Transgenic chinese hamster V79 cell lines which exhibit variable levels ofgpt mutagenesis

1990 ◽  
Vol 16 (1) ◽  
pp. 1-12 ◽  
Author(s):  
Catherine B. Klein ◽  
Toby G. Rossman ◽  
T. R. Skopek
Keyword(s):  
Cell Lines ◽  
Chinese Hamster ◽  
V79 Cell

Heterologous Expression of Mouse Cytochrome P450 2e1 in V79 Cells: Construction and Characterisation of the Cell Line and Comparison with V79 Cell Lines Stably Expressing Rat P450 2E1 and Human P450 2E1

2003 ◽  
Vol 31 (1) ◽  
pp. 21-30 ◽  
Author(s):  
Ulrike Bernauer ◽  
Hansruedi Glatt ◽  
Barbara Heinrich-Hirsch ◽  
Yungang Liu ◽  
Eva Muckel ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Cell Line ◽  
Cell Lines ◽  
Active Form ◽  
Chinese Hamster ◽  
Established Cell Line ◽  
Cytochrome P450 2E1 ◽  
Hydroxylase Activity ◽  
Catalytically Active ◽  
V79 Cell

A V79 Chinese hamster cell line was constructed for stable expression of mouse cytochrome P450 2e1 (Cyp2e1), as an addition to the existing cell battery consisting of cell lines stably expressing rat CYP2E1 and human CYP2E1 (V79 Cell Battery™). The aim was to establish a cell battery that offers the in vitro possibility of investigating species-specific differences in the toxicity and metabolism of chemicals representing substrates for CYP2E1. The newly established cell line (V79m2E1) effectively expressed Cyp2e1 in the catalytically active form. The expression of catalytically active CYP2E1 in V79m2E1 cells was maintained over several months in culture, as demonstrated by Western Blotting and chlorzoxazone (CLX) 6-hydroxylase activity. The cells exhibited CLX 6-hydroxylase activity with a Km of 27.8μM/l and Vmax of 40pmol/mg protein/minute, compared with a Km of 28.2/28.6μM/l and a Vmax of 130/60pmol/mg protein/minute from V79r2E1/V79h2E1 cells. Furthermore, the CYP2E1-dependent mutagenicity of N-nitrosodimethylamine could be demonstrated in the V79m2E1 cells. Therefore, the new cell battery permits the interspecies comparison of CYP2E1-dependent toxicity and of metabolism of chemicals between humans and the two major rodent species — the rat and the mouse — that are usually used in classical toxicity studies.

scholarly journals Differential activation of the hprt gene on the inactive X chromosome in primary and transformed Chinese hamster cells.

1989 ◽  
Vol 9 (4) ◽  
pp. 1635-1641 ◽  
Author(s):  
S G Grant ◽  
R G Worton
Keyword(s):  
Cell Lines ◽  
X Chromosome ◽  
Gene Activation ◽  
Chinese Hamster ◽  
Fibroblast Cell ◽  
Dna Demethylation ◽  
Hprt Gene ◽  
Primary Fibroblast ◽  
Chinese Hamster Cells ◽  
Inactive X Chromosome

We have investigated the genetic activation of the hprt (hypoxanthine-guanine phosphoribosyltransferase) gene located on the inactive X chromosome in primary and transformed female diploid Chinese hamster cells after treatment with the DNA methylation inhibitor 5-azacytidine (5azaCR). Mutants deficient in HPRT were first selected by growth in 6-thioguanine from two primary fibroblast cell lines and from transformed lines derived from them. These HPRT- mutants were then treated with 5azaCR and plated in HAT (hypoxanthine-methotrexate-thymidine) medium to select for cells that had reexpressed the hprt gene on the inactive X chromosome. Contrary to previous results with primary human cells, 5azaCR was effective in activating the hprt gene in primary Chinese hamster fibroblasts at a low but reproducible frequency of 2 x 10(-6) to 7 x 10(-6). In comparison, the frequency in independently derived transformed lines varied from 1 x 10(-5) to 5 x 10(-3), consistently higher than in the nontransformed cells. This increase remained significant when the difference in growth rates between the primary and transformed lines was taken into account. Treatment with 5azaCR was also found to induce transformation in the primary cell lines but at a low frequency of 4 x 10(-7) to 8 x 10(-7), inconsistent with a two-step model of transformation followed by gene activation to explain the derepression of hprt in primary cells. Thus, these results indicate that upon transformation, the hprt gene on the inactive Chinese hamster X chromosome is rendered more susceptible to action by 5azaCR, consistent with a generalized DNA demethylation associated with the transformation event or with an increase in the instability of an underlying primary mechanism of X inactivation.

scholarly journals Affecting HEK293 Cell Growth and Production Performance by Modifying the Expression of Specific Genes

Cells ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1667
Author(s):  
Laura Abaandou ◽  
David Quan ◽  
Joseph Shiloach
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Hek293 Cell ◽  
Chinese Hamster ◽  
Production Performance ◽  
Cellular Processes ◽  
Hek293 Cell Line ◽  
Global Engineering ◽  
Genomic Tools ◽  
Serum Free Suspension

