The vasoactive intestinal peptide-receptor system is involved in human glioblastoma cell migration

Abstract 

Glioblastoma multiforme (GBM) is the most aggressive form of brain tumor in adults. This cancer has an infiltrative nature and the median survival of patients is about one year. Vasoactive intestinal peptide (VIP) belongs to a structurally related family of polypeptides and is a major regulatory factor in the central and peripheral nervous systems. VIP regulates proliferation of astrocytes and of numerous cancer cell lines and modulates migration in prostatic and colonic cancer cell lines. Little is known about the involvement of VIP and its receptors (VIP-receptor system) in proliferation or migration of GBM cells. The effects of VIP, PACAP and of synthetic VIP antagonists were tested in two human GBM cell lines, M059K and M059J, established from two different parts of a single tumor. In these cells, the data revealed that the VIP-receptor system did not affect proliferation but controlled cell migration. Indeed, in M059K cells which express components of the VIP receptor system, the VIP receptor antagonists and a PACAP antibody enhanced migration. The VIP receptor antagonists increased generation of typical migration-associated processes: filopodia and lamellipodia, and activation of Rac1 and Cdc42 GTPases. Reciprocally, in M059J cells which poorly express the VIP-receptor system, treatments with the agonists VIP and PACAP resulted in decreased cell migration. Furthermore, the peptides appeared to act through a subclass of binding sites displaying an uncommon very high affinity for these ligands. Taken together, these observations suggest that components of the VIP-receptor system negatively regulate cell migration, thus showing potential anti-oncogenic properties.

Abbreviations: VIP, vasoactive intestinal peptide, PACAP, pituitary adenylate cyclase-activating polypeptide, DMEM, Dulbecco’s modified Eagle’s medium, GAPDH, glyceraldehyde-3-phosphate dehydrogenase, PCR, polymerase chain reaction, PBS, phosphate-buffered saline, DTT, dithiothreitol, PMSF, phenylmethylsulfonyl fluoride, BSA, bovin serum albumin, ECL, electrochemiluminescence, TFA, trifluoroacetic acid, OD, optical density, MTS, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, DAB, diaminobenzidine, PAK-PBD, p21-activated kinase-PAK binding domain

Keywords: VIP receptors, Glioblastoma cell lines, Proliferation, Migration, Actin cytoskeleton, Small GTPases