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Juglone induces apoptosis of tumor stem-like cells through ROS-p38 pathway in glioblastoma

View Article: PubMed Central - PubMed

ABSTRACT

Background: Juglone is a natural pigment, which has cytotoxic effect against various human tumor cells. However, its cytotoxicity to glioma cells, especially to tumor stem-like cells (TSCs) has not been demonstrated.

Methods: TSCs of glioma were enriched from U87 and two primary cells (SHG62, and SHG66) using serum-free medium supplemented with growth factors, including bFGF, EGF and B27. After treatment of juglone with gradient concentrations (0, 10, 20, and 40 μM), the viability and apoptosis of TSCs were evaluated by WST-8 assay and flow cytometry. Reactive oxygen species (ROS) was labeled by the cell-permeable fluorescent probe and detected with flow cytometry. ROS scavenger (NAC) and p38-MAPK inhibitor (SB203580) were applied to resist the cytotoxic effect. Caspase 9 cleavage and p38 phosphorylation (P-p38) were quantified by western blot. Juglone as well as temozolomide (TMZ) were administrated in intracranial xenografts and MR scan was performed every week to evaluate the anti-tumor effect in vivo.

Results: Juglone could obviously inhibit the proliferation of TSCs in glioma by decreasing cell viability (P < 0.01) and inducing apoptosis (P < 0.01), which was accompanied by increased caspase 9 cleavage in a dose-dependent manner (P < 0.01). In the meantime, juglone could generate ROS significantly and increase p38 phosphorylation (P < 0.01). In addition, pretreatment with ROS scavenger or p38-MAPK inhibitor could reverse juglone-induced cytotoxicity (P < 0.01). More importantly, juglone could also suppress tumor growth in vivo and improve the survival of U87-bearing mice compared with control (P < 0.05), although TMZ seemed to have better effect.

Conclusions: Juglone could inhibit the growth of TSCs in gliomas through the activation of ROS-p38-MAPK pathway in vitro, and the anti-glioma effect was validated in vivo, which offers a potential therapeutic agent to gliomas.

No MeSH data available.


Related in: MedlinePlus

Pretreatment with NAC and SB203580 could reverse juglone-induced inhibition of glioma stem-like cells. a Statistic analysis indicated that NAC or SB203580 could resist juglone-induced cell viability decrease (**P < 0.01). b Statistic analysis indicated that NAC or SB203580 could resist juglone-induced cell death (**P < 0.01). c Flow cytometry showed that NAC or SB203580 pretreatment could reverse the decrease of cell viability and the increase of cell death induced by juglone (**P < 0.01)
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Fig4: Pretreatment with NAC and SB203580 could reverse juglone-induced inhibition of glioma stem-like cells. a Statistic analysis indicated that NAC or SB203580 could resist juglone-induced cell viability decrease (**P < 0.01). b Statistic analysis indicated that NAC or SB203580 could resist juglone-induced cell death (**P < 0.01). c Flow cytometry showed that NAC or SB203580 pretreatment could reverse the decrease of cell viability and the increase of cell death induced by juglone (**P < 0.01)

Mentions: As demonstrated by WST-8 assay (Fig. 4a), juglone (40 μM) treatment could decreased cell viability by 65 ± 2.53% (P < 0.01), whereas NAC and SB203580 could reversed the cytotoxic effect of juglone by 75 ± 3.18% (P < 0.01) and 58 ± 3.92%(P < 0.01). Cell death data (Fig. 4b and c) showed that treatment with juglone (40 μM) increased cell death by 17 ± 3.87% (P < 0.01), while NAC and SB203580 pretreatment could reverse juglone-mediated increases of cell death by 12.4 ± 2.33% (P < 0.01) and 7.1 ± 2.91% (P < 0.01). All those results indicated that ROS-p38-MAPK pathway was involved in the juglone-induced cytotoxicity.Fig. 4


