Mechanisms of Liver Inflammation

描述(由申请人提供)：我的长期职业目标是明确非酒精性脂肪性肝炎的肝脏炎症机制。目前的建议集中在内质网(ER)应激激活的未折叠蛋白反应(UPR)的三个已知分支中的两个在NASH的巨噬细胞生物学调控中的二分作用。在初步实验中，我们观察到巨噬细胞在棕榈酸(PA)处理后被激活；我们将这种现象称为脂激活。脂激活的巨噬细胞经历内质网应激，也会发生细胞死亡，称为脂细胞凋亡。我们观察到在NASH小鼠模型中巨噬细胞聚集，同时PA水平升高。我们的初步观察导致了一个中心假设，即UPR调节肝巨噬细胞中的脂激活或脂凋亡，从而分别引发或减轻脂肪性肝炎的炎症。因此，这项建议的目标是了解：i)UPR的肌醇需要蛋白-1α(IRE1α)/X-box结合蛋白(XBP-1)分支如何通过靶向细胞因子单核细胞趋化蛋白-1(MCP-1)和白介素1β(IL-1β)来促进巨噬细胞的激活和细胞因子的分泌，从而增加肝脏的促炎环境；以及ii)UPR的CHOP分支如何通过减少抗凋亡蛋白Mcl-1和增加促凋亡蛋白Bim来促进巨噬细胞的凋亡，从而增加肝脏的抗炎环境。拟议的实验将分别采用脂毒性和NASH的体外和体内补充模型；以及化学、药理学、分子和遗传学方法来解决特定的目的，以检验假设：i)UPR通过IRE1？/XBP-1信号决定巨噬细胞的募集和激活，ii)通过CHOP的内质网应激促进巨噬细胞的凋亡，以及iii)UPR调节NASH的肝脏炎症。为了解决这些假设，申请者已经擅长巨噬细胞分离、细胞因子分泌的分析、转录调控以及有条件删除IRE1或CHOP的啮齿动物模型。通过K08指导临床科学家研究职业发展奖的资助，申请者将继续接受巨噬细胞生物学和天然免疫学方面的额外培训，以发展这些肝脏炎症关键调节因子的专业知识。申请者建立了一个网络，其中Gregory J.Gores博士是她的主要导师，Peter J.Wettstein博士是免疫学合作者，Randal J.Kaufman博士是普遍定期审议合作者。我们的结果将从机制上深入了解单个UPR组分对巨噬细胞脂激活和脂凋亡的调节，从而确定可以作为治疗干预目标的潜在分子。

DESCRIPTION (provided by applicant): My long term career objective is to define the mechanisms of liver inflammation in nonalcoholic steatohepatitis. The current proposal focuses on the dichotomous role of two of the three known branches of the endoplasmic reticulum (ER) stress-activated unfolded protein response (UPR) in the regulation of macrophage biology in NASH. In preliminary experiments we have observed that macrophages are activated upon treatment with palmitic acid (PA); we have termed this phenomenon lipoactivation. Lipoactivated macrophages undergo ER stress, and also cell death, termed lipoapoptosis. We have observed macrophage accumulation in a mouse model of NASH along with elevated PA levels. Our preliminary observations have led to the central hypothesis that the UPR regulates either lipoactivation or lipoapoptosis in hepatic macrophages thereby instigating or mitigating inflammation, respectively, in steatohepatitis. Therefore, the goals of this proposal are to understand: i) how the Inositol Requiring Protein-1 alpha (IRE1¿)/ X-box binding protein (XBP-1) branch of the UPR increases the proinflammatory milieu in the liver by enhancing macrophage activation and cytokine secretion via targeting the cytokines monocyte chemotactic protein-1 (MCP-1) and interleukin-1beta (IL-1¿), and ii) how the CHOP branch of the UPR increases the antiinflammatory milieu in the liver by enhancing macrophage apoptosis by decreasing the antiapoptotic protein Mcl-1 and increasing the proapoptotic protein Bim. The proposed experiments will employ complementary in vitro and in vivo models of lipotoxicity and NASH, respectively; and chemical, pharmacological, molecular and genetic approaches to address the specific aims to test the hypotheses that: i) The UPR determines macrophage recruitment and activation via IRE1¿/XBP-1 signalling, ii) ER stress via CHOP promotes macrophage apoptosis, and iii) The UPR regulates hepatic inflammation in NASH. To address these hypotheses the applicant has become adept at macrophage isolation, assays of cytokine secretion, transcriptional regulation, and in vivo rodent models of conditional deletion of IRE1¿ or CHOP. With funding through this K08 Mentored Clinical Scientist Research Career Development Award the applicant will pursue additional training in macrophage biology and Innate Immunology to develop expertise in these key regulators of hepatic inflammation. The applicant has established a network consisting of Dr. Gregory J. Gores as her primary mentor, Dr. Peter J. Wettstein as the Immunology collaborator, and Dr. Randal J. Kaufman as the UPR collaborator. Our results will yield mechanistic insights into regulation of macrophage lipoactivation and lipoapoptosis by individual UPR components, thus identifying potential molecules that can be targeted by therapeutic interventions.