Department of Physiology, University of Toronto, Toronto, Ontario M5S 1A8, USA.
J Biol Chem. 2010 Oct 29;285(44):33623-31. doi: 10.1074/jbc.M109.085084. Epub 2010 Aug 13.
The functional impact of adiponectin on pancreatic beta cells is so far poorly understood. Although adiponectin receptors (AdipoR1/2) were identified, their involvement in adiponectin-induced signaling and other molecules involved is not clearly defined. Therefore, we investigated the role of adiponectin in beta cells and the signaling mediators involved. MIN6 beta cells and mouse islets were stimulated with globular (2.5 μg/ml) or full-length (5 μg/ml) adiponectin under serum starvation, and cell viability, proliferation, apoptosis, insulin gene expression, and secretion were measured. Lysates were subjected to Western blot analysis to determine phosphorylation of AMP-activated protein kinase (AMPK), Akt, or ERK. Functional significance of signaling was confirmed using dominant negative mutants or pharmacological inhibitors. Participation of AdipoRs was assessed by overexpression or siRNA. Adiponectin failed to activate AMPK after 10 min or 1- and 24-h stimulation. ERK was significantly phosphorylated after 24-h treatment with adiponectin, whereas Akt was activated at all time points examined. 24-h stimulation with adiponectin significantly increased cell viability by decreasing cellular apoptosis, and this was prevented by dominant negative Akt, wortmannin (PI3K inhibitor), and U0126 (MEK inhibitor). Moreover, adiponectin regulated insulin gene expression and glucose-stimulated insulin secretion, which was also prevented by wortmannin and U0126 treatment. Interestingly, the data also suggest adiponectin-induced changes in Akt and ERK phosphorylation and caspase-3 may occur independent of the level of AdipoR expression. This study demonstrates a lack of AMPK involvement and implicates Akt and ERK in adiponectin signaling, leading to protection against apoptosis and stimulation of insulin gene expression and secretion in pancreatic beta cells.
脂联素对胰岛β细胞的功能影响目前尚不清楚。尽管已经鉴定出脂联素受体（AdipoR1/2），但其在脂联素诱导的信号转导中的作用以及涉及的其他分子尚不清楚。因此，我们研究了脂联素在β细胞中的作用及其涉及的信号转导介质。在血清饥饿的情况下，用球形（2.5 μg/ml）或全长（5 μg/ml）脂联素刺激 MIN6β细胞和小鼠胰岛，测量细胞活力、增殖、凋亡、胰岛素基因表达和分泌。对裂解物进行 Western blot 分析以确定 AMP 激活的蛋白激酶（AMPK）、Akt 或 ERK 的磷酸化。使用显性负突变体或药理学抑制剂来确认信号的功能意义。通过过表达或 siRNA 评估 AdipoRs 的参与。脂联素在 10 分钟或 1 和 24 小时刺激后未能激活 AMPK。ERK 在脂联素处理 24 小时后显著磷酸化，而 Akt 在所有检测时间点均被激活。脂联素刺激 24 小时可通过减少细胞凋亡来显著增加细胞活力，这可被显性负 Akt、wortmannin（PI3K 抑制剂）和 U0126（MEK 抑制剂）所阻止。此外，脂联素还调节胰岛素基因表达和葡萄糖刺激的胰岛素分泌，wortmannin 和 U0126 处理也可阻止这一过程。有趣的是，数据还表明脂联素诱导的 Akt 和 ERK 磷酸化和 caspase-3 的变化可能与 AdipoR 表达水平无关。本研究表明 AMPK 参与脂联素信号转导的程度较低，而 Akt 和 ERK 则参与脂联素信号转导，从而导致对胰岛β细胞凋亡的保护以及对胰岛素基因表达和分泌的刺激。
