Nakao K, Kagawa S, Wakabayashi S, Matsuoka A
Endocrinol Jpn. 1986 Oct;33(5):561-73. doi: 10.1507/endocrj1954.33.561.
Monolayer cultures of the pancreas of the neonatal rat were maintained in TCM 199 medium, supplemented with 5.5 mM glucose, with or without 5 mM 3-amino-3-deoxyglucose, and perifused to examine the changes which occurred in the insulin secretory response during culture. On day 0, B cells showed a monophasic insulin secretion in response to 16.7 mM glucose, whereas in the presence of 200 nM 12-o-tetradecanoyl phorbol-13-acetate, 40 microM lysophosphatidylcholine, 10 microM forskolin or 1 mM 3-isobutyl-1-methylxanthine, the same dose of glucose stimulated insulin secretion in a biphasic fashion. Under culture conditions without 3-amino-3-deoxyglucose, the response to glucose totally disappeared after 7 days, and that to 10 mM of either leucine or 2-ketoisocaproate was as low as that of day 0. In contrast, B cells that had been cultured for 7 days in medium with 3-amino-3-deoxyglucose showed an adult-like biphasic pattern in response to glucose. When stimulated by glucose at a linear gradient concentration running from 0 to 20 mM, the B cells responded to increasing concentrations of glucose in a dose-dependent fashion. Further, the response of cAMP to glucose was increased by adding forskolin or 3-isobutyl-1-methylxanthine, which also enhanced the secretion of insulin under either a step-wise or slow-rise stimulation with glucose. The effect of 12-o-tetradecanoyl phorbol-13-acetate was also outstanding. Likewise, the addition of either leucine or 2-keptoisocaproate induced a striking increase in the secondary phase secretion as well as promoting the rates of glutamine oxidation within the cells. In conclusion, it is suggested that the high response to a wider variety of stimuli may represent the reaction of neonatal B cells to the cultural milieu rather than a process of physiological development, and these effects exhibited by 3-amino-3-deoxyglucose would be related to a change in the constituents of glycoproteins in the cells.
新生大鼠胰腺的单层培养物在补充有5.5 mM葡萄糖的TCM 199培养基中培养，添加或不添加5 mM 3-氨基-3-脱氧葡萄糖，并进行灌流以检查培养过程中胰岛素分泌反应发生的变化。在第0天，B细胞对16.7 mM葡萄糖表现出单相胰岛素分泌，而在存在200 nM 12-O-十四烷酰佛波醇-13-乙酸酯、40 μM溶血磷脂酰胆碱、10 μM福斯高林或1 mM 3-异丁基-1-甲基黄嘌呤的情况下，相同剂量的葡萄糖以双相方式刺激胰岛素分泌。在没有3-氨基-3-脱氧葡萄糖的培养条件下，对葡萄糖的反应在7天后完全消失，对10 mM亮氨酸或2-酮异己酸的反应与第0天一样低。相反，在含有3-氨基-3-脱氧葡萄糖的培养基中培养7天的B细胞对葡萄糖表现出类似成年细胞的双相模式。当受到从0到20 mM的线性梯度浓度葡萄糖刺激时，B细胞以剂量依赖方式对增加的葡萄糖浓度作出反应。此外，添加福斯高林或3-异丁基-1-甲基黄嘌呤可增加cAMP对葡萄糖的反应，这也增强了在葡萄糖的逐步或缓慢升高刺激下胰岛素的分泌。12-O-十四烷酰佛波醇-13-乙酸酯的作用也很显著。同样，添加亮氨酸或2-酮异己酸会导致第二阶段分泌显著增加，并促进细胞内谷氨酰胺氧化速率。总之，有人认为对更广泛的刺激的高反应可能代表新生B细胞对培养环境的反应，而不是生理发育过程，3-氨基-3-脱氧葡萄糖所表现出的这些作用可能与细胞中糖蛋白成分的变化有关。
