit J, Monetti AR, Savino JR, Asthana A, Orlando G. Regenerative medicine technologies applied to transplant medicine. An update. Front Bioeng Biotechnol 2022; 10:1015628. [PMID: 36263358 PMCID: PMC9576214 DOI: 10.3389/fbioe.2022.1015628] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2022] [Accepted: 09/12/2022] [Indexed: 11/13/2022] Open
Abstract
Regenerative medicine (RM) is changing how we think and practice transplant medicine. In regenerative medicine, the aim is to develop and employ methods to regenerate, restore or replace damaged/diseased tissues or organs. Regenerative medicine investigates using tools such as novel technologies or techniques, extracellular vesicles, cell-based therapies, and tissue-engineered constructs to design effective patient-specific treatments. This review illustrates current advancements in regenerative medicine that may pertain to transplant medicine. We highlight progress made and various tools designed and employed specifically for each tissue or organ, such as the kidney, heart, liver, lung, vasculature, gastrointestinal tract, and pancreas. By combing both fields of transplant and regenerative medicine, we can harbor a successful collaboration that would be beneficial and efficacious for the repair and design of de novo engineered whole organs for transplantations.
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Affiliation(s)
- Astgik Petrosyan
- GOFARR Laboratory for Organ Regenerative Research and Cell Therapeutics in Urology, Saban Research Institute, Division of Urology, Children’s Hospital Los Angeles, Los Angeles, CA, United States
| | - Filippo Montali
- Department of General Surgery, di Vaio Hospital, Fidenza, Italy
| | - Andrea Peloso
- Visceral Surgery Division, University Hospitals of Geneva, Geneva, Switzerland
| | - Antonio Citro
- San Raffaele Diabetes Research Institute, IRCCS Ospedale San Raffaele, Milan, Italy
| | - Lori N. Byers
- Wake Forest School of Medicine, Winston Salem, NC, United States
| | | | - Mara Suleiman
- Wake Forest School of Medicine, Winston Salem, NC, United States
- Department of Clinical and Experimental Medicine, University of Pisa, Pisa, Italy
| | - Alice Marchetti
- Wake Forest School of Medicine, Winston Salem, NC, United States
- Department of Pharmaceutical Sciences, Università del Piemonte Orientale, Novara, Italy
| | - Eoin P. Mcneill
- Department of Pediatric Surgery, The University of Texas Health Science Center at Houston McGovern Medical School, Houston, TX, United States
| | - Allison L Speer
- Department of Pediatric Surgery, The University of Texas Health Science Center at Houston McGovern Medical School, Houston, TX, United States
| | - Wai Hoe Ng
- Department of Biomedical Engineering, Carnegie Mellon University, Pittsburgh, PA, United States
| | - Xi Ren
- Department of Biomedical Engineering, Carnegie Mellon University, Pittsburgh, PA, United States
| | - Benedetta Bussolati
- Department of Molecular Biotechnology and Health Sciences, University of Turin, Turin, Italy
| | - Laura Perin
- GOFARR Laboratory for Organ Regenerative Research and Cell Therapeutics in Urology, Saban Research Institute, Division of Urology, Children’s Hospital Los Angeles, Los Angeles, CA, United States
| | - Paolo Di Nardo
- Centro Interdipartimentale per la Medicina Rigenerativa (CIMER), Università Degli Studi di Roma Tor Vergata, Rome, Italy
| | - Vincenzo Cardinale
- Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Rome, Italy
| | - Jerome Duisit
- Department of Plastic, Reconstructive and Aesthetic Surgery, CHU Rennes, University of Rennes I, Rennes, France
| | | | | | - Amish Asthana
- Wake Forest School of Medicine, Winston Salem, NC, United States
| | - Giuseppe Orlando
- Wake Forest School of Medicine, Winston Salem, NC, United States
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13
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Zhang L, Luo W, Liu J, Xu M, Peng Q, Zou W, You J, Shu Y, Zhao P, Wagstaff W, Zhao G, Qin K, Haydon RC, Luu HH, Reid RR, Bi Y, Zhao T, He TC, Fu Z. Modeling lung diseases using reversibly immortalized mouse pulmonary alveolar type 2 cells (imPAC2). Cell Biosci 2022; 12:159. [PMID: 36138472 PMCID: PMC9502644 DOI: 10.1186/s13578-022-00894-4] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/21/2022] [Accepted: 08/30/2022] [Indexed: 12/09/2022] Open
Abstract
BACKGROUND A healthy alveolar epithelium is critical to the gas exchange function of the lungs. As the major cell type of alveolar epithelium, alveolar type 2 (AT2) cells play a critical role in maintaining pulmonary homeostasis by serving as alveolar progenitors during lung injury, inflammation, and repair. Dysregulation of AT2 cells may lead to the development of acute and chronic lung diseases and cancer. The lack of clinically relevant AT2 cell models hampers our ability to understand pulmonary diseases. Here, we sought to establish reversibly immortalized mouse pulmonary alveolar type 2 cells (imPAC2) and investigate their potential in forming alveolar organoids to model pulmonary diseases. METHODS Primary mouse pulmonary alveolar cells (mPACs) were isolated and immortalized with a retroviral expression of SV40 Large T antigen (LTA). Cell proliferation and survival was assessed by crystal violet staining and WST-1 assays. Marker gene expression was assessed by qPCR, Western blotting, and/or immunostaining. Alveolar organoids were generated by using matrigel. Ad-TGF-β1 was used to transiently express TGF-β1. Stable silencing β-catenin or overexpression of mutant KRAS and TP53 was accomplished by using retroviral vectors. Subcutaneous cell implantations were carried out in athymic nude mice. The retrieved tissue masses were subjected to H & E histologic evaluation. RESULTS We immortalized primary mPACs with SV40 LTA to yield the imPACs that were non-tumorigenic and maintained long-term proliferative activity that was reversible by FLP-mediated removal of SV40 LTA. The EpCAM+ AT2-enriched subpopulation (i.e., imPAC2) was sorted out from the imPACs, and was shown to express AT2 markers and form alveolar organoids. Functionally, silencing β-catenin decreased the expression of AT2 markers in imPAC2 cells, while TGF-β1 induced fibrosis-like response by regulating the expression of epithelial-mesenchymal transition markers in the imPAC2 cells. Lastly, concurrent expression of oncogenic KRAS and mutant TP53 rendered the imPAC2 cells a tumor-like phenotype and activated lung cancer-associated pathways. Collectively, our results suggest that the imPAC2 cells may faithfully represent AT2 populations that can be further explored to model pulmonary diseases.
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Affiliation(s)
- Linghuan Zhang
- Stem Cell Biology and Therapy Laboratory, Ministry of Education Key Laboratory of Child Development and Disorders, and the Department of Respiratory Diseases, The Children's Hospital of Chongqing Medical University, Chongqing, 400014, China
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
| | - Wenping Luo
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
- Laboratory Animal Center, Southwest University, Chongqing, 400715, China
| | - Jiang Liu
- Stem Cell Biology and Therapy Laboratory, Ministry of Education Key Laboratory of Child Development and Disorders, and the Department of Respiratory Diseases, The Children's Hospital of Chongqing Medical University, Chongqing, 400014, China
| | - Maozhu Xu
- Stem Cell Biology and Therapy Laboratory, Ministry of Education Key Laboratory of Child Development and Disorders, and the Department of Respiratory Diseases, The Children's Hospital of Chongqing Medical University, Chongqing, 400014, China
| | - Qi Peng
- University-Town Hospital, Chongqing Medical University, Chongqing, 401331, China
| | - Wenjing Zou
- Stem Cell Biology and Therapy Laboratory, Ministry of Education Key Laboratory of Child Development and Disorders, and the Department of Respiratory Diseases, The Children's Hospital of Chongqing Medical University, Chongqing, 400014, China
| | - Jingyi You
- Stem Cell Biology and Therapy Laboratory, Ministry of Education Key Laboratory of Child Development and Disorders, and the Department of Respiratory Diseases, The Children's Hospital of Chongqing Medical University, Chongqing, 400014, China
| | - Yi Shu
- Stem Cell Biology and Therapy Laboratory, Ministry of Education Key Laboratory of Child Development and Disorders, and the Department of Respiratory Diseases, The Children's Hospital of Chongqing Medical University, Chongqing, 400014, China
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
| | - Piao Zhao
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
- Departments of Orthopaedic Surgery and Urology, The First Affiliated Hospital of Chongqing Medical University, Chongqing, 400046, China
| | - William Wagstaff
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
| | - Guozhi Zhao
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
- Departments of Orthopaedic Surgery and Urology, The First Affiliated Hospital of Chongqing Medical University, Chongqing, 400046, China
| | - Kevin Qin
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
- Rosalind Franklin University of Medicine, North Chicago, IL, 60064, USA
| | - Rex C Haydon
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
| | - Hue H Luu
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
| | - Russell R Reid
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
- Department of Surgery, The University of Chicago Medical Center, Chicago, IL, 60637, USA
- Laboratory of Craniofacial Suture Biology and Development, Department of Surgery Section of Plastic Surgery, The University of Chicago Medical Center, Chicago, IL, 60637, USA
| | - Yang Bi
- Stem Cell Biology and Therapy Laboratory, Ministry of Education Key Laboratory of Child Development and Disorders, and the Department of Respiratory Diseases, The Children's Hospital of Chongqing Medical University, Chongqing, 400014, China
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA
| | - Tianyu Zhao
- Chongqing Key Laboratory of Oral Diseases and Biomedical Sciences, the Stomatological Hospital of Chongqing Medical University, Chongqing, 401147, China
| | - Tong-Chuan He
- Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, 5841 South Maryland Avenue, MC3079, Chicago, IL, 60637, USA.
- Department of Surgery, The University of Chicago Medical Center, Chicago, IL, 60637, USA.
| | - Zhou Fu
- Stem Cell Biology and Therapy Laboratory, Ministry of Education Key Laboratory of Child Development and Disorders, and the Department of Respiratory Diseases, The Children's Hospital of Chongqing Medical University, Chongqing, 400014, China.
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Lee H, Jeong SH, Lee H, Kim C, Nam YJ, Kang JY, Song MO, Choi JY, Kim J, Park EK, Baek YW, Lee JH. Analysis of lung cancer-related genetic changes in long-term and low-dose polyhexamethylene guanidine phosphate (PHMG-p) treated human pulmonary alveolar epithelial cells. BMC Pharmacol Toxicol 2022; 23:19. [PMID: 35354498 PMCID: PMC8969249 DOI: 10.1186/s40360-022-00559-5] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2021] [Accepted: 03/21/2022] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND Lung injury elicited by respiratory exposure to humidifier disinfectants (HDs) is known as HD-associated lung injury (HDLI). Current elucidation of the molecular mechanisms related to HDLI is mostly restricted to fibrotic and inflammatory lung diseases. In our previous report, we found that lung tumors were caused by intratracheal instillation of polyhexamethylene guanidine phosphate (PHMG-p) in a rat model. However, the lung cancer-related genetic changes concomitant with the development of these lung tumors have not yet been fully defined. We aimed to discover the effect of long-term exposure of PHMG-p on normal human lung alveolar cells. METHODS We investigated whether PHMG-p could increase distorted homeostasis of oncogenes and tumor-suppressor genes, with long-term and low-dose treatment, in human pulmonary alveolar epithelial cells (HPAEpiCs). Total RNA sequencing was performed with cells continuously treated with PHMG-p and harvested after 35 days. RESULTS After PHMG-p treatment, genes with transcriptional expression changes of more than 2.0-fold or less than 0.5-fold were identified. Within 10 days of exposure, 2 protein-coding and 5 non-coding genes were selected, whereas in the group treated for 27-35 days, 24 protein-coding and 5 non-coding genes were identified. Furthermore, in the long-term treatment group, 11 of the 15 upregulated genes and 9 of the 14 downregulated genes were reported as oncogenes and tumor suppressor genes in lung cancer, respectively. We also found that 10 genes of the selected 24 protein-coding genes were clinically significant in lung adenocarcinoma patients. CONCLUSIONS Our findings demonstrate that long-term exposure of human pulmonary normal alveolar cells to low-dose PHMG-p caused genetic changes, mainly in lung cancer-associated genes, in a time-dependent manner.
