Serum-free endothelial cell culture medium for vascular smooth muscle cells sheet formation
Abstract Background Cell sheet technology has been identified as a promising approach for the construction of tissue-engineered vascular grafts (TEVGs). However, concerns regarding immunogenicity and ethical issues, which are raised by the use of fetal bovine serum (FBS) in traditional culture syste...
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| Language: | English |
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BMC
2025-05-01
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| Series: | Journal of Biological Engineering |
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| Online Access: | https://doi.org/10.1186/s13036-025-00522-y |
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| author | Jing Yang Xuheng Sun Hongjing Jiang Jiandong Li Jierong Liang Zhanyi Lin |
| author_facet | Jing Yang Xuheng Sun Hongjing Jiang Jiandong Li Jierong Liang Zhanyi Lin |
| author_sort | Jing Yang |
| collection | DOAJ |
| description | Abstract Background Cell sheet technology has been identified as a promising approach for the construction of tissue-engineered vascular grafts (TEVGs). However, concerns regarding immunogenicity and ethical issues, which are raised by the use of fetal bovine serum (FBS) in traditional culture systems, limit its potential for clinical translation. Serum-free medium (SFM) has emerged as a safer and more controllable alternative, but further validation is required to determine its effectiveness and superiority in generating high-quality cell sheets. Methods This study systematically compared cell sheets generated under SFM and 10% FBS culture conditions in terms of structure, cellular phenotype, and functional properties. The expression levels of α-SMA and SM22, markers of vascular smooth muscle cells(VSMCs), were evaluated using immunofluorescence staining, qRT-PCR, and Western blot analysis to assess cellular phenotype. Histological staining and mechanical testing were employed to compare the morphology and mechanical properties of the cell sheets, while extracellular matrix (ECM) deposition and biochemical characteristics were also analyzed. Results Under SFM conditions, cells exhibited significantly higher α-SMA and SM22 expression levels (qRT-PCR showed a 1.8-fold and 2-fold increase, respectively; ****p < 0.0001) with clearer cytoskeletal arrangement. Cell sheets formed in SFM displayed comparable area(ns, p > 0.05), thickness(**p < 0.01), and mechanical properties to those cultured in 10% FBS, while ECM deposition was significantly enhanced (collagen content increased by approximately 40%, **p < 0.01). Furthermore, histological analysis revealed that cell sheets generated under SFM conditions were more compact and uniform, exhibiting superior structural organization. Conclusion SFM facilitates the generation of cell sheets that exhibit structural and functional properties analogous to those cultured in FBS. Additionally, SFM promotes cellular phenotype transition and ECM deposition. Consequently, SFM provides a safer, more controllable, and clinically translatable solution for cell sheet construction. Graphical Abstract |
| format | Article |
| id | doaj-art-3b200d14fc734a7cb63a84b5b760a8f5 |
| institution | OA Journals |
| issn | 1754-1611 |
| language | English |
| publishDate | 2025-05-01 |
| publisher | BMC |
| record_format | Article |
| series | Journal of Biological Engineering |
| spelling | doaj-art-3b200d14fc734a7cb63a84b5b760a8f52025-08-20T02:00:09ZengBMCJournal of Biological Engineering1754-16112025-05-0119111310.1186/s13036-025-00522-ySerum-free endothelial cell culture medium for vascular smooth muscle cells sheet formationJing Yang0Xuheng Sun1Hongjing Jiang2Jiandong Li3Jierong Liang4Zhanyi Lin5School of Medicine, South China University of TechnologyGuangdong Cardiovascular InstituteSchool of Medicine, South China University of TechnologyJIHUA LaboratoryJIHUA LaboratorySchool of Medicine, South China University of TechnologyAbstract Background Cell sheet technology has been identified as a promising approach for the construction of tissue-engineered vascular grafts (TEVGs). However, concerns regarding immunogenicity and ethical issues, which are raised by the use of fetal bovine serum (FBS) in traditional culture systems, limit its potential for clinical translation. Serum-free medium (SFM) has emerged as a safer and more controllable alternative, but further validation is required to determine its effectiveness and superiority in generating high-quality cell sheets. Methods This study systematically compared cell sheets generated under SFM and 10% FBS culture conditions in terms of structure, cellular phenotype, and functional properties. The expression levels of α-SMA and SM22, markers of vascular smooth muscle cells(VSMCs), were evaluated using immunofluorescence staining, qRT-PCR, and Western blot analysis to assess cellular phenotype. Histological staining and mechanical testing were employed to compare the morphology and mechanical properties of the cell sheets, while extracellular matrix (ECM) deposition and biochemical characteristics were also analyzed. Results Under SFM conditions, cells exhibited significantly higher α-SMA and SM22 expression levels (qRT-PCR showed a 1.8-fold and 2-fold increase, respectively; ****p < 0.0001) with clearer cytoskeletal arrangement. Cell sheets formed in SFM displayed comparable area(ns, p > 0.05), thickness(**p < 0.01), and mechanical properties to those cultured in 10% FBS, while ECM deposition was significantly enhanced (collagen content increased by approximately 40%, **p < 0.01). Furthermore, histological analysis revealed that cell sheets generated under SFM conditions were more compact and uniform, exhibiting superior structural organization. Conclusion SFM facilitates the generation of cell sheets that exhibit structural and functional properties analogous to those cultured in FBS. Additionally, SFM promotes cellular phenotype transition and ECM deposition. Consequently, SFM provides a safer, more controllable, and clinically translatable solution for cell sheet construction. Graphical Abstracthttps://doi.org/10.1186/s13036-025-00522-ySerum freeCell sheetVascular smooth muscle cellsExtracellular matrixTissue-engineered vascular grafts |
| spellingShingle | Jing Yang Xuheng Sun Hongjing Jiang Jiandong Li Jierong Liang Zhanyi Lin Serum-free endothelial cell culture medium for vascular smooth muscle cells sheet formation Journal of Biological Engineering Serum free Cell sheet Vascular smooth muscle cells Extracellular matrix Tissue-engineered vascular grafts |
| title | Serum-free endothelial cell culture medium for vascular smooth muscle cells sheet formation |
| title_full | Serum-free endothelial cell culture medium for vascular smooth muscle cells sheet formation |
| title_fullStr | Serum-free endothelial cell culture medium for vascular smooth muscle cells sheet formation |
| title_full_unstemmed | Serum-free endothelial cell culture medium for vascular smooth muscle cells sheet formation |
| title_short | Serum-free endothelial cell culture medium for vascular smooth muscle cells sheet formation |
| title_sort | serum free endothelial cell culture medium for vascular smooth muscle cells sheet formation |
| topic | Serum free Cell sheet Vascular smooth muscle cells Extracellular matrix Tissue-engineered vascular grafts |
| url | https://doi.org/10.1186/s13036-025-00522-y |
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