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The impact of tumor nitric oxide production on VEGFA expression and tumor growth in a zebrafish rat glioma xenograft model.

Yousfi N, Pruvot B, Lopez T, Magadoux L, Franche N, Pichon L, Salvadori F, Solary E, Garrido C, Laurens V, Chluba J - PLoS ONE (2015)

Bottom Line: Furthermore, we demonstrated by qRT-PCR that the transplanted glioma cells highly expressed Nos2, Vegfa and Cyclin D1 mRNA.In the xenografted embryos we also found increased zebrafish vegfa expression.We conclude that even if there is a heterogeneous nitric oxide production by the xenografted glioma cells that impacts Vegfa and Cyclin D1 expression levels, our results suggest that reduction of nitric oxide levels by nitric oxide scavenging could be an efficient approach to treat glioma.

View Article: PubMed Central - PubMed

Affiliation: INSERM, UMR 866, 'Equipe Labellisée Ligue Contre le Cancer', Dijon, France; University of Burgundy, UFR SVTE, Dijon, France.

ABSTRACT
To investigate the effect of nitric oxide on tumor development, we established a rat tumor xenograft model in zebrafish embryos. The injected tumor cells formed masses in which nitric oxide production could be detected by the use of the cell-permeant DAF-FM-DA (diaminofluorophore 4-amino-5-methylamino-2'-7'-difluorofluorescein diacetate) and DAR-4M-AM (diaminorhodamine-4M). This method revealed that nitric oxide production could be co-localized with the tumor xenograft in 46% of the embryos. In 85% of these embryos, tumors were vascularized and blood vessels were observed on day 4 post injection. Furthermore, we demonstrated by qRT-PCR that the transplanted glioma cells highly expressed Nos2, Vegfa and Cyclin D1 mRNA. In the xenografted embryos we also found increased zebrafish vegfa expression. Glioma and zebrafish derived Vegfa and tumor Cyclin D1 expression could be down regulated by the nitric oxide scavenger 2-(4-Carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide or CPTIO. We conclude that even if there is a heterogeneous nitric oxide production by the xenografted glioma cells that impacts Vegfa and Cyclin D1 expression levels, our results suggest that reduction of nitric oxide levels by nitric oxide scavenging could be an efficient approach to treat glioma.

No MeSH data available.


Related in: MedlinePlus

The effect of NO on tumor growth.CM-Dil labeled glioma cells were injected into the yolk of zebrafish embryos at 2 dpf and incubated with CPTIO (200μM). Images were taken at 1 dpi and 4 dpi. Images (A-A”‘) and (B-B”‘) show the tumors at 1 dpi and 4 dpi for control and CPTIO treated embryo. At 4 dpi, tumors have increased in size in untreated embryos (A”-A”‘) in comparison to CPTIO treated embryos (B”-B”‘). (C) Percentage of fluorescence intensity of the tumor surfaces was determined by ZEN software (n = 10). Images were taken with an Axio zoom V16 (Zeiss) macroscope, with the same exposure time for all embryos. Error bars represent SEM, *** p <0, 001, (one way ANOVA). (D) QRT-PCR analysis of Cyclin D1 gene expression in untreated and CPTIO treated xenografted embryos. Cyclin D1 expression was decreased upon CPTIO treatment. One representative experiment from four independent experiments is shown.
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pone.0120435.g006: The effect of NO on tumor growth.CM-Dil labeled glioma cells were injected into the yolk of zebrafish embryos at 2 dpf and incubated with CPTIO (200μM). Images were taken at 1 dpi and 4 dpi. Images (A-A”‘) and (B-B”‘) show the tumors at 1 dpi and 4 dpi for control and CPTIO treated embryo. At 4 dpi, tumors have increased in size in untreated embryos (A”-A”‘) in comparison to CPTIO treated embryos (B”-B”‘). (C) Percentage of fluorescence intensity of the tumor surfaces was determined by ZEN software (n = 10). Images were taken with an Axio zoom V16 (Zeiss) macroscope, with the same exposure time for all embryos. Error bars represent SEM, *** p <0, 001, (one way ANOVA). (D) QRT-PCR analysis of Cyclin D1 gene expression in untreated and CPTIO treated xenografted embryos. Cyclin D1 expression was decreased upon CPTIO treatment. One representative experiment from four independent experiments is shown.

Mentions: We assessed the effect of NO on tumor growth by imaging untreated and CPTIO treated xenografted embryos and by comparing tumor size at 1 dpi and 4 dpi in both groups. At 4 dpi, in the untreated group tumors have grown with local invasion of the yolk (Fig. 6A), whereas in the CPTIO group, the tumor size decreased (Fig. 6B). Moreover, at 4 dpi the tumors of CPTIO treated embryos displayed 20% lower fluorescence intensity per tumor surface than those of the control group (Fig. 6C), which is a sign of a loss of tumor cells.