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Affiliation(s)
- Hong Lee
- Medical Science Research Center, Ansan Hospital, Korea University College of Medicine, Ansan-si, Gyeonggi, Republic of Korea
| | - Sang Hoon Jeong
- Medical Science Research Center, Ansan Hospital, Korea University College of Medicine, Ansan-si, Gyeonggi, Republic of Korea
| | - Hyejin Lee
- Medical Science Research Center, Ansan Hospital, Korea University College of Medicine, Ansan-si, Gyeonggi, Republic of Korea
| | - Cherry Kim
- Department of Radiology, Ansan Hospital, Korea University College of Medicine, Ansan-si, Gyeonggi, Republic of Korea
| | - Yoon Jeong Nam
- Medical Science Research Center, Ansan Hospital, Korea University College of Medicine, Ansan-si, Gyeonggi, Republic of Korea
| | - Ja Young Kang
- Medical Science Research Center, Ansan Hospital, Korea University College of Medicine, Ansan-si, Gyeonggi, Republic of Korea
| | - Myeong Ok Song
- Medical Science Research Center, Ansan Hospital, Korea University College of Medicine, Ansan-si, Gyeonggi, Republic of Korea
| | - Jin Young Choi
- Medical Science Research Center, Ansan Hospital, Korea University College of Medicine, Ansan-si, Gyeonggi, Republic of Korea
| | - Jaeyoung Kim
- Medical Science Research Center, Ansan Hospital, Korea University College of Medicine, Ansan-si, Gyeonggi, Republic of Korea
| | - Eun-Kee Park
- Department of Medical Humanities and Social Medicine, College of Medicine, Kosin University, Busan, Republic of Korea
| | - Yong-Wook Baek
- Environmental Health Research Department, Humidifier Disinfectant Health Center, National Institute of Environmental Research, Incheon, Republic of Korea
| | - Ju-Han Lee
- Department of Pathology, Ansan Hospital, Korea University College of Medicine, Ansan-si, Gyeonggi, Republic of Korea.
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15
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Elghoul M, El-Gendy SAA, Alsafy MAM. Ultrastructural investigation of the pneumocytes in piglets that live in a trashed environment. Morphologie 2021; 106:294-299. [PMID: 34933787 DOI: 10.1016/j.morpho.2021.11.003] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/14/2021] [Revised: 11/17/2021] [Accepted: 11/30/2021] [Indexed: 11/15/2022]
Abstract
The present study provides the ultrastructure of the pneumocytes types II and I in piglets living in the trash environment. Samples of the lungs of twelve piglets two months old were used. By light microscopy, the pneumocytes type I were squamous and somewhat flattened cells, while pneumocytes type II was cuboidal to spherical projected within the alveolar lumen and characterized by a spherical nucleus and foamy cytoplasm, it contained vacuolated bodies that were polygonal with variable size manly at the poles of the cell. The electron microscopy investigation showed blood air barrier between the endothelial lining of pneumocytes type I and therefore the endothelial lining of blood capillary and their nucleus were irregular in shape varied from nearly irregular triangular to polygon rough endoplasmic reticulum represented at their cytoplasm. The pneumocytes type II were frequently covered by pneumocytes type I extensions and united to them by a tight junction. It had been characterized by a high number of mitochondria within the cytoplasm and vacuolated bodies encircled the nucleus and at the two extremities of the cell. The lamellar vacuolated bodies were connected to the endoplasmic reticulum membranes and therefore the intravascular macrophages were attached to the endothelial cells within the pulmonary capillaries until two months old piglets. The occurrence of the intravascular macrophages could be attributed to the higher resistance to the respiratory diseases of the piglets.
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Affiliation(s)
- M Elghoul
- Histology and Cytology Department, Faculty of Veterinary Medicine, Alexandria University, Alexandria, Egypt.
| | - S A A El-Gendy
- Anatomy and Embryology Department, Faculty of Veterinary Medicine, Alexandria University, Alexandria, Egypt.
| | - M A M Alsafy
- Anatomy and Embryology Department, Faculty of Veterinary Medicine, Alexandria University, Alexandria, Egypt.
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Bluhmki T, Traub S, Müller AK, Bitzer S, Schruf E, Bammert MT, Leist M, Gantner F, Garnett JP, Heilker R. Functional human iPSC-derived alveolar-like cells cultured in a miniaturized 96‑Transwell air-liquid interface model. Sci Rep 2021; 11:17028. [PMID: 34426605 PMCID: PMC8382767 DOI: 10.1038/s41598-021-96565-4] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2021] [Accepted: 08/11/2021] [Indexed: 02/06/2023] Open
Abstract
In order to circumvent the limited access and donor variability of human primary alveolar cells, directed differentiation of human pluripotent stem cells (hiPSCs) into alveolar-like cells, provides a promising tool for respiratory disease modeling and drug discovery assays. In this work, a unique, miniaturized 96-Transwell microplate system is described where hiPSC-derived alveolar-like cells were cultured at an air-liquid interface (ALI). To this end, hiPSCs were differentiated into lung epithelial progenitor cells (LPCs) and subsequently matured into a functional alveolar type 2 (AT2)-like epithelium with monolayer-like morphology. AT2-like cells cultured at the physiological ALI conditions displayed characteristics of AT2 cells with classical alveolar surfactant protein expressions and lamellar-body like structures. The integrity of the epithelial barriers between the AT2-like cells was confirmed by applying a custom-made device for 96-parallelized transepithelial electric resistance (TEER) measurements. In order to generate an IPF disease-like phenotype in vitro, the functional AT2-like cells were stimulated with cytokines and growth factors present in the alveolar tissue of IPF patients. The cytokines stimulated the secretion of pro-fibrotic biomarker proteins both on the mRNA (messenger ribonucleic acid) and protein level. Thus, the hiPSC-derived and cellular model system enables the recapitulation of certain IPF hallmarks, while paving the route towards a miniaturized medium throughput approach of pharmaceutical drug discovery.
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Affiliation(s)
- Teresa Bluhmki
- Department of Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, 88397, Biberach an der Riss, Germany.
| | - Stefanie Traub
- Trenzyme GmbH, Byk-Gulden-Str. 2, 78467, Constance, Germany
| | | | - Sarah Bitzer
- Department of Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, 88397, Biberach an der Riss, Germany
| | - Eva Schruf
- Department of Immunology & Respiratory Diseases Research, Boehringer Ingelheim Pharma GmbH & Co. KG, 88397, Biberach an der Riss, Germany
| | - Marie-Therese Bammert
- Department of Immunology & Respiratory Diseases Research, Boehringer Ingelheim Pharma GmbH & Co. KG, 88397, Biberach an der Riss, Germany
| | - Marcel Leist
- In-vitro Toxicology and Biomedicine, University of Konstanz, 78457, Constance, Germany
| | - Florian Gantner
- Department of Translational Medicine and Clinical Pharmacology, C. H. Boehringer Sohn AG & Co. KG, 88397, Biberach an der Riss, Germany
| | - James P Garnett
- Department of Immunology & Respiratory Diseases Research, Boehringer Ingelheim Pharma GmbH & Co. KG, 88397, Biberach an der Riss, Germany
| | - Ralf Heilker
- Department of Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, 88397, Biberach an der Riss, Germany
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17
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Ullah A, Qazi J, Rahman L, Kanaras AG, Khan WS, Hussain I, Rehman A. Nanoparticles-assisted delivery of antiviral-siRNA as inhalable treatment for human respiratory viruses: A candidate approach against SARS-COV-2. NANO SELECT 2020; 1:612-621. [PMID: 34485978 PMCID: PMC7675679 DOI: 10.1002/nano.202000125] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/31/2020] [Accepted: 09/25/2020] [Indexed: 12/13/2022] Open
Abstract
The current pandemic of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has challenged healthcare structures across the globe. Although a few therapies are approved by FDA, the search for better treatment options is continuously on rise. Clinical management includes infection prevention and supportive care such as supplemental oxygen and mechanical ventilatory support. Given the urgent nature of the pandemic and the number of companies and researchers developing COVID-19 related therapies, FDA has created an emergency program to move potential treatments with already approved drugs to patients as quickly as possible in parallel to the development of new drugs that must first pass the clinical trials. In this manuscript, we have reviewed the available literature on the use of sequence-specific degradation of viral genome using short-interfering RNA (siRNA) suggesting it as a possible treatment against SARS-CoV-2. Delivery of siRNA can be promoted by the use of FDA approved lipids, polymers or lipid-polymer hybrids. These nanoparticulate systems can be engineered to exhibit increased targetability and formulated as inhalable aerosols.
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Affiliation(s)
- Ata Ullah
- National Institute for Biotechnology and Genetic EngineeringFaisalabadPakistan
| | - Javaria Qazi
- Department of BiotechnologyQuaid‐i‐Azam UniversityIslamabadPakistan
| | - Lutfur Rahman
- National Institute for Biotechnology and Genetic EngineeringFaisalabadPakistan
| | - Antonios G. Kanaras
- Physics and AstronomyInstitute for Life SciencesUniversity of SouthamptonSouthamptonSO171BJUK
| | - Waheed S. Khan
- National Institute for Biotechnology and Genetic EngineeringFaisalabadPakistan
| | - Irshad Hussain
- Department of Chemistry and Chemical EngineeringSBA School of Science & Engineering (SBASSE)Lahore University of Management Sciences (LUMS)LahorePakistan
| | - Asma Rehman
- National Institute for Biotechnology and Genetic EngineeringFaisalabadPakistan
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Parekh KR, Nawroth J, Pai A, Busch SM, Senger CN, Ryan AL. Stem cells and lung regeneration. Am J Physiol Cell Physiol 2020; 319:C675-C693. [PMID: 32783658 PMCID: PMC7654650 DOI: 10.1152/ajpcell.00036.2020] [Citation(s) in RCA: 64] [Impact Index Per Article: 12.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2020] [Revised: 08/03/2020] [Accepted: 08/03/2020] [Indexed: 12/20/2022]
Abstract
The ability to replace defective cells in an airway with cells that can engraft, integrate, and restore a functional epithelium could potentially cure a number of lung diseases. Progress toward the development of strategies to regenerate the adult lung by either in vivo or ex vivo targeting of endogenous stem cells or pluripotent stem cell derivatives is limited by our fundamental lack of understanding of the mechanisms controlling human lung development, the precise identity and function of human lung stem and progenitor cell types, and the genetic and epigenetic control of human lung fate. In this review, we intend to discuss the known stem/progenitor cell populations, their relative differences between rodents and humans, their roles in chronic lung disease, and their therapeutic prospects. Additionally, we highlight the recent breakthroughs that have increased our understanding of these cell types. These advancements include novel lineage-traced animal models and single-cell RNA sequencing of human airway cells, which have provided critical information on the stem cell subtypes, transition states, identifying cell markers, and intricate pathways that commit a stem cell to differentiate or to maintain plasticity. As our capacity to model the human lung evolves, so will our understanding of lung regeneration and our ability to target endogenous stem cells as a therapeutic approach for lung disease.
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Affiliation(s)
- Kalpaj R Parekh
- Department Surgery, Division of Cardiothoracic Surgery, University of Iowa, Iowa City, Iowa
| | - Janna Nawroth
- Hastings Center for Pulmonary Research, Division of Pulmonary, Critical Care and Sleep Medicine, Department of Medicine, University of Southern California, Los Angeles, California
| | - Albert Pai
- Department Surgery, Division of Cardiothoracic Surgery, University of Iowa, Iowa City, Iowa
| | - Shana M Busch
- Hastings Center for Pulmonary Research, Division of Pulmonary, Critical Care and Sleep Medicine, Department of Medicine, University of Southern California, Los Angeles, California
| | - Christiana N Senger
- Hastings Center for Pulmonary Research, Division of Pulmonary, Critical Care and Sleep Medicine, Department of Medicine, University of Southern California, Los Angeles, California
| | - Amy L Ryan
- Hastings Center for Pulmonary Research, Division of Pulmonary, Critical Care and Sleep Medicine, Department of Medicine, University of Southern California, Los Angeles, California
- Department of Stem Cell Biology and Regenerative Medicine, University of Southern California, Los Angeles, California
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19
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Valacchi G, Magnani N, Woodby B, Ferreira SM, Evelson P. Particulate Matter Induces Tissue OxInflammation: From Mechanism to Damage. Antioxid Redox Signal 2020; 33:308-326. [PMID: 32443938 DOI: 10.1089/ars.2019.8015] [Citation(s) in RCA: 43] [Impact Index Per Article: 8.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Significance: Oxidative stress and oxidative damage are central hypothetical mechanisms for the adverse effects of airborne particulate matter (PM). Activation of inflammatory cells capable of generating reactive oxygen and nitrogen species is another proposed damage pathway. Understanding the interplay between these responses can help us understand the adverse health effects attributed to breathing polluted air. Recent Advances: The consequences of PM exposure on different organs are oxidative damage, decreased function, and inflammation, which can lead to the development/exacerbation of proinflammatory disorders. Mitochondrial damage is also an important event in PM-induced cytotoxicity. Critical Issues: Reactive oxygen species (ROS) are generated during phagocytosis of the particles, leading to enhancement of oxidative stress and triggering the inflammatory response. The activation of inflammatory signaling pathways results in the release of cytokines and other mediators, which can further induce ROS production by activating endogenous enzymes, leading to a positive feedback loop, which can aggravate the effects triggered by PM exposure. Future Directions: Further research is required to elucidate the exact mechanisms by which PM exposure results in adverse health effects, in terms of the relationship between the redox responses triggered by the presence of the particles and the inflammation observed in the different organs, so the development/exacerbation of PM-associated health problems can be prevented.
