Limits...
Immunoglobulin E induces VEGF production in mast cells and potentiates their pro-tumorigenic actions through a Fyn kinase-dependent mechanism.

Jiménez-Andrade GY, Ibarra-Sánchez A, González D, Lamas M, González-Espinosa C - J Hematol Oncol (2013)

Bottom Line: Although effects of IgE have been suggested to relay on the low-intensity activation of distinct effector elements of the immune system, such as mast cells (MC), experimental evidence on the role of IgE-induced production of inflammatory mediators on specific pathologies is scarce.In this work, we investigated the effect of monomeric IgE (in the absence of antigen) on the production of VEGF in MC, analyzed if monomeric IgE could exacerbate the pro-tumorigenic properties of that cell type and characterized some of the molecular mechanisms behind the effects of IgE on VEGF production and tumor growth.Our data suggest that monomeric IgE, in the absence of antigen, induces VEGF production in MC and in vivo contributes to melanoma tumor growth through a Fyn kinase-dependent mechanism.

View Article: PubMed Central - HTML - PubMed

Affiliation: Departamento de Farmacobiología, Cinvestav, IPN, Sede Sur, Calzada de los Tenorios 235, Col. Granjas Coapa, Tlalpan CP 14330, Mexico City, Mexico.

ABSTRACT

Background: High concentrations of plasmatic IgE have been related to distinct systemic inflammatory conditions that frequently predispose individuals to hypersensitivity reactions. Although effects of IgE have been suggested to relay on the low-intensity activation of distinct effector elements of the immune system, such as mast cells (MC), experimental evidence on the role of IgE-induced production of inflammatory mediators on specific pathologies is scarce. MC are an important component in tumor microenvironment where they seem to secrete a number of immunomodulatory and angiogenic mediators, such as the Vascular Endothelial Growth Factor (VEGF) by not well-described mechanisms. In this work, we investigated the effect of monomeric IgE (in the absence of antigen) on the production of VEGF in MC, analyzed if monomeric IgE could exacerbate the pro-tumorigenic properties of that cell type and characterized some of the molecular mechanisms behind the effects of IgE on VEGF production and tumor growth.

Methods: For in vitro studies, murine bone marrow-derived mast cells (BMMCs) were used. Pharmacological inhibitors and phosphorylation of key elements controlling VEGF secretion and protein translation were used to characterize the mechanism of VEGF production triggered by IgE.In vivo, the effect of a single i.v. administration of monomeric IgE on B16 melanoma tumor weight, intratumoral blood vessel formation and tumor-associated MC was assessed in four groups of mice: MC-proficient (WT), MC-deficient (Wsh), Wsh reconstituted with MC derived from WT mice (Wsh Rec WT) and Wsh reconstituted with MC derived from Fyn -/- mice (Wsh Rec Fyn -/-).

Results: Monomeric IgE induced VEGF secretion through a Fyn kinase-dependent mechanism and modulated de novo protein synthesis modifying the activity of the translational regulator 4E-BP1 in BMMCs. In vivo, monomeric IgE increased melanoma tumor growth, peritumoral MC and blood vessel numbers in WT but not in Wsh mice. The positive effects of IgE on melanoma tumor growth were reproduced after reconstitution of Wsh mice with WT but not with Fyn -/- BMMCs.

Conclusion: Our data suggest that monomeric IgE, in the absence of antigen, induces VEGF production in MC and in vivo contributes to melanoma tumor growth through a Fyn kinase-dependent mechanism.

Show MeSH

Related in: MedlinePlus

IgE-induced VEGF protein but not mRNA accumulation is affected in Fyn −/− BMMCs. (A and B) Intracellular VEGF accumulation after IgE treatment of MC. Two million WT and Fyn −/− BMMCs were stimulated with 1000 ng of IgE for eight h at 37°C. Cells were collected in a cytospin and slides were incubated in the presence of an anti-VEGF antibody (red) and DAPI (blue). Samples were processed to detect intracellular VEGF by confocal microscopy. A representative picture obtained from at least three independent experiments is shown. Scale bar = 10 μm. Mean fluorescence intensity values of VEGF signal obtained from experiments shown in A (n = 3) are represented in B. *, P < 0.0001 compared with IgE 0 ng; &, P < 0.001 compared with WT. Two way ANOVA post hoc Student-Newman-Keuls. (C and D) VEGF mRNA expression in IgE-stimulated MC. Total mRNA was purified from WT or Fyn −/− BMMCs treated with 1000 ng IgE and VEGF164 mRNA was amplified by RT-PCR. GAPDH amplification is shown as a control. Densitometric quantification of VEGF164 mRNA (n = 2) is shown in D.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3750531&req=5

