Impact of the Use of 2-Phospho-L Ascorbic Acid in the Production of Engineered Stromal Tissue for Regenerative Medicine
Tissue engineering enables autologous reconstruction of human tissues, addressing limitations in tissue availability and immune compatibility. Several tissue engineering techniques, such as self-assembly, rely on or benefit from extracellular matrix (ECM) secretion by fibroblasts to produce biomimet...
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MDPI AG
2025-07-01
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| author | David Brownell Laurence Carignan Reza Alavi Christophe Caneparo Maxime Labroy Todd Galbraith Stéphane Chabaud François Berthod Laure Gibot François Bordeleau Stéphane Bolduc |
| author_facet | David Brownell Laurence Carignan Reza Alavi Christophe Caneparo Maxime Labroy Todd Galbraith Stéphane Chabaud François Berthod Laure Gibot François Bordeleau Stéphane Bolduc |
| author_sort | David Brownell |
| collection | DOAJ |
| description | Tissue engineering enables autologous reconstruction of human tissues, addressing limitations in tissue availability and immune compatibility. Several tissue engineering techniques, such as self-assembly, rely on or benefit from extracellular matrix (ECM) secretion by fibroblasts to produce biomimetic scaffolds. Models have been developed for use in humans, such as skin and corneas. Ascorbic acid (vitamin C, AA) is essential for collagen biosynthesis. However, AA is chemically unstable in culture, with a half-life of 24 h, requiring freshly prepared AA with each change of medium. This study aims to demonstrate the functional equivalence of 2-phospho-L-ascorbate (2PAA), a stable form of AA, for tissue reconstruction. Dermal, vaginal, and bladder stroma were reconstructed by self-assembly using tissue-specific protocols. The tissues were cultured in a medium supplemented with either freshly prepared or frozen AA, or with 2PAA. Biochemical analyses were performed on the tissues to evaluate cell density and tissue composition, including collagen secretion and deposition. Histology and quantitative polarized light microscopy were used to evaluate tissue architecture, and mechanical evaluation was performed both by tensiometry and atomic force microscopy (AFM) to evaluate its macroscopic and cell-scale mechanical properties. The tissues produced by the three ascorbate conditions had similar collagen deposition, architecture, and mechanical properties in each organ-specific stroma. Mechanical characterization revealed tissue-specific differences, with tensile modulus values ranging from 1–5 MPa and AFM-derived apparent stiffness in the 1–2 kPa range, reflecting the nonlinear and scale-dependent behavior of the engineered stroma. The results demonstrate the possibility of substituting AA with 2PAA for tissue engineering. This protocol could significantly reduce the costs associated with tissue production by reducing preparation time and use of materials. This is a crucial factor for any scale-up activity. |
| format | Article |
| id | doaj-art-a11deee798b448d1a3be802afddaba37 |
| institution | DOAJ |
| issn | 2073-4409 |
| language | English |
| publishDate | 2025-07-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Cells |
| spelling | doaj-art-a11deee798b448d1a3be802afddaba372025-08-20T03:08:00ZengMDPI AGCells2073-44092025-07-011414112310.3390/cells14141123Impact of the Use of 2-Phospho-L Ascorbic Acid in the Production of Engineered Stromal Tissue for Regenerative MedicineDavid Brownell0Laurence Carignan1Reza Alavi2Christophe Caneparo3Maxime Labroy4Todd Galbraith5Stéphane Chabaud6François Berthod7Laure Gibot8François Bordeleau9Stéphane Bolduc10Regenerative Medicine Division, Centre de Recherche du CHU de Québec-Université Laval, Québec, QC G1J 5B3, CanadaCentre de Recherche en Organogénèse Expérimentale de L’Université Laval/LOEX, Université Laval, Québec, QC G1J 5B3, CanadaCentre de Recherche en Organogénèse Expérimentale de L’Université Laval/LOEX, Université Laval, Québec, QC G1J 5B3, CanadaRegenerative Medicine Division, Centre de Recherche du CHU de Québec-Université Laval, Québec, QC G1J 5B3, CanadaCentre de Recherche en Organogénèse Expérimentale de L’Université Laval/LOEX, Université Laval, Québec, QC G1J 5B3, CanadaRegenerative