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Caspase-7 deficiency in Chinese hamster ovary cells reduces cell proliferation and viability
Safari, Fatemeh; Farajnia, Safar; Behbahani, Abbas Behzad; Zarredar, Habib; Barekati-Mowahed, Mazyar; Dehghani, Hesam.
  • Safari, Fatemeh; Tabriz University of Medical Sciences. Faculty of Advanced Medical Sciences. Department of Medical Biotechnology. Tabriz. IR
  • Farajnia, Safar; Tabriz University of Medical Sciences. Biotechnology Research Center. Tabriz. IR
  • Behbahani, Abbas Behzad; Shiraz University of Medical Sciences. School of Paramedical Sciences. Diagnostic Laboratory Sciences and Technology Research Center. Shiraz. IR
  • Zarredar, Habib; Tabriz University of Medical Sciences. Tuberculosis and Lung Diseases Research Center. Tabriz. IR
  • Barekati-Mowahed, Mazyar; Case Western Reserve University. School of Medicine. Department of Physiology & Biophysics. Cleveland. US
  • Dehghani, Hesam; Ferdowsi University of Mashhad. Faculty of Veterinary Medicine. Department of Basic Sciences. Mashhad. IR
Biol. Res ; 53: 52, 2020. tab, graf
Article in English | LILACS | ID: biblio-1142419
ABSTRACT

BACKGROUND:

Chinese hamster ovary (CHO) cells are the most commonly used mammalian host cell In the commercial-scale production of biopharmaceutical proteins. Modification of genes involved in apoptosis may improve the productivity of CHO cells. Executive caspases, including caspases 3 and 7, play critical roles in apoptosis. The effects of the ablation of the caspase 7 gene on proliferation and viability of CHO cells remains unknown. In this study, we applied clustered regularly interspaced short palindromic repeat (CRISPR/Cas9) to target caspase 7 gene of CHO K1 cell via all in one and homology targeted integration strategies. Consequently, the effect of caspase 7 deficiency on cell proliferation, viability, and apoptosis was studied by MTT assay and flow cytometry.

RESULTS:

Findings of gel electrophoresis, western blotting, and sequencing confirmed the caspase 7 gene silencing in CHO cells (CHO-KO). Proliferation assay revealed that caspase 7 deficiency in CHO cells resulted in the reduction of proliferation in various CHO-KO clones. Besides, the disruption of caspase 7 had negative effects on cell viability in exposure with NaBu which confirmed by MTT assay. Results of flow cytometry using Anexin V/PI demonstrated that Nabu treatment (11 mM) declined the percentage of live CHO-K1 and CHO-KO cells to 70.3% and 5.79%. These results verified that the CHO-K1 cells were more resistant to apoptosis than CHO-KO, however most of CHO-KO cells undergone early apoptosis (91.9%) which seems to be a fascinating finding.

CONCLUSION:

These results reveal that caspase 7 may be involved in the cell cycle progression of CHO cells. Furthermore, it seems that targeting caspase 7 is not the ideal route as it had previously been imagined within the prevention of apoptosis but the relation between caspase 7 deficiency, cell cycle arrest, and the occurrence of early apoptosis will require more investigation.
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Full text: Available Index: LILACS (Americas) Main subject: Cell Survival / Apoptosis / Cell Proliferation / Caspase 7 Limits: Animals Language: English Journal: Biol. Res Journal subject: Biology Year: 2020 Type: Article Affiliation country: Iran / United States Institution/Affiliation country: Case Western Reserve University/US / Ferdowsi University of Mashhad/IR / Shiraz University of Medical Sciences/IR / Tabriz University of Medical Sciences/IR

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Full text: Available Index: LILACS (Americas) Main subject: Cell Survival / Apoptosis / Cell Proliferation / Caspase 7 Limits: Animals Language: English Journal: Biol. Res Journal subject: Biology Year: 2020 Type: Article Affiliation country: Iran / United States Institution/Affiliation country: Case Western Reserve University/US / Ferdowsi University of Mashhad/IR / Shiraz University of Medical Sciences/IR / Tabriz University of Medical Sciences/IR