The HEK293 cell line has earned its place as a producer of biotherapeutics. In addition to its ease of growth in serum-free suspension culture and its amenability to transfection, this cell line’s most important attribute is its human origin, which makes it suitable to produce biologics intended for human use. At the present time, the growth and production properties of the HEK293 cell line are inferior to those of non-human cell lines, such as the Chinese hamster ovary (CHO) and the murine myeloma NSO cell lines. However, the modification of genes involved in cellular processes, such as cell proliferation, apoptosis, metabolism, glycosylation, secretion, and protein folding, in addition to bioprocess, media, and vector optimization, have greatly improved the performance of this cell line. This review provides a comprehensive summary of important achievements in HEK293 cell line engineering and on the global engineering approaches and functional genomic tools that have been employed to identify relevant genes for targeted engineering.

scholarly journals Recombinant γY278H Fibrinogen Showed Normal Secretion from CHO Cells, but a Corresponding Heterozygous Patient Showed Hypofibrinogenemia

2021 ◽  
Vol 22 (10) ◽  
pp. 5218
Author(s):  
Tomu Kamijo ◽  
Takahiro Kaido ◽  
Masahiro Yoda ◽  
Shinpei Arai ◽  
Kazuyoshi Yamauchi ◽  
...  
Keyword(s):  
Cell Lines ◽  
Cho Cells ◽  
Fibrinogen Level ◽  
Culture Media ◽  
Chinese Hamster ◽  
Plasma Fibrinogen ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cho Cell ◽  
Accelerated Degradation ◽  
Heterozygous Patient

We identified a novel heterozygous hypofibrinogenemia, γY278H (Hiroshima). To demonstrate the cause of reduced plasma fibrinogen levels (functional level: 1.12 g/L and antigenic level: 1.16 g/L), we established γY278H fibrinogen-producing Chinese hamster ovary (CHO) cells. An enzyme-linked immunosorbent assay demonstrated that synthesis of γY278H fibrinogen inside CHO cells and secretion into the culture media were not reduced. Then, we established an additional five variant fibrinogen-producing CHO cell lines (γL276P, γT277P, γT277R, γA279D, and γY280C) and conducted further investigations. We have already established 33 γ-module variant fibrinogen-producing CHO cell lines, including 6 cell lines in this study, but only the γY278H and γT277R cell lines showed disagreement, namely, recombinant fibrinogen production was not reduced but the patients’ plasma fibrinogen level was reduced. Finally, we performed fibrinogen degradation assays and demonstrated that the γY278H and γT277R fibrinogens were easily cleaved by plasmin whereas their polymerization in the presence of Ca2+ and “D:D” interaction was normal. In conclusion, our investigation suggested that patient γY278H showed hypofibrinogenemia because γY278H fibrinogen was secreted normally from the patient’s hepatocytes but then underwent accelerated degradation by plasmin in the circulation.

scholarly journals Chinese hamster ovary cell line DXB-11: chromosomal instability and karyotype heterogeneity

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Victoria I. Turilova ◽  
Tatyana S. Goryachaya ◽  
Tatiana K. Yakovleva
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Chinese Hamster Ovary Cell ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster ◽  
Chromosome 8 ◽  
Ovary Cell ◽  
Differential Staining ◽  
Nucleolar Organizer Regions ◽  
Karyotype Heterogeneity

Abstract Background Chinese hamster ovary cell lines, also known as CHO cells, represent a large family of related, yet quite different, cell lines which are metabolic mutants derived from the original cell line, CHO-ori. Dihydrofolate reductase-deficient DXB-11 cell line, one of the first CHO derivatives, serves as the host cell line for the production of therapeutic proteins. It is generally assumed that DXB-11 is identical to DUKX or CHO-DUK cell lines, but, to our knowledge, DXB-11 karyotype has not been described yet. Results Using differential staining approaches (G-, C-banding and Ag-staining), we presented DXB-11 karyotype and revealed that karyotypes of DXB-11 and CHO-DUK cells have a number of differences. Although the number of chromosomes is equal—20 in each cell line—DXB-11 has normal chromosomes of the 1st and 5th pairs as well as an intact chromosome 8. Besides, in DXB-11 line, chromosome der(Z9) includes the material of chromosomes X and 6, whereas in CHO-DUK it results from the translocation of chromosomes 1 and 6. Ag-positive nucleolar organizer regions were revealed in the long arms of chromosome del(4)(q11q12) and both chromosome 5 homologues, as well as in the short arms of chromosomes 8 and add(8)(q11). Only 19 from 112 (16.96%) DXB-11 cells display identical chromosome complement accepted as the main structural variant of karyotype. The karyotype heterogeneity of all the rest of cells (93, 83.04%) occurs due to clonal and nonclonal additional structural rearrangements of chromosomes. Estimation of the frequency of chromosome involvement in these rearrangements allowed us to reveal that chromosomes 9, der(X)t(X;3;4), del(2)(p21p23), del(2)(q11q22) /Z2, der(4) /Z7, add(6)(p11) /Z8 are the most stable, whereas mar2, probably der(10), is the most unstable chromosome. A comparative analysis of our own and literary data on CHO karyotypes allowed to designate conservative chromosomes, both normal and rearranged, that remain unchanged in different CHO cell lines, as well as variable chromosomes that determine the individuality of karyotypes of CHO derivatives. Conclusion DXB-11and CHO-DUK cell lines differ in karyotypes. The revealed differential instability of DXB-11 chromosomes is likely not incidental and results in karyotype heterogeneity of cell population.

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