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Rac1-regulated endothelial radiation response stimulates extravasation and metastasis that can be blocked by HMG-CoA reductase inhibitors.

Hamalukic M, Huelsenbeck J, Schad A, Wirtz S, Kaina B, Fritz G - PLoS ONE (2011)

Bottom Line: IR-stimulated TC-EC adhesion was blocked by the HMG-CoA reductase inhibitor lovastatin.Glycyrrhizic acid from liquorice root, which acts as a Sialyl-Lewis X mimetic drug, and the Rac1 inhibitor NSC23766 also reduced TC-EC adhesion.To examine the in vivo relevance of these findings, tumorigenic cells were injected into the tail vein of immunodeficient mice followed by total body irradiation (TBI).

View Article: PubMed Central - PubMed

Affiliation: Institute of Toxicology, University Medical Center of the Johannes Gutenberg University Mainz, Mainz, Germany.

ABSTRACT
Radiotherapy (RT) plays a key role in cancer treatment. Although the benefit of ionizing radiation (IR) is well established, some findings raise the possibility that irradiation of the primary tumor not only triggers a killing response but also increases the metastatic potential of surviving tumor cells. Here we addressed the question of whether irradiation of normal cells outside of the primary tumor augments metastasis by stimulating the extravasation of circulating tumor cells. We show that IR exposure of human endothelial cells (EC), tumor cells (TC) or both increases TC-EC adhesion in vitro. IR-stimulated TC-EC adhesion was blocked by the HMG-CoA reductase inhibitor lovastatin. Glycyrrhizic acid from liquorice root, which acts as a Sialyl-Lewis X mimetic drug, and the Rac1 inhibitor NSC23766 also reduced TC-EC adhesion. To examine the in vivo relevance of these findings, tumorigenic cells were injected into the tail vein of immunodeficient mice followed by total body irradiation (TBI). The data obtained show that TBI dramatically enhances tumor cell extravasation and lung metastasis. This pro-metastatic radiation effect was blocked by pre-treating mice with lovastatin, glycyrrhizic acid or NSC23766. TBI of mice prior to tumor cell transplantation also stimulated metastasis, which was again blocked by lovastatin. The data point to a pro-metastatic trans-effect of RT, which likely rests on the endothelial radiation response promoting the extravasation of circulating tumor cells. Administration of the widely used lipid-lowering drug lovastatin prior to irradiation counteracts this process, likely by suppressing Rac1-regulated E-selectin expression following irradiation. The data support the concern that radiation exposure might increase the extravasation of circulating tumor cells and recommend co-administration of lipid-lowering drugs to avoid this adverse effect of ionizing radiation.

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The small GTPase Rac1 and E-selectin ligands are involved in IR-induced TC-EC adhesion in vitro and TBI driven extravasation of metastasis in vivo.A, B: Human colon carcinoma cells (HT29) and/or endothelial cells (HUVEC) were left untreated or were pre-treated with the Sialy-Lewis X mimetic glycyrrhizic acid (2 mM, 30 min) (A) or the Rac1 inhibitor NSC23766 (100 µM, 4 h) (B). 4 h after irradiation (10 Gy), TC-EC adhesion was analyzed as described in methods. *p≤0.05; **p<0.01 (n = 8). C, D: CHO-K1 cells (2×106) were injected into the tail vein of Rag2−/− Balb/c mice that have been pre-treated or not with glycyrrhizic acid (GL), NSC23766 (NSC). 3–4 weeks after TBI (4 Gy), formation of lung metastases was analyzed. C, Representative morphological and histopathological photographs are shown. D, The histogram displays the percent (%) tumor area in lung sections. Con, non-irradiated control; IR, TBI with 4 Gy. ** p≤0.01; *** p≤0.001 (n = 4–8 mice).
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pone-0026413-g005: The small GTPase Rac1 and E-selectin ligands are involved in IR-induced TC-EC adhesion in vitro and TBI driven extravasation of metastasis in vivo.A, B: Human colon carcinoma cells (HT29) and/or endothelial cells (HUVEC) were left untreated or were pre-treated with the Sialy-Lewis X mimetic glycyrrhizic acid (2 mM, 30 min) (A) or the Rac1 inhibitor NSC23766 (100 µM, 4 h) (B). 4 h after irradiation (10 Gy), TC-EC adhesion was analyzed as described in methods. *p≤0.05; **p<0.01 (n = 8). C, D: CHO-K1 cells (2×106) were injected into the tail vein of Rag2−/− Balb/c mice that have been pre-treated or not with glycyrrhizic acid (GL), NSC23766 (NSC). 3–4 weeks after TBI (4 Gy), formation of lung metastases was analyzed. C, Representative morphological and histopathological photographs are shown. D, The histogram displays the percent (%) tumor area in lung sections. Con, non-irradiated control; IR, TBI with 4 Gy. ** p≤0.01; *** p≤0.001 (n = 4–8 mice).