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Affiliation(s)
- Giuseppe Valacchi
- Department of Animal Science, Plants for Human Health Institute, NC Research Campus, NC State University, Kannapolis, North Carolina, USA.,Department of Biomedical and Specialist Surgical Sciences, University of Ferrara, Ferrara, Italy.,Department of Food and Nutrition, Kyung Hee University, Seoul, South Korea
| | - Natalia Magnani
- Universidad de Buenos Aires, Facultad de Farmacia y Bioquímica, Departamento de Química Analítica y Fisicoquímica, Cátedra de Química General e Inorgánica, Buenos Aires, Argentina.,CONICET, Instituto de Bioquímica y Medicina Molecular (IBIMOL), Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina
| | - Brittany Woodby
- Department of Animal Science, Plants for Human Health Institute, NC Research Campus, NC State University, Kannapolis, North Carolina, USA
| | - Sandra María Ferreira
- Universidad de Buenos Aires, Facultad de Farmacia y Bioquímica, Departamento de Química Analítica y Fisicoquímica, Cátedra de Química General e Inorgánica, Buenos Aires, Argentina.,CONICET, Instituto de Bioquímica y Medicina Molecular (IBIMOL), Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina
| | - Pablo Evelson
- Universidad de Buenos Aires, Facultad de Farmacia y Bioquímica, Departamento de Química Analítica y Fisicoquímica, Cátedra de Química General e Inorgánica, Buenos Aires, Argentina.,CONICET, Instituto de Bioquímica y Medicina Molecular (IBIMOL), Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Buenos Aires, Argentina
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20
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Gu C, Zhang Q, Ni D, Xiao QF, Cao LF, Fei CY, Ying Y, Li N, Tao F. Therapeutic Effects of SRT2104 on Lung Injury in Rats with Emphysema via Reduction of Type II Alveolar Epithelial Cell Senescence. COPD 2020; 17:444-451. [PMID: 32722945 DOI: 10.1080/15412555.2020.1797657] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/23/2022]
Abstract
Chronic obstructive pulmonary disease (COPD) is one of the most prevalent and severe diseases worldwide with high societal and health care costs. The pathogenesis of COPD is very complicated, and no curative treatment is available. Cellular senescence promotes the development of COPD. Type II alveolar epithelial cells (AECII) play a momentous role in lung tissue repair and maintenance of alveolar homeostasis. Sirtuin 1 (SIRT1), an antiaging molecule involved in the response to chronic inflammation and oxidative stress, regulates many pathophysiological changes including stress resistance, apoptosis, inflammation, and cellular senescence. This study aimed to investigate whether the pharmacological SIRT1 activator SRT2104 protects against AECII senescence in rats with emphysema. Our findings confirmed that SRT2104 administration reduced the pathological characteristics of emphysema and improved lung function parameters, including pulmonary resistance, pulmonary dynamic compliance, and peak expiratory flow. Moreover, SRT2104 treatment upregulated the expression of surfactant proteins A and C, SIRT1, and forkhead box O 3a (FoxO3a), decreased senescence-associated-β-galactosidase (SA-β-gal) activity, increased SIRT1 deacetylase activity, and downregulated the levels of p53 and p21. Therefore, SRT2104 administration protected against AECII senescence in rats with emphysema via SIRT1/FoxO3a and SIRT1/p53 signaling pathways and may provide a novel potential therapeutic strategy for COPD.
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Affiliation(s)
- Chao Gu
- Department of Respiratory Medicine, The First Hospital of Jiaxing (the Affiliated Hospital of Jiaxing University), Jiaxing, Zhejiang, People's Republic of China
| | - Qi Zhang
- Department of Respiratory Medicine, The First Hospital of Jiaxing (the Affiliated Hospital of Jiaxing University), Jiaxing, Zhejiang, People's Republic of China
| | - Dan Ni
- Department of Respiratory Medicine, The First Hospital of Jiaxing (the Affiliated Hospital of Jiaxing University), Jiaxing, Zhejiang, People's Republic of China
| | - Qin-Feng Xiao
- Department of Respiratory Medicine, The First Hospital of Jiaxing (the Affiliated Hospital of Jiaxing University), Jiaxing, Zhejiang, People's Republic of China
| | - Lin-Feng Cao
- Department of Respiratory Medicine, The First Hospital of Jiaxing (the Affiliated Hospital of Jiaxing University), Jiaxing, Zhejiang, People's Republic of China
| | - Chun-Yuan Fei
- Department of Respiratory Medicine, The First Hospital of Jiaxing (the Affiliated Hospital of Jiaxing University), Jiaxing, Zhejiang, People's Republic of China
| | - Ying Ying
- Department of Respiratory Medicine, The First Hospital of Jiaxing (the Affiliated Hospital of Jiaxing University), Jiaxing, Zhejiang, People's Republic of China
| | - Na Li
- Department of Respiratory Medicine, The First Hospital of Jiaxing (the Affiliated Hospital of Jiaxing University), Jiaxing, Zhejiang, People's Republic of China
| | - Feng Tao
- Department of Respiratory Medicine, The First Hospital of Jiaxing (the Affiliated Hospital of Jiaxing University), Jiaxing, Zhejiang, People's Republic of China
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21
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Xenogeneic cross-circulation for extracorporeal recovery of injured human lungs. Nat Med 2020; 26:1102-1113. [PMID: 32661401 PMCID: PMC9990469 DOI: 10.1038/s41591-020-0971-8] [Citation(s) in RCA: 57] [Impact Index Per Article: 11.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2019] [Accepted: 06/09/2020] [Indexed: 12/15/2022]
Abstract
Patients awaiting lung transplantation face high wait-list mortality, as injury precludes the use of most donor lungs. Although ex vivo lung perfusion (EVLP) is able to recover marginal quality donor lungs, extension of normothermic support beyond 6 h has been challenging. Here we demonstrate that acutely injured human lungs declined for transplantation, including a lung that failed to recover on EVLP, can be recovered by cross-circulation of whole blood between explanted human lungs and a Yorkshire swine. This xenogeneic platform provided explanted human lungs a supportive, physiologic milieu and systemic regulation that resulted in functional and histological recovery after 24 h of normothermic support. Our findings suggest that cross-circulation can serve as a complementary approach to clinical EVLP to recover injured donor lungs that could not otherwise be utilized for transplantation, as well as a translational research platform for immunomodulation and advanced organ bioengineering.
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22
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Sveiven SN, Nordgren TM. Lung-resident mesenchymal stromal cells are tissue-specific regulators of lung homeostasis. Am J Physiol Lung Cell Mol Physiol 2020; 319:L197-L210. [PMID: 32401672 DOI: 10.1152/ajplung.00049.2020] [Citation(s) in RCA: 21] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Until recently, data supporting the tissue-resident status of mesenchymal stromal cells (MSC) has been ambiguous since their discovery in the 1950-60s. These progenitor cells were first discovered as bone marrow-derived adult multipotent cells and believed to migrate to sites of injury, opposing the notion that they are residents of all tissue types. In recent years, however, it has been demonstrated that MSC can be found in all tissues and MSC from different tissues represent distinct populations with differential protein expression unique to each tissue type. Importantly, these cells are efficient mediators of tissue repair, regeneration, and prove to be targets for therapeutics, demonstrated by clinical trials (phase 1-4) for MSC-derived therapies for diseases like graft-versus-host-disease, multiple sclerosis, rheumatoid arthritis, and Crohn's disease. The tissue-resident status of MSC found in the lung is a key feature of their importance in the context of disease and injuries of the respiratory system, since these cells could be instrumental to providing more specific and targeted therapies. Currently, bone marrow-derived MSC have been established in the treatment of disease, including diseases of the lung. However, with lung-resident MSC representing a unique population with a different phenotypic and gene expression pattern than MSC derived from other tissues, their role in remediating lung inflammation and injury could provide enhanced efficacy over bone marrow-derived MSC methods. Through this review, lung-resident MSC will be characterized, using previously published data, by surface markers, gene expression patterns, and compared with bone-marrow MSC to highlight similarities and, importantly, differences in these cell types.
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Affiliation(s)
- Stefanie Noel Sveiven
- Division of Biomedical Sciences, School of Medicine, University of California-Riverside, Riverside, California
| | - Tara M Nordgren
- Division of Biomedical Sciences, School of Medicine, University of California-Riverside, Riverside, California
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23
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Yanagihara T, Chong SG, Vierhout M, Hirota JA, Ask K, Kolb M. Current models of pulmonary fibrosis for future drug discovery efforts. Expert Opin Drug Discov 2020; 15:931-941. [PMID: 32396021 DOI: 10.1080/17460441.2020.1755252] [Citation(s) in RCA: 45] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
INTRODUCTION Pulmonary fibrosis includes several lung disorders characterized by progressive fibrosis, of which idiopathic pulmonary fibrosis (IPF) is a particularly severe form with a median survival time of 3-5 years after diagnosis. Although numerous compounds have shown efficacy in attenuating pulmonary fibrosis using animal models, only a few compounds have shown their beneficial effects for IPF in clinical trials. Thus, there is an emergent need to improve the preclinical development process to better identify, characterize and select clinically useful targets. AREAS COVERED In this review, the authors extensively describe current models of pulmonary fibrosis, including rodent models, ex vivo models, and in vitro models. EXPERT OPINION Based upon our current understanding, improving the identification and characterization of clinically relevant molecules or pathways responsible for progressive fibrotic diseases and use of the appropriate preclinical model system to test these will likely be required to improve the drug development pipeline for pulmonary fibrosis. Combination with appropriate preclinical models with ex vivo (precision-cut lung slices) or in vitro models would be beneficial for high-throughput drug discovery or validation of drug effects.
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Affiliation(s)
- Toyoshi Yanagihara
- Firestone Institute for Respiratory Health, Research Institute at St Joseph's Healthcare, Department of Medicine, McMaster University , Hamilton, ON, Canada.,Research Institute for Diseases of the Chest, Graduate School of Medical Sciences, Kyushu University , Fukuoka, Japan
| | - Sy Giin Chong
- Firestone Institute for Respiratory Health, Research Institute at St Joseph's Healthcare, Department of Medicine, McMaster University , Hamilton, ON, Canada
| | - Megan Vierhout
- Firestone Institute for Respiratory Health, Research Institute at St Joseph's Healthcare, Department of Medicine, McMaster University , Hamilton, ON, Canada
| | - Jeremy A Hirota
- Firestone Institute for Respiratory Health, Research Institute at St Joseph's Healthcare, Department of Medicine, McMaster University , Hamilton, ON, Canada
| | - Kjetil Ask
- Firestone Institute for Respiratory Health, Research Institute at St Joseph's Healthcare, Department of Medicine, McMaster University , Hamilton, ON, Canada
| | - Martin Kolb
- Firestone Institute for Respiratory Health, Research Institute at St Joseph's Healthcare, Department of Medicine, McMaster University , Hamilton, ON, Canada
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24
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Jieming G, Liu C, Yang Y, Mo S, Yang X, Wang J. Inhibitory effects of msFGFR2c on the epithelial-to-mesenchymal transition of AE2 cells in pulmonary fibrosis. Biotechnol Lett 2020; 42:1061-1070. [PMID: 32130565 PMCID: PMC7211205 DOI: 10.1007/s10529-020-02852-x] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/08/2019] [Accepted: 02/26/2020] [Indexed: 11/29/2022]
Abstract
In interstitial fibrosis, alveolar epithelial type II (AE2) cells fail to repair damaged epithelium. However, whether this dysfunction is related to fibroblast growth factor (FGF) signal pathway and how it affects the fibrotic process remains unclear. In our study, the medium of the human foetal lung fibroblast cell line MRC-5 (Med) can induce epithelial-to-mesenchymal transition (EMT) in AE2 cells, we also found that TGF-β in Med can induce FGF-2 and CTGF expression in AE2 cells. TGF-β or CTGF exposure trigger a FGFR2 subtype b to c transition which can be supressed by siRNA-CTGF. All together, since FGFR2IIIc have the highest affinity with FGF-2 in all of the FGFRs, we indicate the activation of FGF2 signal pathway was induced by TGF-β, which is the key component of Med Here, we also find the inhibitory effect of msFGFR2c (S252W mutant of soluble FGFR2IIIc extracellular domain) on EMT of mouse primary AE2 cells in pulmonary fibrotic process. In a bleomycin-induced mouse pulmonary fibrosis model, msFGFR2c alleviate pulmonary fibrosis and suppress the decrease in pro-SPC levels. Thus, msFGFR2c can inhibit EMT-induced fibrosis of AE2 cells via FGF-2 signal and AE2 cells is suggested to play an important role in the lung fibrotic process.
