Caspase-7 deficiency in Chinese hamster ovary cells reduces cell proliferation and viability
Biol. Res
;
53: 52, 2020. tab, graf
Artigo
em Inglês
| 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.
Texto completo:
DisponíveL
Índice:
LILACS (Américas)
Assunto principal:
Sobrevivência Celular
/
Apoptose
/
Proliferação de Células
/
Caspase 7
Limite:
Animais
Idioma:
Inglês
Revista:
Biol. Res
Assunto da revista:
Biologia
Ano de publicação:
2020
Tipo de documento:
Artigo
País de afiliação:
Irã
/
Estados Unidos
Instituição/País de afiliação:
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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