Sphingosine-1-phosphate (S1P) as a check point for proinflammatory cytokine-induced inflammation and death in pancreatic beta-cells during T1DM development – in search for a novel beta-cell specific target to prevent and treat the disease

1型糖尿病（T1DM）是一种具有强烈遗传背景的慢性自身免疫性疾病，导致控制葡萄糖稳态的胰腺胰岛素分泌β细胞逐渐丧失。在T1DM发展过程中，被激活的免疫细胞浸润胰岛，释放促炎细胞因子，诱导应激反应，导致β细胞死亡。β细胞非常容易受到细胞因子毒性的影响，因为它们的抗氧化防御能力较弱，并且促炎细胞因子信号比抗炎细胞因子信号更占优势。炎症的分子机制在β细胞中仍然知之甚少，使新的治疗方法无法产生。最近在个体的血清和pbmc鞘脂谱显著紊乱的风险或新近诊断为T1DM的描述。研究表明，在自身免疫性糖尿病动物模型中，使用芬戈莫德（一种鞘氨醇-1磷酸（S1P）受体1拮抗剂）可以预防糖尿病并减少胰岛浸润。然而，对于S1P在细胞因子诱导的β细胞炎症中的作用，特别是β细胞起源的作用，目前仍缺乏评估。我们的项目旨在缩小这一知识差距。我们将研究β细胞S1P代谢在细胞因子毒性和炎症中的作用，特别关注人类β细胞。我们的初步结果表明，S1P主要由促炎鞘氨酸激酶2 （SK2）产生，该基因在T1DM受试者中发现了多态性，并在某些细胞器中产生S1P。我们的数据表明，即使没有平行的细胞因子暴露，SK2敲低和SK2过表达对细胞因子介导的毒性也有保护作用。此外，我们的初步脂质组学结果表明，大量细胞因子介导的脂质重排可能与炎症有关，并可能与异常的S1P代谢有关。我们将使用一系列现代分子生物学技术和工具来研究S1P是否作为暴露于细胞因子的β细胞的炎症检查点。S1P代谢酶的基因操作对细胞因子介导的β细胞命运的影响将在大鼠和人β细胞、原代胰岛和动物模型中进行研究。我们的目标是揭示细胞因子诱导的新的s1p依赖性炎症机制。利用转基因人类β细胞与来自T1DM个体的pbmc共培养模型，我们将探索胰岛分泌的β细胞来源的S1P在T1DM条件下自身免疫介导的β细胞破坏中的作用。最后，我们将分析非糖尿病和T1DM患者胰腺组织中与炎症和β细胞身份标记平行的s1p代谢酶的丰度。我们的项目旨在确定与细胞因子介导的炎症和β细胞衰竭相关的最相关的S1P代谢酶，作为T1DM中β细胞保护的新药物靶点。

Type 1 diabetes (T1DM) is a chronic autoimmune disease with a strong genetic background, leading to a gradual loss of pancreatic insulin-secreting beta-cells that control glucose homeostasis. Proinflammatory cytokines released by activated immune cells infiltrating pancreatic islets during T1DM development induce a stress response resulting in beta-cell death. Beta-cells are very susceptible to cytokine toxicity due to a weak antioxidative defense and predominance of proinflammatory over antiinflammatory cytokine signaling. The molecular mechanisms of inflammation remain poorly understood in beta-cells, disabling the generation of novel therapeutic approaches. Recently significant disturbances in serum and PBMCs sphingolipid profiles in individuals at risk of or lately diagnosed with T1DM were described. Studies using fingolimod, a sphingosine-1 phosphate (S1P) receptor 1 antagonist, documented diabetes prevention and reduced islet infiltration in animal models of autoimmune diabetes. However, evaluation of S1P contribution, particularly of beta-cell origin, to cytokine-induced inflammation in beta-cells is still missing. Our project aims to close this knowledge gap. We will study the role of beta-cell S1P metabolism in cytokine toxicity and inflammation, with a particular focus on human beta-cells. Our preliminary results show that S1P is predominantly generated by the proinflammatory sphingosine kinase 2 (SK2), of which a gene polymorphism has been identified in T1DM subjects and which generates S1P in certain cellular organelles. Our data demonstrate protection against cytokine-mediated toxicity by SK2 knockdown and a pro-apoptotic effect of SK2 overexpression, even without a parallel cytokine exposure. Moreover, our preliminary lipidomics results suggest substantial cytokine-mediated lipid rearrangements that can be linked to inflammation and are likely related to an aberrant S1P metabolism. We will use an array of modern molecular biology techniques and tools to investigate whether S1P acts as an inflammation check point in beta-cells exposed to cytokines. The effects of genetic manipulations of S1P metabolizing enzymes on cytokine-mediated beta-cell fate will be studied in rat and humanbeta-cells, primary islets and animal models. We aim to uncover novel S1P-dependent inflammation mechanisms induced by cytokines. Using a co-culture model of genetically modified human beta-cells with PBMCs from T1DM individuals we will explore a role of beta-cell derived S1P secreted within islets in the autoimmune-mediated beta-cell destruction under T1DM conditions. Finally, the abundance of S1P-metabolizing enzymes in parallel with inflammation and beta-cell identity markers will be analyzed in pancreatic tissue from nondiabetic and T1DM individuals. Our project aims to identify the most relevant S1P metabolizing enzyme associated with cytokine-mediated inflammation and beta-cell failure as a novel druggable target for beta-cell protection in T1DM.