Hypoxia and transforming growth factor‐beta1 pathway activation promote Chinese Hamster Ovary cell aggregation

Qian, Yueming; Rehmann, Matthew S.; Qian, Nan‐Xin; He, Aiqing; Borys, Michael; Kayne, Paul S.; Li, Zheng Jian

doi:10.1002/bit.26520

Cited by 8 publications

(2 citation statements)

References 34 publications

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“…With regard to extracellular stimuli, the p21 CIP1 can be induced by transforming growth factor beta (TGFβ, encoded by Tgfb ), which acts as a negative paracrine/autocrine growth factor in growth regulation, as shown in Figure A. The TGFβ protein is activated from its latent complex upon sensing various environmental stresses such as ROS, hypoxic conditions, pH, and proteases . As mentioned before, the DO and pH are well controlled during both the ActiCHO and the FortiCHO processes.…”

Section: Resultsmentioning

confidence: 99%

Transcriptome Analysis of CHO Cell Size Increase During a Fed-Batch Process

et al. 2018

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In a Chinese Hamster Ovary (CHO) cell fed-batch process, arrest of cell proliferation and an almost threefold increase in cell size occurred, which is associated with an increase in cell-specific productivity. In this study, transcriptome analysis is performed to identify the molecular mechanisms associated with this. Cell cycle analysis reveals that the cells are arrested mainly in the G /G phase. The cell cycle arrest is associated with significant up-regulation of cyclin-dependent kinases inhibitors (CDKNs) and down-regulation of cyclin-dependent kinases (CDKs) and cyclins. During the cell size increase phase, the gene expression of the upstream pathways of mechanistic target of rapamycin (mTOR), which is related to the extracellular growth factor, cytokine, and amino acid conditions, shows a strongly synchronized pattern to promote the mTOR activity. The downstream genes of mTOR also show a synchronized pattern to stimulate protein translation and lipid synthesis. The results demonstrate that cell cycle inhibition and stimulated mTOR activity at the transcriptome level are related to CHO cell size increase. The cell size increase is related to the extracellular nutrient conditions through a number of cascade pathways, indicating that by rational design of media and feeds, CHO cell size can be manipulated during culture processes, which may further improve cell growth and specific productivity.

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Section: Resultsmentioning

confidence: 99%

Transcriptome Analysis of CHO Cell Size Increase During a Fed-Batch Process

et al. 2018

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“…Traditionally, bioreactor conditions and cell populations within cell cultures are regarded as homogeneous. Only in recent years, inhomogeneities in bioreactor hydrodynamics and culture conditions, such as pH (Brunner et al, 2017;Lara, Galindo, Ramírez, & Palomares, 2006) or dissolved oxygen concentration (Gao et al, 2016;Qian et al, 2018;Xing, Kenty, Li, & Lee, 2009) and their impact on the bioprocess performance were examined. Hence, heterogeneities based on differences in the cell populations, like varying intraclonal protein expression levels (Pilbrough, Munro, & Gray, 2009), distribution of chromosome counts and patterns in CHO cell line generation (Vcelar et al, 2018), or changing subpopulation dynamics (Kacmar & Srienc, 2005) have moved into the focus of research.…”

Section: Introductionmentioning

confidence: 99%

Model‐based identification of cell‐cycle‐dependent metabolism and putative autocrine effects in antibody producing CHO cell culture

Möller

Korte

Pörtner

et al. 2018

Biotech & Bioengineering

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The understanding of cell-cycle-dependent population heterogeneities in mammalian cell culture and their influence on production rates is still limited. Furthermore, metabolic regulations arising from self-expressed signaling factors (autocrine/autoinhibitory factors) have been postulated in the past, but no determination of such effects have been made so far for fast-growing production Chinese hamster ovary (CHO) cells in chemically defined media. In this study, a novel approach combining near-physiological treatment of cells (including synchronization), population resolved mechanistic modeling and statistical analysis was developed to identify population inhomogeneities. Cell-cycle-dependent population dynamics and metabolic regulations due to a putative autocrine factor were examined and their impact on the metabolic rates and antibody production of near-physiologically synchronized CHO DP-12 cell cultures was determined. To achieve this, a population resolved model was extended to describe putative autocrine-dependentt and cell-cycle-related metabolic rates for glucose, glutamine, lactate, ammonia, and antibody production. The model parameters were estimated based on data of two repeated batch cultivations (three batches each), with main substrates in excess and potentially inhibiting waste products (lactate and ammonium) controlled within narrow ranges. Significant changes, due to a putative autocrine factor, were identified for lactate and ammonia formation and antibody production. The cell growth and the uptake of glucose and glutamine were only partially affected by the putative autocrine under the given conditions. The results indicate the presence of a self-expressed autocrine factor and its strong impact on the metabolism of CHO DP-12 cells. Furthermore, glucose and glutamine uptake, as well as the formation of ammonium and antibody were found to be significantly cell-cycle-dependent. The combined approach has a strong potential to improve the understanding of the interplay of population heterogeneities and signal factors in mammalian cell culture.

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Bioreactor Scale‐Up

Bowers

Seamans

et al. 2018

Kirk-Othmer Encyclopedia of Chemical Technology

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Large‐scale bioreactors are used for producing biological products (e.g., monoclonal antibodies, fusion proteins, enzymes) expressed in mammalian cells. Compared with small‐scale bioreactors, they are intrinsically more challenging in mixing and mass transfer and require more optimization. Proper bioreactor scale‐up is needed to ensure productivity and product quality. Here, common approaches and challenges in industrial mammalian cell culture scale‐up are covered. These are different from the scale‐up challenges and methodologies typically used in microbial bioreactors, since cell‐culture bioreactors operate in different agitation power and gas flow regimes. Engineering considerations including mass transfer, mixing, and hydrodynamic stress are presented in detail. Single‐use bioreactors are focused due to their rapid adoption by industry in clinical and commercial production. Though the main body of this article is on suspension cultures, special considerations in scaling‐up microcarrier‐based cultures are also briefly discussed. Finally, new challenges and desired improvements in scaling up high density cultures are presented.

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Hypoxia and transforming growth factor‐beta1 pathway activation promote Chinese Hamster Ovary cell aggregation

Cited by 8 publications

References 34 publications

Transcriptome Analysis of CHO Cell Size Increase During a Fed-Batch Process

Transcriptome Analysis of CHO Cell Size Increase During a Fed-Batch Process

Model‐based identification of cell‐cycle‐dependent metabolism and putative autocrine effects in antibody producing CHO cell culture

Bioreactor Scale‐Up

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