Document Detail


Cryopreservation of alginate-encapsulated recombinant cells for antiangiogenic therapy.
MedLine Citation:
PMID:  15040603     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The potential benefit of continuous local administration of antiangiogenic proteins to CNS tumors in vivo has recently been demonstrated using endostatin-producing recombinant cells encapsulated in alginate beads. Due to the treatment potential of transplanted alginate-encapsulated cells producing therapeutic proteins, we describe a successful method of cryopreservation (CP) of such beads, in which cellular viability, alginate structure, and protein secretion were maintained. Alginate beads containing human embryonic kidney cells (HEK 293 cells) stably transfected with the gene encoding for endostatin were cryopreserved in dimethyl sulfoxide (DMSO) using a slow freezing procedure. Briefly, the DMSO concentration was gradually increased prior to the freezing procedure. The cryotubes were further supercooled to -7.5 degrees C and nucleated. Thereafter, the samples were cooled at a rate of 0.25 degrees C/min and stored in liquid nitrogen. The viability of the encapsulated cells was assessed using confocal microscopy quantification (CLSM) technique and a MTS assay. The cell cycle distribution inside the beads was assessed by DNA flow cytometry and endostatin production was determined by an endostatin-specific ELISA assay, both prior to and after CP. CLSM measurements showed sustained esterase activity in the beads after thawing, with only a slight transient decrease 24 h after CP. The MTS assay verified these findings by displaying similar variations of intracellular dehydrogenase activity. Flow cytometric analyses revealed no cryorelated disturbances in cellular ploidy. Furthermore, ELISA measurements showed a well-preserved endostatin production after CP. In conclusion, this work describes the successful CP of alginate-encapsulated recombinant cells secreting a therapeutic protein. Together with previous published reports, these results further substantiate the feasibility and potential of cell encapsulation therapy in the treatment of malignant tumors.
Authors:
V Stensvaag; T Furmanek; K Lønning; A J A Terzis; R Bjerkvig; T Visted
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Cell transplantation     Volume:  13     ISSN:  0963-6897     ISO Abbreviation:  Cell Transplant     Publication Date:  2004  
Date Detail:
Created Date:  2004-03-25     Completed Date:  2004-10-05     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  9208854     Medline TA:  Cell Transplant     Country:  United States    
Other Details:
Languages:  eng     Pagination:  35-44     Citation Subset:  IM    
Affiliation:
NORLUX Neuro-Oncology, Department of Anatomy and Cell Biology, University of Bergen, Jonas Lies vei 91, 5009 Bergen, Norway.
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MeSH Terms
Descriptor/Qualifier:
Alginates*
Angiogenesis Inhibitors
Capsules
Cell Line
Cell Transplantation / methods*
Cryopreservation / methods
Dimethyl Sulfoxide
Endostatins / genetics
Glucuronic Acid*
Hexuronic Acids*
Humans
Image Processing, Computer-Assisted
Kidney / cytology
Microscopy, Confocal
Transfection
Chemical
Reg. No./Substance:
0/Alginates; 0/Angiogenesis Inhibitors; 0/Capsules; 0/Endostatins; 0/Hexuronic Acids; 576-37-4/Glucuronic Acid; 67-68-5/Dimethyl Sulfoxide; 9005-32-7/alginic acid

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