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Characterization of bimodal cell death of insect cells in a rotating- wall vessel and shaker flask

Publication ,  Journal Article
Cowger, NL; O'Connor, KC; Hammond, TG; Lacks, DJ; Navar, GL
Published in: Biotechnology and Bioengineering
1999

In previous publications, we reported the benefits of a high-aspect rotating-wall vessel (HARV) over conventional bioreactors for insect-cell cultivation in terms of reduced medium requirements and enhanced longevity. To more fully understand the effects that HARV cultivation has on longevity, the present study characterizes the mode and kinetics of Spodoptera frugiperda cell death in this quiescent environment relative to a shaker- flask control. Data from flow cytometry and fluorescence microscopy show a greater accumulation of apoptotic cells in the HARV culture, by a factor of at least 2 at the end of the cultivation period. We present a kinetic model of growth and bimodal cell death. The model is unique for including both apoptosis and necrosis, and further, transition steps within the two pathways. Kinetic constants reveal that total cell death is reduced in the HARV and the accumulation of apoptotic cells in this vessel results from reduced depletion by lysis and secondary necrosis. The ratio of early apoptotic to necrotic cell formation is found independent of cultivation conditions. In the model, apoptosis is only well represented by an integral term, which may indicate its dependence on accumulation of some factor over time; in contrast, necrosis is adequately represented with a first-order term. Cell-cycle analysis shows the percent of tetraploid cells gradually decreases during cultivation in both vessels. For example, between 90% and 70% viability, tetraploid cells in the HARV drop from 43 ± 1% to 24 ± 4%. The data suggests the tetraploid phase as the likely origin for apoptosis in our cultures. Possible mechanisms for these changes in bimodal cell death are discussed, including hydrodynamic forces, cell-cell interactions, waste accumulation, and mass transport. These studies may benefit insect-cell cultivation by increasing our understanding of cell death in culture and providing a means for further enhancing culture longevity.

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Published In

Biotechnology and Bioengineering

DOI

Publication Date

1999

Volume

64

Issue

1

Start / End Page

14 / 26

Related Subject Headings

  • Biotechnology
 

Citation

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Cowger, N. L., O’Connor, K. C., Hammond, T. G., Lacks, D. J., & Navar, G. L. (1999). Characterization of bimodal cell death of insect cells in a rotating- wall vessel and shaker flask. Biotechnology and Bioengineering, 64(1), 14–26. https://doi.org/10.1002/(SICI)1097-0290(19990705)64:1<14::AID-BIT2>3.0.CO;2-R
Cowger, N. L., K. C. O’Connor, T. G. Hammond, D. J. Lacks, and G. L. Navar. “Characterization of bimodal cell death of insect cells in a rotating- wall vessel and shaker flask.” Biotechnology and Bioengineering 64, no. 1 (1999): 14–26. https://doi.org/10.1002/(SICI)1097-0290(19990705)64:1<14::AID-BIT2>3.0.CO;2-R.
Cowger NL, O’Connor KC, Hammond TG, Lacks DJ, Navar GL. Characterization of bimodal cell death of insect cells in a rotating- wall vessel and shaker flask. Biotechnology and Bioengineering. 1999;64(1):14–26.
Cowger, N. L., et al. “Characterization of bimodal cell death of insect cells in a rotating- wall vessel and shaker flask.” Biotechnology and Bioengineering, vol. 64, no. 1, 1999, pp. 14–26. Scival, doi:10.1002/(SICI)1097-0290(19990705)64:1<14::AID-BIT2>3.0.CO;2-R.
Cowger NL, O’Connor KC, Hammond TG, Lacks DJ, Navar GL. Characterization of bimodal cell death of insect cells in a rotating- wall vessel and shaker flask. Biotechnology and Bioengineering. 1999;64(1):14–26.

Published In

Biotechnology and Bioengineering

DOI

Publication Date

1999

Volume

64

Issue

1

Start / End Page

14 / 26

Related Subject Headings

  • Biotechnology