Perfusion Enhances Functions of Bone Marrow Stromal Cells in Three-Dimensional Culture
Perfusion of medium through three-dimensional (3D) collagen sponges enhanced viability and function of cocultivated marrow stromal and hematopoietic cell lines. Cells of the murine bone marrow stromal cell line GPIa were cultured in novel 3D collagen sponges, made from pepsin-digested bovine skin. S...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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SAGE Publishing
1998-05-01
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| Series: | Cell Transplantation |
| Online Access: | https://doi.org/10.1177/096368979800700310 |
| _version_ | 1818260750861336576 |
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| author | Julie Glowacki Ph.D. Shuichi Mizuno Joel S. Greenberger |
| author_facet | Julie Glowacki Ph.D. Shuichi Mizuno Joel S. Greenberger |
| author_sort | Julie Glowacki Ph.D. |
| collection | DOAJ |
| description | Perfusion of medium through three-dimensional (3D) collagen sponges enhanced viability and function of cocultivated marrow stromal and hematopoietic cell lines. Cells of the murine bone marrow stromal cell line GPIa were cultured in novel 3D collagen sponges, made from pepsin-digested bovine skin. Static cultures of sponges were maintained in dishes with media changes every other day. Perfused sponges were contained in a glass column with medium flow set at 1.3 mL/min. In some sponges, the 32D cl3 c-fms m (CRX-1) hematopoietic progenitor cell line was added 7 days after GPIa cells. At 7 and 16 days, light microscopic evaluation showed poor viability of cells in static sponge cultures. In perfused sponge cultures, there was greater cellularity throughout the sponge and abundant accumulation of metachromatic extracellular matrix surrounding GPIa cells. Chondroitin 6-sulfate and heparan sulfate were identified as components of the matrix by immunohistochemical methods. DNA synthesis was evaluated by 15-h exposure of cultures to bromodeoxyuridine (BrdU), with subsequent immunohistochemical localization with monoclonal anti-BrdU antibody. Cells positive for BrdU were identified at the outer surfaces of both static and perfused sponges; however, positive cells were also seen throughout the internal areas of the sponges that were perfused. These results suggest that better nutrient exchange occurred in perfused sponges. In static cocultures of GPIa and CRX-1 cells, there was no detectable viability of the IL-3–dependent CRX-1 cells; however, under perfused conditions, CRX-1 cells flourished within the sponges as documented by BrdU incorporation. Thus, medium perfusion enhanced GPIa stromal cell line viability and function in 3D collagen sponge cultures, as demonstrated by BrdU incorporation, matrix production, and support of CRX-1 cells. This novel culture system may be useful for examining the interactions of bone marrow stromal cells with extracellular matrix molecules, soluble and matrix-bound factors, and with other cell types. |
| first_indexed | 2024-12-12T18:36:18Z |
| format | Article |
| id | doaj.art-165862c031944df780cb588ba878faa3 |
| institution | Directory Open Access Journal |
| issn | 0963-6897 1555-3892 |
| language | English |
| last_indexed | 2024-12-12T18:36:18Z |
| publishDate | 1998-05-01 |
| publisher | SAGE Publishing |
| record_format | Article |
| series | Cell Transplantation |
| spelling | doaj.art-165862c031944df780cb588ba878faa32022-12-22T00:15:47ZengSAGE PublishingCell Transplantation0963-68971555-38921998-05-01710.1177/096368979800700310Perfusion Enhances Functions of Bone Marrow Stromal Cells in Three-Dimensional CultureJulie Glowacki Ph.D.0Shuichi Mizuno1Joel S. Greenberger2Skeletal Biology Research Center, Massachusetts General Hospital, Harvard School of Dental Medicine, Boston, MA 02129Orthopedic Research, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115Department of Radiation Oncology, University of Pittsburgh Cancer Institute, and University of Pittsburgh Medical Center, Pittsburgh, PA 15213Perfusion of medium through three-dimensional (3D) collagen sponges enhanced viability and function of cocultivated marrow stromal and hematopoietic cell lines. Cells of the murine bone marrow stromal cell line GPIa were cultured in novel 3D collagen sponges, made from pepsin-digested bovine skin. Static cultures of sponges were maintained in dishes with media changes every other day. Perfused sponges were contained in a glass column with medium flow set at 1.3 mL/min. In some sponges, the 32D cl3 c-fms m (CRX-1) hematopoietic progenitor cell line was added 7 days after GPIa cells. At 7 and 16 days, light microscopic evaluation showed poor viability of cells in static sponge cultures. In perfused sponge cultures, there was greater cellularity throughout the sponge and abundant accumulation of metachromatic extracellular matrix surrounding GPIa cells. Chondroitin 6-sulfate and heparan sulfate were identified as components of the matrix by immunohistochemical methods. DNA synthesis was evaluated by 15-h exposure of cultures to bromodeoxyuridine (BrdU), with subsequent immunohistochemical localization with monoclonal anti-BrdU antibody. Cells positive for BrdU were identified at the outer surfaces of both static and perfused sponges; however, positive cells were also seen throughout the internal areas of the sponges that were perfused. These results suggest that better nutrient exchange occurred in perfused sponges. In static cocultures of GPIa and CRX-1 cells, there was no detectable viability of the IL-3–dependent CRX-1 cells; however, under perfused conditions, CRX-1 cells flourished within the sponges as documented by BrdU incorporation. Thus, medium perfusion enhanced GPIa stromal cell line viability and function in 3D collagen sponge cultures, as demonstrated by BrdU incorporation, matrix production, and support of CRX-1 cells. This novel culture system may be useful for examining the interactions of bone marrow stromal cells with extracellular matrix molecules, soluble and matrix-bound factors, and with other cell types.https://doi.org/10.1177/096368979800700310 |
| spellingShingle | Julie Glowacki Ph.D. Shuichi Mizuno Joel S. Greenberger Perfusion Enhances Functions of Bone Marrow Stromal Cells in Three-Dimensional Culture Cell Transplantation |
| title | Perfusion Enhances Functions of Bone Marrow Stromal Cells in Three-Dimensional Culture |
| title_full | Perfusion Enhances Functions of Bone Marrow Stromal Cells in Three-Dimensional Culture |
| title_fullStr | Perfusion Enhances Functions of Bone Marrow Stromal Cells in Three-Dimensional Culture |
| title_full_unstemmed | Perfusion Enhances Functions of Bone Marrow Stromal Cells in Three-Dimensional Culture |
| title_short | Perfusion Enhances Functions of Bone Marrow Stromal Cells in Three-Dimensional Culture |
| title_sort | perfusion enhances functions of bone marrow stromal cells in three dimensional culture |
| url | https://doi.org/10.1177/096368979800700310 |
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