Mentions: Sialyl-Lewis X structures are known as major counter-ligands of the endothelial adhesion molecule E-selectin [25], [50]. They have been shown to be of importance for the progression of colon cancer [23]. To examine whether these tetrasaccharide carbohydrates are involved in IR-stimulated TC-EC adhesion in vitro and metastasis in vivo, we made use of the Sialyl-Lewis X mimetic glycyrrhizic acid (GL) [51], which is the major sweet tasting compound of liquorice root. As shown in Fig. 5A, glycyrrhizic acid largely blocked IR-induced binding of tumor cell to endothelial cells, indicating that E-selectin and its ligand Sialyl-LewisX are important for radiation-promoted TC-EC adhesion in vitro. Next, we investigated the involvement of the E-selectin counter-ligand Sialyl-Lewis X on TBI-driven extravasation and metastasis in vivo. Fully in line with our in vitro data we observed that pre-treatment of mice with GL largely attenuated the TBI-induced extravasation and formation of lung metastases (Fig. 5C). The data support the concept that E-selectin and its ligands are essential for TBI-stimulated tumor cell extravasation and metastasis.


Rac1-regulated endothelial radiation response stimulates extravasation and metastasis that can be blocked by HMG-CoA reductase inhibitors.

Hamalukic M, Huelsenbeck J, Schad A, Wirtz S, Kaina B, Fritz G - PLoS ONE (2011)

The small GTPase Rac1 and E-selectin ligands are involved in IR-induced TC-EC adhesion in vitro and TBI driven extravasation of metastasis in vivo.A, B: Human colon carcinoma cells (HT29) and/or endothelial cells (HUVEC) were left untreated or were pre-treated with the Sialy-Lewis X mimetic glycyrrhizic acid (2 mM, 30 min) (A) or the Rac1 inhibitor NSC23766 (100 µM, 4 h) (B). 4 h after irradiation (10 Gy), TC-EC adhesion was analyzed as described in methods. *p≤0.05; **p<0.01 (n = 8). C, D: CHO-K1 cells (2×106) were injected into the tail vein of Rag2−/− Balb/c mice that have been pre-treated or not with glycyrrhizic acid (GL), NSC23766 (NSC). 3–4 weeks after TBI (4 Gy), formation of lung metastases was analyzed. C, Representative morphological and histopathological photographs are shown. D, The histogram displays the percent (%) tumor area in lung sections. Con, non-irradiated control; IR, TBI with 4 Gy. ** p≤0.01; *** p≤0.001 (n = 4–8 mice).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3198428&req=5

