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Growth of recombinant fibroblasts in alginate microcapsules
Author(s) -
Chang P. L.,
Hortelano G.,
Awrey D. E.,
Tse M.
Publication year - 1994
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.260431005
Subject(s) - recombinant dna , chemistry , biochemistry , microbiology and biotechnology , biology , gene
To develop a novel strategy of nonautologous somatic gene therapy, we now demonstrate the feasibility of culturing genetically modified fibroblasts within an immunoprotective environment and the optimal conditions required for their continued survival in vitro. When mouse Ltk − fibroblasts transfected with the human growth hormone gene were enclosed within permselective microcapsules fabricated from alginate–polylysine–alginate, they continued to secrete human growth hormone at the same rates as the nonencapsulated cells. They also continued to proliferate in vitro for at least 1 month even though their viability gradually declined to about 50%. The viability can be improved by controlling for (a) temperature during encapsulation, (b) duration of treatment with polylysine, (c) duration of liquefying the core alginate with sodium citrate, and (d) cell density at the time of encapsulation. The best conditions leading to improved survival and maximum proliferation of cells within the microcapsules were obtained by encapsulating the cells at 4 to 10°C instead of room temperature, coating the microspheres with polylysine for 6 to 10 min instead of 20 min, liquefying the core alginate by treating with citrate for 20 min instead of 6 to 10 min, and using a concentration of 2 × 10 6 cells/mL of alginate for encapsulation. Under such conditions, normally adherent and genetically engineered mouse fibroblasts survived and proliferated optimally within the microcapsule environment. The encapsulated fibroblasts maintained their level of transgene expression while recombinant gene products such as human growth hormone could diffuse through the microcapsule membrane without impediment. The demonstration that genetically modified fibroblasts can survive and continue to deliver recombinant gene products from within these microcapsules and the optimization for their maximal viability and growth within microcapsules should increase the potential for success in using such microencapsulated recombinant cells for somatic gene therapy. © 1994 John Wiley & Sons, Inc.