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Mild hypothermia upregulates myc and xbp1s expression and improves anti-TNF? production in CHO cells.

ABSTRACT: Chinese hamster ovary (CHO) cells are the most frequently used host for commercial production of therapeutic proteins. However, their low protein productivity in culture is the main hurdle to overcome. Mild hypothermia has been established as an effective strategy to enhance protein specific productivity, although the causes of such improvement still remain unclear. The self-regulation of global transcriptional regulatory factors, such as Myc and XBP1s, seems to be involved in increased the recombinant protein production at low temperature. This study evaluated the impact of low temperature in CHO cell cultures on myc and xbp1s expression and their effects on culture performance and cell metabolism. Two anti-TNF? producing CHO cell lines were selected considering two distinct phenotypes: i.e. maximum cell growth, (CN1) and maximum specific anti-TNF? production (CN2), and cultured at 37, 33 and 31°C in a batch system. Low temperature led to an increase in the cell viability, the expression of the recombinant anti-TNF? and the production of anti-TNF? both in CN1 and CN2. The higher production of anti-TNF? in CN2 was mainly associated with the large expression of anti-TNF?. Under mild hypothermia myc and xbp1s expression levels were directly correlated to the maximal viable cell density and the specific anti-TNF? productivity, respectively. Moreover, cells showed a simultaneous metabolic shift from production to consumption of lactate and from consumption to production of glutamine, which were exacerbated by reducing culture temperature and coincided with the increased anti-TNF? production. Our current results provide new insights of the regulation of myc and xbp1s in CHO cells at low temperature, and suggest that the presence and magnitude of the metabolic shift might be a relevant metabolic marker of productive cell line.

SUBMITTER: Torres M

PROVIDER: S-EPMC5864046 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Mild hypothermia upregulates myc and xbp1s expression and improves anti-TNFα production in CHO cells.

Torres Mauro M Zúñiga Roberto R Gutierrez Matias M Vergara Mauricio M Collazo Norberto N Reyes Juan J Berrios Julio J Aguillon Juan Carlos JC Molina Maria Carmen MC Altamirano Claudia C Altamirano Claudia C

PloS one 20180322 3

Chinese hamster ovary (CHO) cells are the most frequently used host for commercial production of therapeutic proteins. However, their low protein productivity in culture is the main hurdle to overcome. Mild hypothermia has been established as an effective strategy to enhance protein specific productivity, although the causes of such improvement still remain unclear. The self-regulation of global transcriptional regulatory factors, such as Myc and XBP1s, seems to be involved in increased the reco ...[more]

PMID: 29566086

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Project description:Bone marrow mesenchymal stem cells (BMSCs) are used as a great promising choice for the treatment of cerebral ischemia. Herein, we discuss the neuroprotective effects of the combination of BMSCs transplantation and mild hypothermia (MH) in an ischemia-reperfusion rat model. First, BMSCs were isolated using density gradient centrifugation and the adherent screening method, followed by culture, identification and labeling with DAPI. Second, adult male SD rats were divided into 5 groups: sham group (surgery without blockage of middle cerebral artery), model group (middle cerebral artery occlusion (MCAO) was established 2h prior to reperfusion), BMSCs group (injection of BMSCs via the lateral ventricle 24h after MCAO), MH group (mild hypothermia for 3h immediately after MCAO) and combination therapy group (combination of BMSCs and MH). Finally, the modified neurological severity score (mNSS) test was performed to assess behavioral function at different time points (before MCAO, before transplantation, at day 1, day 5 and day 10 after transplantation). After that, the brain was subjected to TTC staining, and the homing and angiogenesis were evaluated by immumofluorescence and immunohistochemistry. Immunofluorescence staining and Western Blot analysis were performed to calculate the percentage of the infarct area and explore glial fibrillary acidic protein (GFAP) and vascular endothelial growth factor (VEGF). Our results showed that the combination therapy significantly decreased mNSS scores (P<0.01) and reduced the percentage of the infarct area (P<0.01) than a single treatment. Moreover, the expression of GFAP and VEGF increased significantly in the combination therapy group (at day 5, day 10 after transplantation; at all time points after transplantation, respectively) compared to the single treatment groups. Taken together, it was suggested that the combination of BMSCs transplantation and MH can significantly reduce the percentage of the infarct area and improve functional recovery by promoting homing and angiogenesis, which may be a beneficial treatment for cerebral ischemia.

Dataset Information

Mild hypothermia upregulates myc and xbp1s expression and improves anti-TNF? production in CHO cells.

Publications

Mild hypothermia upregulates myc and xbp1s expression and improves anti-TNFα production in CHO cells.

Similar Datasets

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