Cruzat Vinicius Fernandes, Keane Kevin Noel, Scheinpflug Anita Lavarda, Cordeiro Robson, Soares Mario J, Newsholme Philip
School of Biomedical SciencesDirectorate of NutritionDietetics and Food Technology, School of Public Health, Curtin Health Innovation Research Institute of Ageing and Chronic Disease - Curtin University, GPO Box U1987, Perth, Western Australia, Australia 6845
School of Biomedical SciencesDirectorate of NutritionDietetics and Food Technology, School of Public Health, Curtin Health Innovation Research Institute of Ageing and Chronic Disease - Curtin University, GPO Box U1987, Perth, Western Australia, Australia 6845.
J Endocrinol. 2015 Mar;224(3):261-71. doi: 10.1530/JOE-14-0677. Epub 2014 Dec 30.
Obesity-associated diabetes and concomitant inflammation may compromise pancreatic β-cell integrity and function. l-glutamine and l-alanine are potent insulin secretagogues, with antioxidant and cytoprotective properties. Herein, we studied whether the dipeptide l-alanyl-l-glutamine (Ala-Gln) could exert protective effects via sirtuin 1/HUR (SIRT1/HUR) signalling in β-cells, against detrimental responses following ex vivo stimulation with inflammatory mediators derived from macrophages (IMMs). The macrophages were derived from blood obtained from obese subjects. Macrophages were exposed (or not) to lipopolysaccharide (LPS) to generate a pro-inflammatory cytokine cocktail. The cytokine profile was determined following analysis by flow cytometry. Insulin-secreting BRIN-BD11 β-cells were exposed to IMMs and then cultured with or without Ala-Gln for 24 h. Chronic insulin secretion, the l-glutamine-glutathione (GSH) axis, and the level of insulin receptor β (IR-β), heat shock protein 70 (HSP70), SIRT1/HUR, CCAAT-enhancer-binding protein homologous protein (CHOP) and cytochrome c oxidase IV (COX IV) were evaluated. Concentrations of cytokines, including interleukin 1β (IL1β), IL6, IL10 and tumour necrosis factor alpha (TNFα) in the IMMs, were higher following exposure to LPS. Subsequently, when β-cells were exposed to IMMs, chronic insulin secretion, and IR-β and COX IV levels were decreased, but these effects were partially or fully attenuated by the addition of Ala-Gln. The glutamine-GSH axis and HSP70 levels, which were compromised by IMMs, were also restored by Ala-Gln, possibly due to protection of SIRT1/HUR levels, and a reduction of CHOP expression. Using an ex vivo inflammatory approach, we have demonstrated Ala-Gln-dependent β-cell protection mediated by coordinated effects on the glutamine-GSH axis, and the HSP pathway, maintenance of mitochondrial metabolism and stimulus-secretion coupling essential for insulin release.
肥胖相关的糖尿病和伴随的炎症可能会损害胰腺β细胞的完整性和功能。L-谷氨酰胺和L-丙氨酸是有效的胰岛素促分泌剂，具有抗氧化和细胞保护特性。在此，我们研究了二肽L-丙氨酰-L-谷氨酰胺（Ala-Gln）是否能通过β细胞中的沉默调节蛋白1/ HuR（SIRT1/HuR）信号传导发挥保护作用，以对抗用源自巨噬细胞的炎症介质（IMMs）进行体外刺激后的有害反应。巨噬细胞来源于肥胖受试者的血液。将巨噬细胞暴露（或不暴露）于脂多糖（LPS）以产生促炎细胞因子混合物。通过流式细胞术分析后确定细胞因子谱。将分泌胰岛素的BRIN-BD11β细胞暴露于IMMs，然后在有或没有Ala-Gln的情况下培养24小时。评估慢性胰岛素分泌、L-谷氨酰胺-谷胱甘肽（GSH）轴以及胰岛素受体β（IR-β）、热休克蛋白70（HSP70）、SIRT1/HuR、CCAAT增强子结合蛋白同源蛋白（CHOP）和细胞色素c氧化酶IV（COX IV）的水平。暴露于LPS后，IMMs中细胞因子的浓度，包括白细胞介素1β（IL1β）、IL6、IL10和肿瘤坏死因子α（TNFα）更高。随后，当β细胞暴露于IMMs时，慢性胰岛素分泌以及IR-β和COX IV水平降低，但添加Ala-Gln可部分或完全减弱这些作用。IMMs损害的谷氨酰胺-GSH轴和HSP70水平也通过Ala-Gln得以恢复，这可能是由于对SIRT1/HuR水平的保护以及CHOP表达的降低。使用体外炎症方法，我们证明了Ala-Gln依赖的β细胞保护作用，其通过对谷氨酰胺-GSH轴、HSP途径、线粒体代谢的维持以及胰岛素释放所必需的刺激-分泌偶联的协同作用介导。
