College of Animal Science and Technology, Northwest A&F University, Yangling 712100, China.
College of Animal Science and Technology, Northwest A&F University, Yangling 712100, China
Biosci Rep. 2018 Dec 11;38(6). doi: 10.1042/BSR20181455. Print 2018 Dec 21.
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 α-amylase activity in media decreased significantly (<0.05), while further increase in leucine concentration did not show any decrease in α-amylase activity. Addition of leucine inhibited (<0.05) the expression of AF and CCKR, and decreased the activity of proteasome (<0.05) by 76%, 63%, 24%, 7%, and 9%, respectively. Correlation analysis results showed α-amylase secretion was negatively correlated with leucine concentration (<0.01), and positively correlated with proteasome activity (<0.01) and the expression of CCK1R (<0.01) and AF (<0.05). The biggest regression coefficient was showed between α-amylase activity and proteasome (0.7699, <0.001). After inhibition of proteasome by MG-132, low dosage leucine decreased (<0.05) the activity of proteasome and α-amylase, as well as the expression of CCK1R. In conclusion, we demonstrated that the high-concentration leucine induced decrease in α-amylase release was mainly by decreasing proteasome activity.
本研究旨在探讨亮氨酸是否通过控制抗分泌因子 (AF) 和胆囊收缩素受体 (CCKR) 的表达以及胰腺腺泡细胞蛋白酶体活性来影响奶牛胰腺外分泌。从新生荷斯坦公牛犊牛中分离胰腺腺泡细胞，并用 Dulbecco's modified Eagle's medium/nutrient mixture F12 Ham's liquid (DMEM/F12) 进行培养。亮氨酸剂量有 6 个处理，分别为 0（对照）、0.23、0.45、1.35、4.05 和 12.15 mM。培养 3 h 后收集样品进行后续分析。随着亮氨酸浓度从 0 增加到 1.35 mM，培养基中 α-淀粉酶的活性显著降低（<0.05），而进一步增加亮氨酸浓度并未导致 α-淀粉酶活性降低。添加亮氨酸抑制（<0.05）AF 和 CCKR 的表达，并使蛋白酶体活性降低（<0.05）分别为 76%、63%、24%、7%和 9%。相关性分析结果表明，α-淀粉酶分泌与亮氨酸浓度呈负相关（<0.01），与蛋白酶体活性呈正相关（<0.01），与 CCK1R 表达呈正相关（<0.01），与 AF 表达呈正相关（<0.05）。α-淀粉酶活性与蛋白酶体之间的回归系数最大（0.7699，<0.001）。用 MG-132 抑制蛋白酶体后，低剂量亮氨酸降低（<0.05）蛋白酶体和 α-淀粉酶活性以及 CCK1R 的表达。总之，我们证明高浓度亮氨酸诱导的 α-淀粉酶释放减少主要是通过降低蛋白酶体活性来实现的。
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