A 3D collagen microsphere culture system for GDNF-secreting HEK293 cells with enhanced protein productivity.
Mammalian cell culture technology has been used for decades in mass production of therapeutic proteins. However, unrestricted cell proliferation usually results in low-protein productivity. Controlled proliferation technologies such as metabolism intervention and genetic manipulation are therefore applied to enhance the productivity. Nevertheless, these strategies induced growth arrest with reduced viability and increased apoptosis. In this study, we report a new controlled proliferation technology by encapsulating human embryonic kidney (HEK) 293 cells over-expressing glial-derived neurotrophic factor (GDNF) in 3D collagen microspheres for extended culture. We investigated the viability, proliferation, cell cycle and GDNF productivity of HEK293 cells in microspheres as compared to monolayer culture. This system provides a physiologically relevant tissue-like environment for cells to grow and exerts proliferation control throughout the culture period without compromising the viability. A significant increase in the production rate of GDNF was found in the 3D microsphere system comparing with the monolayer culture. GDNF productivity was also significantly affected by the initial cell number and the serum concentration. The secreted GDNF was still bioactive as it induced neurite extension in PC12 cells. In summary, the 3D collagen microsphere system presents a cost-effective controlled growth technology for protein production in pharmaceutical manufacturing.
Wong
Hoi-Ling
HL
Medical Engineering Program, Department of Mechanical Engineering, Faculty of Engineering, The University of Hong Kong, Pokfulam Road, Hong Kong.
Wang
Ming-Xi
MX
Cheung
Pik-To
PT
Yao
Kwok-Ming
KM
Chan
Barbara Pui
BP
D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line; D024042:Collagen Type I; D051100:Glial Cell Line-Derived Neurotrophic Factor / Q000096:biosynthesis / Q000378:metabolism*; D006801:Humans; D007668:Kidney / Q000166:cytology* / Q000378:metabolism*; D051379:Mice; D008863:Microspheres; D016716:PC12 Cells; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism*
D024042:Collagen Type I; C495040:Gdnf protein, mouse; D051100:Glial Cell Line-Derived Neurotrophic Factor; D011994:Recombinant Proteins