Effects of lorglumide on growth and invasion of human pancreatic cancer cell line Mia PaCa-2 in vitro through the cholecystokinin-cholecystokinin-1 receptor pathway

Abstract

Background: Cholecystokinin (CCK) has been found to be a growth stimulant through its special receptor pathway, especially for gastrointestinal malignancies. Although the CCK-1 receptor has been shown to be highly expressed in resected human pancreatic cancer samples, its role is less clear. Objective: The aim of this in vitro study was to investigate the CCK-1 receptor expression and the function of the CCK-CCK-1 receptor pathway in the human pancreatic adenocarcinoma cell line, Mia PaCa-2. Methods: The expression of the CCK-1 receptor in Mia PaCa-2 cells was detected by reverse-transcriptase polymerase chain reaction and flow cytometry. CCK-1 receptor agonist CCK-8S (the major transmitter form of CCK) and antagonist lorglumide were cultured respectively with Mia PaCa-2. Three groups were created for this study: CCK-8S group (Mia PaCa-2 cells treated with CCK-8S), lorglumide group (Mia PaCa-2 cells treated with lorglumide), and the control group (Mia PaCa-2 cells alone). Investigators were blinded to group designation. 3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and flow cytometry were used to detect the cell growth, cell cycle, and apoptosis. Apoptosis index rate was measured by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling. Cell invasion ability was observed by invasion assay. Expression of matrix metalloproteinase-2 (MMP-2) was measured by Western blotting. Results: Mia PaCa-2 cells were found to express the CCK-1 receptor. Compared with the control group (70.2% [1.5%]), CCK-8S was associated with significant mean (SD) cell proliferation (85.1% [1.7%]; P = 0.039), and the ratio in the S stage of the cell cycle increased significantly (50.5% [1.7%] vs 42.2% [1.4%]; P = 0.021). CCK-8S was also associated with increased Mia PaCa-2 cell invasion ability (123.8 [1.7] vs 102.1 [5.8]; P = 0.005 vs control). Compared with the control group, lorglumide was associated with significantly inhibited cell growth (52.1% [1.8%]; P = 0.002) and cell invasion (77.6% [1.2%]; P = 0.003). Lorglumide also induced G0/G1 cell cycle arrest and apoptosis (27.1% [3–5%] vs 3–7% [0.6%]; P = 0.003 vs control). The change of invasion ability appeared to be mediated by MMP-2 expression, which was upregulated by CCK-8S and downregulated by lorglumide. Conclusion: The findings of this in vitro study suggest that CCK may exert a trophic action on the Mia PaCa-2 cell line, while lorglumide inhibited the cell growth and invasion.