Cold storage of human precision-cut lung slices in TiProtec preserves cellular composition and transcriptional responses and enables on-demand mechanistic studies
Abstract Background Human precision-cut lung slices (hPCLS) are a unique platform for functional, mechanistic, and drug discovery studies in the field of respiratory research. However, tissue availability, generation, and cultivation time represent important challenges for their usage. Therefore, th...
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| Format: | Article |
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BMC
2025-02-01
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| Series: | Respiratory Research |
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| Online Access: | https://doi.org/10.1186/s12931-025-03132-w |
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| author | M. Camila Melo-Narvaez Fee Gölitz Eshita Jain Janine Gote-Schniering Mircea Gabriel Stoleriu Wilhelm Bertrams Bernd Schmeck Ali Önder Yildirim Ursula Rauen Timo Wille Mareike Lehmann |
| author_facet | M. Camila Melo-Narvaez Fee Gölitz Eshita Jain Janine Gote-Schniering Mircea Gabriel Stoleriu Wilhelm Bertrams Bernd Schmeck Ali Önder Yildirim Ursula Rauen Timo Wille Mareike Lehmann |
| author_sort | M. Camila Melo-Narvaez |
| collection | DOAJ |
| description | Abstract Background Human precision-cut lung slices (hPCLS) are a unique platform for functional, mechanistic, and drug discovery studies in the field of respiratory research. However, tissue availability, generation, and cultivation time represent important challenges for their usage. Therefore, the present study evaluated the efficacy of a specifically designed tissue preservation solution, TiProtec, complete or in absence (-) of iron chelators, for long-term cold storage of hPCLS. Methods hPCLS were generated from peritumor control tissues and stored in DMEM/F-12, TiProtec, or TiProtec (-) for up to 28 days. Viability, metabolic activity, and tissue structure were determined. Moreover, bulk-RNA sequencing was used to study transcriptional changes, regulated signaling pathways, and cellular composition after cold storage. Induction of cold storage-associated senescence was determined by transcriptomics and immunofluorescence (IF). Finally, cold-stored hPCLS were exposed to a fibrotic cocktail and early fibrotic changes were assessed by RT-qPCR and IF. Results Here, we found that TiProtec preserves the viability, metabolic activity, transcriptional profile, as well as cellular composition of hPCLS for up to 14 days. Cold storage did not significantly induce cellular senescence in hPCLS. Moreover, TiProtec downregulated pathways associated with cell death, inflammation, and hypoxia while activating pathways protective against oxidative stress. Cold-stored hPCLS remained responsive to fibrotic stimuli and upregulated extracellular matrix-related genes such as fibronectin and collagen 1 as well as alpha-smooth muscle actin, a marker for myofibroblasts. Conclusions Optimized long-term cold storage of hPCLS preserves their viability, metabolic activity, transcriptional profile, and cellular composition for up to 14 days, specifically in TiProtec. Finally, our study demonstrated that cold-stored hPCLS can be used for on-demand mechanistic studies relevant for respiratory research. Graphical Abstract |
| format | Article |
| id | doaj-art-0011bbccb2c245f5866a719cf7b83bd2 |
| institution | DOAJ |
| issn | 1465-993X |
| language | English |
| publishDate | 2025-02-01 |
| publisher | BMC |
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| series | Respiratory Research |
| spelling | doaj-art-0011bbccb2c245f5866a719cf7b83bd22025-08-20T03:13:14ZengBMCRespiratory Research1465-993X2025-02-0126111910.1186/s12931-025-03132-wCold storage of human precision-cut lung slices in TiProtec preserves cellular composition and transcriptional responses and enables on-demand mechanistic studiesM. Camila Melo-Narvaez0Fee Gölitz1Eshita Jain2Janine Gote-Schniering3Mircea Gabriel Stoleriu4Wilhelm Bertrams5Bernd Schmeck6Ali Önder Yildirim7Ursula Rauen8Timo Wille9Mareike Lehmann10Comprehensive Pneumology Center with the CPC-M bioArchive and Institute of Lung Health and Immunity, Helmholtz Center Munich, German Center for Lung Research (DZL)Bundeswehr Institute of Pharmacology and ToxicologyComprehensive Pneumology Center with the CPC-M bioArchive and Institute of Lung Health and Immunity, Helmholtz Center Munich, German Center for Lung Research (DZL)Department of Rheumatology and Immunology, Department of Pulmonary Medicine, Allergology and Clinical Immunology, Inselspital, Bern University Hospital, University of BernComprehensive Pneumology Center with the CPC-M