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Paeonol oxime inhibits bFGF-induced angiogenesis and reduces VEGF levels in fibrosarcoma cells.

Lee HJ, Kim SA, Lee HJ, Jeong SJ, Han I, Jung JH, Lee EO, Zhu S, Chen CY, Kim SH - PLoS ONE (2010)

Bottom Line: The treatment with PO at 12.5 microg/ml reduced the levels of phosphorylated AKT and VEGF expression (intracellular and extracelluar) in HT-1080 cells.Consistently, immunefluorescence imaging analysis revealed that PO treatment attenuated AKT phosphorylation in HT-1080 cells.Taken together, these results suggest that PO inhibits bFGF-induced angiogenesis in HUVECs and decreased the levels of PI3K, phospho-AKT and VEGF in HT-1080 cells.

View Article: PubMed Central - PubMed

Affiliation: College of Oriental Medicine, Kyung Hee University, Seoul, Republic of Korea.

ABSTRACT

Background: We previously reported the anti-angiogenic activity of paeonol isolated from Moutan Cortex. In the present study, we investigated the negative effect of paeonol oxime (PO, a paeonol derivative) on basic fibroblast growth factor (bFGF)-mediated angiogenesis in human umbilical vein endothelial cells (HUVECs) (including tumor angiogenesis) and pro-survival activity in HT-1080 fibrosarcoma cell line.

Methodology/principal findings: We showed that PO (IC(50) = 17.3 microg/ml) significantly inhibited bFGF-induced cell proliferation, which was achieved with higher concentrations of paeonol (IC(50) over 200 microg). The treatment with PO blocked bFGF-stimulated migration and in vitro capillary differentiation (tube formation) in a dose-dependent manner. Furthermore, PO was able to disrupt neovascularization in vivo. Interestingly, PO (25 microg/ml) decreased the cell viability of HT-1080 fibrosarcoma cells but not that of HUVECs. The treatment with PO at 12.5 microg/ml reduced the levels of phosphorylated AKT and VEGF expression (intracellular and extracelluar) in HT-1080 cells. Consistently, immunefluorescence imaging analysis revealed that PO treatment attenuated AKT phosphorylation in HT-1080 cells.

Conclusions/significance: Taken together, these results suggest that PO inhibits bFGF-induced angiogenesis in HUVECs and decreased the levels of PI3K, phospho-AKT and VEGF in HT-1080 cells.

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Related in: MedlinePlus

PO inhibits the tube formation of HUVECs induced by culture supernatants of HT-1080 cells.Tube formation assay was performed using growth factor reduced Matrigel. (A) Cells were fixed with Diff-Quick solution and photographed randomly under an Axiovert S 100 light microscope at ×100 magnification. (B) Tube networks were quantified using NIH Scion image program. All data were expressed as mean ± S.D. The statistically significant differences between control and PO treated groups were calculated by the Student's t-test. ###, p<0.001 versus untreated control in M199 medium. ***, p<0.001 versus culture supernatant from HT-1080.
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pone-0012358-g006: PO inhibits the tube formation of HUVECs induced by culture supernatants of HT-1080 cells.Tube formation assay was performed using growth factor reduced Matrigel. (A) Cells were fixed with Diff-Quick solution and photographed randomly under an Axiovert S 100 light microscope at ×100 magnification. (B) Tube networks were quantified using NIH Scion image program. All data were expressed as mean ± S.D. The statistically significant differences between control and PO treated groups were calculated by the Student's t-test. ###, p<0.001 versus untreated control in M199 medium. ***, p<0.001 versus culture supernatant from HT-1080.

Mentions: To further confirm that the effects of PO on HT-1080 cells are due to its action on angiogenic properties, in vitro tube formation assay was carried out in the cells with or without the treatment with PO for 24 h (Fig. 6). The culture medium (CM) from HT-1080 cells induced HUVEC tube formation compared with that from untreated control cells (lanes 1 and 4). In contrast, less than half of PO-treated cells grown in CM (>47%) were able to induce HUVEC tube formation, than that cultured in CM alone (lanes 4 and 5), suggesting that PO is a potent anti-angiogenic agent. LY294002 was also prevented the tube formation (lane 6). VEGF and bFGF served as positive controls (lanes 2 and 3). However, PO had no significant effect on HT-1080 mediated cell proliferation and VEGF secretion in HUVECs (data not shown).


