A perfusion-independent high-throughput method to isolate liver sinusoidal endothelial cells
Abstract Liver sinusoidal endothelial cells (LSECs) critically regulate homeostatic liver function and liver pathogenesis. However, the isolation of LSECs remains a major technological bottleneck in studying molecular mechanisms governing LSEC functions. Current techniques to isolate LSECs, relying...
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Nature Portfolio
2025-01-01
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| Series: | Communications Biology |
| Online Access: | https://doi.org/10.1038/s42003-025-07458-5 |
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| author | Anna Babin-Ebell Gonçalves Yifang Mao Tinja Baljkas Felix Wiedmann Larissa Eis Franziska Pilz Manuel Winkler Sina W. Kürschner-Zacharias Marlene Hoffarth Charlotta Funaya Réza Shahidi Cyrill Géraud Chi-Chung Wu Constanze Schmidt Sergij Goerdt Philipp-Sebastian Reiners-Koch Mahak Singhal |
| author_facet | Anna Babin-Ebell Gonçalves Yifang Mao Tinja Baljkas Felix Wiedmann Larissa Eis Franziska Pilz Manuel Winkler Sina W. Kürschner-Zacharias Marlene Hoffarth Charlotta Funaya Réza Shahidi Cyrill Géraud Chi-Chung Wu Constanze Schmidt Sergij Goerdt Philipp-Sebastian Reiners-Koch Mahak Singhal |
| author_sort | Anna Babin-Ebell Gonçalves |
| collection | DOAJ |
| description | Abstract Liver sinusoidal endothelial cells (LSECs) critically regulate homeostatic liver function and liver pathogenesis. However, the isolation of LSECs remains a major technological bottleneck in studying molecular mechanisms governing LSEC functions. Current techniques to isolate LSECs, relying on perfusion-dependent liver digestion, are cumbersome with limited throughput. We here describe a perfusion-independent high-throughput procedure to isolate LSECs with high purity. Indifferently from previous perfusion-independent approaches, chopped liver tissue was incubated in the digestion mix for 30 minutes with intermittent mixing with a serological pipette. This led to the safeguarding of LSEC integrity and yielded 10 ± 1.0 million LSECs per adult mouse liver, which is far higher than previous perfusion-independent protocols and comparable yield to established perfusion-dependent protocols for isolating LSECs. Combining magnetic and fluorescence-activated cell sorting (FACS), LSECs from different zones of the hepatic sinusoid can now be isolated in high numbers in less than two hours for downstream applications including proteomics. Our protocol enables the isolation of LSECs from fibrotic liver tissues from mice and healthy liver tissues from higher vertebrate species (pigs), where traditional perfusion-based digestion protocols have very limited application. In conclusion, these technical advancements reduce post-mortem changes in the LSEC state and aid in reliable investigation of LSEC functions. |
| format | Article |
| id | doaj-art-fe5f4f0a2fe844b59898f9f8b579f4ee |
| institution | DOAJ |
| issn | 2399-3642 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Communications Biology |
| spelling | doaj-art-fe5f4f0a2fe844b59898f9f8b579f4ee2025-08-20T02:40:36ZengNature PortfolioCommunications Biology2399-36422025-01-018111010.1038/s42003-025-07458-5A perfusion-independent high-throughput method to isolate liver sinusoidal endothelial cellsAnna Babin-Ebell Gonçalves0Yifang Mao1Tinja Baljkas2Felix Wiedmann3Larissa Eis4Franziska Pilz5Manuel Winkler6Sina W. Kürschner-Zacharias7Marlene Hoffarth8Charlotta Funaya9Réza Shahidi10Cyrill Géraud11Chi-Chung Wu12Constanze Schmidt13Sergij Goerdt14Philipp-Sebastian Reiners-Koch15Mahak Singhal16AngioRhythms in Health and Disease, European Center for Angioscience (ECAS), Medical Faculty Mannheim, Heidelberg UniversityAngioRhythms in Health and Disease, European Center for Angioscience (ECAS), Medical Faculty Mannheim, Heidelberg UniversityDepartment of Dermatology, Venereology and Allergology, Center of Excellence in Dermatology, University Medical Center and Medical Faculty Mannheim, Heidelberg UniversityDepartment of Cardiology, University Medical Center and Medical Faculty Heidelberg, Heidelberg UniversityElectron Microscopy Core Facility, Heidelberg UniversityAngioRhythms in Health and Disease, European Center for Angioscience (ECAS), Medical