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Affiliation(s)
- Guo Jieming
- Tai Shan College, Shandong University, Jinan, 250000, China
| | - Chuan Liu
- Institute of Biomedicine, National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, 510632, China
| | - Yin Yang
- Institute of Biomedicine, National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, 510632, China
| | - Shanyi Mo
- Institute of Biomedicine, National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, 510632, China
| | - Xuesong Yang
- Department of Pediatrics and Neonatology, Institute of Fetal-Preterm Labor Medicine, The First Affiliated Hospital, Jinan University, Guangzhou, 510630, China
| | - Ju Wang
- Institute of Biomedicine, National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, 510632, China.
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25
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Chu SG, Poli De Frias S, Sakairi Y, Kelly RS, Chase R, Konishi K, Blau A, Tsai E, Tsoyi K, Padera RF, Sholl LM, Goldberg HJ, Mallidi HR, Camp PC, El-Chemaly SY, Perrella MA, Choi AMK, Washko GR, Raby BA, Rosas IO. Biobanking and cryopreservation of human lung explants for omic analysis. Eur Respir J 2020; 55:13993003.01635-2018. [PMID: 31699836 DOI: 10.1183/13993003.01635-2018] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2018] [Accepted: 10/10/2019] [Indexed: 02/05/2023]
Affiliation(s)
- Sarah G Chu
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.,These authors contributed equally to the manuscript
| | - Sergio Poli De Frias
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.,These authors contributed equally to the manuscript
| | - Yuichi Sakairi
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Rachel S Kelly
- Channing Division of Network Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Robert Chase
- Channing Division of Network Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Kazuhisa Konishi
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Ashley Blau
- Partners Biobank and Translational Genomics Core, Harvard Medical School, Boston, MA, USA
| | - Ellen Tsai
- Partners Biobank and Translational Genomics Core, Harvard Medical School, Boston, MA, USA
| | - Konstantin Tsoyi
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Robert F Padera
- Dept of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Lynette M Sholl
- Dept of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Hilary J Goldberg
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Hari R Mallidi
- Division of Thoracic Surgery, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Phillip C Camp
- Division of Thoracic Surgery, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Souheil Y El-Chemaly
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Mark A Perrella
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.,Dept of Pediatric Newborn Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | | | - George R Washko
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Benjamin A Raby
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.,Channing Division of Network Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.,Division of Pulmonary and Respiratory Diseases, Boston Children's Hospital, Harvard Medical School, Boston, MA, USA
| | - Ivan O Rosas
- Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
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26
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Yin B, Chan CKW, Liu S, Hong H, Wong SHD, Lee LKC, Ho LWC, Zhang L, Leung KCF, Choi PCL, Bian L, Tian XY, Chan MN, Choi CHJ. Intrapulmonary Cellular-Level Distribution of Inhaled Nanoparticles with Defined Functional Groups and Its Correlations with Protein Corona and Inflammatory Response. ACS NANO 2019; 13:14048-14069. [PMID: 31725257 DOI: 10.1021/acsnano.9b06424] [Citation(s) in RCA: 44] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/10/2023]
Abstract
Concerns over the health risks associated with airborne exposure to ultrafine particles [PM0.1, or nanoparticles (NPs)] call for a comprehensive understanding in the interactions of inhaled NPs along their respiratory journey. We prepare a collection of polyethylene glycol-coated gold nanoparticles that bear defined functional groups commonly identified in atmospheric particulates (Au@PEG-X NPs, where X = OCH3, COOH, NH2, OH, or C12H25). Regardless of the functional group, these ∼50 nm NPs remain colloidally stable following aerosolization and incubation in bronchoalveolar lavage fluid (BALF), without pronouncedly crossing the air-blood barrier. The type of BALF proteins adhered onto the NPs is similar, but the composition of protein corona depends on functional group. By subjecting Balb/c mice to inhalation of Au@PEG-X NPs for 6 h, we demonstrate that the intrapulmonary distribution of NPs among the various types of cells (both found in BALF and isolated from the lavaged lung) and the acute inflammatory responses induced by inhalation are sensitive to the functional group of NPs and postinhalation period (0, 24, or 48 h). By evaluating the pairwise correlations between the three variables of "lung-nano" interactions (protein corona, intrapulmonary cellular-level distribution, and inflammatory response), we reveal strong statistical correlations between the (1) fractions of albumin or carbonyl reductase bound to NPs, (2) associations of inhaled NPs to neutrophils in BALF or macrophages in the lavaged lung, and (3) level of total protein in BALF. Our results provide insights into the effect of functional group on lung-nano interactions and health risks associated with inhalation of PM0.1.
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Affiliation(s)
| | | | | | | | | | | | | | | | - Ken Cham-Fai Leung
- Department of Chemistry , Hong Kong Baptist University , Kowloon , Hong Kong
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27
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Suzuki T, Ota C, Fujino N, Tando Y, Suzuki S, Yamada M, Kondo T, Okada Y, Kubo H. Improving the viability of tissue-resident stem cells using an organ-preservation solution. FEBS Open Bio 2019; 9:2093-2104. [PMID: 31642604 PMCID: PMC6886303 DOI: 10.1002/2211-5463.12748] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/15/2019] [Revised: 09/02/2019] [Accepted: 10/22/2019] [Indexed: 12/24/2022] Open
Abstract
Human clinical specimens are a valuable source of tissue‐resident stem cells, but such cells need to be collected immediately after tissue collection. To extend the timescale for collection from fresh human samples, we developed a new extracellular fluid (ECF)‐type preservation solution based on a high‐sodium and low‐potassium solution containing low‐molecular‐weight dextran and glucose, which is used for preservation of organs for transplantation. In this study, we compared the preservation of tissue‐resident stem cells using our ECF solution with that using three other solutions: PBS, Dulbecco’s modified Eagle’s medium and Euro‐Collins solution. These solutions represent a common buffer, a common culture medium and a benchmark organ‐preservation solution, respectively. Lung tissues were removed from mice and preserved for 72 h under low‐temperature conditions. Of the solutions tested, only preservation in the ECF‐type solution could maintain the proliferation and differentiation capacity of mouse lung tissue‐resident stem cells. In addition, the ECF solution could preserve the viability and proliferation of human alveolar epithelial progenitor cells when stored for more than 7 days at 4 °C. The mean viability of human alveolar type II cells at 2, 5, 8 and 14 days of low‐temperature preservation was 90.9%, 84.8%, 85.7% and 66.3%, respectively, with no significant differences up to 8 days. Overall, our findings show that use of our ECF‐type preservation solution may maintain the viability and function of tissue‐resident stem cells. Use of this preservation solution may facilitate the investigation of currently unobtainable human tissue specimens for human stem cell biology. Here, we describe a newly developed extracellular fluid‐type organ preservation solution that maintained the viability of human lung stem/progenitor cells, such as alveolar type II cells, during 7‐day refrigerated preservation after the collection of lung specimens in local hospitals. This ready‐to‐use solution may be suitable for the transport of human clinical specimens from hospitals to scientific and bioengineering laboratories.![]()
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Affiliation(s)
- Takaya Suzuki
- Department of Thoracic Surgery, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Japan
| | - Chiharu Ota
- Department of Pediatrics, Tohoku University Hospital, Sendai, Japan
| | - Naoya Fujino
- Department of Respiratory Medicine, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Yukiko Tando
- Cell Resource Center for Biomedical Research, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Japan
| | - Satoshi Suzuki
- Department of Thoracic Surgery, Japanese Red Cross Ishinomaki Hospital, Ishinomaki, Japan
| | - Mitsuhiro Yamada
- Department of Respiratory Medicine, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Takashi Kondo
- Department of Thoracic Surgery, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Japan
| | - Yoshinori Okada
- Department of Thoracic Surgery, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Japan
| | - Hiroshi Kubo
- Department of Advanced Preventive Medicine for Infectious Disease, Tohoku University Graduate School of Medicine, Sendai, Japan
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28
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Wang S, Yu J, Kane MA, Moise AR. Modulation of retinoid signaling: therapeutic opportunities in organ fibrosis and repair. Pharmacol Ther 2019; 205:107415. [PMID: 31629008 DOI: 10.1016/j.pharmthera.2019.107415] [Citation(s) in RCA: 29] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2019] [Accepted: 09/17/2019] [Indexed: 02/08/2023]
Abstract
The vitamin A metabolite, retinoic acid, is an important signaling molecule during embryonic development serving critical roles in morphogenesis, organ patterning and skeletal and neural development. Retinoic acid is also important in postnatal life in the maintenance of tissue homeostasis, while retinoid-based therapies have long been used in the treatment of a variety of cancers and skin disorders. As the number of people living with chronic disorders continues to increase, there is great interest in extending the use of retinoid therapies in promoting the maintenance and repair of adult tissues. However, there are still many conflicting results as we struggle to understand the role of retinoic acid in the multitude of processes that contribute to tissue injury and repair. This review will assess our current knowledge of the role retinoic acid signaling in the development of fibroblasts, and their transformation to myofibroblasts, and of the potential use of retinoid therapies in the treatment of organ fibrosis.
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Affiliation(s)
- Suya Wang
- Department of Cardiology, Boston Children's Hospital, 300 Longwood Avenue, Boston, MA, 02115, USA
| | - Jianshi Yu
- Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland, Baltimore, MD, 21201, USA
| | - Maureen A Kane
- Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland, Baltimore, MD, 21201, USA.
| | - Alexander R Moise
- Medical Sciences Division, Northern Ontario School of Medicine, Sudbury, ON P3E 2C6, Canada; Departments of Chemistry and Biochemistry, and Biology and Biomolecular Sciences Program, Laurentian University, Sudbury, ON, P3E 2C6, Canada.
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29
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Comber EM, Palchesko RN, Ng WH, Ren X, Cook KE. De novo lung biofabrication: clinical need, construction methods, and design strategy. Transl Res 2019; 211:1-18. [PMID: 31103468 DOI: 10.1016/j.trsl.2019.04.008] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/01/2019] [Revised: 04/10/2019] [Accepted: 04/25/2019] [Indexed: 01/22/2023]
Abstract
Chronic lung disease is the 4th leading cause of death in the United States. Due to a shortage of donor lungs, alternative approaches to support failing, native lungs have been attempted, including mechanical ventilation and various forms of artificial lungs. However, each of these support methods causes significant complications when used for longer than a few days and are thus not capable of long-term support. For artificial lungs, complications arise due to interactions between the artificial materials of the device and the blood of the recipient. A potential new approach is the fabrication of lungs from biological materials, such that the gas exchange membranes provide a more biomimetic blood-contacting interface. Recent advancements with three-dimensional, soft-tissue biofabrication methods and the engineering of thin, basement membranes demonstrate the potential of fabricating a lung scaffold from extracellular matrix materials. This scaffold could then be seeded with endothelial and epithelial cells, matured within a bioreactor, and transplanted. In theory, this fully biological lung could provide improved, long-term biocompatibility relative to artificial lungs, but significant work is needed to perfect the organ design and construction methods. Like artificial lungs, biofabricated lungs do not need to follow the shape and structure of a native lung, allowing for simpler manufacture. However, various functional requirements must still be met, including stable, efficient gas exchange for a period of years. Design decisions depend on the disease state, how the organ is implanted, and the latest biofabrication methods available in a rapidly evolving field.
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Affiliation(s)
- Erica M Comber
- Department of Biomedical Engineering, Carnegie Mellon University, Pittsburgh, Pennsylvania.
| | - Rachelle N Palchesko
- Department of Biomedical Engineering, Carnegie Mellon University, Pittsburgh, Pennsylvania
| | - Wai Hoe Ng
- Department of Biomedical Engineering, Carnegie Mellon University, Pittsburgh, Pennsylvania
| | - Xi Ren
- Department of Biomedical Engineering, Carnegie Mellon University, Pittsburgh, Pennsylvania
| | - Keith E Cook
- Department of Biomedical Engineering, Carnegie Mellon University, Pittsburgh, Pennsylvania
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30
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Lee D, Chambers M. A bilayer tissue culture model of the bovine alveolus. F1000Res 2019; 8:357. [PMID: 31448101 PMCID: PMC6685456 DOI: 10.12688/f1000research.18696.1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 03/27/2019] [Indexed: 08/02/2024] Open
Abstract
The epithelial lining of the lung is often the first point of interaction between the host and inhaled pathogens, allergens and medications. Epithelial cells are therefore the main focus of studies which aim to shed light on host-pathogen interactions, to dissect the mechanisms of local host immunity and study toxicology. If these studies are not to be conducted exclusively in vivo, it is imperative that in vitro models are developed with a high in vitro- in vivo correlation. We describe here a co-culture bilayer model of the bovine alveolus, designed to overcome some of the limitations encountered with mono-culture and live animal models. Our system includes bovine pulmonary arterial endothelial cells (BPAECs) seeded onto a permeable membrane in 24 well Transwell format. The BPAECs are overlaid with immortalised bovine alveolar type II epithelial cells and the bilayer cultured at air-liquid interface for 14 days before use; in our case to study host-mycobacterial interactions. Characterisation of novel cell lines and the bilayer model have provided compelling evidence that immortalised bovine alveolar type II cells are an authentic substitute for primary alveolar type II cells and their culture as a bilayer in conjunction with BPAECs provides a physiologically relevant in vitro model of the bovine alveolus. The bilayer model may be used to study dynamic intracellular and extracellular host-pathogen interactions, using proteomics, genomics, live cell imaging, in-cell ELISA and confocal microscopy. The model presented in this article enables other researchers to establish an in vitro model of the bovine alveolus that is easy to set up, malleable and serves as a comparable alternative to in vivo models, whilst allowing study of early host-pathogen interactions, currently not feasible in vivo. The model therefore achieves one of the 3Rs objectives in that it replaces the use of animals in research of bovine respiratory diseases.
