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Suppressive effects of vascular endothelial growth factor-B on tumor growth in a mouse model of pancreatic neuroendocrine tumorigenesis.

Albrecht I, Kopfstein L, Strittmatter K, Schomber T, Falkevall A, Hagberg CE, Lorentz P, Jeltsch M, Alitalo K, Eriksson U, Christofori G, Pietras K - PLoS ONE (2010)

Bottom Line: Ectopic expression of VEGF-B in the insulin-producing β-cells of the pancreas did not alter the abundance or architecture of the islets of Langerhans.No differences in vascular density, perfusion or immune cell infiltration upon altered Vegfb gene dosage were noted.Taken together, our results illustrate the differences in biological function between members of the VEGF family, and highlight the necessity of in-depth functional studies of VEGF-B to fully understand the effects of VEGFR-1 inhibitors currently used in the clinic.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedicine, Institute of Biochemistry and Genetics, University of Basel, Basel, Switzerland.

ABSTRACT

Background: The family of vascular endothelial growth factors (VEGF) contains key regulators of blood and lymph vessel development, including VEGF-A, -B, -C, -D, and placental growth factor. The role of VEGF-B during physiological or pathological angiogenesis has not yet been conclusively delineated. Herein, we investigate the function of VEGF-B by the generation of mouse models of cancer with transgenic expression of VEGF-B or homozygous deletion of Vegfb.

Methodology/principal findings: Ectopic expression of VEGF-B in the insulin-producing β-cells of the pancreas did not alter the abundance or architecture of the islets of Langerhans. The vasculature from transgenic mice exhibited a dilated morphology, but was of similar density as that of wildtype mice. Unexpectedly, we found that transgenic expression of VEGF-B in the RIP1-Tag2 mouse model of pancreatic neuroendocrine tumorigenesis retarded tumor growth. Conversely, RIP1-Tag2 mice deficient for Vegfb presented with larger tumors. No differences in vascular density, perfusion or immune cell infiltration upon altered Vegfb gene dosage were noted. However, VEGF-B acted to increase blood vessel diameter both in normal pancreatic islets and in RIP1-Tag2 tumors.

Conclusions/significance: Taken together, our results illustrate the differences in biological function between members of the VEGF family, and highlight the necessity of in-depth functional studies of VEGF-B to fully understand the effects of VEGFR-1 inhibitors currently used in the clinic.

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Characterization of the phenotype of tumors from RIP1-Tag2; RIP1-VEGFB mice.A) RT-PCR analysis of VEGF-R1 expression in GLP1R+ β-tumor-cells and CD31+ tumor-derived blood-endothelial cells (BEC) isolated from 12 weeks old RIP1-Tag2 mice. B) Pancreatic tumor sections of control RIP1-Tag2 (left) and RIP1-Tag2; RIP1-VEGFB (right) mice were stained for human VEGF-B (red) to detect transgene expression. Nuclei were counterstained with DAPI. T =  Tumor, E =  Exocrine pancreas. Scale bar: 100 µm. C) Tumor incidence (left) and volumes (right) of RIP1-Tag2 (N =  36) and RIP1-Tag2; RIP1-VEGFB (N =  38) mice were determined at the age of 12 weeks. Single points represent the total tumor volume (or tumor number) per mouse as indicated. * P  =  0.0149 (Student's t-test). D) Tumor cell proliferation (left) and apoptosis (right) in RIP1-Tag2 and RIP1-Tag2; RIP1-VEGFB mice was determined by counting the number of BrdU and TUNEL positive tumor cells in a total of 7 to 10 microscopic fields (magnification 400×) per mouse.
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pone-0014109-g002: Characterization of the phenotype of tumors from RIP1-Tag2; RIP1-VEGFB mice.A) RT-PCR analysis of VEGF-R1 expression in GLP1R+ β-tumor-cells and CD31+ tumor-derived blood-endothelial cells (BEC) isolated from 12 weeks old RIP1-Tag2 mice. B) Pancreatic tumor sections of control RIP1-Tag2 (left) and RIP1-Tag2; RIP1-VEGFB (right) mice were stained for human VEGF-B (red) to detect transgene expression. Nuclei were counterstained with DAPI. T =  Tumor, E =  Exocrine pancreas. Scale bar: 100 µm. C) Tumor incidence (left) and volumes (right) of RIP1-Tag2 (N =  36) and RIP1-Tag2; RIP1-VEGFB (N =  38) mice were determined at the age of 12 weeks. Single points represent the total tumor volume (or tumor number) per mouse as indicated. * P  =  0.0149 (Student's t-test). D) Tumor cell proliferation (left) and apoptosis (right) in RIP1-Tag2 and RIP1-Tag2; RIP1-VEGFB mice was determined by counting the number of BrdU and TUNEL positive tumor cells in a total of 7 to 10 microscopic fields (magnification 400×) per mouse.