pone-0026413-g005: The small GTPase Rac1 and E-selectin ligands are involved in IR-induced TC-EC adhesion in vitro and TBI driven extravasation of metastasis in vivo.A, B: Human colon carcinoma cells (HT29) and/or endothelial cells (HUVEC) were left untreated or were pre-treated with the Sialy-Lewis X mimetic glycyrrhizic acid (2 mM, 30 min) (A) or the Rac1 inhibitor NSC23766 (100 µM, 4 h) (B). 4 h after irradiation (10 Gy), TC-EC adhesion was analyzed as described in methods. *p≤0.05; **p<0.01 (n = 8). C, D: CHO-K1 cells (2×106) were injected into the tail vein of Rag2−/− Balb/c mice that have been pre-treated or not with glycyrrhizic acid (GL), NSC23766 (NSC). 3–4 weeks after TBI (4 Gy), formation of lung metastases was analyzed. C, Representative morphological and histopathological photographs are shown. D, The histogram displays the percent (%) tumor area in lung sections. Con, non-irradiated control; IR, TBI with 4 Gy. ** p≤0.01; *** p≤0.001 (n = 4–8 mice).
Mentions: Sialyl-Lewis X structures are known as major counter-ligands of the endothelial adhesion molecule E-selectin [25], [50]. They have been shown to be of importance for the progression of colon cancer [23]. To examine whether these tetrasaccharide carbohydrates are involved in IR-stimulated TC-EC adhesion in vitro and metastasis in vivo, we made use of the Sialyl-Lewis X mimetic glycyrrhizic acid (GL) [51], which is the major sweet tasting compound of liquorice root. As shown in Fig. 5A, glycyrrhizic acid largely blocked IR-induced binding of tumor cell to endothelial cells, indicating that E-selectin and its ligand Sialyl-LewisX are important for radiation-promoted TC-EC adhesion in vitro. Next, we investigated the involvement of the E-selectin counter-ligand Sialyl-Lewis X on TBI-driven extravasation and metastasis in vivo. Fully in line with our in vitro data we observed that pre-treatment of mice with GL largely attenuated the TBI-induced extravasation and formation of lung metastases (Fig. 5C). The data support the concept that E-selectin and its ligands are essential for TBI-stimulated tumor cell extravasation and metastasis.

Bottom Line: IR-stimulated TC-EC adhesion was blocked by the HMG-CoA reductase inhibitor lovastatin.Glycyrrhizic acid from liquorice root, which acts as a Sialyl-Lewis X mimetic drug, and the Rac1 inhibitor NSC23766 also reduced TC-EC adhesion.To examine the in vivo relevance of these findings, tumorigenic cells were injected into the tail vein of immunodeficient mice followed by total body irradiation (TBI).

View Article: PubMed Central - PubMed

Affiliation: Institute of Toxicology, University Medical Center of the Johannes Gutenberg University Mainz, Mainz, Germany.

ABSTRACT
Radiotherapy (RT) plays a key role in cancer treatment. Although the benefit of ionizing radiation (IR) is well established, some findings raise the possibility that irradiation of the primary tumor not only triggers a killing response but also increases the metastatic potential of surviving tumor cells. Here we addressed the question of whether irradiation of normal cells outside of the primary tumor augments metastasis by stimulating the extravasation of circulating tumor cells. We show that IR exposure of human endothelial cells (EC), tumor cells (TC) or both increases TC-EC adhesion in vitro. IR-stimulated TC-EC adhesion was blocked by the HMG-CoA reductase inhibitor lovastatin. Glycyrrhizic acid from liquorice root, which acts as a Sialyl-Lewis X mimetic drug, and the Rac1 inhibitor NSC23766 also reduced TC-EC adhesion. To examine the in vivo relevance of these findings, tumorigenic cells were injected into the tail vein of immunodeficient mice followed by total body irradiation (TBI). The data obtained show that TBI dramatically enhances tumor cell extravasation and lung metastasis. This pro-metastatic radiation effect was blocked by pre-treating mice with lovastatin, glycyrrhizic acid or NSC23766. TBI of mice prior to tumor cell transplantation also stimulated metastasis, which was again blocked by lovastatin. The data point to a pro-metastatic trans-effect of RT, which likely rests on the endothelial radiation response promoting the extravasation of circulating tumor cells. Administration of the widely used lipid-lowering drug lovastatin prior to irradiation counteracts this process, likely by suppressing Rac1-regulated E-selectin expression following irradiation. The data support the concern that radiation exposure might increase the extravasation of circulating tumor cells and recommend co-administration of lipid-lowering drugs to avoid this adverse effect of ionizing radiation.

Show MeSH
Related in: MedlinePlus