ANTI INFLAMMATORY AND ANTIFIBROTIC ACTIONS OF THYMOSIN BETA 4 IN ALD

胸腺肽 4 在 ALD 中的抗炎和抗纤维化作用

• 批准号: 8609964
• 负责人: RAJ M LAKSHMAN
• 金额: $ 14.96万
• 依托单位: GEORGE WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-06-01 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8609964
• 关键词: ADD-1 protein; Acetaldehyde; Aftercare; Alcoholic Liver Diseases; Alcohols; Aldehydes; Anti-Inflammatory Agents; Anti-inflammatory; Appearance; Applications Grants; Architecture; Biochemical; Biochemistry; Chemistry; Chronic; Clinical Treatment; Clinical Trials; Collagen; Collagen Gene; Dissociation; Down-Regulation; Endotoxins; Epigenetic Process; Ethanol; Extracellular Matrix Proteins; Eye; Fetoprotein; Fibronectins; Fibrosis; Future; Gene Proteins; Generations; Genes; Goals; Health; Heart; Hepatic; Hepatic Fibrogenesis; Hepatic Stellate Cell; Hepatic Tissue; Hepatocyte; In Vitro; Inflammation; Inflammatory; Injury; Lead; Lipids; Liver; Liver diseases; Mediating; Metabolic; Methyl-CpG-Binding Protein 2; Modeling; Molecular; Molecular Biology; Mus; Myofibroblast; Natural regeneration; Oxidative Stress; PPAR gamma; Peptides; Pericytes; Peroxisome Proliferator-Activated Receptors; Phenotype; Phosphorylation; Platelet-Derived Growth Factor; Platelet-Derived Growth Factor beta Receptor; Process; Property; Proteins; Reactive Oxygen Species; Reporting; Role; Serum; Serum Markers; Signal Transduction; Site; Skin; Smooth Muscle Actin Staining Method; Techniques; Testing; Therapeutic Agents; Thymosin; Time; Tissues; Tumor Necrosis Factor-alpha; Up-Regulation; Vitamin A; adduct; base; cytokine; fibrogenesis; in vitro Model; in vivo; innovation; liver inflammation; liver injury; mouse model; novel; novel therapeutics; overexpression; preclinical study; prevent; receptor; stellate cell; thymosin beta(4); transdifferentiation

DESCRIPTION (provided by applicant): Based on the well accepted two-hit model for ALD, Chronic Ethanol (EtOH)/LPS generated endotoxins activate NFκB that up regulates TNFα, and IL1β the potent proinflammatory cytokines causing hepatocyte injury. Further, EtOH induced Cyp2E1 and ADH lead to metabolic generation of acetaldehyde and increased reactive oxygen species (ROS), which activate quiescent HSC to myofibroblasts resulting in up regulation of fibrogenic genes, platelet derived growth factor β-receptor (PDGFβr), α-smooth muscle actin (αSMA) and extracellular matrix proteins (ECM), collagen I, III, and fibronectin and epigenetic repressor gene, methyl-CpG binding protein 2 (MeCP2). In contrast, adipogenic genes, peroxisome proliferator-activated receptor γ (PPARγ), and sterol regulatory element-binding protein 1c (SREBP1c) are suppressed resulting in the loss of their vitamin A stores and their transdifferentiation from quiescent lipid storing phenotype to active myofibroblastic phenotype. Significantly, thymosin β4 (Tβ4), a bioactive peptide, is reported to prevent inflammation and fibrosis in many extra-hepatic tissues. Based on these, PI hypothesizes that Tβ4's anti-inflammatory and anti-fibrogenic actions against EtOH/LPS liver injury are mediated by (i) inhibiting the activation of NFκB by blocking the phosphorylation and dissociation of IκB and thereby prevent the up regulation of TNFα, and IL1β the potent proinflammatory cytokines and consequent liver injury, and (ii) suppressing the up regulated MeCP2, that coordinately reverses (a) the down regulated adipogenic genes and (b) up regulated fibrogenic genes and thereby prevent the trans-differentiation of HSC from lipid-storing pericytes to myofibroblasts. We also hypothesize that Tβ4 elicits its above actions by overexpressing miR132 that suppresses MeCP2 overexpression caused by EtOH/LPS. PI has the following encouraging preliminary results in the EtOH/LPS mouse model to reassure the feasibilities of his novel exploratory approaches: Tβ4 protects against EtOH/LPS induced 1. Up-regulation of NFκB signaling cascade, pIκB, TNFα, IL1β and consequent serum markers for liver injury; 2. up regulation of hepatic MeCP2, PDGFβr, αSMA, Col1α1 & proteins; and 3. down-regulation of adipogenic gene, PPARγ. As a result, we propose that Tβ4 blocks the (i) activation of NFκB, TNFα and inflammatory cascade and (ii) transdifferentiation of quiescent HSC to fibrogenic HSC; yet, Tβ4 maintains hepatocyte regeneration. Thus, the major goals of our innovative proposal are to accomplish the following specific aims: Specific Aim 1. What are the possible mechanism/s of action/s of Tβ4 "Pre- and Post-treatment to protect/alleviate" EtOH/LPS-mediated up regulation of NFκB signaling cascade, TNFα & IL1β, and consequent serum and liver markers for liver injury? Specific Aim 2: What are the possible mechanism/s of action/s of Tβ4 "Pre- and Post-treatment to protect/alleviate" against EtOH/LPS-mediated (a) up regulation of hepatic fibrogenic genes and their products? and (b) down regulation of adipogenic genes and their products? Are there corresponding morphological changes in liver cell architecture as well as in HSC phenotype? PI plans to approach using established EtOH/LPS two-hit mouse in vivo & in vitro model utilizing biochemistry, molecular biology, immuno- & histo-chemistry techniques. PI has a strong molecular biology and biochemical group that has the expertise in all aspects of this proposal. This may lead to novel therapy for ALD.

