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 – 摘要/摘要 项目 1 竞争性更新解决了未满足的临床和科学需求，以增强供体肝脏供应 通过器官“返老还童”。我们报道了肝脏 CEACAM1（由 CC1 基因编码；CD66a）决定 供体肝脏质量，并防止小鼠和人类的早期 OLT 损伤。 CC1 mRNA 进行选择性剪接 (AS) 生成剪接变体，其特征是包含（CC1-L；长）或排除（CC1-S；短）外显子 7. 假设：肝细胞 CC1-S 充当细胞保护哨兵，而 OLT 浸润受体中的 CC1-L 衍生的中性粒细胞，调节肝脏红外炎症并微调其分辨率。 目标 1：描述肝 CC1-S 亚型促进 IR- 细胞保护的机制 强调鼠标 OLT。假设：反义寡聚体吗啉代 (MO) 增强肝 CC1 的 AS 在供体小鼠肝脏中产生 CC1-S 亚型，在 HIF-1α 的控制下刺激细胞保护 通过阻断 p-p38（在冷红外应激下），同时促进 GPX4 并靶向铁死亡（在热红外应激下）。 在这里，我们将确定 1.1：CC1-AS 是否会加速急性和急性期受损的肝功能恢复。 IR 炎症的消退阶段。 1.2：CC1-S通过调节肝细胞死亡途径来控制IRI-OLT。 1.3：CC1-S 介导的细胞保护作用在热肝脏 IR 应激与冷肝脏 IR 应激下受 HIF-1α 信号传导控制。 目标 2：定义中性粒细胞 CC1-L 亚型发挥抗炎和抗炎作用的机制 IR 应激小鼠 OLT 中的细胞保护功能。假设：受体 CC1-L 中性粒细胞抵抗急性 IRI-OLT，通过协调 N1/N2 极化刺激炎症消退，从而改善肝脏 稳态/再生重塑。我们将研究2.1：受体CC1-L免疫细胞是否缺陷 加剧肝脏IRI-OLT。 2.2：CC1-L 极化 N2 中性粒细胞以促进抗炎表型/ 改善 OLT 结果。 2.3：CC1-L 熟练的中性粒细胞极化 M2 巨噬细胞/促进肝自噬。 目标 3：阐明肝脏 CC1-S 亚型使废弃人类肝脏恢复活力的机制 进行常温机器保存（NMP）。假设：NMP，辅以 HIF-1α – CC1- S轴修饰剂，通过减轻铁死亡同时促进废弃人类肝脏的功能 自噬。我们将把新兴的 NMP 策略与辅助 CC1 干预结合起来，以评估是否 3.1: MOs 条件下的人源化 CC1 转基因小鼠 (huCC1-Tg) 模拟 CC1-AS 对肝脏 IR 的影响 正常小鼠的压力，但可以转化为人类肝脏。 3.2：CC1-S亚型与HIF-1α协同作用 改善肝功能。 3.3：CC1-S 与 HIF-1α 协同作用，增强人肝脏的细胞保护作用。 与 PPG 整合：项目 1 补充了评估体内平衡机制的研究 项目 2 中 IRI-OLT 的分辨率。项目 1 中的目标 1-2 将通过人类 OLT 活检筛查进行验证 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 （行政）、核心 B（小鼠和人类 OLT 手术）和核心 C（计算和生物统计学）。