The Unfolded Protein Response in Sterol Cytoxicity

描述(由申请人提供)： 细胞对高水平甾醇的反应对疾病过程和正常生理有重要意义。在一种情况下，动脉粥样硬化巨噬细胞(MFS)积累的游离胆固醇(FC)导致MF死亡，这可能会促进斑块不稳定和急性临床事件，如心肌梗死。在另一种情况下，肠道上皮细胞(IECS)暴露在高水平的潜在有毒的饮食和胆汁胆固醇以及饮食中的植物类固醇中，因此机制是必要的，以防止肠道功能障碍。PI最近发现，内质网(ER)中一个复杂的信号转导途径--未折叠蛋白反应(UPR)--在培养的MRS对FC的反应中起着关键作用。UPR首先触发一系列适应性反应，但如果适应性反应失败，则会诱导细胞凋亡。因此，UPR最初促进FC负载的MFS存活，但在长期FC负载后促进细胞凋亡。由于UPR的一个独特分支定位于胃肠道，并且由于UPR诱导的保护IECs免受类固醇细胞毒性的关键酶，UPR也可能在保护IECs免受高水平类固醇的影响中发挥重要作用。在此背景下，本提案的总体目标是研究UPR在与动脉粥样瘤MFS和IECS积聚过量甾醇相关的适应性和病理生理过程中的作用和机制。这一目标将通过：(A)确定两个关键的UPR基因--一个参与适应性反应，另一个参与细胞凋亡--的零突变是否影响apoE/-小鼠动脉粥样硬化模型中的皮损性MF死亡和斑块不稳定；(B)确定暴露于过量饮食和植物类固醇的小鼠的IECS是否诱导UPR；UPR是否诱导已知有助于保护IECs免受类固醇伤害的蛋白质；以及UPR的破坏是否导致肠道功能障碍；以及(C)通过检验以下假设来研究FC是如何诱导UPR的：Fc通过抑制肌浆/内质网钙ATPase或通过激活ER钙通道来消耗内质网钙，内质网钙是UPR的已知和有效的诱导剂。MF的研究将增加我们对斑块如何变得脆弱的理解，并将提出涉及急性血管事件的新的治疗靶点，而肠道研究将揭示肠道类固醇生理学的一个新的重要方面。

DESCRIPTION (provided by applicant): The responses of cells to high levels of sterols has important implications for disease processes and normal physiology. In one setting, the accumulation of free cholesterol (FC) by atheroma macrophages (Mfs) leads to Mf death, which likely promotes plaque instability and acute clinical events, like myocardial infarction. In another setting, intestinal epithelial cells (IECs) are exposed to high levels of potentially toxic dietary and biliary cholesterol and dietary plant sterols, and so mechanisms are necessary to prevent intestinal dysfunction. The PI has recently discovered that a complex signal transduction pathway in the endoplasmic reticulum (ER)--the Unfolded Protein Response (UPR)--plays a critical role in the response of cultured Mrs to FC. The UPR first triggers a set of adaptive response, but then induces apoptosis if the adaptive responses fail. As such, the UPR initially promotes survival in FC-Ioaded Mfs, but then promotes apoptosis after prolonged FC loading. Because a unique branch of the UPR is localized to the gastrointestinal tract, and because a critical enzyme that protects IECs from sterol cytotoxicity is induced by the UPR, the UPR may also play an important role in the protection of IECs from high levels of sterols. In this context, the overall objective of this proposal is to investigate the roles and mechanisms of the UPR in adaptive and pathophysiological processes associated with the accumulation of excess sterols by atheroma Mfs and IECs. This objective will be met by: (a) determining if null mutations in two key UPR genes--one involved in the adaptive response and the other in apoptosis--affect lesional Mf death and plaque instability in the apoE-/- mouse model of atherosclerosis; (b) determining if the UPR is induced in IECs of mice exposed to excess dietary and plant sterols; if proteins known to help protect IECs from sterols are induced by the UPR; and if disruption of the UPR leads to intestinal dysfunction; and (c) investigating how FC induces the UPR by testing the hypothesis that FC enrichment of the ER membrane depletes ER calcium, which is a well-known and potent inducer of the UPR, by inhibiting the sarco/endoplasmic reticulum calcium ATPase or by activating ER calcium channels. The Mf studies will add to our understanding of how plaques become vulnerable and will suggest novel therapeutic targets involved in acute vascular events, and the intestinal studies will reveal a new and important aspect of intestinal-sterol physiology.