描述（由申请方提供）：基于广泛接受的ALD两次打击模型，慢性乙醇（EtOH）/LPS产生的内毒素激活NFκB B，从而上调TNFα和IL 1 β（导致肝细胞损伤的强效促炎细胞因子）。此外，EtOH诱导的Cyp 2 E1和ADH导致乙醛的代谢产生和活性氧（ROS）的增加，其将静止的HSC活化为肌成纤维细胞，导致纤维化基因、血小板衍生生长因子β受体（PDGFβr）、α-平滑肌肌动蛋白（αSMA）和细胞外基质蛋白（ECM）、胶原蛋白I、III、纤连蛋白和表观遗传阻遏基因的上调，甲基-CpG结合蛋白2（MeCP 2）。相反，脂肪形成基因、过氧化物酶体增殖物激活受体γ（PPARγ）和固醇调节元件结合蛋白1c（SREBP 1c）受到抑制，导致其维生素A储存丧失，并从静止的脂质储存表型转分化为活跃的成肌纤维细胞表型。胸腺素β4（thymosin β4，Tβ4）是一种具有生物活性的多肽，被认为可以预防肝外组织的炎症和纤维化。基于这些，PI推测Tβ4对EtOH/LPS肝损伤的抗炎和抗纤维化作用是通过（i）通过阻断IκB的磷酸化和解离来抑制NFκB B的活化，从而防止TNFα和IL 1 β的上调以及随后的肝损伤，和（ii）抑制上调的MeCP 2，其协同逆转（a）下调的脂肪形成基因和（B）上调的纤维形成基因，从而阻止HSC从储存脂质的周细胞转分化为肌成纤维细胞。我们还推测Tβ4通过过表达miR 132抑制EtOH/LPS引起的MeCP 2过表达而发挥上述作用。PI在EtOH/LPS小鼠模型中获得了以下令人鼓舞的初步结果，以保证他的新探索方法的特性：Tβ4保护EtOH/LPS诱导的1。NFκB信号通路、pIκB、TNFα、IL 1 β及相应的肝损伤血清标志物的上调; 2.上调肝脏MeCP 2、PDGFβr、αSMA、Col 1 α 1蛋白;成脂基因PPARγ表达下调。因此，我们认为Tβ4阻断了（i）NFκB、TNFα和炎性级联反应的激活，以及（ii）静止HSC向纤维化HSC的转分化;然而，Tβ4维持了肝细胞再生。因此，我们的创新提案的主要目标是实现以下具体目标：具体目标1。Tβ4“保护/减轻”EtOH/LPS介导的NFκB信号级联、TNFα和IL 1 β上调的“治疗前和治疗后”可能的作用机制是什么，以及随后的肝损伤血清和肝脏标志物？具体目标二：Tβ4“治疗前和治疗后保护/缓解”EtOH/LPS介导的（a）肝纤维化基因及其产物上调的可能作用机制是什么？和（B）成脂基因及其产物的下调？肝细胞结构和HSC表型是否有相应的形态学变化？PI计划利用生物化学、分子生物学、免疫和组织化学技术，使用已建立的EtOH/LPS两次打击小鼠体内和体外模型进行研究。PI有一个强大的分子生物学和生物化学小组，在这个提案的各个方面都有专业知识。这可能导致ALD的新疗法。