CEACAM1 Alternative Splicing in Liver Ischemia-Reperfusion Injury

CEACAM1 选择性剪接在肝脏缺血再灌注损伤中的作用

• 批准号: 10622462
• 负责人: Jerzy W Kupiec-Weglinski
• 金额: $ 54.09万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10622462
• 关键词: Acceleration; Acute; Address; Advocate; Alternative Splicing; Anti-Inflammatory Agents; Autophagocytosis; Biological Markers; Biometry; Biopsy; Brain Death; Cardiac Death; Cell Culture System; Cell Death; Cells; Clinical; Complement; Complement component C1; Cryopreservation; Cytoplasm; Cytoprotection; Data; Exclusion; Exons; Genes; Hepatic; Hepatic Tissue; Hepatocyte; Human; Immune; Impairment; Infiltration; Inflammation; Injury; Intervention; Ischemia; Life; Liver; Lymphocyte; MAP Kinase Gene; Macrophage; Mediating; Messenger RNA; Mucosal Immunity; Mus; Myelogenous; Natural Immunity; Natural regeneration; Operative Surgical Procedures; Organ; Organ Donor; Organ Preservation; Organ Transplantation; Outcome; Pathway interactions; Patients; Phase; Phenotype; Protein Isoforms; RNA Splicing; Recovery; Regulation; Rejuvenation; Reperfusion Injury; Reporting; Resistance; Resolution; Sentinel; Signal Transduction; Sterility; Stress; Tissues; Transgenic Mice; Transplantation; Transplantation Surgery; Variant; Waiting Lists; Warm Ischemia; carcinoembryonic antigen-related cell adhesion molecules; clinically relevant; end stage liver disease; experimental study; functional improvement; immunoregulation; improved; insight; liver function; liver inflammation; liver ischemia; liver transplantation; mortality; mouse model; neutrophil; novel; novel therapeutics; p38 Mitogen Activated Protein Kinase; preservation; prevent; programs; regenerative; repaired; screening; standard of care; success; synergism; transplant model

PROJECT 1 – SUMMARY/ABSTRACT Project 1 competitive renewal addresses the unmet clinical and scientific needs to enhance donor liver supply through organ “rejuvenation.” We reported that hepatic CEACAM1 (encoded by CC1 gene; CD66a) dictates donor liver quality, and prevents early OLT injury in mice and humans. CC1 mRNA undergoes alternative splicing (AS) to generate splice variants, characterized by the inclusion (CC1-L; long) or exclusion (CC1-S; short) of exon 7. Hypothesis: Hepatocyte CC1-S functions as a cytoprotective sentinel, while CC1-L in OLT-infiltrating recipient- derived neutrophils, regulates liver IR-inflammation and fine-tunes its resolution. Aim 1: Delineate mechanisms by which hepatic CC1-S isoform promotes cellular protection in IR- stressed mouse OLT. Hypothesis: Antisense oligomer morpholinos (MOs)-enforced AS of hepatic CC1 generates CC1-S isoform in the donor mouse liver, which under the control of HIF-1α, stimulates cytoprotection by blocking p-p38 (under cold IR-stress) while promoting GPX4 and targeting ferroptosis (under warm IR-stress). Here, we will ascertain whether 1.1: CC1-AS accelerates otherwise impaired hepatic recovery in the acute and the resolution phase of IR-inflammation. 1.2: CC1-S controls IRI-OLT by regulating hepatic cell death pathways. 1.3: CC1-S-mediated cytoprotection is controlled by HIF-1α signaling under warm vs. cold hepatic IR-stress. Aim 2: Define mechanisms by which neutrophil CC1-L isoform exerts anti-inflammatory and cytoprotective functions in IR-stressed mouse OLT. Hypothesis: Recipient CC1-L neutrophils counteract acute IRI-OLT, and stimulate inflammation resolution by orchestrating N1/N2 polarization, with resultant liver homeostatic/regenerative remodeling. We will study whether 2.1: Recipient CC1-L deficient immune cells exacerbate hepatic IRI-OLT. 2.2: CC1-L polarizes N2 neutrophils to promote an anti-inflammatory phenotype/ improve OLT outcomes. 2.3: CC1-L proficient neutrophils polarize M2 macrophages/promote hepatic autophagy. Aim 3: Elucidate mechanisms by which hepatic CC1-S isoform rejuvenates discarded human livers subjected to normothermic machine preservation (NMP). Hypothesis: NMP, supplemented with HIF-1α – CC1- S axis modifiers, improves the function of discarded human livers by mitigating ferroptosis while promoting autophagy. We will incorporate the emerging NMP strategy with adjunctive CC1-intervention to assess whether 3.1: MOs-conditioned humanized CC1 transgenic mice (huCC1-Tg) mimic the effects of CC1-AS upon liver IR- stress in normal mice but be translatable to the human liver. 3.2: CC1-S isoform synergizes with HIF-1α to improve liver function. 3.3: CC1-S synergizes with HIF-1α to enhance cytoprotection in human livers. Integration with PPG: Project 1 complements studies assessing homeostatic mechanisms in the resolution of IRI-OLT in Project 2. Aim 1-2 in Project 1 will be validated by the screening of human OLT biopsies to accelerate assessments of clinical innate immune phenotypes in Project 3. Project 1 is dependent on Core A (Administrative), Core B (Mouse and Human OLT Surgery), and Core C (Computational and Biostatistics).