Mentions: The consequences of VEGF-B expression on tumor angiogenesis was assessed in the RIP1-Tag2 mouse model of islet cell carcinoma; a model that has been widely used to study tumor angiogenesis [21], [22], [23], [24], [25], [26], [27]. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) revealed that VEGF-B is readily detectable in normal β-islets of the mouse pancreas, and maintained at similar levels during the progression through hyperplastic islets and angiogenic islets into overt carcinomas in RIP1-Tag2 mice (data not shown). A cell line established from a RIP1-Tag2 tumor, β-TC3 [28], did not express the bona fide receptor for VEGF-B, VEGFR-1, and was not affected in its growth rate by VEGF-B (data not shown). Moreover, tumorous β-cells isolated from RIP1-Tag2 tumors did not express VEGFR-1 mRNA, in contrast to isolated blood endothelial cells from the same tumors (Figure 2a), making it likely that potential effects of transgenic expression of VEGF-B on tumor progression in RIP1-Tag2 mice are caused by paracrine stimulation. Double-transgenic RIP1-Tag2; RIP1-VEGFB mice expressed VEGF-B protein in pancreatic islets at high levels throughout the tumor progression pathway, as determined by immunostaining for human VEGF-B (Figure 2b). Moreover, tumors from RIP1-Tag2; RIP1-VEGFB mice contained abundant levels of human VEGF-B mRNA, as assessed by qRT-PCR, and protein, as assessed by ELISA (Figure S3a–b). No compensatory change was noted in the expression of mouse VEGF-B upon transgenic expression of human VEGF-B (Figure S3a).


Suppressive effects of vascular endothelial growth factor-B on tumor growth in a mouse model of pancreatic neuroendocrine tumorigenesis.

Albrecht I, Kopfstein L, Strittmatter K, Schomber T, Falkevall A, Hagberg CE, Lorentz P, Jeltsch M, Alitalo K, Eriksson U, Christofori G, Pietras K - PLoS ONE (2010)

Characterization of the phenotype of tumors from RIP1-Tag2; RIP1-VEGFB mice.A) RT-PCR analysis of VEGF-R1 expression in GLP1R+ β-tumor-cells and CD31+ tumor-derived blood-endothelial cells (BEC) isolated from 12 weeks old RIP1-Tag2 mice. B) Pancreatic tumor sections of control RIP1-Tag2 (left) and RIP1-Tag2; RIP1-VEGFB (right) mice were stained for human VEGF-B (red) to detect transgene expression. Nuclei were counterstained with DAPI. T =  Tumor, E =  Exocrine pancreas. Scale bar: 100 µm. C) Tumor incidence (left) and volumes (right) of RIP1-Tag2 (N =  36) and RIP1-Tag2; RIP1-VEGFB (N =  38) mice were determined at the age of 12 weeks. Single points represent the total tumor volume (or tumor number) per mouse as indicated. * P  =  0.0149 (Student's t-test). D) Tumor cell proliferation (left) and apoptosis (right) in RIP1-Tag2 and RIP1-Tag2; RIP1-VEGFB mice was determined by counting the number of BrdU and TUNEL positive tumor cells in a total of 7 to 10 microscopic fields (magnification 400×) per mouse.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2991338&req=5