Medicine Division, Centre de Recherche du CHU de Québec-Université Laval, Québec, QC G1J 5B3, CanadaRegenerative Medicine Division, Centre de Recherche du CHU de Québec-Université Laval, Québec, QC G1J 5B3, CanadaRegenerative Medicine Division, Centre de Recherche du CHU de Québec-Université Laval, Québec, QC G1J 5B3, CanadaUniversité de Toulouse, CNRS UMR 5623, Laboratoire Softmat, 31062 Toulouse, FranceCentre de Recherche en Organogénèse Expérimentale de L’Université Laval/LOEX, Université Laval, Québec, QC G1J 5B3, CanadaRegenerative Medicine Division, Centre de Recherche du CHU de Québec-Université Laval, Québec, QC G1J 5B3, CanadaTissue engineering enables autologous reconstruction of human tissues, addressing limitations in tissue availability and immune compatibility. Several tissue engineering techniques, such as self-assembly, rely on or benefit from extracellular matrix (ECM) secretion by fibroblasts to produce biomimetic scaffolds. Models have been developed for use in humans, such as skin and corneas. Ascorbic acid (vitamin C, AA) is essential for collagen biosynthesis. However, AA is chemically unstable in culture, with a half-life of 24 h, requiring freshly prepared AA with each change of medium. This study aims to demonstrate the functional equivalence of 2-phospho-L-ascorbate (2PAA), a stable form of AA, for tissue reconstruction. Dermal, vaginal, and bladder stroma were reconstructed by self-assembly using tissue-specific protocols. The tissues were cultured in a medium supplemented with either freshly prepared or frozen AA, or with 2PAA. Biochemical analyses were performed on the tissues to evaluate cell density and tissue composition, including collagen secretion and deposition. Histology and quantitative polarized light microscopy were used to evaluate tissue architecture, and mechanical evaluation was performed both by tensiometry and atomic force microscopy (AFM) to evaluate its macroscopic and cell-scale mechanical properties. The tissues produced by the three ascorbate conditions had similar collagen deposition, architecture, and mechanical properties in each organ-specific stroma. Mechanical characterization revealed tissue-specific differences, with tensile modulus values ranging from 1–5 MPa and AFM-derived apparent stiffness in the 1–2 kPa range, reflecting the nonlinear and scale-dependent behavior of the engineered stroma. The results demonstrate the possibility of substituting AA with 2PAA for tissue engineering. This protocol could significantly reduce the costs associated with tissue production by reducing preparation time and use of materials. This is a crucial factor for any scale-up activity.https://www.mdpi.com/2073-4409/14/14/1123tissue engineeringextracellular matrixsodium L-ascorbate2-phospo-L-ascorbatehuman primary fibroblasts |
| spellingShingle | David Brownell Laurence Carignan Reza Alavi Christophe Caneparo Maxime Labroy Todd Galbraith Stéphane Chabaud François Berthod Laure Gibot François Bordeleau Stéphane Bolduc Impact of the Use of 2-Phospho-L Ascorbic Acid in the Production of Engineered Stromal Tissue for Regenerative Medicine Cells tissue engineering extracellular matrix sodium L-ascorbate 2-phospo-L-ascorbate human primary fibroblasts |
| title | Impact of the Use of 2-Phospho-L Ascorbic Acid in the Production of Engineered Stromal Tissue for Regenerative Medicine |
| title_full | Impact of the Use of 2-Phospho-L Ascorbic Acid in the Production of Engineered Stromal Tissue for Regenerative Medicine |
| title_fullStr | Impact of the Use of 2-Phospho-L Ascorbic Acid in the Production of Engineered Stromal Tissue for Regenerative Medicine |
| title_full_unstemmed | Impact of the Use of 2-Phospho-L Ascorbic Acid in the Production of Engineered Stromal Tissue for Regenerative Medicine |
| title_short | Impact of the Use of 2-Phospho-L Ascorbic Acid in the Production of Engineered Stromal Tissue for Regenerative Medicine |
| title_sort | impact of the use of 2 phospho l ascorbic acid in the production of engineered stromal tissue for regenerative medicine |
| topic | tissue engineering extracellular matrix sodium L-ascorbate 2-phospo-L-ascorbate human primary fibroblasts |
| url | https://www.mdpi.com/2073-4409/14/14/1123 |
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