The Notch signaling pathway is related to neurovascular progression of pancreatic cancer.
OBJECTIVE
To analyze the potential role of the Notch signaling pathway in pancreatic cancer angiogenesis and invasion.


BACKGROUND
Angiogenesis, pain, and early neuroinvasion are clinical features of pancreatic cancer. Blood vessels and nerves develop together and use common routes through the organism. The Notch pathway (Notch-1/4, Jagged-1/2, Delta-1) appears crucial in this process. The current study analyzed the Notch pathway in pancreatic cancer and characterized its angiogenic and invasive effects.


METHODS
Five PaCa cell lines were cultured for the in vitro experiments. Real-time quantitative RT-PCR was done to quantify mRNA expression in 31 human PaCa specimens, and immunohistochemistry was used to localize protein expression within tumor specimens. Activation of the Notch signaling was done by transfection of PaCa cells with a constitutive active Notch-1 mutant (Notch-IC). Overexpression of Jagged and Delta was achieved by transfection of full-length cDNA. Spheroid assays were used to study angiogenesis and ELISAs to measure VEGF, bFGF, and angiogenin expression. Matrigel invasion assays were used to analyze tumor cell invasion.


RESULTS
Notch-3 and Notch-4 mRNA were significantly (P < 0.001) overexpressed in PaCa. Immunohistochemistry revealed protein accumulation of Notch-1 as well. All ligands were significantly up-regulated. A positive immunosignal of ligands was seen in nerves, blood vessels, and ductal tumor cells. Transfection of PaCa cells with the constitutive active Notch-IC mutant and with Jagged-1 revealed increased levels for VEGF. Concomitantly, recombinant Jagged-1 increased sprouting of endothelial cells in the spheroid assay.


CONCLUSIONS
The Notch pathway most likely regulates neurovascular development in pancreatic cancer. Activation of this signaling pathway by constitutive Notch-1 mutants and by Jagged-1 causes an angiogenic and invasive tumor phenotype. Specific blockade of Notch signaling may therefore be beneficial for patients with pancreatic cancer.
Büchler
Peter
P
Department of General Surgery, University of Heidelberg, Im Neuenheimer Feld 110, 69120 Heidelberg, Germany.
Gazdhar
Amiq
A
Schubert
Mario
M
Giese
Nathalia
N
Reber
Howard A
HA
Hines
Oscar J
OJ
Giese
Thomas
T
Ceyhan
Güralp O
GO
Müller
Michael
M
Büchler
Markus W
MW
Friess
Helmut
H
D018450:Disease Progression; D005260:Female; D015870:Gene Expression; D006801:Humans; D007124:Immunoenzyme Techniques; D066298:In Vitro Techniques; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D008297:Male; D009389:Neovascularization, Pathologic; D010190:Pancreatic Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D014162:Transfection; D014407:Tumor Cells, Cultured
D036341:Intercellular Signaling Peptides and Proteins; D012333:RNA, Messenger