bioArchive and Institute of Lung Health and Immunity, Helmholtz Center Munich, German Center for Lung Research (DZL)Institute for Lung Research, Philipps-University Marburg, German Center for Lung Research (DZL)Institute for Lung Research, Philipps-University Marburg, German Center for Lung Research (DZL)Comprehensive Pneumology Center with the CPC-M bioArchive and Institute of Lung Health and Immunity, Helmholtz Center Munich, German Center for Lung Research (DZL)Institute of Physiological Chemistry, University Hospital EssenBundeswehr Institute of Pharmacology and ToxicologyComprehensive Pneumology Center with the CPC-M bioArchive and Institute of Lung Health and Immunity, Helmholtz Center Munich, German Center for Lung Research (DZL)Abstract Background Human precision-cut lung slices (hPCLS) are a unique platform for functional, mechanistic, and drug discovery studies in the field of respiratory research. However, tissue availability, generation, and cultivation time represent important challenges for their usage. Therefore, the present study evaluated the efficacy of a specifically designed tissue preservation solution, TiProtec, complete or in absence (-) of iron chelators, for long-term cold storage of hPCLS. Methods hPCLS were generated from peritumor control tissues and stored in DMEM/F-12, TiProtec, or TiProtec (-) for up to 28 days. Viability, metabolic activity, and tissue structure were determined. Moreover, bulk-RNA sequencing was used to study transcriptional changes, regulated signaling pathways, and cellular composition after cold storage. Induction of cold storage-associated senescence was determined by transcriptomics and immunofluorescence (IF). Finally, cold-stored hPCLS were exposed to a fibrotic cocktail and early fibrotic changes were assessed by RT-qPCR and IF. Results Here, we found that TiProtec preserves the viability, metabolic activity, transcriptional profile, as well as cellular composition of hPCLS for up to 14 days. Cold storage did not significantly induce cellular senescence in hPCLS. Moreover, TiProtec downregulated pathways associated with cell death, inflammation, and hypoxia while activating pathways protective against oxidative stress. Cold-stored hPCLS remained responsive to fibrotic stimuli and upregulated extracellular matrix-related genes such as fibronectin and collagen 1 as well as alpha-smooth muscle actin, a marker for myofibroblasts. Conclusions Optimized long-term cold storage of hPCLS preserves their viability, metabolic activity, transcriptional profile, and cellular composition for up to 14 days, specifically in TiProtec. Finally, our study demonstrated that cold-stored hPCLS can be used for on-demand mechanistic studies relevant for respiratory research. Graphical Abstracthttps://doi.org/10.1186/s12931-025-03132-wHuman precision-cut lung slices (hPCLS)3RLong-term cold storageTissue preservationFibrosisHuman lung models |
| spellingShingle | M. Camila Melo-Narvaez Fee Gölitz Eshita Jain Janine Gote-Schniering Mircea Gabriel Stoleriu Wilhelm Bertrams Bernd Schmeck Ali Önder Yildirim Ursula Rauen Timo Wille Mareike Lehmann Cold storage of human precision-cut lung slices in TiProtec preserves cellular composition and transcriptional responses and enables on-demand mechanistic studies Respiratory Research Human precision-cut lung slices (hPCLS) 3R Long-term cold storage Tissue preservation Fibrosis Human lung models |
| title | Cold storage of human precision-cut lung slices in TiProtec preserves cellular composition and transcriptional responses and enables on-demand mechanistic studies |
| title_full | Cold storage of human precision-cut lung slices in TiProtec preserves cellular composition and transcriptional responses and enables on-demand mechanistic studies |
| title_fullStr | Cold storage of human precision-cut lung slices in TiProtec preserves cellular composition and transcriptional responses and enables on-demand mechanistic studies |
| title_full_unstemmed | Cold storage of human precision-cut lung slices in TiProtec preserves cellular composition and transcriptional responses and enables on-demand mechanistic studies |
| title_short | Cold storage of human precision-cut lung slices in TiProtec preserves cellular composition and transcriptional responses and enables on-demand mechanistic studies |
| title_sort | cold storage of human precision cut lung slices in tiprotec preserves cellular composition and transcriptional responses and enables on demand mechanistic studies |
| topic | Human precision-cut lung slices (hPCLS) 3R Long-term cold storage Tissue preservation Fibrosis Human lung models |
| url | https://doi.org/10.1186/s12931-025-03132-w |
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