pone-0014109-g002: Characterization of the phenotype of tumors from RIP1-Tag2; RIP1-VEGFB mice.A) RT-PCR analysis of VEGF-R1 expression in GLP1R+ β-tumor-cells and CD31+ tumor-derived blood-endothelial cells (BEC) isolated from 12 weeks old RIP1-Tag2 mice. B) Pancreatic tumor sections of control RIP1-Tag2 (left) and RIP1-Tag2; RIP1-VEGFB (right) mice were stained for human VEGF-B (red) to detect transgene expression. Nuclei were counterstained with DAPI. T =  Tumor, E =  Exocrine pancreas. Scale bar: 100 µm. C) Tumor incidence (left) and volumes (right) of RIP1-Tag2 (N =  36) and RIP1-Tag2; RIP1-VEGFB (N =  38) mice were determined at the age of 12 weeks. Single points represent the total tumor volume (or tumor number) per mouse as indicated. * P  =  0.0149 (Student's t-test). D) Tumor cell proliferation (left) and apoptosis (right) in RIP1-Tag2 and RIP1-Tag2; RIP1-VEGFB mice was determined by counting the number of BrdU and TUNEL positive tumor cells in a total of 7 to 10 microscopic fields (magnification 400×) per mouse.
Mentions: The consequences of VEGF-B expression on tumor angiogenesis was assessed in the RIP1-Tag2 mouse model of islet cell carcinoma; a model that has been widely used to study tumor angiogenesis [21], [22], [23], [24], [25], [26], [27]. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) revealed that VEGF-B is readily detectable in normal β-islets of the mouse pancreas, and maintained at similar levels during the progression through hyperplastic islets and angiogenic islets into overt carcinomas in RIP1-Tag2 mice (data not shown). A cell line established from a RIP1-Tag2 tumor, β-TC3 [28], did not express the bona fide receptor for VEGF-B, VEGFR-1, and was not affected in its growth rate by VEGF-B (data not shown). Moreover, tumorous β-cells isolated from RIP1-Tag2 tumors did not express VEGFR-1 mRNA, in contrast to isolated blood endothelial cells from the same tumors (Figure 2a), making it likely that potential effects of transgenic expression of VEGF-B on tumor progression in RIP1-Tag2 mice are caused by paracrine stimulation. Double-transgenic RIP1-Tag2; RIP1-VEGFB mice expressed VEGF-B protein in pancreatic islets at high levels throughout the tumor progression pathway, as determined by immunostaining for human VEGF-B (Figure 2b). Moreover, tumors from RIP1-Tag2; RIP1-VEGFB mice contained abundant levels of human VEGF-B mRNA, as assessed by qRT-PCR, and protein, as assessed by ELISA (Figure S3a–b). No compensatory change was noted in the expression of mouse VEGF-B upon transgenic expression of human VEGF-B (Figure S3a).

Bottom Line: Ectopic expression of VEGF-B in the insulin-producing β-cells of the pancreas did not alter the abundance or architecture of the islets of Langerhans.No differences in vascular density, perfusion or immune cell infiltration upon altered Vegfb gene dosage were noted.Taken together, our results illustrate the differences in biological function between members of the VEGF family, and highlight the necessity of in-depth functional studies of VEGF-B to fully understand the effects of VEGFR-1 inhibitors currently used in the clinic.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedicine, Institute of Biochemistry and Genetics, University of Basel, Basel, Switzerland.

ABSTRACT

Background: The family of vascular endothelial growth factors (VEGF) contains key regulators of blood and lymph vessel development, including VEGF-A, -B, -C, -D, and placental growth factor. The role of VEGF-B during physiological or pathological angiogenesis has not yet been conclusively delineated. Herein, we investigate the function of VEGF-B by the generation of mouse models of cancer with transgenic expression of VEGF-B or homozygous deletion of Vegfb.

Methodology/principal findings: Ectopic expression of VEGF-B in the insulin-producing β-cells of the pancreas did not alter the abundance or architecture of the islets of Langerhans. The vasculature from transgenic mice exhibited a dilated morphology, but was of similar density as that of wildtype mice. Unexpectedly, we found that transgenic expression of VEGF-B in the RIP1-Tag2 mouse model of pancreatic neuroendocrine tumorigenesis retarded tumor growth. Conversely, RIP1-Tag2 mice deficient for Vegfb presented with larger tumors. No differences in vascular density, perfusion or immune cell infiltration upon altered Vegfb gene dosage were noted. However, VEGF-B acted to increase blood vessel diameter both in normal pancreatic islets and in RIP1-Tag2 tumors.

Conclusions/significance: Taken together, our results illustrate the differences in biological function between members of the VEGF family, and highlight the necessity of in-depth functional studies of VEGF-B to fully understand the effects of VEGFR-1 inhibitors currently used in the clinic.

Show MeSH
Related in: MedlinePlus