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A novel peptide to enhance recombinant BMP-2 production in mammalian cell cultures.

Zhou AJ, Clokie CM, Peel SA - BMC Proc (2011)

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Oral and Maxillofacial Surgery, Faculty of Dentistry, University of Toronto, Toronto, Ontario Canada, M5G 1G6.

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Furin-specific siRNA was used to knock down the furin expression in CHO-BMP2 cells and its effect on the rhBMP-2 production was determined... Stably transfected CHO-BMP2 cells secreted 36 kDa rhBMP-2 dimers that were biologically active... In 24 h cell cultures, IND-1 treated cells produced significantly greater amounts of proBMP-2 (≥ 10-fold, P <0.001) and mature BMP-2 (≥ 3-fold, P <0.001) in their conditioned medium (Figure 1)... In long-term CHO-BMP2 culture, IND-1 continued to increase the yields of BMP-2 (≥ 50%) and proBMP-2 (≥ 2-fold) without affecting cell growth or viability... IND-1 treatment had no effect on the mRNA level of BMP-2 and furin, indicating IND-1 affects rhBMP-2 yield post-transcriptionally... While IND-1 was taken up by the cells and inhibited PC activity when added directly to the cell lysates, cells cultured with IND-1 showed no changes in their PC activity at doses 50 times higher than required to affect BMP-2 yields... Furthermore, knockdown of furin at both the mRNA (≥ 80%, P <0.001) and the protein level (≥ 70%, P <0.001), did not affect rhBMP-2 yields... These results suggest that furin inhibition is most likely not the mechanism by which IND-1 enhances rhBMP-2 yields... The addition of a novel peptide IND-1 to the cell culture medium significantly enhanced the yields of both pro- and mature BMP-2 in stably transfected CHO and HEK cell lines.

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After 24 h incubation with or without IND-1, the amount of proBMP-2 and mature BMP-2 proteins in (A) CHO and (B) HEK conditioned media were measured by proBMP-2 and BMP-2 ELISA. Data are presented as mean ± SEM (n = 3 experiments) (**P <0.01, ***P <0.001).
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Figure 1: After 24 h incubation with or without IND-1, the amount of proBMP-2 and mature BMP-2 proteins in (A) CHO and (B) HEK conditioned media were measured by proBMP-2 and BMP-2 ELISA. Data are presented as mean ± SEM (n = 3 experiments) (**P <0.01, ***P <0.001).

Mentions: Stably transfected CHO-BMP2 cells secreted 36 kDa rhBMP-2 dimers that were biologically active. In 24 h cell cultures, IND-1 treated cells produced significantly greater amounts of proBMP-2 (≥ 10-fold, P <0.001) and mature BMP-2 (≥ 3-fold, P <0.001) in their conditioned medium (Figure 1). In long-term CHO-BMP2 culture, IND-1 continued to increase the yields of BMP-2 (≥ 50%) and proBMP-2 (≥ 2-fold) without affecting cell growth or viability.


A novel peptide to enhance recombinant BMP-2 production in mammalian cell cultures.

Zhou AJ, Clokie CM, Peel SA - BMC Proc (2011)

After 24 h incubation with or without IND-1, the amount of proBMP-2 and mature BMP-2 proteins in (A) CHO and (B) HEK conditioned media were measured by proBMP-2 and BMP-2 ELISA. Data are presented as mean ± SEM (n = 3 experiments) (**P <0.01, ***P <0.001).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3285006&req=5

Figure 1: After 24 h incubation with or without IND-1, the amount of proBMP-2 and mature BMP-2 proteins in (A) CHO and (B) HEK conditioned media were measured by proBMP-2 and BMP-2 ELISA. Data are presented as mean ± SEM (n = 3 experiments) (**P <0.01, ***P <0.001).
Mentions: Stably transfected CHO-BMP2 cells secreted 36 kDa rhBMP-2 dimers that were biologically active. In 24 h cell cultures, IND-1 treated cells produced significantly greater amounts of proBMP-2 (≥ 10-fold, P <0.001) and mature BMP-2 (≥ 3-fold, P <0.001) in their conditioned medium (Figure 1). In long-term CHO-BMP2 culture, IND-1 continued to increase the yields of BMP-2 (≥ 50%) and proBMP-2 (≥ 2-fold) without affecting cell growth or viability.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Oral and Maxillofacial Surgery, Faculty of Dentistry, University of Toronto, Toronto, Ontario Canada, M5G 1G6.

AUTOMATICALLY GENERATED EXCERPT
Please rate it.

Furin-specific siRNA was used to knock down the furin expression in CHO-BMP2 cells and its effect on the rhBMP-2 production was determined... Stably transfected CHO-BMP2 cells secreted 36 kDa rhBMP-2 dimers that were biologically active... In 24 h cell cultures, IND-1 treated cells produced significantly greater amounts of proBMP-2 (≥ 10-fold, P <0.001) and mature BMP-2 (≥ 3-fold, P <0.001) in their conditioned medium (Figure 1)... In long-term CHO-BMP2 culture, IND-1 continued to increase the yields of BMP-2 (≥ 50%) and proBMP-2 (≥ 2-fold) without affecting cell growth or viability... IND-1 treatment had no effect on the mRNA level of BMP-2 and furin, indicating IND-1 affects rhBMP-2 yield post-transcriptionally... While IND-1 was taken up by the cells and inhibited PC activity when added directly to the cell lysates, cells cultured with IND-1 showed no changes in their PC activity at doses 50 times higher than required to affect BMP-2 yields... Furthermore, knockdown of furin at both the mRNA (≥ 80%, P <0.001) and the protein level (≥ 70%, P <0.001), did not affect rhBMP-2 yields... These results suggest that furin inhibition is most likely not the mechanism by which IND-1 enhances rhBMP-2 yields... The addition of a novel peptide IND-1 to the cell culture medium significantly enhanced the yields of both pro- and mature BMP-2 in stably transfected CHO and HEK cell lines.

No MeSH data available.


Related in: MedlinePlus