Juglone induces apoptosis of tumor stem-like cells through ROS-p38 pathway in glioblastoma
Pretreatment with NAC and SB203580 could reverse juglone-induced inhibition of glioma stem-like cells. a Statistic analysis indicated that NAC or SB203580 could resist juglone-induced cell viability decrease (**P < 0.01). b Statistic analysis indicated that NAC or SB203580 could resist juglone-induced cell death (**P < 0.01). c Flow cytometry showed that NAC or SB203580 pretreatment could reverse the decrease of cell viability and the increase of cell death induced by juglone (**P < 0.01)
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Related In: Results  -  Collection

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Fig4: Pretreatment with NAC and SB203580 could reverse juglone-induced inhibition of glioma stem-like cells. a Statistic analysis indicated that NAC or SB203580 could resist juglone-induced cell viability decrease (**P < 0.01). b Statistic analysis indicated that NAC or SB203580 could resist juglone-induced cell death (**P < 0.01). c Flow cytometry showed that NAC or SB203580 pretreatment could reverse the decrease of cell viability and the increase of cell death induced by juglone (**P < 0.01)
Mentions: As demonstrated by WST-8 assay (Fig. 4a), juglone (40 μM) treatment could decreased cell viability by 65 ± 2.53% (P < 0.01), whereas NAC and SB203580 could reversed the cytotoxic effect of juglone by 75 ± 3.18% (P < 0.01) and 58 ± 3.92%(P < 0.01). Cell death data (Fig. 4b and c) showed that treatment with juglone (40 μM) increased cell death by 17 ± 3.87% (P < 0.01), while NAC and SB203580 pretreatment could reverse juglone-mediated increases of cell death by 12.4 ± 2.33% (P < 0.01) and 7.1 ± 2.91% (P < 0.01). All those results indicated that ROS-p38-MAPK pathway was involved in the juglone-induced cytotoxicity.Fig. 4

View Article: PubMed Central - PubMed

ABSTRACT

Background: Juglone is a natural pigment, which has cytotoxic effect against various human tumor cells. However, its cytotoxicity to glioma cells, especially to tumor stem-like cells (TSCs) has not been demonstrated.

Methods: TSCs of glioma were enriched from U87 and two primary cells (SHG62, and SHG66) using serum-free medium supplemented with growth factors, including bFGF, EGF and B27. After treatment of juglone with gradient concentrations (0, 10, 20, and 40&nbsp;&mu;M), the viability and apoptosis of TSCs were evaluated by WST-8 assay and flow cytometry. Reactive oxygen species (ROS) was labeled by the cell-permeable fluorescent probe and detected with flow cytometry. ROS scavenger (NAC) and p38-MAPK inhibitor (SB203580) were applied to resist the cytotoxic effect. Caspase 9 cleavage and p38 phosphorylation (P-p38) were quantified by western blot. Juglone as well as temozolomide (TMZ) were administrated in intracranial xenografts and MR scan was performed every week to evaluate the anti-tumor effect in vivo.

Results: Juglone could obviously inhibit the proliferation of TSCs in glioma by decreasing cell viability (P&thinsp;&lt;&thinsp;0.01) and inducing apoptosis (P&thinsp;&lt;&thinsp;0.01), which was accompanied by increased caspase 9 cleavage in a dose-dependent manner (P&thinsp;&lt;&thinsp;0.01). In the meantime, juglone could generate ROS significantly and increase p38 phosphorylation (P&thinsp;&lt;&thinsp;0.01). In addition, pretreatment with ROS scavenger or p38-MAPK inhibitor could reverse juglone-induced cytotoxicity (P&thinsp;&lt;&thinsp;0.01). More importantly, juglone could also suppress tumor growth in vivo and improve the survival of U87-bearing mice compared with control (P&thinsp;&lt;&thinsp;0.05), although TMZ seemed to have better effect.

Conclusions: Juglone could inhibit the growth of TSCs in gliomas through the activation of ROS-p38-MAPK pathway in vitro, and the anti-glioma effect was validated in vivo, which offers a potential therapeutic agent to gliomas.

No MeSH data available.


Related in: MedlinePlus