Background: Various alcoholic beverages have different effects on pancreatic enzyme secretion in vivo and in vitro. Recently we demonstrated that beer dose-dependently induces amylase release of rat pancreatic acinar cells, whereas pure ethanol and other alcoholic beverages have no effect. The aims of this study were to: (1) investigate the involved signaling pathways in the beer-induced enzyme secretion of rat pancreatic acinar cells and (2) characterize the responsible nonalcoholic compounds from beer.
Methods: Rat pancreatic AR4-2J cells were differentiated by dexamethasone treatment for 72 hours. After incubation of cells with 1 to 10% (v/v) beer (containing 4.7% v/v ethanol) in the absence or presence of the maximal effective concentration of cholecystokinin (CCK) (100 nM) for 60 minutes, protein secretion was measured using amylase activity assay. To study the involved signaling pathways, cells were pretreated with selective inhibitors or the fluorescent dye Fura2/AM for 15 and 30 minutes, respectively. To characterize the responsible compounds, beer was distilled, lyophilized, dialyzed, or treated with proteases prior stimulation of the cells. Extract of barley was prepared by boiling the crop and subsequent filtration.
Results: Stimulation with 5% and 10% beer (v/v) significantly (p < 0.001) increased maximally CCK-induced amylase by 55 ± 25% and 56 ± 37%, respectively. By using selective antagonists, we found that inhibition of phospholipase C (PLC) and inositol 1,4,5-trisphosphate-receptor binding reduced beer-induced amylase release, whereas inhibition of protein kinase C, adenylate cyclase, and protein kinase A had no significant effect. Using the fluorescent Ca2+ indicator Fura-2/AM revealed that beer induces an increase of cytosolic free Ca2+ concentration. Stimulation of AR4-2J cells with preproducts of beer and fermented glucose indicated that the stimulatory substances from beer derived from barley and are not produced during alcoholic fermentation. Furthermore, the stimulants from beer are thermostable, nonvolatile substances with a molecular weight higher than 15 kDa.
Conclusions: Beer-induced enzyme secretion of AR4-2J cells is, at least in part, mediated by the activation of PLC and subsequent Ca2+ release from internal stores. However, the additive effect of beer on CCK-induced amylase release suggests that additional signaling pathways are involved. The yet unknown stimulants of pancreatic enzyme secretion originate from barley and their stimulatory potential is maintained during the process of malting and brewing.