Figure 2: IgE-induced VEGF protein but not mRNA accumulation is affected in Fyn −/− BMMCs. (A and B) Intracellular VEGF accumulation after IgE treatment of MC. Two million WT and Fyn −/− BMMCs were stimulated with 1000 ng of IgE for eight h at 37°C. Cells were collected in a cytospin and slides were incubated in the presence of an anti-VEGF antibody (red) and DAPI (blue). Samples were processed to detect intracellular VEGF by confocal microscopy. A representative picture obtained from at least three independent experiments is shown. Scale bar = 10 μm. Mean fluorescence intensity values of VEGF signal obtained from experiments shown in A (n = 3) are represented in B. *, P < 0.0001 compared with IgE 0 ng; &, P < 0.001 compared with WT. Two way ANOVA post hoc Student-Newman-Keuls. (C and D) VEGF mRNA expression in IgE-stimulated MC. Total mRNA was purified from WT or Fyn −/− BMMCs treated with 1000 ng IgE and VEGF164 mRNA was amplified by RT-PCR. GAPDH amplification is shown as a control. Densitometric quantification of VEGF164 mRNA (n = 2) is shown in D.

Mentions: In order to investigate the role of Fyn kinase in the IgE-dependent VEGF synthesis, BMMCs derived from WT or Fyn −/− mice were incubated at 37°C in the presence of vehicle or IgE (1000 ng/ml) for 8 hours. Cells were collected using a cytospin centrifuge and processed to detect VEGF by fluorescent immunostaining. Figures 2A and B show that low VEGF positive signal was observed in WT BMMCs and IgE addition increased intracellular VEGF. In contrast, VEGF immunoreactivity was significantly lower in Fyn −/− BMMCs in both basal and IgE-stimulated conditions.


Immunoglobulin E induces VEGF production in mast cells and potentiates their pro-tumorigenic actions through a Fyn kinase-dependent mechanism.

Jiménez-Andrade GY, Ibarra-Sánchez A, González D, Lamas M, González-Espinosa C - J Hematol Oncol (2013)

IgE-induced VEGF protein but not mRNA accumulation is affected in Fyn −/− BMMCs. (A and B) Intracellular VEGF accumulation after IgE treatment of MC. Two million WT and Fyn −/− BMMCs were stimulated with 1000 ng of IgE for eight h at 37°C. Cells were collected in a cytospin and slides were incubated in the presence of an anti-VEGF antibody (red) and DAPI (blue). Samples were processed to detect intracellular VEGF by confocal microscopy. A representative picture obtained from at least three independent experiments is shown. Scale bar = 10 μm. Mean fluorescence intensity values of VEGF signal obtained from experiments shown in A (n = 3) are represented in B. *, P < 0.0001 compared with IgE 0 ng; &, P < 0.001 compared with WT. Two way ANOVA post hoc Student-Newman-Keuls. (C and D) VEGF mRNA expression in IgE-stimulated MC. Total mRNA was purified from WT or Fyn −/− BMMCs treated with 1000 ng IgE and VEGF164 mRNA was amplified by RT-PCR. GAPDH amplification is shown as a control. Densitometric quantification of VEGF164 mRNA (n = 2) is shown in D.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750531&req=5

