Impact of cyclooxygenase 2-dependent generated prostaglandin E2 and other prostanoids in the development of diet-induced non-alcoholic steatohepatitis (NASH)

世界范围内的肥胖症和随之而来的疾病，如代谢综合征和2型糖尿病的患病率正在迅速增加。非酒精性脂肪性肝病（nonalcoholic fatty liver disease，NAFLD）是代谢综合征的肝脏表现，是功能性肝病的主要原因，以肝脏脂质过度积聚为特征。一些患者发展为严重形式的非酒精性脂肪性肝炎（NASH），其中脂肪变性伴有炎症和纤维化，这导致肝硬化、肝癌和器官衰竭的风险增加。除了细胞因子外，前列腺素E2（PGE 2）等前列腺素在炎症调节中起主要作用。前列腺素是在炎症过程中依赖环氧合酶2（Cox 2）合成的生物活性脂质，并且可以起到放大或减弱炎症的作用。在项目HE-7032/1-1的范围内，建立了诱导NASH的饮食，其在野生型小鼠中诱导肝脂肪变性伴炎症和纤维化以及肥胖和胰岛素抵抗，因此在代谢综合征中表现出人NASH的所有临床参数。在这些小鼠以及NASH患者中，Cox 2和mPGES 1（PGE 2合成中的Cox 2下游酶）的肝脏表达增强，表明PGE 2形成局部增加。PGE 2可抑制促炎细胞因子TNFα的表达。在喂食NASH诱导饲料的mPGES 1缺陷小鼠的肝脏中，与野生型小鼠相比，TNFα的诱导增加，导致更明显的肝细胞凋亡。与预期相反，mPGES 1缺乏仅降低肝脏PGE 2水平，但不足以完全消除饮食诱导的肝脏PGE 2生成。因此，基因型之间肝脏PGE 2水平的微小差异不会导致更严重的饮食诱导的肝脏炎症。然而，在与Cox 2抑制剂吲哚美辛孵育的原代巨噬细胞中，炎症介导的PGE 2产生增加被完全消除。后续项目将关注Cox 2依赖性产生的前列腺素如PGE 2在饮食诱导的NASH发展中的影响。巨噬细胞、肝细胞或两种细胞类型中具有Cox 2特异性缺失的小鼠以及它们各自的对照将接受标准和NASH诱导饮食，并将表征NASH的发展和进展。来自个体基因型的巨噬细胞群体和肝细胞的平行体外实验将有助于确定Cox 2在细胞水平上的促炎或抗炎作用。此外，将在人肝探针中验证结果。该项目的第二部分将研究饮食诱导NASH小鼠肝脏中不同巨噬细胞群体的异质性。这些不同的巨噬细胞应被分离和表征其对前列腺素类如PGE 2的敏感性。

The worldwide prevalence of obesity and the ensuing diseases like the metabolic syndrome and type2 diabetes is increasing rapidly. Non-alcoholic fatty liver disease (NAFLD), which is characterized by an excessive hepatic lipid accumulation, is defined as the hepatic manifestation of the metabolic syndrome and the main reason for functional liver diseases. Some patients develop the severe form of non-alcoholic steatohepatitis (NASH), where steatosis isaccompanied by inflammation and fibrosis, which results in an increased risk for cirrhosis, liver cancer and organ failure. Beside cytokines, prostaglandins like prostaglandin E2 (PGE2) play a Major role in the regulation of the inflammation. Prostaglandins are bioactive lipids that are synthesized cyclooxygenase 2 (Cox2)-dependently during inflammatory processes and can act to amplify or to attenuate inflammation. Within the scope of the project HE-7032/1-1 a NASHinducing diet was established, which induced hepatic steatosis with inflammation and fibrosis as well as obesity and insulin resistance in wildtype mice and thus exhibited all clinical parameters of human NASH in the metabolic syndrome. In these mice, as well as in patients with NASH, the hepatic expression of Cox2 and mPGES1, the Cox2- downstream enzyme in PGE2-synthesis, was enhanced, indicating a local increase in PGE2 formation. PGE2 can inhibit the expression of the pro-inflammatory cytokine TNFα. In livers of mPGES1-deficient mice fed a NASH-inducing diet, the induction of TNFα was increased compared to wild type mice, resulting in more pronounced hepatocyte apoptosis. Against the expectations, a deficiency in mPGES1 only lowered the hepatic PGE2 levels but was not sufficient to completely eliminate the diet-induced hepatic PGE2 generation. Consequently, the minor differences in hepatic PGE2 levels between the genotypes did not result in a more severe diet-induced hepatic inflammation. However, in primary macrophages incubated with the Cox2-inhibitor indomethacin the inflammation-mediated increase in PGE2 production was completely abrogated. The follow-up project will focus on theimpact of the Cox2-dependently generated prostanoids like PGE2 in the development of diet-induced NASH. Mice with a specific deletion of Cox2 in macrophages, hepatocytes or both cell types as well as their respective controls will receive standard and NASH-inducing diets and will be characterized regarding the development and progression of NASH. Parallel in vitro experiments with the macrophage populations and hepatocytes from the individual genotypes will help to determine the pro- or anti-inflammatory role of Cox2 on a cellular level. Additionally, the results will be validated in human liver probes. A second part of the project will investigate the heterogeneity of different macrophage populations in mice livers with diet-induced NASH. These distinct macrophages should be isolated and characterized regarding their sensitivity to prostanoids like PGE2.