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Affiliation(s)
- Diane Lee
- School of Veterinary Medicine, University of Surrey, Guildford, Surrey, GU2 7AL, UK
| | - Mark Chambers
- School of Veterinary Medicine, University of Surrey, Guildford, Surrey, GU2 7AL, UK
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31
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Abstract
The epithelial lining of the lung is often the first point of interaction between the host and inhaled pathogens, allergens and medications. Epithelial cells are therefore the main focus of studies which aim to shed light on host-pathogen interactions, to dissect the mechanisms of local host immunity and study toxicology. If these studies are not to be conducted exclusively in vivo, it is imperative that in vitro models are developed with a high in vitro- in vivo correlation. We describe here a co-culture model of the bovine alveolus, designed to overcome some of the limitations encountered with mono-culture and live animal models. Our system includes bovine pulmonary arterial endothelial cells (BPAECs) seeded onto a permeable membrane in 24 well Transwell format. The BPAECs are overlaid with immortalised bovine alveolar type II epithelial cells and cultured at air-liquid interface for 14 days before use; in our case to study host-mycobacterial interactions. Characterisation of novel cell lines and the co-culture model have provided compelling evidence that immortalised bovine alveolar type II cells are an authentic substitute for primary alveolar type II cells and their co-culture with BPAECs provides a physiologically relevant in vitro model of the bovine alveolus. The co-culture model may be used to study dynamic intracellular and extracellular host-pathogen interactions, using proteomics, genomics, live cell imaging, in-cell ELISA and confocal microscopy. The model presented in this article enables other researchers to establish an in vitro model of the bovine alveolus that is easy to set up, malleable and serves as a comparable alternative to in vivo models, whilst allowing study of early host-pathogen interactions, currently not feasible in vivo. The model therefore achieves one of the 3Rs objectives in that it replaces the use of animals in research of bovine respiratory diseases.
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Affiliation(s)
- Diane Lee
- School of Veterinary Medicine, University of Surrey, Guildford, Surrey, GU2 7AL, UK
| | - Mark Chambers
- School of Veterinary Medicine, University of Surrey, Guildford, Surrey, GU2 7AL, UK
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32
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Lee DF, Chambers MA. Isolation of Alveolar Type II Cells from Adult Bovine Lung. ACTA ACUST UNITED AC 2019; 80:e71. [PMID: 30875462 DOI: 10.1002/cptx.71] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023]
Abstract
Alveolar type II (ATII) cells play a key role as part of the distal lung epithelium, including in the innate immune response and as self-renewing progenitors to replace alveolar type I (ATI) cells during epithelial regeneration. Their secretion of surfactant protein helps maintain homeostasis and exerts protective, antimicrobial properties. ATII cells remain difficult to study, partly due to inefficient and expensive isolation methods, a propensity to differentiate into ATI cells, and susceptibility to fibroblast contamination. Published methods of isolation often require specialized technology, negatively impacting the development of in vitro models of disease, including bovine tuberculosis. Presented here is a simple and cost-effective method for generation of bovine primary ATII cells. These cells exhibit an ATII phenotype in 2D and 3D culture and are conducive to further study of the role of ATII cells in bovine respiratory diseases. © 2019 by John Wiley & Sons, Inc.
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Affiliation(s)
- Diane Frances Lee
- School of Veterinary Medicine, University of Surrey, Guildford, Surrey, United Kingdom
| | - Mark Andrew Chambers
- School of Veterinary Medicine, University of Surrey, Guildford, Surrey, United Kingdom
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33
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Yoon YS, Jin M, Sin DD. Accelerated lung aging and chronic obstructive pulmonary disease. Expert Rev Respir Med 2019; 13:369-380. [PMID: 30735057 DOI: 10.1080/17476348.2019.1580576] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/12/2023]
Abstract
INTRODUCTION The prevalence of chronic obstructive pulmonary disease (COPD) increases exponentially with aging. Its pathogenesis, however, is not well known and aside from smoking cessation, there are no disease-modifying treatments for this disease. Areas covered: COPD is associated with accelerating aging and aging-related diseases. In this review, we will discuss the hallmarks of aging including genomic instability, telomere attrition, epigenetic alteration, loss of proteostasis, mitochondrial dysfunction, deregulated nutrient sensing, cellular senescence, stem cell exhaustion, and altered intercellular communication, which may be involved in COPD pathogenesis. Expert commentary: COPD and the aging process share similar molecular and cellular changes. Aging-related molecular pathways may represent novel therapeutic targets and biomarkers for COPD.
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Affiliation(s)
- Young Soon Yoon
- a Centre for Heart and Lung Innovation , St. Paul's Hospital & University of British Columbia , Vancouver , BC , Canada.,b Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine , Dongguk University Ilsan Hospital , Goyang , South Korea
| | - Minhee Jin
- a Centre for Heart and Lung Innovation , St. Paul's Hospital & University of British Columbia , Vancouver , BC , Canada
| | - Don D Sin
- a Centre for Heart and Lung Innovation , St. Paul's Hospital & University of British Columbia , Vancouver , BC , Canada.,c Division of Respiratory Medicine (Department of Medicine) , University of British Columbia , Vancouver , BC , Canada
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34
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Coppolino I, Ruggeri P, Nucera F, Cannavò MF, Adcock I, Girbino G, Caramori G. Role of Stem Cells in the Pathogenesis of Chronic Obstructive Pulmonary Disease and Pulmonary Emphysema. COPD 2018; 15:536-556. [DOI: 10.1080/15412555.2018.1536116] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/29/2022]
Affiliation(s)
- Irene Coppolino
- Dipartimento di Scienze Biomediche, Unità Operativa Complessa di Pneumologia, Odontoiatriche e delle Immagini Morfologiche e Funzionali (BIOMORF), Università degli Studi di Messina, Messina, Italy
| | - Paolo Ruggeri
- Dipartimento di Scienze Biomediche, Unità Operativa Complessa di Pneumologia, Odontoiatriche e delle Immagini Morfologiche e Funzionali (BIOMORF), Università degli Studi di Messina, Messina, Italy
| | - Francesco Nucera
- Dipartimento di Scienze Biomediche, Unità Operativa Complessa di Pneumologia, Odontoiatriche e delle Immagini Morfologiche e Funzionali (BIOMORF), Università degli Studi di Messina, Messina, Italy
| | - Mario Francesco Cannavò
- Dipartimento di Scienze Biomediche, Unità Operativa Complessa di Pneumologia, Odontoiatriche e delle Immagini Morfologiche e Funzionali (BIOMORF), Università degli Studi di Messina, Messina, Italy
| | - Ian Adcock
- Airways Disease Section, National Heart and Lung Institute, Royal Brompton Hospital Biomedical Research Unit, Imperial College, London, UK
| | - Giuseppe Girbino
- Dipartimento di Scienze Biomediche, Unità Operativa Complessa di Pneumologia, Odontoiatriche e delle Immagini Morfologiche e Funzionali (BIOMORF), Università degli Studi di Messina, Messina, Italy
| | - Gaetano Caramori
- Dipartimento di Scienze Biomediche, Unità Operativa Complessa di Pneumologia, Odontoiatriche e delle Immagini Morfologiche e Funzionali (BIOMORF), Università degli Studi di Messina, Messina, Italy
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35
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Lee DF, Salguero FJ, Grainger D, Francis RJ, MacLellan-Gibson K, Chambers MA. Isolation and characterisation of alveolar type II pneumocytes from adult bovine lung. Sci Rep 2018; 8:11927. [PMID: 30093682 PMCID: PMC6085293 DOI: 10.1038/s41598-018-30234-x] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/01/2018] [Accepted: 07/23/2018] [Indexed: 12/14/2022] Open
Abstract
Alveolar type II (ATII) cells play a key role as part of the distal lung epithelium, including roles in the innate immune response and as self-renewing progenitors to replace alveolar type I (ATI) cells during regeneration of the alveolar epithelium. Their secretion of surfactant protein helps to maintain homeostasis in the distal lung and exert protective, antimicrobial properties. Despite the cell's crucial roles, they remain difficult to study, in part due to inefficient and expensive isolation methods, a propensity to differentiate into alveolar type I cells in culture and susceptibility to fibroblast overgrowth from primary isolations. Published methods of isolation often require specialist technology, negatively impacting the development of in vitro models of disease, including bovine tuberculosis (BTB), a serious re-emerging disease in both animals and humans worldwide. We present here a simple and cost-effective method that may be utilised in the generation of bovine primary ATII cells. These exhibit an ATII phenotype in 2D and 3D culture in our studies and are conducive to further study of the role of ATII cells in bovine respiratory diseases.
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Affiliation(s)
- Diane Frances Lee
- School of Veterinary Medicine, University of Surrey, Daphne Jackson Road, Guildford, GU2 7AL, England.
| | - Francisco Javier Salguero
- School of Veterinary Medicine, University of Surrey, Daphne Jackson Road, Guildford, GU2 7AL, England
| | - Duncan Grainger
- School of Veterinary Medicine, University of Surrey, Daphne Jackson Road, Guildford, GU2 7AL, England
| | - Robert James Francis
- National Institute of Biological Standards and Control, Blanche Lane, South Mimms, Potters Bar, UK EN6 3QG, England
| | - Kirsty MacLellan-Gibson
- National Institute of Biological Standards and Control, Blanche Lane, South Mimms, Potters Bar, UK EN6 3QG, England
| | - Mark Andrew Chambers
- School of Veterinary Medicine, University of Surrey, Daphne Jackson Road, Guildford, GU2 7AL, England
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36
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Bandyopadhyay G, Huyck HL, Misra RS, Bhattacharya S, Wang Q, Mereness J, Lillis J, Myers JR, Ashton J, Bushnell T, Cochran M, Holden-Wiltse J, Katzman P, Deutsch G, Whitsett JA, Xu Y, Mariani TJ, Pryhuber GS. Dissociation, cellular isolation, and initial molecular characterization of neonatal and pediatric human lung tissues. Am J Physiol Lung Cell Mol Physiol 2018; 315:L576-L583. [PMID: 29975103 DOI: 10.1152/ajplung.00041.2018] [Citation(s) in RCA: 29] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
Human lung morphogenesis begins by embryonic life and continues after birth into early childhood to form a complex organ with numerous morphologically and functionally distinct cell types. Pulmonary organogenesis involves dynamic changes in cell proliferation, differentiation, and migration of specialized cells derived from diverse embryonic lineages. Studying the molecular and cellular processes underlying formation of the fully functional lung requires isolating distinct pulmonary cell populations during development. We now report novel methods to isolate four major pulmonary cell populations from pediatric human lung simultaneously. Cells were dissociated by protease digestion of neonatal and pediatric lung and isolated on the basis of unique cell membrane protein expression patterns. Epithelial, endothelial, nonendothelial mesenchymal, and immune cells were enriched by fluorescence-activated cell sorting. Dead cells and erythrocytes were excluded by 7-aminoactinomycin D uptake and glycophorin-A (CD235a) expression, respectively. Leukocytes were identified by membrane CD45 (protein tyrosine phosphatase, receptor type C), endothelial cells by platelet endothelial cell adhesion molecule-1 (CD31) and vascular endothelial cadherin (CD144), and both were isolated. Thereafter, epithelial cell adhesion molecule (CD326)-expressing cells were isolated from the endothelial- and immune cell-depleted population to enrich epithelial cells. Cells lacking these membrane markers were collected as "nonendothelial mesenchymal" cells. Quantitative RT-PCR and RNA sequencing analyses of population specific transcriptomes demonstrate the purity of the subpopulations of isolated cells. The method efficiently isolates major human lung cell populations that we announce are now available through the National Heart, Lung, and Blood Institute Lung Molecular Atlas Program (LungMAP) for their further study.
