Cancer Medicine – University Cancer Center Hamburg (UCCH)
Beschreibung:
Background: Receptor Activator of Nuclear Factor kappa-B (RANK) and its ligand (RANKL) are well-described players in osteometabolism, immunomodulation and carcinogenesis. Their roles in human Pancreatic Ductal Adenocarcinoma (PDAC) have not been described so far.
Methods: Protein expression was analyzed via Westernblots and qrtPCR. A lentivrial si-RNA mediated knockdown of RANK was performed in two pancreatic carcinoma cell lines BxPC3(BxPc3 RANK-KO) and Panc-1(Panc-1RANK-KO). Proliferation rates of cell lines were determined by established MTT assays. Migration rates were measured by transwell migration assays. Destribution of RANK and RANKL expression was analyzed via immunohistochemical staining (IHC) in malignant and precancerous tissue specimes of 34 PDAC patients.
Results: RANKL, RANK, and Cyclooxygenase2 (COX2) were expressed in PDAC cell lines on mRNA- and protein level. In contrast to BxPC3 , Panc-1 showed high levels of RANK but low levels of COX2 expression. COX2 is inducible by soluble RANKL (sRANKL) stimulation in Panc-1 and BxPC3 cells (BxPC3>Panc-1). BxPC3 cells increased proliferation rate significantly compared to a non-malignant reference pancreas cell line (HPDE) after sRANKL stimulation. Panc-1RANK-KO and BxPc3 RANK-KO cells did not respond to sRANKL stimulation. Migration rate of BxPC3 cells decreased after sRANKL stimulation. IHC staining of PDAC specimens showed positivity for RANKL and RANK. RANK showed signs of suborganelle positive staining; in contrast, it was expressed membranously in pancreatic intraepithelial neoplasia(PANIN). RANKL showed positivity in Fibroblasts and PDAC tissue in IHC.
Conclusions: We provide evidence for RANK- and RANKL expression in PDAC and the stromal compartment, respectively. The RANK-signaling pathway becomes activated upon RANKL stimulation, resulting in target gene expression, e.g. COX2. RANK-signaling seems to be a key player in COX2 dependent proliferation and migration in human PDAC cells. Also, RANK is cytoplasmic expressed in human PDAC tissue and membranous expressed in PANINs; thus RANK might be involved in carcinogenesis . RANKL shows signs of expression in PDAC cells and fibroblast indicating tumor-stroma interaction.