Faculty Mannheim, Heidelberg UniversityDepartment of Dermatology, Venereology and Allergology, Center of Excellence in Dermatology, University Medical Center and Medical Faculty Mannheim, Heidelberg UniversityDepartment of Dermatology, Venereology and Allergology, Center of Excellence in Dermatology, University Medical Center and Medical Faculty Mannheim, Heidelberg UniversityAngioRhythms in Health and Disease, European Center for Angioscience (ECAS), Medical Faculty Mannheim, Heidelberg UniversityElectron Microscopy Core Facility, Heidelberg UniversityElectron Microscopy Core Facility, Heidelberg UniversityDepartment of Dermatology, Venereology and Allergology, Center of Excellence in Dermatology, University Medical Center and Medical Faculty Mannheim, Heidelberg UniversityHelmholtz-Institute for Translational AngioCardioScience (HI-TAC) of the Max Delbrück Center for Molecular Medicine in the Helmholtz Association (MDC) at Heidelberg UniversityDepartment of Cardiology, University Medical Center and Medical Faculty Heidelberg, Heidelberg UniversityDepartment of Dermatology, Venereology and Allergology, Center of Excellence in Dermatology, University Medical Center and Medical Faculty Mannheim, Heidelberg UniversityDepartment of Dermatology, Venereology and Allergology, Center of Excellence in Dermatology, University Medical Center and Medical Faculty Mannheim, Heidelberg UniversityAngioRhythms in Health and Disease, European Center for Angioscience (ECAS), Medical Faculty Mannheim, Heidelberg UniversityAbstract Liver sinusoidal endothelial cells (LSECs) critically regulate homeostatic liver function and liver pathogenesis. However, the isolation of LSECs remains a major technological bottleneck in studying molecular mechanisms governing LSEC functions. Current techniques to isolate LSECs, relying on perfusion-dependent liver digestion, are cumbersome with limited throughput. We here describe a perfusion-independent high-throughput procedure to isolate LSECs with high purity. Indifferently from previous perfusion-independent approaches, chopped liver tissue was incubated in the digestion mix for 30 minutes with intermittent mixing with a serological pipette. This led to the safeguarding of LSEC integrity and yielded 10 ± 1.0 million LSECs per adult mouse liver, which is far higher than previous perfusion-independent protocols and comparable yield to established perfusion-dependent protocols for isolating LSECs. Combining magnetic and fluorescence-activated cell sorting (FACS), LSECs from different zones of the hepatic sinusoid can now be isolated in high numbers in less than two hours for downstream applications including proteomics. Our protocol enables the isolation of LSECs from fibrotic liver tissues from mice and healthy liver tissues from higher vertebrate species (pigs), where traditional perfusion-based digestion protocols have very limited application. In conclusion, these technical advancements reduce post-mortem changes in the LSEC state and aid in reliable investigation of LSEC functions.https://doi.org/10.1038/s42003-025-07458-5 |
| spellingShingle | Anna Babin-Ebell Gonçalves Yifang Mao Tinja Baljkas Felix Wiedmann Larissa Eis Franziska Pilz Manuel Winkler Sina W. Kürschner-Zacharias Marlene Hoffarth Charlotta Funaya Réza Shahidi Cyrill Géraud Chi-Chung Wu Constanze Schmidt Sergij Goerdt Philipp-Sebastian Reiners-Koch Mahak Singhal A perfusion-independent high-throughput method to isolate liver sinusoidal endothelial cells Communications Biology |
| title | A perfusion-independent high-throughput method to isolate liver sinusoidal endothelial cells |
| title_full | A perfusion-independent high-throughput method to isolate liver sinusoidal endothelial cells |
| title_fullStr | A perfusion-independent high-throughput method to isolate liver sinusoidal endothelial cells |
| title_full_unstemmed | A perfusion-independent high-throughput method to isolate liver sinusoidal endothelial cells |
| title_short | A perfusion-independent high-throughput method to isolate liver sinusoidal endothelial cells |
| title_sort | perfusion independent high throughput method to isolate liver sinusoidal endothelial cells |
| url | https://doi.org/10.1038/s42003-025-07458-5 |
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