A Novel Fibroblast Reporter Cell Line for in vitro Studies of Pulmonary Fibrosis
Idiopathic pulmonary fibrosis (IPF) is a fatal disease of the lower respiratory tract with restricted therapeutic options. Repetitive injury of the bronchoalveolar epithelium leads to activation of pulmonary fibroblasts, differentiation into myofibroblasts and excessive extracellular matrix (ECM) de...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2020-10-01
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| Series: | Frontiers in Physiology |
| Subjects: | |
| Online Access: | https://www.frontiersin.org/article/10.3389/fphys.2020.567675/full |
| _version_ | 1818890409200320512 |
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| author | Julia Nemeth Annika Schundner Karsten Quast Veronika E. Winkelmann Manfred Frick |
| author_facet | Julia Nemeth Annika Schundner Karsten Quast Veronika E. Winkelmann Manfred Frick |
| author_sort | Julia Nemeth |
| collection | DOAJ |
| description | Idiopathic pulmonary fibrosis (IPF) is a fatal disease of the lower respiratory tract with restricted therapeutic options. Repetitive injury of the bronchoalveolar epithelium leads to activation of pulmonary fibroblasts, differentiation into myofibroblasts and excessive extracellular matrix (ECM) deposition resulting in aberrant wound repair. However, detailed molecular and cellular mechanisms underlying initiation and progression of fibrotic changes are still elusive. Here, we report the generation of a representative fibroblast reporter cell line (10-4ABFP) to study pathophysiological mechanisms of IPF in high throughput or high resolution in vitro live cell assays. To this end, we immortalized primary fibroblasts isolated from the distal lung of Sprague-Dawley rats. Molecular and transcriptomic characterization identified clone 10-4A as a matrix fibroblast subpopulation. Mechanical or chemical stimulation induced a reversible fibrotic state comparable to effects observed in primary isolated fibroblasts. Finally, we generated a reporter cell line (10-4ABFP) to express nuclear blue fluorescent protein (BFP) under the promotor of the myofibroblast marker alpha smooth muscle actin (Acta2) using CRISPR/Cas9 technology. We evaluated the suitability of 10-4ABFP as reporter tool in plate reader assays. In summary, the 10-4ABFP cell line provides a novel tool to study fibrotic processes in vitro to gain new insights into the cellular and molecular processes involved in fibrosis formation and propagation. |
| first_indexed | 2024-12-19T17:24:27Z |
| format | Article |
| id | doaj.art-6c75d968925a4f5998087629576e6e89 |
| institution | Directory Open Access Journal |
| issn | 1664-042X |
| language | English |
| last_indexed | 2024-12-19T17:24:27Z |
| publishDate | 2020-10-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Physiology |
| spelling | doaj.art-6c75d968925a4f5998087629576e6e892022-12-21T20:12:36ZengFrontiers Media S.A.Frontiers in Physiology1664-042X2020-10-011110.3389/fphys.2020.567675567675A Novel Fibroblast Reporter Cell Line for in vitro Studies of Pulmonary FibrosisJulia Nemeth0Annika Schundner1Karsten Quast2Veronika E. Winkelmann3Manfred Frick4Institute of General Physiology, Ulm University, Ulm, GermanyInstitute of General Physiology, Ulm University, Ulm, GermanyBoehringer Ingelheim Pharma GmbH & Co. KG, Biberach, GermanyInstitute of General Physiology, Ulm University, Ulm, GermanyInstitute of General Physiology, Ulm University, Ulm, GermanyIdiopathic pulmonary fibrosis (IPF) is a fatal disease of the lower respiratory tract with restricted therapeutic options. Repetitive injury of the bronchoalveolar epithelium leads to activation of pulmonary fibroblasts, differentiation into myofibroblasts and excessive extracellular matrix (ECM) deposition resulting in aberrant wound repair. However, detailed molecular and cellular mechanisms underlying initiation and progression of fibrotic changes are still elusive. Here, we report the generation of a representative fibroblast reporter cell line (10-4ABFP) to study pathophysiological mechanisms of IPF in high throughput or high resolution in vitro live cell assays. To this end, we immortalized primary fibroblasts isolated from the distal lung of Sprague-Dawley rats. Molecular and transcriptomic characterization identified clone 10-4A as a matrix fibroblast subpopulation. Mechanical or chemical stimulation induced a reversible fibrotic state comparable to effects observed in primary isolated fibroblasts. Finally, we generated a reporter cell line (10-4ABFP) to express nuclear blue fluorescent protein (BFP) under the promotor of the myofibroblast marker alpha smooth muscle actin (Acta2) using CRISPR/Cas9 technology. We evaluated the suitability of 10-4ABFP as reporter tool in plate reader assays. In summary, the 10-4ABFP cell line provides a novel tool to study fibrotic processes in vitro to gain new insights into the cellular and molecular processes involved in fibrosis formation and propagation.https://www.frontiersin.org/article/10.3389/fphys.2020.567675/fullidiopathic pulmonary fibrosislungmyofibroblastTGF-βextracellular matrixalpha smooth muscle actin |
| spellingShingle | Julia Nemeth Annika Schundner Karsten Quast Veronika E. Winkelmann Manfred Frick A Novel Fibroblast Reporter Cell Line for in vitro Studies of Pulmonary Fibrosis Frontiers in Physiology idiopathic pulmonary fibrosis lung myofibroblast TGF-β extracellular matrix alpha smooth muscle actin |
| title | A Novel Fibroblast Reporter Cell Line for in vitro Studies of Pulmonary Fibrosis |
| title_full | A Novel Fibroblast Reporter Cell Line for in vitro Studies of Pulmonary Fibrosis |
| title_fullStr | A Novel Fibroblast Reporter Cell Line for in vitro Studies of Pulmonary Fibrosis |
| title_full_unstemmed | A Novel Fibroblast Reporter Cell Line for in vitro Studies of Pulmonary Fibrosis |
| title_short | A Novel Fibroblast Reporter Cell Line for in vitro Studies of Pulmonary Fibrosis |
| title_sort | novel fibroblast reporter cell line for in vitro studies of pulmonary fibrosis |
| topic | idiopathic pulmonary fibrosis lung myofibroblast TGF-β extracellular matrix alpha smooth muscle actin |
| url | https://www.frontiersin.org/article/10.3389/fphys.2020.567675/full |
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