Inhibitory effect of high leucine concentration on α-amylase secretion by pancreatic acinar cells: possible key factor of proteasome

The present study aimed to investigate whether leucine affects the pancreatic exocrine by controlling the antisecretory factor (AF) and cholecystokinin receptor (CCKR) expression as well as the proteasome activity in pancreatic acinar cells of dairy calves. The pancreatic acinar cells were isolated from newborn Holstein bull calves and cultured using the Dulbecco's modified Eagle's medium/nutrient mixture F12 Ham's liquid (DMEM/F12). There were six treatments of leucine dosage including 0 (control), 0.23, 0.45, 1.35, 4.05, and 12.15 mM, respectively. After culture for 3 h, the samples were collected for subsequent analysis. As the leucine concentration increased from 0 to 1.35 mM, the a-amylase activity in media decreased significantly (P<0.05), while further increase in leucine concentration did not show any decrease in alpha-amylase activity. Addition of leucine inhibited (P<0.05) the expression of AF and CCKR, and decreased the activity of proteasome (P<0.05) by 76%, 63%, 24%, 7%, and 9%, respectively. Correlation analysis results showed alpha-amylase secretion was negatively correlated with leucine concentration (P<0.01), and positively correlated with proteasome activity (P<0.01) and the expression of CCK1R (P<0.01) and AF (P<0.05). The biggest regression coefficient was showed between alpha-amylase activity and proteasome (0.7699, P<0.001). After inhibition of proteasome byMG-132, low dosage leucine decreased (P<0.05) the activity of proteasome and alpha-amylase, as well as the expression of CCK1R. In conclusion, we demonstrated that the high-concentration leucine induced decrease in alpha-amylase release was mainly by decreasing proteasome activity.