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Phosphatase inhibitors with anti-angiogenic effect in vitro.

Sylvest L, Bendiksen CD, Houen G - APMIS (2010)

Bottom Line: Historically, intracellular phosphatases have been associated with the downregulation of signaling pathways, and kinases with their upregulation, but lately, the phospatases have also been coupled to positive signaling, which is why inhibition of phosphatases has become associated with anti-tumorigenic and anti-angiogenic effects.The results obtained in this work reveal several agents with anti-angiogenic potential and give a strong indication that phosphatase inhibition is linked to anti-angiogenic activity.By looking at the morphological results, it was seen that most of the inhibitors impaired proliferation and elongation of the endothelial cells, which still had a differentiated appearance.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical Biochemistry and Immunology, Statens Serum Institut, Copenhagen, Denmark.

ABSTRACT
Levamisole has previously been identified as an inhibitor of angiogenesis in vitro and in vivo, but the mechanism behind the anti-angiogenic behavior has not yet been established. However, one known effect of levamisole is the inhibition of alkaline phosphatase, and this fact encouraged us to test other phosphatase inhibitors for their anti-angiogenic effects by using the same method as used to identify levamisole: an ELISA-based co-culture angiogenesis assay giving quantitative and qualitative results. Historically, intracellular phosphatases have been associated with the downregulation of signaling pathways, and kinases with their upregulation, but lately, the phospatases have also been coupled to positive signaling, which is why inhibition of phosphatases has become associated with anti-tumorigenic and anti-angiogenic effects. The results obtained in this work reveal several agents with anti-angiogenic potential and give a strong indication that phosphatase inhibition is linked to anti-angiogenic activity. An apparent disruption of endothelial tube formation was seen for seven of eight phosphatase inhibitors tested in the angiogenesis assay. By looking at the morphological results, it was seen that most of the inhibitors impaired proliferation and elongation of the endothelial cells, which still had a differentiated appearance. One inhibitor, PTP inhibitor IV, seemed to impair endothelial cell differentiation and induced the same morphology as when cells were treated with levamisole, although at a 200 times lower concentration than that of levamisole. Hence, our work points out compounds with a potential that may be of use in the search for new medical products for the treatment of malignant tumors, or other conditions where angiogenesis plays a central role.

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Related in: MedlinePlus

The phosphatase inhibitors NSC87877, clodronate and ibandronate impair HUVEC proliferation. ELISA results showing the inhibitory effect of 0.2 mM NSC87877, 1 mM clodronate and 0.1 mM ibandronate on HUVEC (CD31) and NHDF (dsDNA) grown in co-culture. Data are presented as relative absorbance (tested compound/milliQ water), from three to four independent experiments ± SD. Each experiment was performed in duplicate, n = 6–8.
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fig01: The phosphatase inhibitors NSC87877, clodronate and ibandronate impair HUVEC proliferation. ELISA results showing the inhibitory effect of 0.2 mM NSC87877, 1 mM clodronate and 0.1 mM ibandronate on HUVEC (CD31) and NHDF (dsDNA) grown in co-culture. Data are presented as relative absorbance (tested compound/milliQ water), from three to four independent experiments ± SD. Each experiment was performed in duplicate, n = 6–8.

Mentions: The ELISA results for all the tested compounds are illustrated as relative absorbances compared to control in Figs. 1 and 3. The effect of phosphatase inhibitors dissolved in milliQ water is shown in Fig. 1 and is normalized to a control where 10% milliQ water was added to the medium instead of 10% phosphatase inhibitor. All the inhibitors had a strong reducing effect on the endothelial cell proliferation (CD31) with a relative absorbance around 0.4, hence below 0.8, which is the limit according to earlier findings regarding the inhibitory effect on the endothelial cells (23). The relative absorbance measured with dsDNA antibody, indicating the proliferation of fibroblasts, shows that NSC87877 and clodronate do not affect the fibroblasts (relative absorbance higher than 0.8). Ibandronate, however, has the same inhibitory effect on the fibroblasts as on the endothelial cells.