项目1 -总结/摘要 项目1竞争性更新解决了未满足的临床和科学需求，以提高供体肝脏供应 通过器官“再生”。我们报道了肝脏CEACAM 1（由CC 1基因编码; CD 66 a）决定了 供体肝脏质量，并防止小鼠和人类的早期奥尔特损伤。CC 1 mRNA经历选择性剪接 (AS)以产生剪接变体，其特征在于包含（CC 1-L;长）或排除（CC 1-S;短）外显子 7.假设：肝细胞CC 1-S起细胞保护性哨兵的作用，而在OLT浸润的受体中，CC 1-L起细胞保护性哨兵的作用。 衍生的中性粒细胞，调节肝脏IR炎症并微调其分辨率。 目的1：阐明肝脏CC 1-S亚型促进IR-1细胞保护作用的机制。 应激小鼠奥尔特。假设：反义寡聚体吗啉代（MO）-强制肝CC 1的AS 在供体小鼠肝脏中产生CC 1-S亚型，其在HIF-1α的控制下，刺激细胞保护作用 通过阻断p-p38（在冷IR应激下）同时促进GPX 4和靶向铁凋亡（在热IR应激下）。 在此，我们将确定1.1：CC 1-AS是否加速急性和慢性肝损伤中受损的肝脏恢复， IR炎症的消退阶段。1.2：CC 1-S通过调节肝细胞死亡途径控制IRI-OLT。 1.3：CC 1-S介导的细胞保护作用在热与冷肝IR应激下由HIF-1α信号传导控制。 目的2：确定中性粒细胞CC 1-L亚型发挥抗炎作用的机制， IR应激小鼠奥尔特的细胞保护功能。假设：CDCC 1-L中性粒细胞抵消急性 IRI-OLT，并通过协调N1/N2极化刺激炎症消退， 稳态/再生重塑。我们将研究是否2.1：CC 1-L缺陷型免疫细胞 加重肝IRI-OLT。2.2：CC 1-L使N2中性粒细胞极化以促进抗炎表型/ 改善奥尔特结果。2.3：CC 1-L熟练的嗜中性粒细胞/巨噬细胞/促进肝自噬。 目的3：阐明肝CC 1-S亚型使废弃人肝再生的机制 进行常温机器保存（NMP）。假设：NMP，补充有HIF-1α -CC 1- S轴修饰剂，通过减轻铁凋亡同时促进 自噬我们将把新兴的NMP策略与连续性CC 1干预结合起来，以评估 3.1：MO条件化的人源化CC 1转基因小鼠（huCC 1-Tg）模拟CC 1-AS对肝脏IR的影响。 但是可以转移到人类的肝脏。3.2：CC 1-S同种型与HIF-1α协同作用， 改善肝功能。3.3：CC 1-S与HIF-1α协同增强人肝脏的细胞保护作用。 与PPG的整合：项目1补充了评估体内稳态机制的研究， 解决方案2中的IRI-OLT。项目1中的目标1-2将通过人类奥尔特活检的筛选来验证 加速项目3中临床先天免疫表型的评估。项目1依赖于核心A （管理）、核心B（小鼠和人类奥尔特手术）和核心C（计算和生物统计学）。