title: A fibrous-bed bioreactor for continuous production of developmental endothelial locus-1 by osteosarcoma cells.
abstract: Genetically engineered human osteosarcoma cells containing developmental endothelial locus-1 (del-1) gene were studied for production of Del-1, a protein that has the properties of an extracellular matrix protein and can regulate vascular morphogenesis and remodeling. Del-1 has been studied as a potential anti-angiogenesis drug targeting solid tumors. In this study, osteosarcoma cells were cultured in a fibrous-bed bioreactor (FBB) to continuously produce Del-1. The FBB was constructed by packing a polyester fibrous matrix into a 1.5-l spinner flask. The effects of media composition, including the serum content in the medium, and dilution rate on cell growth, metabolism, and Del-1 production were studied. A gradual reduction of serum content from 10% (v/v) to 0.5% (v/v) caused no loss in Del-1 production. However, the production of Del-1 decreased significantly in a serum-free medium, suggesting some nutrients present in the serum were important to culture viability and Del-1 production. The continuous FBB culture was stable for long-term production of Del-1, with a higher Del-1 titer than that normally obtained in T-flask cultures and overall productivity similar to the total production from 300 25-cm(2) T-flasks. Reducing geneticin in the medium from 250 microg ml(-1) to zero at later culturing stages had no significant effect on Del-1 production. The FBB was operated for a period of more than 4 months without any notable degeneration, and reached a final cell density of 3 x 10(8) cells ml(-1) of packing volume with >90% cell viability. The good reactor performance can be attributed to the three-dimensional environment provided by the fibrous matrix that allows for efficient mass transfer and cell immobilization and growth. Scanning electron microscopic and confocal scanning laser microscopic studies of the cell-matrix showed that cells formed large aggregates in the fibrous matrix and cell density was relatively uniform in the matrix.
authors.affiliation: Department of Chemical Engineering, The Ohio State University, 140 West 19th Avenue, Columbus, OH 43210, USA.
authors.forename: Chunnuan
authors.initials: C
authors.lastname: Chen
authors.forename: Yu Liang
authors.initials: YL
authors.lastname: Huang
authors.forename: Shang-Tian
authors.initials: ST
authors.lastname: Yang
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D019149:Bioreactors; D001798:Blood Proteins / Q000494:pharmacology; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins / Q000096:biosynthesis*; D015815:Cell Adhesion Molecules; D002452:Cell Count; D003470:Culture Media / Q000494:pharmacology; D005839:Gentamicins / Q000494:pharmacology; D005973:Glutamine / Q000378:metabolism; D006801:Humans; D007201:Indicator Dilution Techniques; D008855:Microscopy, Electron, Scanning; D008954:Models, Biological; D012516:Osteosarcoma; D011091:Polyesters / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure
chemical_list: D000900:Anti-Bacterial Agents; D001798:Blood Proteins; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D015815:Cell Adhesion Molecules; D003470:Culture Media; C110183:EDIL3 protein, human; D005839:Gentamicins; D011091:Polyesters; D005973:Glutamine; C010680:antibiotic G 418