Phosphatase inhibitors with anti-angiogenic effect in vitro.

Sylvest L, Bendiksen CD, Houen G - APMIS (2010)

The phosphatase inhibitors NSC87877, clodronate and ibandronate impair HUVEC proliferation. ELISA results showing the inhibitory effect of 0.2 mM NSC87877, 1 mM clodronate and 0.1 mM ibandronate on HUVEC (CD31) and NHDF (dsDNA) grown in co-culture. Data are presented as relative absorbance (tested compound/milliQ water), from three to four independent experiments ± SD. Each experiment was performed in duplicate, n = 6–8.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2806050&req=5

fig01: The phosphatase inhibitors NSC87877, clodronate and ibandronate impair HUVEC proliferation. ELISA results showing the inhibitory effect of 0.2 mM NSC87877, 1 mM clodronate and 0.1 mM ibandronate on HUVEC (CD31) and NHDF (dsDNA) grown in co-culture. Data are presented as relative absorbance (tested compound/milliQ water), from three to four independent experiments ± SD. Each experiment was performed in duplicate, n = 6–8.
Mentions: The ELISA results for all the tested compounds are illustrated as relative absorbances compared to control in Figs. 1 and 3. The effect of phosphatase inhibitors dissolved in milliQ water is shown in Fig. 1 and is normalized to a control where 10% milliQ water was added to the medium instead of 10% phosphatase inhibitor. All the inhibitors had a strong reducing effect on the endothelial cell proliferation (CD31) with a relative absorbance around 0.4, hence below 0.8, which is the limit according to earlier findings regarding the inhibitory effect on the endothelial cells (23). The relative absorbance measured with dsDNA antibody, indicating the proliferation of fibroblasts, shows that NSC87877 and clodronate do not affect the fibroblasts (relative absorbance higher than 0.8). Ibandronate, however, has the same inhibitory effect on the fibroblasts as on the endothelial cells.

Bottom Line: Historically, intracellular phosphatases have been associated with the downregulation of signaling pathways, and kinases with their upregulation, but lately, the phospatases have also been coupled to positive signaling, which is why inhibition of phosphatases has become associated with anti-tumorigenic and anti-angiogenic effects.The results obtained in this work reveal several agents with anti-angiogenic potential and give a strong indication that phosphatase inhibition is linked to anti-angiogenic activity.By looking at the morphological results, it was seen that most of the inhibitors impaired proliferation and elongation of the endothelial cells, which still had a differentiated appearance.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical Biochemistry and Immunology, Statens Serum Institut, Copenhagen, Denmark.

ABSTRACT
Levamisole has previously been identified as an inhibitor of angiogenesis in vitro and in vivo, but the mechanism behind the anti-angiogenic behavior has not yet been established. However, one known effect of levamisole is the inhibition of alkaline phosphatase, and this fact encouraged us to test other phosphatase inhibitors for their anti-angiogenic effects by using the same method as used to identify levamisole: an ELISA-based co-culture angiogenesis assay giving quantitative and qualitative results. Historically, intracellular phosphatases have been associated with the downregulation of signaling pathways, and kinases with their upregulation, but lately, the phospatases have also been coupled to positive signaling, which is why inhibition of phosphatases has become associated with anti-tumorigenic and anti-angiogenic effects. The results obtained in this work reveal several agents with anti-angiogenic potential and give a strong indication that phosphatase inhibition is linked to anti-angiogenic activity. An apparent disruption of endothelial tube formation was seen for seven of eight phosphatase inhibitors tested in the angiogenesis assay. By looking at the morphological results, it was seen that most of the inhibitors impaired proliferation and elongation of the endothelial cells, which still had a differentiated appearance. One inhibitor, PTP inhibitor IV, seemed to impair endothelial cell differentiation and induced the same morphology as when cells were treated with levamisole, although at a 200 times lower concentration than that of levamisole. Hence, our work points out compounds with a potential that may be of use in the search for new medical products for the treatment of malignant tumors, or other conditions where angiogenesis plays a central role.

Show MeSH
Related in: MedlinePlus