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Affiliation(s)
- Gautam Bandyopadhyay
- Division of Neonatology, Department of Pediatrics, University of Rochester Medical Center , Rochester, New York
| | - Heidie L Huyck
- Division of Neonatology, Department of Pediatrics, University of Rochester Medical Center , Rochester, New York
| | - Ravi S Misra
- Division of Neonatology, Department of Pediatrics, University of Rochester Medical Center , Rochester, New York
| | - Soumyaroop Bhattacharya
- Division of Neonatology, Department of Pediatrics, University of Rochester Medical Center , Rochester, New York.,Program in Pediatric Molecular and Personalized Medicine, Department of Pediatrics, University of Rochester Medical Center, Rochester, New York
| | - Qian Wang
- Division of Neonatology, Department of Pediatrics, University of Rochester Medical Center , Rochester, New York.,Program in Pediatric Molecular and Personalized Medicine, Department of Pediatrics, University of Rochester Medical Center, Rochester, New York
| | - Jared Mereness
- Division of Neonatology, Department of Pediatrics, University of Rochester Medical Center , Rochester, New York.,Program in Pediatric Molecular and Personalized Medicine, Department of Pediatrics, University of Rochester Medical Center, Rochester, New York
| | - Jacquelyn Lillis
- University of Rochester Genomics Research Center, University of Rochester Medical Center , Rochester, New York
| | - Jason R Myers
- University of Rochester Genomics Research Center, University of Rochester Medical Center , Rochester, New York
| | - John Ashton
- University of Rochester Genomics Research Center, University of Rochester Medical Center , Rochester, New York
| | - Timothy Bushnell
- University of Rochester Flow Cytometry Core Facility, University of Rochester Medical Center , Rochester, New York
| | - Matthew Cochran
- University of Rochester Flow Cytometry Core Facility, University of Rochester Medical Center , Rochester, New York
| | - Jeanne Holden-Wiltse
- University of Rochester Biocomputational Center, University of Rochester Medical Center , Rochester, New York
| | - Philip Katzman
- Department of Pathology, University of Rochester Medical Center , Rochester, New York
| | - Gail Deutsch
- Department of Pathology, Seattle Children's Hospital, University of Washington , Seattle, Washington
| | - Jeffrey A Whitsett
- Division of Neonatology, Perinatal and Pulmonary Biology Cincinnati Children's Hospital Medical Center , Cincinnati, Ohio
| | - Yan Xu
- Division of Neonatology, Perinatal and Pulmonary Biology Cincinnati Children's Hospital Medical Center , Cincinnati, Ohio
| | - Thomas J Mariani
- Division of Neonatology, Department of Pediatrics, University of Rochester Medical Center , Rochester, New York.,Program in Pediatric Molecular and Personalized Medicine, Department of Pediatrics, University of Rochester Medical Center, Rochester, New York
| | - Gloria S Pryhuber
- Division of Neonatology, Department of Pediatrics, University of Rochester Medical Center , Rochester, New York
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37
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Taylor DA, Sampaio LC, Ferdous Z, Gobin AS, Taite LJ. Decellularized matrices in regenerative medicine. Acta Biomater 2018; 74:74-89. [PMID: 29702289 DOI: 10.1016/j.actbio.2018.04.044] [Citation(s) in RCA: 195] [Impact Index Per Article: 27.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2017] [Revised: 04/19/2018] [Accepted: 04/23/2018] [Indexed: 01/04/2023]
Abstract
Of all biologic matrices, decellularized extracellular matrix (dECM) has emerged as a promising tool used either alone or when combined with other biologics in the fields of tissue engineering or regenerative medicine - both preclinically and clinically. dECM provides a native cellular environment that combines its unique composition and architecture. It can be widely obtained from native organs of different species after being decellularized and is entitled to provide necessary cues to cells homing. In this review, the superiority of the macro- and micro-architecture of dECM is described as are methods by which these unique characteristics are being harnessed to aid in the repair and regeneration of organs and tissues. Finally, an overview of the state of research regarding the clinical use of different matrices and the common challenges faced in using dECM are provided, with possible solutions to help translate naturally derived dECM matrices into more robust clinical use. STATEMENT OF SIGNIFICANCE Ideal scaffolds mimic nature and provide an environment recognized by cells as proper. Biologically derived matrices can provide biological cues, such as sites for cell adhesion, in addition to the mechanical support provided by synthetic matrices. Decellularized extracellular matrix is the closest scaffold to nature, combining unique micro- and macro-architectural characteristics with an equally unique complex composition. The decellularization process preserves structural integrity, ensuring an intact vasculature. As this multifunctional structure can also induce cell differentiation and maturation, it could become the gold standard for scaffolds.
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38
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Heinzelmann K, Lehmann M, Gerckens M, Noskovičová N, Frankenberger M, Lindner M, Hatz R, Behr J, Hilgendorff A, Königshoff M, Eickelberg O. Cell-surface phenotyping identifies CD36 and CD97 as novel markers of fibroblast quiescence in lung fibrosis. Am J Physiol Lung Cell Mol Physiol 2018; 315:L682-L696. [PMID: 29952218 DOI: 10.1152/ajplung.00439.2017] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023] Open
Abstract
Fibroblasts play an important role in lung homeostasis and disease. In lung fibrosis, fibroblasts adopt a proliferative and migratory phenotype, with increased expression of α-smooth muscle actin (αSMA) and enhanced secretion of extracellular matrix components. Comprehensive profiling of fibroblast heterogeneity is limited because of a lack of specific cell-surface markers. We have previously profiled the surface proteome of primary human lung fibroblasts. Here, we sought to define and quantify a panel of cluster of differentiation (CD) markers in primary human lung fibroblasts and idiopathic pulmonary fibrosis (IPF) lung tissue, using immunofluorescence and FACS analysis. Fibroblast function was assessed by analysis of replicative senescence. We observed the presence of distinct fibroblast phenotypes in vivo, characterized by various combinations of Desmin, αSMA, CD36, or CD97 expression. Most markers demonstrated stable expression over passages in vitro, but significant changes were observed for CD36, CD54, CD82, CD106, and CD140a. Replicative senescence of fibroblasts was observed from passage 10 onward. CD36- and CD97-positive but αSMA-negative cells were present in remodeled areas of IPF lungs. Transforming growth factor (TGF)-β treatment induced αSMA and collagen I expression but repressed CD36 and CD97 expression. We identified a panel of stable surface markers in human lung fibroblasts, applicable for positive-cell isolation directly from lung tissue. TGF-β exposure represses CD36 and CD97 expression, despite increasing αSMA expression; we therefore identified complex surface protein changes during fibroblast-myofibroblast activation. Coexistence of quiescence and activated fibroblast subtypes in the IPF lung suggests dynamic remodeling of fibroblast activation upon subtle changes to growth factor exposure in local microenvironmental niches.
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Affiliation(s)
- Katharina Heinzelmann
- Comprehensive Pneumology Center, University Hospital of the Ludwig-Maximilians University, Munich and Helmholtz Zentrum München, Member of the Comprehensive Pneumology Center-Munich BioArchive, Member of the German Center for Lung Research , Munich , Germany
| | - Mareike Lehmann
- Comprehensive Pneumology Center, University Hospital of the Ludwig-Maximilians University, Munich and Helmholtz Zentrum München, Member of the Comprehensive Pneumology Center-Munich BioArchive, Member of the German Center for Lung Research , Munich , Germany
| | - Michael Gerckens
- Comprehensive Pneumology Center, University Hospital of the Ludwig-Maximilians University, Munich and Helmholtz Zentrum München, Member of the Comprehensive Pneumology Center-Munich BioArchive, Member of the German Center for Lung Research , Munich , Germany
| | - Nina Noskovičová
- Comprehensive Pneumology Center, University Hospital of the Ludwig-Maximilians University, Munich and Helmholtz Zentrum München, Member of the Comprehensive Pneumology Center-Munich BioArchive, Member of the German Center for Lung Research , Munich , Germany
| | - Marion Frankenberger
- Comprehensive Pneumology Center, University Hospital of the Ludwig-Maximilians University, Munich and Helmholtz Zentrum München, Member of the Comprehensive Pneumology Center-Munich BioArchive, Member of the German Center for Lung Research , Munich , Germany
| | - Michael Lindner
- Comprehensive Pneumology Center, University Hospital of the Ludwig-Maximilians University, Munich and Helmholtz Zentrum München, Member of the Comprehensive Pneumology Center-Munich BioArchive, Member of the German Center for Lung Research , Munich , Germany.,Thoraxchirurgisches Zentrum München, Asklepios Fachkliniken München-Gauting, Munich , Germany
| | - Rudolf Hatz
- Thoraxchirurgisches Zentrum München, Asklepios Fachkliniken München-Gauting, Munich , Germany.,Thoraxchirurgisches Zentrum, Klinik für Allgemeine-, Viszeral-, Transplantations-, Gefäss- und Thoraxchirurgie, Klinikum Grosshadern, Ludwig-Maximilians-Universität, Munich , Germany
| | - Jürgen Behr
- Thoraxchirurgisches Zentrum München, Asklepios Fachkliniken München-Gauting, Munich , Germany.,Medizinische Klinik und Poliklinik V, Klinikum der Ludwig-Maximilians-Universität, Munich , Germany
| | - Anne Hilgendorff
- Comprehensive Pneumology Center, University Hospital of the Ludwig-Maximilians University, Munich and Helmholtz Zentrum München, Member of the Comprehensive Pneumology Center-Munich BioArchive, Member of the German Center for Lung Research , Munich , Germany.,Department of Neonatology, Perinatal Center Grosshadern, Ludwig-Maximilians University , Munich , Germany.,Center for Comprehensive Developmental Care, Dr. von Haunersches Children's Hospital University Hospital Ludwig-Maximilians University , Munich , Germany
| | - Melanie Königshoff
- Comprehensive Pneumology Center, University Hospital of the Ludwig-Maximilians University, Munich and Helmholtz Zentrum München, Member of the Comprehensive Pneumology Center-Munich BioArchive, Member of the German Center for Lung Research , Munich , Germany.,Division of Respiratory Sciences and Critical Care Medicine, University of Colorado , Denver, Colorado
| | - Oliver Eickelberg
- Comprehensive Pneumology Center, University Hospital of the Ludwig-Maximilians University, Munich and Helmholtz Zentrum München, Member of the Comprehensive Pneumology Center-Munich BioArchive, Member of the German Center for Lung Research , Munich , Germany.,Division of Respiratory Sciences and Critical Care Medicine, University of Colorado , Denver, Colorado
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39
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Wu J, Wang Y, Liu G, Jia Y, Yang J, Shi J, Dong J, Wei J, Liu X. Characterization of air-liquid interface culture of A549 alveolar epithelial cells. ACTA ACUST UNITED AC 2017; 51:e6950. [PMID: 29267508 PMCID: PMC5731333 DOI: 10.1590/1414-431x20176950] [Citation(s) in RCA: 60] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/14/2017] [Accepted: 09/27/2017] [Indexed: 12/24/2022]
Abstract
Alveolar epithelia play an essential role in maintaining the integrity and homeostasis of lungs, in which alveolar epithelial type II cells (AECII) are a cell type with stem cell potential for epithelial injury repair and regeneration. However, mechanisms behind the physiological and pathological roles of alveolar epithelia in human lungs remain largely unknown, partially owing to the difficulty of isolation and culture of primary human AECII cells. In the present study, we aimed to characterize alveolar epithelia generated from A549 lung adenocarcinoma cells that were cultured in an air-liquid interface (ALI) state. Morphological analysis demonstrated that A549 cells could reconstitute epithelial layers in ALI cultures as evaluated by histochemistry staining and electronic microscopy. Immunofluorescent staining further revealed an expression of alveolar epithelial type I cell (AECI) markers aquaporin-5 protein (AQP-5), and AECII cell marker surfactant protein C (SPC) in subpopulations of ALI cultured cells. Importantly, molecular analysis further revealed the expression of AQP-5, SPC, thyroid transcription factor-1, zonula occludens-1 and Mucin 5B in A549 ALI cultures as determined by both immunoblotting and quantitative RT-PCR assay. These results suggest that the ALI culture of A549 cells can partially mimic the property of alveolar epithelia, which may be a feasible and alternative model for investigating roles and mechanisms of alveolar epithelia in vitro.
