Non-viral endostatin plasmid transfection of mesenchymal stem cells via collagen scaffolds |
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| Authors: | Xiao-Dan Sun Lily Jeng Catherine Bolliet Bjorn R. Olsen Myron Spector |
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| Affiliation: | 1. Tissue Engineering, VA Boston Healthcare System, Mail Stop: 151 Research, 150 S. Huntington Ave., Boston, MA 02130, USA;2. Laboratory of Advanced Materials, Department of Materials Science and Engineering, Tsinghua University, Beijing 100084, China;3. Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA;4. Department of Developmental Biology, Harvard School of Dental Medicine, Boston, MA 02115, USA;5. Department of Orthopaedic Surgery, Brigham and Women''s Hospital, Harvard Medical School, Boston, MA 02115, USA;1. Regenerative Medicine Research Institute, Universitat Internacional de Catalunya, Barcelona, Spain;2. Grup d’Enginyeria de Materials (GEMAT), Institut Químic de Sarrià, Universitat Ramon Llull, Barcelona, Spain;3. Centro de Investigación Biomédica en Red en Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Zaragoza, Spain;1. Department of Physiology, Favaloro University, Buenos Aires, Argentina;2. Department of Pathology, Favaloro University, Buenos Aires, Argentina;3. Bio Sidus S.A., Buenos Aires, Argentina;1. State Key Laboratory of Military Stomatology, Department of Prosthetic Dentistry, School of Stomatology, The Fourth Military Medical University, No. 145 West Changle Road, Xi''an 710032, China;2. The Second Artillery Engineering University, No. 2 Tongxin Road, Xi''an 710025, China;3. State Key Laboratory of Military Stomatology, Department of Periodontology, School of Stomatology, The Fourth Military Medical University, No. 145 West Changle Road, Xi''an 710032, China;4. Department of Orthopedics, The First Affiliated Hospital of CPLA General Hospital, Beijing 100048, China;1. State Key Laboratory of Military Stomatology, Shaanxi Key Laboratory of Stomatology, Department of Periodontology, School of Stomatology, Fourth Military Medical University, No. 145 West Changle Road, Xi’an 710032, China;2. College of Marine Life Science, Ocean University of China, No. 5 Yushan Road, Qingdao 266003, Shandong Province, China;3. Qingdao First Sanatorium, Jinan Military Region, No. 27 West Hong Kong Road, Qingdao 266071, Shandong Province, China;4. State Key Laboratory of Military Stomatology, Department of Prosthetic Dentistry, School of Stomatology, Fourth Military Medical University, No. 145 West Changle Road, Xi’an 710032, China |
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| Abstract: | Angiogenesis is critical in the early stage of reparative processes and tissue regeneration, but the persistence of a vascular network may interfere with later transformation/maturation in naturally avascular tissues such as articular cartilage. Our supposition is that the timed delivery of an anti-angiogenic factor in cartilage tissue engineering may facilitate the formation of hyaline cartilage by inducing the regression of vascularization. To this end our overall goal is to prepare an off-the-shelf scaffold containing the gene for a potent anti-angiogenic factor. The objective of this study was to investigate the use of a type I/III collagen scaffold for the non-viral transfection of marrow stromal cells (MSCs, also referred to as mesenchymal stem cells) with the plasmid encoding endostatin. Caprine MSCs were transfected by the naked plasmid alone and plasmid incorporated into a cationic lipid complex in three experiments: 1) cells were transfected in monolayer; 2) monolayer-transfected cells were grown in a collagen sponge-like scaffold; and 3) non-transfected cells were grown in a collagen scaffold containing the naked plasmid and endostatin lipoplex. Independent variables were the passage number of the cells and the plasmid loading. The amount of endostatin released by the cells into the medium was measured using an ELISA. The results demonstrated the overexpression of endostatin by MSCs growing in the endostatin lipoplex-supplemented collagen scaffolds. Endostatin released by the cell-seeded scaffolds reached a peak of 13 ng/ml for scaffolds incorporating as little as 20 μg of plasmid, at the 3-day collection period ending 5 days post-seeding. The accumulated endostatin synthesis over a 2-week period began to achieve what may be a therapeutic level. MSCs transfected with the endostatin gene in monolayer continued to express the gene when grown in the collagen scaffolds. The results demonstrate the promise of the non-viral delivery of the gene for this potent anti-angiogenic protein to MSCs via a collagen scaffold. |
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