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Endothelial cell-derived interleukin-6 regulates tumor growth.

Neiva KG, Warner KA, Campos MS, Zhang Z, Moren J, Danciu TE, Nör JE - BMC Cancer (2014)

Bottom Line: In search for a possible mechanism for the activation of the STAT3 signaling pathway, we observed that silencing interleukin (IL)-6 in tumor-associated endothelial cells inhibited STAT3 phosphorylation in tumor cells.Notably, tumors vascularized with IL-6-silenced endothelial cells showed lower intratumoral microvessel density, lower tumor cell proliferation, and slower growth than tumors vascularized with control endothelial cells.Collectively, these results demonstrate that IL-6 secreted by endothelial cells enhance tumor growth, and suggest that cancer patients might benefit from targeted approaches that block signaling events initiated by endothelial cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Angiogenesis Research Laboratory, Department of Cariology, Restorative Sciences, and Endodontics, University of Michigan School of Dentistry, Ann Arbor, Michigan 48109-1078, USA. jenor@umich.edu.

ABSTRACT

Background: Endothelial cells play a complex role in the pathobiology of cancer. This role is not limited to the making of blood vessels to allow for influx of oxygen and nutrients required for the high metabolic demands of tumor cells. Indeed, it has been recently shown that tumor-associated endothelial cells secrete molecules that enhance tumor cell survival and cancer stem cell self-renewal. The hypothesis underlying this work is that specific disruption of endothelial cell-initiated signaling inhibits tumor growth.

Methods: Conditioned medium from primary human dermal microvascular endothelial cells (HDMEC) stably transduced with silencing RNA for IL-6 (or controls) was used to evaluate the role of endothelial-derived IL-6 on the activation of key signaling pathways in tumor cells. In addition, these endothelial cells were co-transplanted with tumor cells into immunodefficient mice to determine the impact of endothelial cell-derived IL-6 on tumor growth and angiogenesis.

Results: We observed that tumor cells adjacent to blood vessels show strong phosphorylation of STAT3, a key mediator of tumor progression. In search for a possible mechanism for the activation of the STAT3 signaling pathway, we observed that silencing interleukin (IL)-6 in tumor-associated endothelial cells inhibited STAT3 phosphorylation in tumor cells. Notably, tumors vascularized with IL-6-silenced endothelial cells showed lower intratumoral microvessel density, lower tumor cell proliferation, and slower growth than tumors vascularized with control endothelial cells.

Conclusions: Collectively, these results demonstrate that IL-6 secreted by endothelial cells enhance tumor growth, and suggest that cancer patients might benefit from targeted approaches that block signaling events initiated by endothelial cells.

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Related in: MedlinePlus

STAT3 phosphorylation induced by endothelial cell-secreted factors is independent of Akt and ERK phosphorylation. Western blot for phosphorylated and total STAT3, Akt, and ERK in HeLa serum-starved overnight and exposed to A, HDMEC conditioned medium (CM) or unconditioned medium (EBM) for the indicated time points. In addition, HeLa were pre-incubated for 1 to 2 hours with B, 20 μM Stattic; C, 20 μM LY294002; or D, 20 μM U0126, and then exposed to HDMEC CM or EBM in presence of the specific inhibitor for the indicated time points.
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Figure 2: STAT3 phosphorylation induced by endothelial cell-secreted factors is independent of Akt and ERK phosphorylation. Western blot for phosphorylated and total STAT3, Akt, and ERK in HeLa serum-starved overnight and exposed to A, HDMEC conditioned medium (CM) or unconditioned medium (EBM) for the indicated time points. In addition, HeLa were pre-incubated for 1 to 2 hours with B, 20 μM Stattic; C, 20 μM LY294002; or D, 20 μM U0126, and then exposed to HDMEC CM or EBM in presence of the specific inhibitor for the indicated time points.

Mentions: To explore the interdependence of molecular signaling events initiated by endothelial cells on tumor cells, we exposed HeLa to HDMEC CM in the presence of chemical inhibitors of STAT3, Akt, or ERK pathways and analyzed the interdependency of the phosphorylation events. To establish the baseline for these experiments, we exposed HeLa to HDMEC CM and analyzed phosphorylation of STAT3, Akt, and ERK with a detailed time course up to 1 hour (Figure 2A). We observed that HDMEC CM induces first ERK phosphorylation (with strong activation as early as 1 minute, persisting until 15 minutes, and decreasing at 30 minutes), followed by STAT3 and Akt (increasing until 15 minutes, and maintaining activation for up to 1 hour) (Figure 2A). When we inhibited STAT3 phosphorylation using the chemical inhibitor Stattic, we did not observe significant changes in phosphorylation of Akt or ERK (Figure 2B). However, when we inhibited Akt phosphorylation using the PI3K inhibitor LY294002 we observed an increase in ERK phosphorylation levels (maintaining strong phosphorylation for up to 1 hour), while phosphorylation levels of STAT3 did not change (Figure 2C). Similarly, when we inhibited ERK phosphorylation using the MEK1/2 inhibitor U0126 we observed increased Akt phosphorylation (maintaining strong phosphorylation for up to 1 hour), whereas phosphorylation levels of STAT3 remained unchanged (Figure 2D).


