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Resveratrol Inhibits the Invasion of Glioblastoma-Initiating Cells via Down-Regulation of the PI3K/Akt/NF-κB Signaling Pathway.

Jiao Y, Li H, Liu Y, Guo A, Xu X, Qu X, Wang S, Zhao J, Li Y, Cao Y - Nutrients (2015)

Bottom Line: Resveratrol (RES) is a natural compound that exhibits anti-invasion properties in multiple tumor cell lines.Invasive behaviors, including the adhesion, invasion and migration of GICs, were determined by tumor invasive assays in vitro and in vivo.The activity of matrix metalloproteinases (MMPs) was measured by the gelatin zymography assay.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurosurgery, Beijing Tiantan Hospital, Capital Medical University, Beijing 100050, China. shengxiongyuming@163.com.

ABSTRACT
Invasion and metastasis of glioblastoma-initiating cells (GICs) are thought to be responsible for the progression and recurrence of glioblastoma multiforme (GBM). A safe drug that can be applied during the rest period of temozolomide (TMZ) maintenance cycles would greatly improve the prognosis of GBM patients by inhibiting GIC invasion. Resveratrol (RES) is a natural compound that exhibits anti-invasion properties in multiple tumor cell lines. The current study aimed to evaluate whether RES can inhibit GIC invasion in vitro and in vivo. GICs were identified using CD133 and Nestin immunofluorescence staining and tumorigenesis in non-obese diabetic severe combined immunodeficient (NOD/SCID) mice. Invasive behaviors, including the adhesion, invasion and migration of GICs, were determined by tumor invasive assays in vitro and in vivo. The activity of matrix metalloproteinases (MMPs) was measured by the gelatin zymography assay. Western blotting analysis and immunofluorescence staining were used to determine the expression of signaling effectors in GICs. We demonstrated that RES suppressed the adhesion, invasion and migration of GICs in vitro and in vivo. Moreover, we proved that RES inhibited the invasion of GICs via the inhibition of PI3K/Akt/NF-κB signal transduction and the subsequent suppression of MMP-2 expression.

No MeSH data available.


Related in: MedlinePlus

PI3K/AKT signaling pathway is involved in the inhibitory effect of RES on NF-κB activation. (A) GICs (412) were treated with DMSO (vehicle) and 5 μM, 10 μM, and 20 μM RES for 48 h. Total expression and phosphorylation of Akt and mTOR were analyzed by western blotting using anti-Akt, anti-p-Akt, anti-mTOR, anti-p-mTOR, antibodies. β-actin was used as a loading control; (B) Total expression and phosphorylation of JNK, ERK, and p38 were analyzed by western blotting using anti-JNK, anti-p-JNK, anti-ERK, anti-p-ERK, anti-p38 anti-p-p38 antibodies. β-actin was used as a loading control; (C) GICs (412) were pre-treated with IGF-1 (200 ng·mL−1) or wortmannin (5 μM) for 30 min and then incubated in the presence or absence of RES (10 μM). GICs were then subjected to cell invasion assay. Then, randomly chosen fields were photographed (200×), and the number of cells that migrated to the lower surface was calculated as a percentage of invasion. Data are shown as the mean ± SD of three independent experiments by Student’s t-test. * p < 0.05, ** p < 0.01; (D) GICs were pre-treated with IGF-1 (200 ng·mL−1) or wormannin (5 μM) for 30 min and incubated in the presence or absence of RES (10 μM) for 48 h, and then, the cell lysates were subjected to western blotting with anti-p-Akt and anti-Akt antibodies. β-actin was used as a loading control. The CM were subjected to gelatin zymography assay to determine the activity of MMP-2.
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nutrients-07-04383-f006: PI3K/AKT signaling pathway is involved in the inhibitory effect of RES on NF-κB activation. (A) GICs (412) were treated with DMSO (vehicle) and 5 μM, 10 μM, and 20 μM RES for 48 h. Total expression and phosphorylation of Akt and mTOR were analyzed by western blotting using anti-Akt, anti-p-Akt, anti-mTOR, anti-p-mTOR, antibodies. β-actin was used as a loading control; (B) Total expression and phosphorylation of JNK, ERK, and p38 were analyzed by western blotting using anti-JNK, anti-p-JNK, anti-ERK, anti-p-ERK, anti-p38 anti-p-p38 antibodies. β-actin was used as a loading control; (C) GICs (412) were pre-treated with IGF-1 (200 ng·mL−1) or wortmannin (5 μM) for 30 min and then incubated in the presence or absence of RES (10 μM). GICs were then subjected to cell invasion assay. Then, randomly chosen fields were photographed (200×), and the number of cells that migrated to the lower surface was calculated as a percentage of invasion. Data are shown as the mean ± SD of three independent experiments by Student’s t-test. * p < 0.05, ** p < 0.01; (D) GICs were pre-treated with IGF-1 (200 ng·mL−1) or wormannin (5 μM) for 30 min and incubated in the presence or absence of RES (10 μM) for 48 h, and then, the cell lysates were subjected to western blotting with anti-p-Akt and anti-Akt antibodies. β-actin was used as a loading control. The CM were subjected to gelatin zymography assay to determine the activity of MMP-2.

Mentions: It was reported that the MAPK and PI3K/AKT pathways can regulate fundamental cellular processes by the induction of IKK-dependent NF-κB activation [19,20]. In our study, RES significantly reduced the phosphorylation of Akt and mTOR in GICs without affecting the total level (Figure 6A). However, RES had little effect on the MAPK signaling pathway, which was supported by the unchanged phosphorylation of the ERK1/2, p38, and JNK pathways (Figure 6B).


