3D Spheroids — a Cellular Model for Studying the Effects of Hypoxia on the Epicardial Microenvironment
Fundamental research in recent years has allowed us to reassess the molecular and cellular mechanisms of cardiac ontogenesis and its repair after damage. The epicardium, the outer, tightly adjoining layer of the cardiac wall formed by epicardial mesothelial cells, collagen and elastic fibers, has gai...
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Federal Research and Clinical Center of Intensive Care Medicine and Rehabilitology, Moscow, Russia
2023-02-01
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| Series: | Общая реаниматология |
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| Online Access: | https://www.reanimatology.com/rmt/article/view/2292 |
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| author | K. V. Dergilev Z. I. Tsokolaeva I. B. Beloglazova D. O. Traktuev M. T. Rasulova E. V. Parfenova |
| author_facet | K. V. Dergilev Z. I. Tsokolaeva I. B. Beloglazova D. O. Traktuev M. T. Rasulova E. V. Parfenova |
| author_sort | K. V. Dergilev |
| collection | DOAJ |
| description | Fundamental research in recent years has allowed us to reassess the molecular and cellular mechanisms of cardiac ontogenesis and its repair after damage. The epicardium, the outer, tightly adjoining layer of the cardiac wall formed by epicardial mesothelial cells, collagen and elastic fibers, has gained special relevance as an important participant of reparative processes. Better insight into poorly understood epicardial function is challenged due to anatomical issues and lack of relevant cellular models.The aim of this study was to develop a spheroid 3D model of the epicardial microenvironment and determine responses of spheroids to hypoxia.Materials and methods. Spheroids were harvested in V-shaped culture dishes with a low adhesion coating. Immunofluorescent staining of cryosections, histological methods and real-time PCR were used for characterization of cultured spheroids.Results. We demonstrated that cultivation of cells under low adhesion conditions in V-shaped culture dishes resulted in the formation of spheroids with an average size of 136+21 µm and cell viability rates of over 98%. The cells in the spheroids cultured under normoxic conditions formed tight junctions and were characterized by a low level of proliferation and the ability to synthesize extracellular matrix proteins. Under hypoxia cells in the spheroids showed partial loss of intercellular contacts, acquired a spindle shape, started to express HIF1a, SNAIL, COL1Al and accumulate collagen. All these features demonstrated the activation of mesothelial(endothelial)-mesenchymal transition strongly resembling epicardial cellular responses to ischemia in vivo.Conclusion. An epicardial spheroid cell culture model suitable for study cellular responses to hypoxic environment was developed. This model can be used to clarify mechanisms regulating epicardial microenvironment and test new targeted candidate drugs. |
| format | Article |
| id | doaj-art-45abb70fae994e5d8b006c42550bc745 |
| institution | DOAJ |
| issn | 1813-9779 2411-7110 |
| language | English |
| publishDate | 2023-02-01 |
| publisher | Federal Research and Clinical Center of Intensive Care Medicine and Rehabilitology, Moscow, Russia |
| record_format | Article |
| series | Общая реаниматология |
| spelling | doaj-art-45abb70fae994e5d8b006c42550bc7452025-08-20T03:18:33ZengFederal Research and Clinical Center of Intensive Care Medicine and Rehabilitology, Moscow, RussiaОбщая реаниматология1813-97792411-71102023-02-01191434910.15360/1813-9779-2023-1-229218893D Spheroids — a Cellular Model for Studying the Effects of Hypoxia on the Epicardial MicroenvironmentK. V. Dergilev0Z. I. Tsokolaeva1I. B. Beloglazova2D. O. Traktuev3M. T. Rasulova4E. V. Parfenova5Acad. Chazov National Medical Research Center for Cardiology, Experimental Cardiology InstituteAcad. Chazov National Medical Research Center for Cardiology, Experimental Cardiology Institute; V. A. Negovsky Research Institute of General Reanimatology, Federal Research and Clinical Center of Intensive Care Medicine and RehabilitologyAcad. Chazov National Medical Research Center for Cardiology, Experimental Cardiology InstituteRegenerative Medicine Department, Center for Cardiovascular Medicine, Florida Medical InstituteFergana Medical Institute for Public HealthAcad. Chazov National Medical Research Center for Cardiology, Experimental Cardiology InstituteFundamental research in recent years has allowed us to reassess the molecular and cellular mechanisms of cardiac ontogenesis and its repair after damage. The epicardium, the outer, tightly adjoining layer of the cardiac wall formed by epicardial mesothelial cells, collagen and elastic fibers, has gained special relevance as an important participant of reparative processes. Better insight into poorly understood epicardial function is challenged due to anatomical issues and lack of relevant cellular models.The aim of this study was to develop a spheroid 3D model of the epicardial microenvironment and determine responses of spheroids to hypoxia.Materials and methods. Spheroids were harvested in V-shaped culture dishes with a low adhesion coating. Immunofluorescent staining of cryosections, histological methods and real-time PCR were used for characterization of cultured spheroids.Results. We demonstrated that cultivation of cells under low adhesion conditions in V-shaped culture dishes resulted in the formation of spheroids with an average size of 136+21 µm and cell viability rates of over 98%. The cells in the spheroids cultured under normoxic conditions formed tight junctions and were characterized by a low level of proliferation and the ability to synthesize extracellular matrix proteins. Under hypoxia cells in the spheroids showed partial loss of intercellular contacts, acquired a spindle shape, started to express HIF1a, SNAIL, COL1Al and accumulate collagen. All these features demonstrated the activation of mesothelial(endothelial)-mesenchymal transition strongly resembling epicardial cellular responses to ischemia in vivo.Conclusion. An epicardial spheroid cell culture model suitable for study cellular responses to hypoxic environment was developed. This model can be used to clarify mechanisms regulating epicardial microenvironment and test new targeted candidate drugs.https://www.reanimatology.com/rmt/article/view/2292spheroidhypoxiacardiac repair |
| spellingShingle | K. V. Dergilev Z. I. Tsokolaeva I. B. Beloglazova D. O. Traktuev M. T. Rasulova E. V. Parfenova 3D Spheroids — a Cellular Model for Studying the Effects of Hypoxia on the Epicardial Microenvironment Общая реаниматология spheroid hypoxia cardiac repair |
| title | 3D Spheroids — a Cellular Model for Studying the Effects of Hypoxia on the Epicardial Microenvironment |
| title_full | 3D Spheroids — a Cellular Model for Studying the Effects of Hypoxia on the Epicardial Microenvironment |
| title_fullStr | 3D Spheroids — a Cellular Model for Studying the Effects of Hypoxia on the Epicardial Microenvironment |
| title_full_unstemmed | 3D Spheroids — a Cellular Model for Studying the Effects of Hypoxia on the Epicardial Microenvironment |
| title_short | 3D Spheroids — a Cellular Model for Studying the Effects of Hypoxia on the Epicardial Microenvironment |
| title_sort | 3d spheroids a cellular model for studying the effects of hypoxia on the epicardial microenvironment |
| topic | spheroid hypoxia cardiac repair |
| url | https://www.reanimatology.com/rmt/article/view/2292 |
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