Paeonol oxime inhibits bFGF-induced angiogenesis and reduces VEGF levels in fibrosarcoma cells.

Lee HJ, Kim SA, Lee HJ, Jeong SJ, Han I, Jung JH, Lee EO, Zhu S, Chen CY, Kim SH - PLoS ONE (2010)

PO inhibits the tube formation of HUVECs induced by culture supernatants of HT-1080 cells.Tube formation assay was performed using growth factor reduced Matrigel. (A) Cells were fixed with Diff-Quick solution and photographed randomly under an Axiovert S 100 light microscope at ×100 magnification. (B) Tube networks were quantified using NIH Scion image program. All data were expressed as mean ± S.D. The statistically significant differences between control and PO treated groups were calculated by the Student's t-test. ###, p<0.001 versus untreated control in M199 medium. ***, p<0.001 versus culture supernatant from HT-1080.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2925949&req=5

pone-0012358-g006: PO inhibits the tube formation of HUVECs induced by culture supernatants of HT-1080 cells.Tube formation assay was performed using growth factor reduced Matrigel. (A) Cells were fixed with Diff-Quick solution and photographed randomly under an Axiovert S 100 light microscope at ×100 magnification. (B) Tube networks were quantified using NIH Scion image program. All data were expressed as mean ± S.D. The statistically significant differences between control and PO treated groups were calculated by the Student's t-test. ###, p<0.001 versus untreated control in M199 medium. ***, p<0.001 versus culture supernatant from HT-1080.
Mentions: To further confirm that the effects of PO on HT-1080 cells are due to its action on angiogenic properties, in vitro tube formation assay was carried out in the cells with or without the treatment with PO for 24 h (Fig. 6). The culture medium (CM) from HT-1080 cells induced HUVEC tube formation compared with that from untreated control cells (lanes 1 and 4). In contrast, less than half of PO-treated cells grown in CM (>47%) were able to induce HUVEC tube formation, than that cultured in CM alone (lanes 4 and 5), suggesting that PO is a potent anti-angiogenic agent. LY294002 was also prevented the tube formation (lane 6). VEGF and bFGF served as positive controls (lanes 2 and 3). However, PO had no significant effect on HT-1080 mediated cell proliferation and VEGF secretion in HUVECs (data not shown).

Bottom Line: The treatment with PO at 12.5 microg/ml reduced the levels of phosphorylated AKT and VEGF expression (intracellular and extracelluar) in HT-1080 cells.Consistently, immunefluorescence imaging analysis revealed that PO treatment attenuated AKT phosphorylation in HT-1080 cells.Taken together, these results suggest that PO inhibits bFGF-induced angiogenesis in HUVECs and decreased the levels of PI3K, phospho-AKT and VEGF in HT-1080 cells.

View Article: PubMed Central - PubMed

Affiliation: College of Oriental Medicine, Kyung Hee University, Seoul, Republic of Korea.

ABSTRACT

Background: We previously reported the anti-angiogenic activity of paeonol isolated from Moutan Cortex. In the present study, we investigated the negative effect of paeonol oxime (PO, a paeonol derivative) on basic fibroblast growth factor (bFGF)-mediated angiogenesis in human umbilical vein endothelial cells (HUVECs) (including tumor angiogenesis) and pro-survival activity in HT-1080 fibrosarcoma cell line.

Methodology/principal findings: We showed that PO (IC(50) = 17.3 microg/ml) significantly inhibited bFGF-induced cell proliferation, which was achieved with higher concentrations of paeonol (IC(50) over 200 microg). The treatment with PO blocked bFGF-stimulated migration and in vitro capillary differentiation (tube formation) in a dose-dependent manner. Furthermore, PO was able to disrupt neovascularization in vivo. Interestingly, PO (25 microg/ml) decreased the cell viability of HT-1080 fibrosarcoma cells but not that of HUVECs. The treatment with PO at 12.5 microg/ml reduced the levels of phosphorylated AKT and VEGF expression (intracellular and extracelluar) in HT-1080 cells. Consistently, immunefluorescence imaging analysis revealed that PO treatment attenuated AKT phosphorylation in HT-1080 cells.

Conclusions/significance: Taken together, these results suggest that PO inhibits bFGF-induced angiogenesis in HUVECs and decreased the levels of PI3K, phospho-AKT and VEGF in HT-1080 cells.

Show MeSH
Related in: MedlinePlus