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Affiliation(s)
- J Wu
- College of Clinical Medicine, Ningxia Medical University, Yinchuan, Ningxia, China.,Institute of Human Stem Cell Research, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China
| | - Y Wang
- College of Clinical Medicine, Ningxia Medical University, Yinchuan, Ningxia, China.,Institute of Human Stem Cell Research, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China
| | - G Liu
- College of Clinical Medicine, Ningxia Medical University, Yinchuan, Ningxia, China.,Institute of Human Stem Cell Research, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China
| | - Y Jia
- Institute of Human Stem Cell Research, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China
| | - J Yang
- Institute of Human Stem Cell Research, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China.,Center of Laboratory Medicine, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China
| | - J Shi
- Institute of Human Stem Cell Research, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China.,Center of Laboratory Medicine, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China
| | - J Dong
- Department of Pathology, Ningxia Medical University, Yinchuan, Ningxia, China
| | - J Wei
- College of Clinical Medicine, Ningxia Medical University, Yinchuan, Ningxia, China.,Institute of Human Stem Cell Research, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China.,Center of Laboratory Medicine, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China
| | - X Liu
- College of Clinical Medicine, Ningxia Medical University, Yinchuan, Ningxia, China.,Institute of Human Stem Cell Research, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China.,Center of Laboratory Medicine, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, China.,College of Life Science, Ningxia University, Yinchuan, Ningxia, China
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40
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Phenotypic screening identifies Axl kinase as a negative regulator of an alveolar epithelial cell phenotype. J Transl Med 2017; 97:1047-1062. [PMID: 28553934 DOI: 10.1038/labinvest.2017.52] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2017] [Revised: 04/02/2017] [Accepted: 04/05/2017] [Indexed: 12/11/2022] Open
Abstract
Loss of epithelial barrier integrity is implicated in a number of human lung diseases. However, the molecular pathways underlying this process are poorly understood. In a phenotypic screen, we identified Axl kinase as a negative regulator of epithelial phenotype and function. Furthermore, suppression of Axl activity by a small molecule kinase inhibitor or downregulation of Axl expression by small interfering RNA led to: (1) the increase in epithelial surfactant protein expression; (2) a cell morphology transition from front-rear polarity to cuboidal shape; (3) the cytoskeletal re-organization resulting in decreased cell mobility; and (4) the acquisition of epithelial junctions. Loss of Axl activity reduced activation of the Axl canonical pathway members, Akt and extracellular signal-regulated kinase-1/2 and resulted in the loss of gene expression of a unique profile of epithelial-to-mesenchymal transition transcription factors including SNAI2, HOXA5, TBX2 or TBX3. Finally, we observed that Axl was activated in hyperplasia of epithelial cells in idiopathic pulmonary fibrosis where epithelial barrier integrity was lost. These results suggest that the Axl kinase signaling pathway is associated with the loss integrity of alveolar epithelium in pathological remodeling of human lung diseases.
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41
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Li Y, Shi X, Yang L, Mou Y, Li Y, Dang R, Li C. Hypoxia promotes the skewed differentiation of umbilical cord mesenchymal stem cells toward type II alveolar epithelial cells by regulating microRNA-145. Gene 2017; 630:68-75. [PMID: 28789953 DOI: 10.1016/j.gene.2017.08.006] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2016] [Revised: 07/19/2017] [Accepted: 08/04/2017] [Indexed: 01/26/2023]
Abstract
Mesenchymal stem cells (MSCs) are well recognized for their ability to differentiate into type II alveolar epithelial (ATII) cells in damaged lungs, which is critical for reepithelization and recovery in acute lung injury (ALI). However, the high level of transforming growth factor-β (TGF-β) commonly seen in injured lung tissues is also able to induce MSCs to differentiate into fibroblast-like cells. In this study, we found that hypoxia could promote umbilical cord mesenchymal stem cells (UCMSCs) differentiation into ATII cells rather than into fibroblast-like cells, and this effect was mainly mediated by microRNA-145 (miR-145), which could induce the inhibition of TGF-β signaling by targeting TGF-β receptor II (TGFβRII). Clarifying the function of hypoxia in the fate determination of MSCs is important for improving stem cell-based therapies for ALI.
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Affiliation(s)
- Yang Li
- Department of Thoracic Surgery, The First Hospital of Jilin University, Changchun 130000, Jilin Province, China
| | - Xu Shi
- Central Laboratory, The First Hospital of Jilin University, Changchun 130000, Jilin Province, China
| | - Liming Yang
- Department of Nephropathy, The First Hospital of Jilin University, Changchun 130000, Jilin Province, China
| | - Yan Mou
- Department of Dermatology, The Second Hospital of Jilin University, Changchun 130000, Jilin Province, China
| | - Yingbo Li
- Central Laboratory, The First Hospital of Jilin University, Changchun 130000, Jilin Province, China
| | - Rongjing Dang
- Central Laboratory, The First Hospital of Jilin University, Changchun 130000, Jilin Province, China
| | - Changyuan Li
- Department of Thoracic Surgery, The First Hospital of Jilin University, Changchun 130000, Jilin Province, China.
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42
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de Aguiar Vallim TQ, Lee E, Merriott DJ, Goulbourne CN, Cheng J, Cheng A, Gonen A, Allen RM, Palladino END, Ford DA, Wang T, Baldán Á, Tarling EJ. ABCG1 regulates pulmonary surfactant metabolism in mice and men. J Lipid Res 2017; 58:941-954. [PMID: 28264879 DOI: 10.1194/jlr.m075101] [Citation(s) in RCA: 37] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2017] [Revised: 03/03/2017] [Indexed: 12/27/2022] Open
Abstract
Idiopathic pulmonary alveolar proteinosis (PAP) is a rare lung disease characterized by accumulation of surfactant. Surfactant synthesis and secretion are restricted to epithelial type 2 (T2) pneumocytes (also called T2 cells). Clearance of surfactant is dependent upon T2 cells and macrophages. ABCG1 is highly expressed in both T2 cells and macrophages. ABCG1-deficient mice accumulate surfactant, lamellar body-loaded T2 cells, lipid-loaded macrophages, B-1 lymphocytes, and immunoglobulins, clearly demonstrating that ABCG1 has a critical role in pulmonary homeostasis. We identify a variant in the ABCG1 promoter in patients with PAP that results in impaired activation of ABCG1 by the liver X receptor α, suggesting that ABCG1 basal expression and/or induction in response to sterol/lipid loading is essential for normal lung function. We generated mice lacking ABCG1 specifically in either T2 cells or macrophages to determine the relative contribution of these cell types on surfactant lipid homeostasis. These results establish a critical role for T2 cell ABCG1 in controlling surfactant and overall lipid homeostasis in the lung and in the pathogenesis of human lung disease.
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Affiliation(s)
- Thomas Q de Aguiar Vallim
- Department of Medicine, University of California Los Angeles, Los Angeles, CA 90095.,Department of Biological Chemistry, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA 90095.,Molecular Biology Institute, University of California Los Angeles, Los Angeles, CA 90095.,Johnson Comprehensive Cancer Center, University of California Los Angeles, Los Angeles, CA 90095
| | - Elinor Lee
- Department of Medicine, University of California Los Angeles, Los Angeles, CA 90095.,Division of Pulmonary and Critical Care Medicine, University of California Los Angeles, Los Angeles, CA 90095
| | - David J Merriott
- Department of Biological Chemistry, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA 90095
| | | | - Joan Cheng
- Department of Biological Chemistry, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA 90095
| | - Angela Cheng
- Department of Biological Chemistry, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA 90095
| | - Ayelet Gonen
- Department of Medicine, University of California San Diego, La Jolla, CA 92093
| | - Ryan M Allen
- Edward A. Doisy Department of Biochemistry and Molecular Biology, Saint Louis University, St. Louis, MO 63104
| | - Elisa N D Palladino
- Edward A. Doisy Department of Biochemistry and Molecular Biology, Saint Louis University, St. Louis, MO 63104.,Center for Cardiovascular Research, School of Medicine, Saint Louis University, St. Louis, MO 63104
| | - David A Ford
- Edward A. Doisy Department of Biochemistry and Molecular Biology, Saint Louis University, St. Louis, MO 63104.,Center for Cardiovascular Research, School of Medicine, Saint Louis University, St. Louis, MO 63104
| | - Tisha Wang
- Department of Medicine, University of California Los Angeles, Los Angeles, CA 90095.,Division of Pulmonary and Critical Care Medicine, University of California Los Angeles, Los Angeles, CA 90095
| | - Ángel Baldán
- Edward A. Doisy Department of Biochemistry and Molecular Biology, Saint Louis University, St. Louis, MO 63104
| | - Elizabeth J Tarling
- Department of Medicine, University of California Los Angeles, Los Angeles, CA 90095 .,Molecular Biology Institute, University of California Los Angeles, Los Angeles, CA 90095.,Johnson Comprehensive Cancer Center, University of California Los Angeles, Los Angeles, CA 90095
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43
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Chimenti I, Pagano F, Angelini F, Siciliano C, Mangino G, Picchio V, De Falco E, Peruzzi M, Carnevale R, Ibrahim M, Biondi‐Zoccai G, Messina E, Frati G. Human Lung Spheroids as In Vitro Niches of Lung Progenitor Cells with Distinctive Paracrine and Plasticity Properties. Stem Cells Transl Med 2017; 6:767-777. [PMID: 28297570 PMCID: PMC5442776 DOI: 10.5966/sctm.2015-0374] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2015] [Accepted: 08/09/2016] [Indexed: 02/05/2023] Open
Abstract
Basic and translational research on lung biology has discovered multiple progenitor cell types, specialized or facultative, responsible for turnover, renewal, and repair. Isolation of populations of resident lung progenitor cells (LPCs) has been described by multiple protocols, and some have been successfully applied to healthy human lung tissue. We aimed at understanding how different cell culture conditions may affect, in vitro, the phenotype of LPCs to create an ideal niche-like microenvironment. The influence of different substrates (i.e., fibronectin, gelatin, laminin) and the impact of a three-dimensional/two-dimensional (3D/2D) culture switch on the biology of LPCs isolated as lung spheroids (LSs) from normal adult human lung biopsy specimens were investigated. We applied a spheroid culture system as the selective/inductive step for progenitor cell culture, as described in many biological systems. The data showed a niche-like proepithelial microenvironment inside the LS, highly sensitive to the 3D culture system and significantly affecting the phenotype of adult LPCs more than culture substrate. LSs favor epithelial phenotypes and LPC maintenance and contain cells more responsive to specific commitment stimuli than 2D monolayer cultures, while secreting a distinctive set of paracrine factors. We have shown for the first time, to our knowledge, how culture as 3D LSs can affect LPC epithelial phenotype and produce strong paracrine signals with a distinctive secretomic profile compared with 2D monolayer conditions. These findings suggest novel approaches to maintain ex vivo LPCs for basic and translational studies. Stem Cells Translational Medicine 2017;6:767-777.
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Affiliation(s)
- Isotta Chimenti
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
| | - Francesca Pagano
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
| | - Francesco Angelini
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
| | - Camilla Siciliano
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
| | - Giorgio Mangino
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
| | - Vittorio Picchio
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
| | - Elena De Falco
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
| | - Mariangela Peruzzi
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
| | - Roberto Carnevale
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
| | - Mohsen Ibrahim
- Department of Medical‐Surgical Science and Translational Medicine, “La Sapienza” University of Rome, Rome, Italy
| | - Giuseppe Biondi‐Zoccai
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
- Department of AngioCardioNeurology, Istituto di Ricovero e Cura a Carattere Scientifico Neuromed, Pozzilli, Italy
| | - Elisa Messina
- Department of Pediatrics and Neuropsychiatry, “Umberto I” Hospital, Rome, Italy
| | - Giacomo Frati
- Department of Medical‐Surgical Sciences and Biotechnology, “Sapienza” University of Rome, Rome, Italy
- Department of AngioCardioNeurology, Istituto di Ricovero e Cura a Carattere Scientifico Neuromed, Pozzilli, Italy
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Abstract
Chronic obstructive pulmonary disease (COPD) is regarded as a disease of accelerated lung aging. This affliction shows all of the hallmarks of aging, including telomere shortening, cellular senescence, activation of PI3 kinase-mTOR signaling, impaired autophagy, mitochondrial dysfunction, stem cell exhaustion, epigenetic changes, abnormal microRNA profiles, immunosenescence, and a low-grade chronic inflammation (inflammaging). Many of these pathways are driven by chronic exogenous and endogenous oxidative stress. There is also a reduction in antiaging molecules, such as sirtuins and Klotho, which further accelerate the aging process. COPD is associated with several comorbidities (multimorbidity), such as cardiovascular and metabolic diseases, that share the same pathways of accelerated aging. Understanding these mechanisms has helped identify several novel therapeutic targets, and several drugs and dietary interventions are now in development to treat multimorbidity.
