Roomp Kirsten, Kristinsson Hjalti, Schvartz Domitille, Ubhayasekera Kumari, Sargsyan Ernest, Manukyan Levon, Chowdhury Azazul, Manell Hannes, Satagopam Venkata, Groebe Karlfried, Schneider Reinhard, Bergquist Jonas, Sanchez Jean-Charles, Bergsten Peter
Luxembourg Centre for Systems Biomedicine, University of Luxembourg, Esch-Belval, Luxembourg.
Department of Medical Cell Biology, Uppsala University, Uppsala, Sweden.
PLoS One. 2017 Apr 27;12(4):e0176391. doi: 10.1371/journal.pone.0176391. eCollection 2017.
Studies on the pathophysiology of type 2 diabetes mellitus (T2DM) have linked the accumulation of lipid metabolites to the development of beta-cell dysfunction and impaired insulin secretion. In most in vitro models of T2DM, rodent islets or beta-cell lines are used and typically focus is on specific cellular pathways or organs. Our aim was to, firstly, develop a combined lipidomics and proteomics approach for lipotoxicity in isolated human islets and, secondly, investigate if the approach could delineate novel and/ or confirm reported mechanisms of lipotoxicity. To this end isolated human pancreatic islets, exposed to chronically elevated palmitate concentrations for 0, 2 and 7 days, were functionally characterized and their levels of multiple targeted lipid and untargeted protein species determined. Glucose-stimulated insulin secretion from the islets increased on day 2 and decreased on day 7. At day 7 islet insulin content decreased and the proinsulin to insulin content ratio doubled. Amounts of cholesterol, stearic acid, C16 dihydroceramide and C24:1 sphingomyelin, obtained from the lipidomic screen, increased time-dependently in the palmitate-exposed islets. The proteomic screen identified matching changes in proteins involved in lipid biosynthesis indicating up-regulated cholesterol and lipid biosynthesis in the islets. Furthermore, proteins associated with immature secretory granules were decreased when palmitate exposure time was increased despite their high affinity for cholesterol. Proteins associated with mature secretory granules remained unchanged. Pathway analysis based on the protein and lipid expression profiles implicated autocrine effects of insulin in lipotoxicity. Taken together the study demonstrates that combining different omics approaches has potential in mapping of multiple simultaneous cellular events. However, it also shows that challenges exist for effectively combining lipidomics and proteomics in primary cells. Our findings provide insight into how saturated fatty acids contribute to islet cell dysfunction by affecting the granule maturation process and confirmation in human islets of some previous findings from rodent islet and cell-line studies.
对2型糖尿病（T2DM）病理生理学的研究已将脂质代谢产物的积累与β细胞功能障碍及胰岛素分泌受损的发展联系起来。在大多数T2DM体外模型中，使用的是啮齿动物胰岛或β细胞系，且通常聚焦于特定的细胞途径或器官。我们的目标，一是开发一种用于分离的人胰岛脂毒性研究的脂质组学和蛋白质组学联合方法，二是研究该方法是否能够描绘出新的和/或证实已报道的脂毒性机制。为此，将分离的人胰岛暴露于慢性升高的棕榈酸浓度下0、2和7天，对其进行功能表征，并测定多种靶向脂质和非靶向蛋白质种类的水平。胰岛的葡萄糖刺激胰岛素分泌在第2天增加，在第7天减少。在第7天，胰岛胰岛素含量降低，胰岛素原与胰岛素含量的比值翻倍。脂质组学筛选得到的胆固醇、硬脂酸、C16二氢神经酰胺和C24:1鞘磷脂的含量在暴露于棕榈酸的胰岛中随时间增加。蛋白质组学筛选确定了参与脂质生物合成的蛋白质的相应变化，表明胰岛中胆固醇和脂质生物合成上调。此外，尽管与胆固醇具有高亲和力，但随着棕榈酸暴露时间增加，与未成熟分泌颗粒相关的蛋白质减少。与成熟分泌颗粒相关的蛋白质保持不变。基于蛋白质和脂质表达谱的通路分析表明胰岛素的自分泌作用与脂毒性有关。综上所述，该研究表明结合不同的组学方法在描绘多个同时发生的细胞事件方面具有潜力。然而，它也表明在原代细胞中有效结合脂质组学和蛋白质组学存在挑战。我们的研究结果深入了解了饱和脂肪酸如何通过影响颗粒成熟过程导致胰岛细胞功能障碍，并在人胰岛中证实了先前啮齿动物胰岛和细胞系研究的一些发现。