Resveratrol Inhibits the Invasion of Glioblastoma-Initiating Cells via Down-Regulation of the PI3K/Akt/NF-κB Signaling Pathway.

Jiao Y, Li H, Liu Y, Guo A, Xu X, Qu X, Wang S, Zhao J, Li Y, Cao Y - Nutrients (2015)

PI3K/AKT signaling pathway is involved in the inhibitory effect of RES on NF-κB activation. (A) GICs (412) were treated with DMSO (vehicle) and 5 μM, 10 μM, and 20 μM RES for 48 h. Total expression and phosphorylation of Akt and mTOR were analyzed by western blotting using anti-Akt, anti-p-Akt, anti-mTOR, anti-p-mTOR, antibodies. β-actin was used as a loading control; (B) Total expression and phosphorylation of JNK, ERK, and p38 were analyzed by western blotting using anti-JNK, anti-p-JNK, anti-ERK, anti-p-ERK, anti-p38 anti-p-p38 antibodies. β-actin was used as a loading control; (C) GICs (412) were pre-treated with IGF-1 (200 ng·mL−1) or wortmannin (5 μM) for 30 min and then incubated in the presence or absence of RES (10 μM). GICs were then subjected to cell invasion assay. Then, randomly chosen fields were photographed (200×), and the number of cells that migrated to the lower surface was calculated as a percentage of invasion. Data are shown as the mean ± SD of three independent experiments by Student’s t-test. * p < 0.05, ** p < 0.01; (D) GICs were pre-treated with IGF-1 (200 ng·mL−1) or wormannin (5 μM) for 30 min and incubated in the presence or absence of RES (10 μM) for 48 h, and then, the cell lysates were subjected to western blotting with anti-p-Akt and anti-Akt antibodies. β-actin was used as a loading control. The CM were subjected to gelatin zymography assay to determine the activity of MMP-2.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4488790&req=5

nutrients-07-04383-f006: PI3K/AKT signaling pathway is involved in the inhibitory effect of RES on NF-κB activation. (A) GICs (412) were treated with DMSO (vehicle) and 5 μM, 10 μM, and 20 μM RES for 48 h. Total expression and phosphorylation of Akt and mTOR were analyzed by western blotting using anti-Akt, anti-p-Akt, anti-mTOR, anti-p-mTOR, antibodies. β-actin was used as a loading control; (B) Total expression and phosphorylation of JNK, ERK, and p38 were analyzed by western blotting using anti-JNK, anti-p-JNK, anti-ERK, anti-p-ERK, anti-p38 anti-p-p38 antibodies. β-actin was used as a loading control; (C) GICs (412) were pre-treated with IGF-1 (200 ng·mL−1) or wortmannin (5 μM) for 30 min and then incubated in the presence or absence of RES (10 μM). GICs were then subjected to cell invasion assay. Then, randomly chosen fields were photographed (200×), and the number of cells that migrated to the lower surface was calculated as a percentage of invasion. Data are shown as the mean ± SD of three independent experiments by Student’s t-test. * p < 0.05, ** p < 0.01; (D) GICs were pre-treated with IGF-1 (200 ng·mL−1) or wormannin (5 μM) for 30 min and incubated in the presence or absence of RES (10 μM) for 48 h, and then, the cell lysates were subjected to western blotting with anti-p-Akt and anti-Akt antibodies. β-actin was used as a loading control. The CM were subjected to gelatin zymography assay to determine the activity of MMP-2.
Mentions: It was reported that the MAPK and PI3K/AKT pathways can regulate fundamental cellular processes by the induction of IKK-dependent NF-κB activation [19,20]. In our study, RES significantly reduced the phosphorylation of Akt and mTOR in GICs without affecting the total level (Figure 6A). However, RES had little effect on the MAPK signaling pathway, which was supported by the unchanged phosphorylation of the ERK1/2, p38, and JNK pathways (Figure 6B).

Bottom Line: Resveratrol (RES) is a natural compound that exhibits anti-invasion properties in multiple tumor cell lines.Invasive behaviors, including the adhesion, invasion and migration of GICs, were determined by tumor invasive assays in vitro and in vivo.The activity of matrix metalloproteinases (MMPs) was measured by the gelatin zymography assay.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurosurgery, Beijing Tiantan Hospital, Capital Medical University, Beijing 100050, China. shengxiongyuming@163.com.

ABSTRACT
Invasion and metastasis of glioblastoma-initiating cells (GICs) are thought to be responsible for the progression and recurrence of glioblastoma multiforme (GBM). A safe drug that can be applied during the rest period of temozolomide (TMZ) maintenance cycles would greatly improve the prognosis of GBM patients by inhibiting GIC invasion. Resveratrol (RES) is a natural compound that exhibits anti-invasion properties in multiple tumor cell lines. The current study aimed to evaluate whether RES can inhibit GIC invasion in vitro and in vivo. GICs were identified using CD133 and Nestin immunofluorescence staining and tumorigenesis in non-obese diabetic severe combined immunodeficient (NOD/SCID) mice. Invasive behaviors, including the adhesion, invasion and migration of GICs, were determined by tumor invasive assays in vitro and in vivo. The activity of matrix metalloproteinases (MMPs) was measured by the gelatin zymography assay. Western blotting analysis and immunofluorescence staining were used to determine the expression of signaling effectors in GICs. We demonstrated that RES suppressed the adhesion, invasion and migration of GICs in vitro and in vivo. Moreover, we proved that RES inhibited the invasion of GICs via the inhibition of PI3K/Akt/NF-κB signal transduction and the subsequent suppression of MMP-2 expression.

No MeSH data available.


Related in: MedlinePlus