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The tumor suppressor semaphorin 3B triggers a prometastatic program mediated by interleukin 8 and the tumor microenvironment.

Rolny C, Capparuccia L, Casazza A, Mazzone M, Vallario A, Cignetti A, Medico E, Carmeliet P, Comoglio PM, Tamagnone L - J. Exp. Med. (2008)

Bottom Line: Biol. 600:118-131).By investigating experimental tumor models, we confirmed that SEMA3B expression inhibited tumor growth, whereas metastatic dissemination was surprisingly increased.We found that SEMA3B induced the production of interleukin (IL) 8 by tumor cells by activating the p38-mitogen-activated protein kinase pathway in a neuropilin 1-dependent manner.

View Article: PubMed Central - PubMed

Affiliation: Institute for Cancer Research and Treatment (IRCC), University of Turin, School of Medicine, 10060 Candiolo, Italy.

ABSTRACT
Semaphorins are a large family of evolutionarily conserved morphogenetic molecules originally identified for their repelling role in axonal guidance. Intriguingly, semaphorins have recently been implicated in cancer progression (Neufeld, G., T. Lange, A. Varshavsky, and O. Kessler. 2007. Adv. Exp. Med. Biol. 600:118-131). In particular, semaphorin 3B (SEMA3B) is considered a putative tumor suppressor, and yet we found that it is expressed at high levels in many invasive and metastatic human cancers. By investigating experimental tumor models, we confirmed that SEMA3B expression inhibited tumor growth, whereas metastatic dissemination was surprisingly increased. We found that SEMA3B induced the production of interleukin (IL) 8 by tumor cells by activating the p38-mitogen-activated protein kinase pathway in a neuropilin 1-dependent manner. Silencing the expression of endogenous SEMA3B in tumor cells impaired IL-8 transcription. The release of IL-8, in turn, induced the recruitment of tumor-associated macrophages and metastatic dissemination to the lung, which could be rescued by blocking IL-8 with neutralizing antibodies. In conclusion, we report that SEMA3B exerts unexpected functions in cancer progression by fostering a prometastatic environment through elevated IL-8 secretion and recruitment of macrophages coupled to the suppression of tumor growth.

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SEMA3B knock down in tumor cells affects IL-8 production and monocyte recruitment. (A) RNA was collected from LoVo, A549, HeLa, and Sk-Mel-2 cells transduced with shRNA against SEMA3B (to suppress gene expression; shSEMA3B) and unrelated sequences (shCTR). Real-time PCR was performed using Taqman probes for SEMA3B (results shown at the bottom) and IL-8 (in the graph). Expression levels were normalized to controls for each cell line and are represented as fold changes. Data shown are the mean ± SEM. (B) Monocyte migration to CM derived from Sk-Mel-2 melanoma cells transduced with shRNAs. IL-8 neutralizing antibodies (MAB208) or unrelated control antibodies (anti-CD19) were included in the lower chamber. Knock down of SEMA3B expression in Sk-Mel-2 cells abolished IL-8–dependent monocyte-attracting activity in the CM. The number of migrated monocytes represents the mean ± SEM of two independent experiments. **, P < 0.01.
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fig6: SEMA3B knock down in tumor cells affects IL-8 production and monocyte recruitment. (A) RNA was collected from LoVo, A549, HeLa, and Sk-Mel-2 cells transduced with shRNA against SEMA3B (to suppress gene expression; shSEMA3B) and unrelated sequences (shCTR). Real-time PCR was performed using Taqman probes for SEMA3B (results shown at the bottom) and IL-8 (in the graph). Expression levels were normalized to controls for each cell line and are represented as fold changes. Data shown are the mean ± SEM. (B) Monocyte migration to CM derived from Sk-Mel-2 melanoma cells transduced with shRNAs. IL-8 neutralizing antibodies (MAB208) or unrelated control antibodies (anti-CD19) were included in the lower chamber. Knock down of SEMA3B expression in Sk-Mel-2 cells abolished IL-8–dependent monocyte-attracting activity in the CM. The number of migrated monocytes represents the mean ± SEM of two independent experiments. **, P < 0.01.

Mentions: Consistent with our evidence that SEMA3B regulates IL-8 production, we asked whether IL-8 levels might be affected upon knocking down endogenous SEMA3B in cancer cells. We thus stably transduced various tumor cells expressing medium to high levels of SEMA3B (i.e., A549, HeLa, LoVo, and Sk-Mel-2 cells) with constructs expressing short hairpin RNAs (shRNAs) targeting SEMA3B (shSEMA3B) or control unrelated sequences (shCTR). Importantly, we found that the knock down of SEMA3B expression led to reduced levels of IL-8 mRNA (Fig. 6 A). Furthermore, three different shRNA sequences for SEMA3B were tested with consistent results, ruling out off-target effects (Fig. S5, available at http://www.jem.org/cgi/content/full/jem.20072509/DC1).