The impact of tumor nitric oxide production on VEGFA expression and tumor growth in a zebrafish rat glioma xenograft model.

Yousfi N, Pruvot B, Lopez T, Magadoux L, Franche N, Pichon L, Salvadori F, Solary E, Garrido C, Laurens V, Chluba J - PLoS ONE (2015)

The effect of NO on tumor growth.CM-Dil labeled glioma cells were injected into the yolk of zebrafish embryos at 2 dpf and incubated with CPTIO (200μM). Images were taken at 1 dpi and 4 dpi. Images (A-A”‘) and (B-B”‘) show the tumors at 1 dpi and 4 dpi for control and CPTIO treated embryo. At 4 dpi, tumors have increased in size in untreated embryos (A”-A”‘) in comparison to CPTIO treated embryos (B”-B”‘). (C) Percentage of fluorescence intensity of the tumor surfaces was determined by ZEN software (n = 10). Images were taken with an Axio zoom V16 (Zeiss) macroscope, with the same exposure time for all embryos. Error bars represent SEM, *** p <0, 001, (one way ANOVA). (D) QRT-PCR analysis of Cyclin D1 gene expression in untreated and CPTIO treated xenografted embryos. Cyclin D1 expression was decreased upon CPTIO treatment. One representative experiment from four independent experiments is shown.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4359111&req=5

pone.0120435.g006: The effect of NO on tumor growth.CM-Dil labeled glioma cells were injected into the yolk of zebrafish embryos at 2 dpf and incubated with CPTIO (200μM). Images were taken at 1 dpi and 4 dpi. Images (A-A”‘) and (B-B”‘) show the tumors at 1 dpi and 4 dpi for control and CPTIO treated embryo. At 4 dpi, tumors have increased in size in untreated embryos (A”-A”‘) in comparison to CPTIO treated embryos (B”-B”‘). (C) Percentage of fluorescence intensity of the tumor surfaces was determined by ZEN software (n = 10). Images were taken with an Axio zoom V16 (Zeiss) macroscope, with the same exposure time for all embryos. Error bars represent SEM, *** p <0, 001, (one way ANOVA). (D) QRT-PCR analysis of Cyclin D1 gene expression in untreated and CPTIO treated xenografted embryos. Cyclin D1 expression was decreased upon CPTIO treatment. One representative experiment from four independent experiments is shown.
Mentions: We assessed the effect of NO on tumor growth by imaging untreated and CPTIO treated xenografted embryos and by comparing tumor size at 1 dpi and 4 dpi in both groups. At 4 dpi, in the untreated group tumors have grown with local invasion of the yolk (Fig. 6A), whereas in the CPTIO group, the tumor size decreased (Fig. 6B). Moreover, at 4 dpi the tumors of CPTIO treated embryos displayed 20% lower fluorescence intensity per tumor surface than those of the control group (Fig. 6C), which is a sign of a loss of tumor cells.

Bottom Line: Furthermore, we demonstrated by qRT-PCR that the transplanted glioma cells highly expressed Nos2, Vegfa and Cyclin D1 mRNA.In the xenografted embryos we also found increased zebrafish vegfa expression.We conclude that even if there is a heterogeneous nitric oxide production by the xenografted glioma cells that impacts Vegfa and Cyclin D1 expression levels, our results suggest that reduction of nitric oxide levels by nitric oxide scavenging could be an efficient approach to treat glioma.

View Article: PubMed Central - PubMed

Affiliation: INSERM, UMR 866, 'Equipe Labellisée Ligue Contre le Cancer', Dijon, France; University of Burgundy, UFR SVTE, Dijon, France.

ABSTRACT
To investigate the effect of nitric oxide on tumor development, we established a rat tumor xenograft model in zebrafish embryos. The injected tumor cells formed masses in which nitric oxide production could be detected by the use of the cell-permeant DAF-FM-DA (diaminofluorophore 4-amino-5-methylamino-2'-7'-difluorofluorescein diacetate) and DAR-4M-AM (diaminorhodamine-4M). This method revealed that nitric oxide production could be co-localized with the tumor xenograft in 46% of the embryos. In 85% of these embryos, tumors were vascularized and blood vessels were observed on day 4 post injection. Furthermore, we demonstrated by qRT-PCR that the transplanted glioma cells highly expressed Nos2, Vegfa and Cyclin D1 mRNA. In the xenografted embryos we also found increased zebrafish vegfa expression. Glioma and zebrafish derived Vegfa and tumor Cyclin D1 expression could be down regulated by the nitric oxide scavenger 2-(4-Carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide or CPTIO. We conclude that even if there is a heterogeneous nitric oxide production by the xenografted glioma cells that impacts Vegfa and Cyclin D1 expression levels, our results suggest that reduction of nitric oxide levels by nitric oxide scavenging could be an efficient approach to treat glioma.

No MeSH data available.


Related in: MedlinePlus