Natalicchio Annalisa, Biondi Giuseppina, Marrano Nicola, Labarbuta Rossella, Tortosa Federica, Spagnuolo Rosaria, D'Oria Rossella, Carchia Emanuele, Leonardini Anna, Cignarelli Angelo, Perrini Sebastio, Laviola Luigi, Giorgino Francesco
Department of Emergency and Organ Transplantation (A.N., G.B., N.M., R.L., F.T., R.S., R.D., A.L., A.C., S.P., L.L., F.G.), Section of Internal Medicine, Endocrinology, Andrology, and Metabolic Diseases, University of Bari Aldo Moro, I-70124 Bari, Italy; and IRGS Biogem (E.C.), I-83031 Ariano Irpino, Avellino, Italy.
Endocrinology. 2016 Jun;157(6):2243-58. doi: 10.1210/en.2015-2003. Epub 2016 Apr 1.
The effects of prolonged exposure of pancreatic β-cells to high saturated fatty acids on glucagon-like peptide-1 (GLP-1) action were investigated. Murine islets, human pancreatic 1.1B4 cells, and rat INS-1E cells were exposed to palmitate for 24 hours. mRNA and protein expression/phosphorylation were measured by real-time RT-PCR and immunoblotting, respectively. Specific short interfering RNAs were used to knockdown expression of the GLP-1 receptor (Glp1r) and Srebf1. Insulin release was assessed with a specific ELISA. Exposure of murine islets, as well as of human and INS-1E β-cells, to palmitate reduced the ability of exendin-4 to augment insulin mRNA levels, protein content, and release. In addition, palmitate blocked exendin-4-stimulated cAMP-response element-binding protein and v-akt murine thymoma viral oncogene homolog phosphorylation, whereas phosphorylation of MAPK-ERK kinase-1/2 and ERK-1/2 was not altered. Similarly, RNA interference-mediated suppression of Glp1r expression prevented exendin-4-induced cAMP-response element-binding protein and v-akt murine thymoma viral oncogene homolog phosphorylation, but did not impair exendin-4 stimulation of MAPK-ERK kinase-1/2 and ERK-1/2. Both islets from mice fed a high fat diet and human and INS-1E β-cells exposed to palmitate showed reduced GLP-1 receptor and pancreatic duodenal homeobox-1 (PDX-1) and increased sterol regulatory element-binding protein (SREBP-1C) mRNA and protein levels. Furthermore, suppression of SREBP-1C protein expression prevented the reduction of PDX-1 and GLP-1 receptor levels and restored exendin-4 signaling and action. Finally, treatment of INS-1E cells with metformin for 24 h resulted in inhibition of SREBP-1C expression, increased PDX-1 and GLP-1 receptor levels, consequently, enhancement of exendin-4-induced insulin release. Palmitate impairs exendin-4 effects on β-cells by reducing PDX-1 and GLP-1 receptor expression and signaling in a SREBP-1C-dependent manner. Metformin counteracts the impairment of GLP-1 receptor signaling induced by palmitate.
研究了胰腺β细胞长期暴露于高饱和脂肪酸对胰高血糖素样肽-1（GLP-1）作用的影响。将小鼠胰岛、人胰腺1.1B4细胞和大鼠INS-1E细胞暴露于棕榈酸24小时。分别通过实时逆转录聚合酶链反应（RT-PCR）和免疫印迹法检测mRNA和蛋白质表达/磷酸化水平。使用特异性短发夹RNA敲低GLP-1受体（Glp1r）和固醇调节元件结合蛋白1（Srebf1）的表达。用特异性酶联免疫吸附测定（ELISA）评估胰岛素释放。将小鼠胰岛以及人和INS-1Eβ细胞暴露于棕榈酸会降低艾塞那肽-4增加胰岛素mRNA水平、蛋白质含量和释放的能力。此外，棕榈酸阻断了艾塞那肽-4刺激的环磷酸腺苷反应元件结合蛋白（CREB）和v-akt小鼠胸腺瘤病毒癌基因同源物（Akt）的磷酸化，而丝裂原活化蛋白激酶激酶1/2（MEK1/2）和细胞外信号调节激酶1/2（ERK1/2）的磷酸化未改变。同样，RNA干扰介导的Glp1r表达抑制可阻止艾塞那肽-4诱导的CREB和Akt磷酸化，但不损害艾塞那肽-4对MEK1/2和ERK1/2的刺激。喂食高脂饮食小鼠的胰岛以及暴露于棕榈酸的人和INS-1Eβ细胞均显示GLP-1受体和胰腺十二指肠同源盒-1（PDX-1）减少，固醇调节元件结合蛋白1C（SREBP-1C）mRNA和蛋白质水平增加。此外，抑制SREBP-1C蛋白表达可防止PDX-1和GLP-1受体水平降低，并恢复艾塞那肽-4信号传导和作用。最后，用二甲双胍处理INS-1E细胞24小时可导致SREBP-1C表达受到抑制，PDX-1和GLP-1受体水平增加，从而增强艾塞那肽-4诱导的胰岛素释放。棕榈酸通过以SREBP-1C依赖性方式降低PDX-1和GLP-1受体表达及信号传导来损害艾塞那肽-4对β细胞的作用。二甲双胍可抵消棕榈酸诱导的GLP-1受体信号传导损伤。