The tumor suppressor semaphorin 3B triggers a prometastatic program mediated by interleukin 8 and the tumor microenvironment.

Rolny C, Capparuccia L, Casazza A, Mazzone M, Vallario A, Cignetti A, Medico E, Carmeliet P, Comoglio PM, Tamagnone L - J. Exp. Med. (2008)

SEMA3B knock down in tumor cells affects IL-8 production and monocyte recruitment. (A) RNA was collected from LoVo, A549, HeLa, and Sk-Mel-2 cells transduced with shRNA against SEMA3B (to suppress gene expression; shSEMA3B) and unrelated sequences (shCTR). Real-time PCR was performed using Taqman probes for SEMA3B (results shown at the bottom) and IL-8 (in the graph). Expression levels were normalized to controls for each cell line and are represented as fold changes. Data shown are the mean ± SEM. (B) Monocyte migration to CM derived from Sk-Mel-2 melanoma cells transduced with shRNAs. IL-8 neutralizing antibodies (MAB208) or unrelated control antibodies (anti-CD19) were included in the lower chamber. Knock down of SEMA3B expression in Sk-Mel-2 cells abolished IL-8–dependent monocyte-attracting activity in the CM. The number of migrated monocytes represents the mean ± SEM of two independent experiments. **, P < 0.01.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2373847&req=5

fig6: SEMA3B knock down in tumor cells affects IL-8 production and monocyte recruitment. (A) RNA was collected from LoVo, A549, HeLa, and Sk-Mel-2 cells transduced with shRNA against SEMA3B (to suppress gene expression; shSEMA3B) and unrelated sequences (shCTR). Real-time PCR was performed using Taqman probes for SEMA3B (results shown at the bottom) and IL-8 (in the graph). Expression levels were normalized to controls for each cell line and are represented as fold changes. Data shown are the mean ± SEM. (B) Monocyte migration to CM derived from Sk-Mel-2 melanoma cells transduced with shRNAs. IL-8 neutralizing antibodies (MAB208) or unrelated control antibodies (anti-CD19) were included in the lower chamber. Knock down of SEMA3B expression in Sk-Mel-2 cells abolished IL-8–dependent monocyte-attracting activity in the CM. The number of migrated monocytes represents the mean ± SEM of two independent experiments. **, P < 0.01.
Mentions: Consistent with our evidence that SEMA3B regulates IL-8 production, we asked whether IL-8 levels might be affected upon knocking down endogenous SEMA3B in cancer cells. We thus stably transduced various tumor cells expressing medium to high levels of SEMA3B (i.e., A549, HeLa, LoVo, and Sk-Mel-2 cells) with constructs expressing short hairpin RNAs (shRNAs) targeting SEMA3B (shSEMA3B) or control unrelated sequences (shCTR). Importantly, we found that the knock down of SEMA3B expression led to reduced levels of IL-8 mRNA (Fig. 6 A). Furthermore, three different shRNA sequences for SEMA3B were tested with consistent results, ruling out off-target effects (Fig. S5, available at http://www.jem.org/cgi/content/full/jem.20072509/DC1).

Bottom Line: Biol. 600:118-131).By investigating experimental tumor models, we confirmed that SEMA3B expression inhibited tumor growth, whereas metastatic dissemination was surprisingly increased.We found that SEMA3B induced the production of interleukin (IL) 8 by tumor cells by activating the p38-mitogen-activated protein kinase pathway in a neuropilin 1-dependent manner.

View Article: PubMed Central - PubMed

Affiliation: Institute for Cancer Research and Treatment (IRCC), University of Turin, School of Medicine, 10060 Candiolo, Italy.

ABSTRACT
Semaphorins are a large family of evolutionarily conserved morphogenetic molecules originally identified for their repelling role in axonal guidance. Intriguingly, semaphorins have recently been implicated in cancer progression (Neufeld, G., T. Lange, A. Varshavsky, and O. Kessler. 2007. Adv. Exp. Med. Biol. 600:118-131). In particular, semaphorin 3B (SEMA3B) is considered a putative tumor suppressor, and yet we found that it is expressed at high levels in many invasive and metastatic human cancers. By investigating experimental tumor models, we confirmed that SEMA3B expression inhibited tumor growth, whereas metastatic dissemination was surprisingly increased. We found that SEMA3B induced the production of interleukin (IL) 8 by tumor cells by activating the p38-mitogen-activated protein kinase pathway in a neuropilin 1-dependent manner. Silencing the expression of endogenous SEMA3B in tumor cells impaired IL-8 transcription. The release of IL-8, in turn, induced the recruitment of tumor-associated macrophages and metastatic dissemination to the lung, which could be rescued by blocking IL-8 with neutralizing antibodies. In conclusion, we report that SEMA3B exerts unexpected functions in cancer progression by fostering a prometastatic environment through elevated IL-8 secretion and recruitment of macrophages coupled to the suppression of tumor growth.

Show MeSH
Related in: MedlinePlus