Endothelial cell-derived interleukin-6 regulates tumor growth.

Neiva KG, Warner KA, Campos MS, Zhang Z, Moren J, Danciu TE, Nör JE - BMC Cancer (2014)

STAT3 phosphorylation induced by endothelial cell-secreted factors is independent of Akt and ERK phosphorylation. Western blot for phosphorylated and total STAT3, Akt, and ERK in HeLa serum-starved overnight and exposed to A, HDMEC conditioned medium (CM) or unconditioned medium (EBM) for the indicated time points. In addition, HeLa were pre-incubated for 1 to 2 hours with B, 20 μM Stattic; C, 20 μM LY294002; or D, 20 μM U0126, and then exposed to HDMEC CM or EBM in presence of the specific inhibitor for the indicated time points.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4016552&req=5

Figure 2: STAT3 phosphorylation induced by endothelial cell-secreted factors is independent of Akt and ERK phosphorylation. Western blot for phosphorylated and total STAT3, Akt, and ERK in HeLa serum-starved overnight and exposed to A, HDMEC conditioned medium (CM) or unconditioned medium (EBM) for the indicated time points. In addition, HeLa were pre-incubated for 1 to 2 hours with B, 20 μM Stattic; C, 20 μM LY294002; or D, 20 μM U0126, and then exposed to HDMEC CM or EBM in presence of the specific inhibitor for the indicated time points.
Mentions: To explore the interdependence of molecular signaling events initiated by endothelial cells on tumor cells, we exposed HeLa to HDMEC CM in the presence of chemical inhibitors of STAT3, Akt, or ERK pathways and analyzed the interdependency of the phosphorylation events. To establish the baseline for these experiments, we exposed HeLa to HDMEC CM and analyzed phosphorylation of STAT3, Akt, and ERK with a detailed time course up to 1 hour (Figure 2A). We observed that HDMEC CM induces first ERK phosphorylation (with strong activation as early as 1 minute, persisting until 15 minutes, and decreasing at 30 minutes), followed by STAT3 and Akt (increasing until 15 minutes, and maintaining activation for up to 1 hour) (Figure 2A). When we inhibited STAT3 phosphorylation using the chemical inhibitor Stattic, we did not observe significant changes in phosphorylation of Akt or ERK (Figure 2B). However, when we inhibited Akt phosphorylation using the PI3K inhibitor LY294002 we observed an increase in ERK phosphorylation levels (maintaining strong phosphorylation for up to 1 hour), while phosphorylation levels of STAT3 did not change (Figure 2C). Similarly, when we inhibited ERK phosphorylation using the MEK1/2 inhibitor U0126 we observed increased Akt phosphorylation (maintaining strong phosphorylation for up to 1 hour), whereas phosphorylation levels of STAT3 remained unchanged (Figure 2D).

Bottom Line: In search for a possible mechanism for the activation of the STAT3 signaling pathway, we observed that silencing interleukin (IL)-6 in tumor-associated endothelial cells inhibited STAT3 phosphorylation in tumor cells.Notably, tumors vascularized with IL-6-silenced endothelial cells showed lower intratumoral microvessel density, lower tumor cell proliferation, and slower growth than tumors vascularized with control endothelial cells.Collectively, these results demonstrate that IL-6 secreted by endothelial cells enhance tumor growth, and suggest that cancer patients might benefit from targeted approaches that block signaling events initiated by endothelial cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Angiogenesis Research Laboratory, Department of Cariology, Restorative Sciences, and Endodontics, University of Michigan School of Dentistry, Ann Arbor, Michigan 48109-1078, USA. jenor@umich.edu.

ABSTRACT

Background: Endothelial cells play a complex role in the pathobiology of cancer. This role is not limited to the making of blood vessels to allow for influx of oxygen and nutrients required for the high metabolic demands of tumor cells. Indeed, it has been recently shown that tumor-associated endothelial cells secrete molecules that enhance tumor cell survival and cancer stem cell self-renewal. The hypothesis underlying this work is that specific disruption of endothelial cell-initiated signaling inhibits tumor growth.

Methods: Conditioned medium from primary human dermal microvascular endothelial cells (HDMEC) stably transduced with silencing RNA for IL-6 (or controls) was used to evaluate the role of endothelial-derived IL-6 on the activation of key signaling pathways in tumor cells. In addition, these endothelial cells were co-transplanted with tumor cells into immunodefficient mice to determine the impact of endothelial cell-derived IL-6 on tumor growth and angiogenesis.

Results: We observed that tumor cells adjacent to blood vessels show strong phosphorylation of STAT3, a key mediator of tumor progression. In search for a possible mechanism for the activation of the STAT3 signaling pathway, we observed that silencing interleukin (IL)-6 in tumor-associated endothelial cells inhibited STAT3 phosphorylation in tumor cells. Notably, tumors vascularized with IL-6-silenced endothelial cells showed lower intratumoral microvessel density, lower tumor cell proliferation, and slower growth than tumors vascularized with control endothelial cells.

Conclusions: Collectively, these results demonstrate that IL-6 secreted by endothelial cells enhance tumor growth, and suggest that cancer patients might benefit from targeted approaches that block signaling events initiated by endothelial cells.

Show MeSH
Related in: MedlinePlus