Figure 2: IgE-induced VEGF protein but not mRNA accumulation is affected in Fyn −/− BMMCs. (A and B) Intracellular VEGF accumulation after IgE treatment of MC. Two million WT and Fyn −/− BMMCs were stimulated with 1000 ng of IgE for eight h at 37°C. Cells were collected in a cytospin and slides were incubated in the presence of an anti-VEGF antibody (red) and DAPI (blue). Samples were processed to detect intracellular VEGF by confocal microscopy. A representative picture obtained from at least three independent experiments is shown. Scale bar = 10 μm. Mean fluorescence intensity values of VEGF signal obtained from experiments shown in A (n = 3) are represented in B. *, P < 0.0001 compared with IgE 0 ng; &, P < 0.001 compared with WT. Two way ANOVA post hoc Student-Newman-Keuls. (C and D) VEGF mRNA expression in IgE-stimulated MC. Total mRNA was purified from WT or Fyn −/− BMMCs treated with 1000 ng IgE and VEGF164 mRNA was amplified by RT-PCR. GAPDH amplification is shown as a control. Densitometric quantification of VEGF164 mRNA (n = 2) is shown in D.
Mentions: In order to investigate the role of Fyn kinase in the IgE-dependent VEGF synthesis, BMMCs derived from WT or Fyn −/− mice were incubated at 37°C in the presence of vehicle or IgE (1000 ng/ml) for 8 hours. Cells were collected using a cytospin centrifuge and processed to detect VEGF by fluorescent immunostaining. Figures 2A and B show that low VEGF positive signal was observed in WT BMMCs and IgE addition increased intracellular VEGF. In contrast, VEGF immunoreactivity was significantly lower in Fyn −/− BMMCs in both basal and IgE-stimulated conditions.

Bottom Line: Although effects of IgE have been suggested to relay on the low-intensity activation of distinct effector elements of the immune system, such as mast cells (MC), experimental evidence on the role of IgE-induced production of inflammatory mediators on specific pathologies is scarce.In this work, we investigated the effect of monomeric IgE (in the absence of antigen) on the production of VEGF in MC, analyzed if monomeric IgE could exacerbate the pro-tumorigenic properties of that cell type and characterized some of the molecular mechanisms behind the effects of IgE on VEGF production and tumor growth.Our data suggest that monomeric IgE, in the absence of antigen, induces VEGF production in MC and in vivo contributes to melanoma tumor growth through a Fyn kinase-dependent mechanism.

View Article: PubMed Central - HTML - PubMed

Affiliation: Departamento de Farmacobiología, Cinvestav, IPN, Sede Sur, Calzada de los Tenorios 235, Col. Granjas Coapa, Tlalpan CP 14330, Mexico City, Mexico.

ABSTRACT

Background: High concentrations of plasmatic IgE have been related to distinct systemic inflammatory conditions that frequently predispose individuals to hypersensitivity reactions. Although effects of IgE have been suggested to relay on the low-intensity activation of distinct effector elements of the immune system, such as mast cells (MC), experimental evidence on the role of IgE-induced production of inflammatory mediators on specific pathologies is scarce. MC are an important component in tumor microenvironment where they seem to secrete a number of immunomodulatory and angiogenic mediators, such as the Vascular Endothelial Growth Factor (VEGF) by not well-described mechanisms. In this work, we investigated the effect of monomeric IgE (in the absence of antigen) on the production of VEGF in MC, analyzed if monomeric IgE could exacerbate the pro-tumorigenic properties of that cell type and characterized some of the molecular mechanisms behind the effects of IgE on VEGF production and tumor growth.

Methods: For in vitro studies, murine bone marrow-derived mast cells (BMMCs) were used. Pharmacological inhibitors and phosphorylation of key elements controlling VEGF secretion and protein translation were used to characterize the mechanism of VEGF production triggered by IgE.In vivo, the effect of a single i.v. administration of monomeric IgE on B16 melanoma tumor weight, intratumoral blood vessel formation and tumor-associated MC was assessed in four groups of mice: MC-proficient (WT), MC-deficient (Wsh), Wsh reconstituted with MC derived from WT mice (Wsh Rec WT) and Wsh reconstituted with MC derived from Fyn -/- mice (Wsh Rec Fyn -/-).

Results: Monomeric IgE induced VEGF secretion through a Fyn kinase-dependent mechanism and modulated de novo protein synthesis modifying the activity of the translational regulator 4E-BP1 in BMMCs. In vivo, monomeric IgE increased melanoma tumor growth, peritumoral MC and blood vessel numbers in WT but not in Wsh mice. The positive effects of IgE on melanoma tumor growth were reproduced after reconstitution of Wsh mice with WT but not with Fyn -/- BMMCs.

Conclusion: Our data suggest that monomeric IgE, in the absence of antigen, induces VEGF production in MC and in vivo contributes to melanoma tumor growth through a Fyn kinase-dependent mechanism.

Show MeSH
Related in: MedlinePlus