NOVEL MODULATORS OF ALCOHOL INDUCED METABOLIC AND LIVER INJURY

描述(申请人提供)：原理：慢性乙醇诱导的沉默调节基因1(SIRT1)和过氧化物酶体增殖物受体辅助激活因子1(PGC1)1的下调以及PGC12的上调影响肝脏的脂质氧化和脂肪生成，导致脂肪肝和随后的损伤。结果表明，日粮低-3-脂肪酸(？3FA)，但不高-3-脂肪酸(？3FA)显著降低血清和肝脏的血脂，而大豆蛋白(SP)具有类似的降脂作用。因此，证明这两种新型调节剂可能通过调控相反的PGC11和PGC12信号通路基因来减轻酒精性脂肪肝和酒精性脂肪肝的损伤具有临床意义。初步研究：我们的理论基础是基于我们的出版记录(过去5年中有12篇出版物，1篇综述文章，1本书和21篇摘要)，以及强大的初步数据总结如下：Low？3FA&SP通过以下方式对抗慢性酒精的作用：(A)降低血脂伴随着脂肪肝的减少；(B)减弱肝脏PGC12、SREBP1c及其靶致脂途径基因的mRNAs上调；(C)防止肝脏SIRT1和PGC11及其靶脂肪酸氧化途径基因的mRNAs下调；(D)防止肝脏乙酰PGC11的增加；(E)恢复受损的肝脏脂肪酸氧化和糖异生；及(F)恢复因慢性酒精治疗而降低的肝脏PO4AMPK水平。(G)通过Western blotts，我们证实了慢性酒精引起的PGC11和PGC12信号通路基因的mRNAs水平以及相应的酶蛋白水平的变化。(H)PGC11-KO小鼠肝脏脂质评分显著升高，表明PGC11在酒精性脂肪肝中起关键作用。(I)PGC12似乎发挥了次要作用，因为PGC12-KO小鼠仍然发生酒精性脂肪肝，尽管没有PGC11-KO小鼠那么明显，可能是因为乙醇介导了对PGC11的强烈抑制。具体目标：基于这些理论基础和初步发现，我们将解决以下具体目标：具体目标1.？3FA和SP对慢性乙醇所致的血浆和肝脂、脂蛋白、AST、ALT升高的影响及其与肝组织病理学的关系。具体目的2.？3FA和SP对乙醇诱导的肝脂合成辅活化子、成脂基因和从头脂肪合成速率上调的作用机制。具体目的3.？3FA和SP对乙醇诱导的肝脂代谢辅活化子、脂氧化基因和脂肪酸氧化率下调的作用机制。方法：Pi和他的团队将通过使用啮齿动物作为实验模型来实现这些特定的目标，他们在分子生物学、免疫和组织化学以及生化方法方面具有专业知识。PI还将使用PGC11-KO和PGC12-KOS小鼠模型来最终证明它们在乙醇、低3FA和SP通过这些信号通路在引发/预防酒精性脂肪肝和损伤中的作用的特异性。

DESCRIPTION (provided by applicant): Rationale: Chronic ethanol-induced down regulations of silence regulator gene 1 (SIRT1) and peroxisome proliferator receptor coactivator 1 (PGC1) 1 and up-regulation of PGC12 affect hepatic lipid oxidation and lipogenesis leading to fatty liver and consequent injury. We show that dietary low ?3-fatty acids (?3FA), but not high ?3FA, significantly decreased serum and liver lipids, while Soy Proteins (SP) had similar lipid lowering effects. Therefore, it is clinically relevant to prove that these two novel modulators attenuate alcohol-induced fatty liver and injury potentially by regulating these opposing PGC11 and PGC12 signaling pathway genes. Preliminary Studies: Our rationales are based on our publication records (12 Publications, 1 Review article, 1 book & 21 Abstracts in the past 5 years), and on strong preliminary data that are summarized below: Low ?3FA & SP opposed the actions of chronic alcohol by (a) Reducing serum lipids with concomitant decreased fatty liver; (b) Attenuating the up-regulation of hepatic mRNAs of PGC12, SREBP1c and their target lipogenic pathway genes; (c) Preventing the down-regulation of hepatic mRNAs of SIRT1 and PGC11 and their target fatty acid oxidation pathway genes; (d) Preventing the hepatic increase in acetyl-PGC11, its inactive form; (e) Restoring impaired hepatic fatty acid oxidation and gluconeogenesis; And (f) Restoring the hepatic level of PO4AMPK that was decreased by chronic alcohol treatment. (g) By Western blots, we demonstrated concomitant changes in levels of mRNAs of PGC11 and PGC12 signaling pathway genes and the corresponding enzyme proteins caused by chronic alcohol. (h) That PGC11 plays a critical role in alcohol- induced fatty liver was demonstrated by dramatic increase in hepatic lipid score in PGC11-KO mice. (i) PGC12 seems to play a secondary role because PGC12-KO mice still developed alcoholic fatty liver albeit less pronounced than in PGC11-KO mice, presumably because of ethanol-mediated strong inhibition of PGC11. Specific aims: Based on these rationales and preliminary findings we will address the following specific aims: Specific Aim 1. Effects of ?3FA and SP on Chronic Ethanol-induced Increases in Plasma and Hepatic Lipids and Lipoproteins, AST, ALT and Their Correlation with Liver Histopathology. Specific Aim 2. Mechanisms of Actions of ?3FA and SP on Ethanol-induced Up-regulation of Hepatic Lipid Anabolic Transcriptional Coactivators, Lipogenic Genes and de novo Lipid Synthetic Rates. Specific Aim 3. Mechanisms of Actions of ?3FA & SP on Ethanol-induced Down-regulation of Hepatic Lipid Catabolic Transcriptional Coactivators, Lipid Oxidizing Genes & Fatty Acid Oxidation Rates. Methods of Approach: PI and his team will accomplish these specific aims by using rodents as experimental models with their expertise in molecular biology, immuno- and histo-chemistry and biochemical approaches. PI will also use PGC11-KO and PGC12-KOs mouse models to conclusively prove their specificities in the actions of ethanol, Low 3FA & SP via these signaling pathways in causing/preventing alcoholic fatty liver and injury.