Exendin-4 protects pancreatic beta cells from palmitate-induced apoptosis by interfering with GPR40 and the MKK4/7 stress kinase signalling pathway
Exendin-4 protects pancreatic beta cells from palmitate-induced apoptosis by interfering with GPR40 and the MKK4/7 stress kinase signalling pathway
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DOI：
10.1007/s00125-013-3028-4
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发表时间：
2013-11-01
期刊：
影响因子：
8.2
通讯作者：
Giorgino, Francesco
中科院分区：
文献类型：
--
作者：
Natalicchio, Annalisa;Labarbuta, Rossella;Giorgino, Francesco
本研究探讨了exendin-4对NEFA诱导的β细胞凋亡的保护作用机制，并在人和小鼠胰岛、大鼠胰岛素分泌型INS-1 E细胞和小鼠胰高血糖素分泌型α-TC 1 -6细胞中评价了exendin-4和棕榈酸酯的作用。mRNA和蛋白质表达/磷酸化分别通过实时RT-PCR和免疫印迹或免疫荧光法测量。使用针对Ib 1和Gpr 40的小干扰（si）RNA。通过两个独立的测定定量细胞凋亡。暴露于exendin-4的人类和小鼠原代胰岛和INS-1 E细胞，但不是α-TC 1 -6细胞，抑制应激激酶，c-Jun N-末端激酶（JNK）和p38丝裂原活化蛋白激酶（MAPK）的磷酸化，并防止响应棕榈酸的细胞凋亡。Exendin-4增加了内源性JNK阻断剂胰岛脑1（IB 1）的蛋白含量;然而，siRNA介导的IB 1减少并不损害Exendin-4抑制JNK和防止细胞凋亡的能力。Exendin-4降低G蛋白偶联受体40（GPR 40）的表达，并抑制棕榈酸诱导的丝裂原活化激酶激酶（MKK）4和MKK 7的磷酸化。exendin-4的作用在蛋白激酶A（PKA）抑制剂H89和KT 5720的存在下被废除。GPR 40的敲除以及特异性GPR 40拮抗剂的使用导致棕榈酸诱导的JNK和p38 MAPK磷酸化和细胞凋亡减少。Exendin-4通过降低GPR 40的表达和抑制MKK 7-和MKK 4-依赖的应激激酶JNK和p38 MAPK的磷酸化，以PKA依赖的方式抵消棕榈酸酯在β细胞中的促凋亡作用。
The mechanisms of the protective effects of exendin-4 on NEFA-induced beta cell apoptosis were investigated.The effects of exendin-4 and palmitate were evaluated in human and murine islets, rat insulin-secreting INS-1E cells and murine glucagon-secreting alpha-TC1-6 cells. mRNA and protein expression/phosphorylation were measured by real-time RT-PCR and immunoblotting or immunofluorescence, respectively. Small interfering (si)RNAs for Ib1 and Gpr40 were used. Cell apoptosis was quantified by two independent assays. Insulin release was assessed with an insulin ELISA.Exposure of human and murine primary islets and INS-1E cells, but not alpha-TC1-6 cells, to exendin-4 inhibited phosphorylation of the stress kinases, c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK), and prevented apoptosis in response to palmitate. Exendin-4 increased the protein content of islet-brain 1 (IB1), an endogenous JNK blocker; however, siRNA-mediated reduction of IB1 did not impair the ability of exendin-4 to inhibit JNK and prevent apoptosis. Exendin-4 reduced G-protein-coupled receptor 40 (GPR40) expression and inhibited palmitate-induced phosphorylation of mitogen-activated kinase kinase (MKK)4 and MKK7. The effects of exendin-4 were abrogated in the presence of the protein kinase A (PKA) inhibitors, H89 and KT5720. Knockdown of GPR40, as well as use of a specific GPR40 antagonist, resulted in diminished palmitate-induced JNK and p38 MAPK phosphorylation and apoptosis. Furthermore, inhibition of JNK and p38 MAPK activity prevented palmitate-induced apoptosis.Exendin-4 counteracts the proapoptotic effects of palmitate in beta cells by reducing GPR40 expression and inhibiting MKK7- and MKK4-dependent phosphorylation of the stress kinases, JNK and p38 MAPK, in a PKA-dependent manner.