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Affiliation(s)
- Peter J. Barnes
- National Heart and Lung Institute, Imperial College, London SW3 6LY, United Kingdom
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45
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Ozawa C, Horiguchi M, Akita T, Oiso Y, Abe K, Motomura T, Yamashita C. Pulmonary Administration of GW0742, a High-Affinity Peroxisome Proliferator-Activated Receptor Agonist, Repairs Collapsed Alveoli in an Elastase-Induced Mouse Model of Emphysema. Biol Pharm Bull 2017; 39:778-85. [PMID: 27150147 DOI: 10.1248/bpb.b15-00909] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Pulmonary emphysema is a disease in which lung alveoli are irreversibly damaged, thus compromising lung function. Our previous study revealed that all-trans-retinoic acid (ATRA) induces the differentiation of human lung alveolar epithelial type 2 progenitor cells and repairs the alveoli of emphysema model mice. ATRA also reportedly has the ability to activate peroxisome proliferator-activated receptor (PPAR) β/δ. A selective PPARβ/δ ligand has been reported to induce the differentiation of human keratinocytes during wound repair. Here, we demonstrate that treatment using a high-affinity PPARβ/δ agonist, GW0742, reverses the lung tissue damage induced by elastase in emphysema-model mice and improves respiratory function. Mice treated with elastase, which collapsed their alveoli, were then treated with either 10% dimethyl sulfoxide (DMSO) in saline (control group) or GW0742 (1.0 mg/kg twice a week) by pulmonary administration. Treatment with GW0742 for 2 weeks increased the in vivo expression of surfactant proteins A and D, which are known alveolar type II epithelial cell markers. GW0742 treatment also shortened the average distance between alveolar walls in the lungs of emphysema model mice, compared with a control group treated with 10% DMSO in saline. Treatment with GW0742 for 3 weeks also improved tissue elastance (cm H2O/mL), as well as the ratio of the forced expiratory volume in the first 0.05 s to the forced vital capacity (FEV 0.05/FVC). In each of these experiments, GW0742 treatment reversed the damage caused by elastase. In conclusion, PPARβ/δ agonists are potential therapeutic agents for pulmonary emphysema.
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Affiliation(s)
- Chihiro Ozawa
- Department of Pharmaceutics and Drug Delivery, Faculty of Pharmaceutical Sciences, Tokyo University of Science
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46
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New Role of Adult Lung c-kit + Cells in a Mouse Model of Airway Hyperresponsiveness. Mediators Inflamm 2016; 2016:3917471. [PMID: 28090152 PMCID: PMC5206449 DOI: 10.1155/2016/3917471] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/22/2016] [Revised: 10/05/2016] [Accepted: 11/10/2016] [Indexed: 12/16/2022] Open
Abstract
Structural changes contribute to airway hyperresponsiveness and airflow obstruction in asthma. Emerging evidence points to the involvement of c-kit+ cells in lung homeostasis, although their potential role in asthma is unknown. Our aim was to isolate c-kit+ cells from normal mouse lungs and to test whether these cells can interfere with hallmarks of asthma in an animal model. Adult mouse GFP-tagged c-kit+ cells, intratracheally delivered in the ovalbumin-induced airway hyperresponsiveness, positively affected airway remodeling and improved airway function. In bronchoalveolar lavage fluid of cell-treated animals, a reduction in the number of inflammatory cells and in IL-4, IL-5, and IL-13 release, along with an increase of IL-10, was observed. In MSC-treated mice, the macrophage polarization to M2-like subset may explain, at least in part, the increment in the level of anti-inflammatory cytokine IL-10. After in vitro stimulation of c-kit+ cells with proinflammatory cytokines, the indoleamine 2,3-dioxygenase and TGFβ were upregulated. These data, together with the increased apoptosis of inflammatory cells in vivo, indicate that c-kit+ cells downregulate immune response in asthma by influencing local environment, possibly by cell-to-cell contact combined to paracrine action. In conclusion, intratracheally administered c-kit+ cells reduce inflammation, positively modulate airway remodeling, and improve function. These data document previously unrecognized properties of c-kit+ cells, able to impede pathophysiological features of experimental airway hyperresponsiveness.
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47
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Lung remodeling associated with recovery from acute lung injury. Cell Tissue Res 2016; 367:495-509. [DOI: 10.1007/s00441-016-2521-8] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/29/2016] [Accepted: 09/29/2016] [Indexed: 12/18/2022]
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48
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Yamada M, Fujino N, Ichinose M. Inflammatory responses in the initiation of lung repair and regeneration: their role in stimulating lung resident stem cells. Inflamm Regen 2016; 36:15. [PMID: 29259688 PMCID: PMC5725654 DOI: 10.1186/s41232-016-0020-7] [Citation(s) in RCA: 33] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2016] [Accepted: 06/10/2016] [Indexed: 12/12/2022] Open
Abstract
The lungs are the primary organs for respiration, the process by which carbon dioxide and oxygen are exchanged. The alveolus, which is the site of gas exchange in the lungs, consists of multiple cell types including alveolar epithelial cells, lung capillary endothelial cells and fibroblasts. Because of their complexity, lung parenchymal cells including epithelial lineage have been thought to have a lower rate of cellular turnover in adult lung. However, accumulating observations suggest that the turnover of parenchymal cells in adult lungs is essential for maintaining homeostasis during the steady state as well as for the repair and regeneration after lung injury. After lung injury by harmful pathogens, inflammation occurs to protect the host. Although excessive inflammation damages lung tissue, inflammatory cells are essential for regeneration because they remove harmful pathogens as well as debris derived from apoptotic and necrotic cells. In addition, subsets of inflammatory cells, especially phagocytic monocytes, produce cytokines and growth factors to resolve inflammation and promote tissue regeneration by stimulating tissue-resident stem cells. Recent advances in the biology of lung-resident stem cells, especially those addressing epithelial lineage, have revealed that there are several cellular populations capable of self-renewal that can differentiate into airway and/or alveolar epithelial cells. A part of these populations does not exist in the steady state but emerges after lung injury, suggesting that signals induced by inflammation may play an important role in initiating the proliferation and differentiation of lung stem or progenitor cells. Understanding the interaction between inflammatory responses and tissue-resident stem cells would help elucidate the pathogenesis of inflammatory lung diseases and promote the discovery of new therapeutic targets.
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Affiliation(s)
- Mitsuhiro Yamada
- Department of Respiratory Medicine, Graduate School of Medicine, Tohoku University, 1-1 Seiryo-machi, Aoba-ku, Sendai, 980-8574 Japan
| | - Naoya Fujino
- Department of Respiratory Medicine, Graduate School of Medicine, Tohoku University, 1-1 Seiryo-machi, Aoba-ku, Sendai, 980-8574 Japan
| | - Masakazu Ichinose
- Department of Respiratory Medicine, Graduate School of Medicine, Tohoku University, 1-1 Seiryo-machi, Aoba-ku, Sendai, 980-8574 Japan
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49
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Ando K, Fujino N, Mitani K, Ota C, Okada Y, Kondo T, Mizobuchi T, Kurihara M, Suzuki K, Hoshika Y, Ebana H, Kobayashi E, Takahashi K, Kubo H, Seyama K. Isolation of individual cellular components from lung tissues of patients with lymphangioleiomyomatosis. Am J Physiol Lung Cell Mol Physiol 2016; 310:L899-908. [PMID: 27016587 DOI: 10.1152/ajplung.00365.2015] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2015] [Accepted: 03/19/2016] [Indexed: 01/30/2023] Open
Abstract
Lymphangioleiomyomatosis (LAM) is a rare neoplastic disease entailing cystic destruction of the lungs and progressive respiratory failure. LAM lungs are histologically characterized by the proliferation of smooth muscle-like cells (LAM cells) and an abundance of lymphatic vessels. To elucidate the pathophysiological processes of LAM, cell-type-specific analyses are required. However, no method exists for isolating the individual types of cells in LAM lesions. Therefore, we established a fluorescence-activated cell sorting (FACS)-based method for the direct isolation of LAM cells and other various cellular components from LAM-affected lung tissue. We obtained LAM-affected lung tissue from resections or transplant recipients and prepared single-cell suspensions. FACS, immunohistochemical, and molecular analysis were used cooperatively to isolate HMB45-positive LAM cells with tuberous sclerosis complex (TSC) 2 loss of heterozygosity (LOH). Using a combination of antibodies against an epithelial cell adhesion molecule (EpCAM) and podoplanin, we fractionated CD45-negative lung cells into three groups: lymphatic endothelial cells (LEC) (EpCAM(-)/podoplanin(hi) subset), alveolar type II cells (EpCAM(hi)/podoplanin(-) subset), and mesenchymal cells (EpCAM(-)/podoplanin(-/low) subset). During subsequent analysis of HMB45 expression, as a LAM-specific marker, we clearly identified LAM cells in the mesenchymal cell population. We then discovered that CD90(+)/CD34(-) cells in the mesenchymal cell population are not only positive for HBM45 but also had TSC2 LOH. These isolated cells were viable and subsequently amenable to cell culture. This method enables us to isolate LAM cells and other cellular components, including LAM-associated LEC, from LAM-affected lung tissues, providing new research opportunities in this field.
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Affiliation(s)
- Katsutoshi Ando
- Division of Respiratory Medicine, Juntendo University Faculty of Medicine and Graduate School of Medicine, Tokyo, Japan;
| | - Naoya Fujino
- Department of Advanced Preventive Medicine for Infectious Disease, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Keiko Mitani
- Division of Human Pathology, Juntendo University Faculty of Medicine and Graduate School of Medicine, Tokyo, Japan
| | - Chiharu Ota
- Department of Advanced Preventive Medicine for Infectious Disease, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Yoshinori Okada
- Department of Thoracic Surgery, Institute of Development, Aging, and Cancer, Tohoku University Graduate School of Medicine, Miyagi, Japan
| | - Takashi Kondo
- Department of Thoracic Surgery, Institute of Development, Aging, and Cancer, Tohoku University Graduate School of Medicine, Miyagi, Japan
| | - Teruaki Mizobuchi
- Pneumothorax Research Center and Division of Thoracic Surgery, Nissan Tamagawa Hospital, Tokyo, Japan; and
| | - Masatoshi Kurihara
- Pneumothorax Research Center and Division of Thoracic Surgery, Nissan Tamagawa Hospital, Tokyo, Japan; and
| | - Kenji Suzuki
- Division of Thoracic Surgery, Juntendo University Faculty of Medicine and Graduate School of Medicine, Bunkyo-Ku, Tokyo, Japan
| | - Yoshito Hoshika
- Division of Respiratory Medicine, Juntendo University Faculty of Medicine and Graduate School of Medicine, Tokyo, Japan
| | - Hiroki Ebana
- Division of Respiratory Medicine, Juntendo University Faculty of Medicine and Graduate School of Medicine, Tokyo, Japan
| | - Etsuko Kobayashi
- Division of Respiratory Medicine, Juntendo University Faculty of Medicine and Graduate School of Medicine, Tokyo, Japan
| | - Kazuhisa Takahashi
- Division of Respiratory Medicine, Juntendo University Faculty of Medicine and Graduate School of Medicine, Tokyo, Japan
| | - Hiroshi Kubo
- Department of Advanced Preventive Medicine for Infectious Disease, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Kuniaki Seyama
- Division of Respiratory Medicine, Juntendo University Faculty of Medicine and Graduate School of Medicine, Tokyo, Japan
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50
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An official American Thoracic Society workshop report: stem cells and cell therapies in lung biology and diseases. Ann Am Thorac Soc 2016; 12:S79-97. [PMID: 25897748 DOI: 10.1513/annalsats.201502-086st] [Citation(s) in RCA: 22] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
The University of Vermont College of Medicine and the Vermont Lung Center, in collaboration with the NHLBI, Alpha-1 Foundation, American Thoracic Society, European Respiratory Society, International Society for Cell Therapy, and the Pulmonary Fibrosis Foundation, convened a workshop, "Stem Cells and Cell Therapies in Lung Biology and Lung Diseases," held July 29 to August 1, 2013 at the University of Vermont. The conference objectives were to review the current understanding of the role of stem and progenitor cells in lung repair after injury and to review the current status of cell therapy and ex vivo bioengineering approaches for lung diseases. These are all rapidly expanding areas of study that both provide further insight into and challenge traditional views of mechanisms of lung repair after injury and pathogenesis of several lung diseases. The goals of the conference were to summarize the current state of the field, discuss and debate current controversies, and identify future research directions and opportunities for both basic and translational research in cell-based therapies for lung diseases. This conference was a follow-up to four previous biennial conferences held at the University of Vermont in 2005, 2007, 2009, and 2011. Each of those conferences, also sponsored by the National Institutes of Health, American Thoracic Society, and Respiratory Disease Foundations, has been important in helping guide research and funding priorities. The major conference recommendations are summarized at the end of the report and highlight both the significant progress and major challenges in these rapidly progressing fields.
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