Lcn10 in Sepsis-Induced Vascular Leakage and Heart Failure

Lcn10 在脓毒症引起的血管渗漏和心力衰竭中的作用

• 批准号: 10532242
• 负责人: Guo-Chang Fan
• 金额: $ 66.61万
• 依托单位: UNIVERSITY OF CINCINNATI
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-12-01 至 2025-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10532242
• 关键词: Actin-Binding Protein; Actins; Acute Respiratory Distress Syndrome; Adherens Junction; Animals; Antibiotic Therapy; Antisepsis; Blood Vessels; Blood capillaries; Cardiac; Cardiac Edema; Cardiac Myocytes; Cardiovascular system; Cause of Death; Cell physiology; Cells; Cessation of life; Coronary; Critical Care; Critical Illness; Cytoskeleton; Data; Development; Edema; Endothelial Cells; Endothelium; Endotoxins; Extravasation; Fibroblasts; Future; Heart; Heart Injuries; Heart failure; Homologous Gene; Human; In Vitro; Inflammatory; Injections; Intensive Care Units; Intercellular Junctions; Knock-out; Knockout Mice; Knowledge; Leucocytic infiltrate; Link; Liquid substance; Lung; Measures; Mediating; Mediator; Microfilaments; Mission; Monitor; Mus; Myocardial; Myocardial dysfunction; Operative Surgical Procedures; Organ failure; Partner in relationship; Pathway interactions; Patients; Permeability; Phosphoric Monoester Hydrolases; Physiology; Play; Protein Dephosphorylation; Protein Secretion; Proteins; Public Health; Pulmonary Edema; RNA analysis; Recombinants; Regulation; Research; Resistance; Resuscitation; Role; Sepsis; Septic Shock; Small Interfering RNA; Survival Rate; Testing; Therapeutic; Tight Junctions; Time; Tissues; Transgenic Mice; Translating; United States National Institutes of Health; Up-Regulation; Vascular Permeabilities; Wild Type Mouse; Work; animal mortality; autocrine; cardiac depression; cecal ligation puncture; cell type; cofilin; comparison control; disability; heart function; improved; in vivo; knock-down; member; monolayer; mortality; mouse model; novel; novel therapeutics; overexpression; polymicrobial sepsis; potential biomarker; pre-clinical; septic; septic patients; survival outcome; tool; transcriptome sequencing; translational study

Vascular hyperpermeability is well-recognized to be responsible for sepsis-triggered organ failure and patient mortality. Despite decades of intensive study, there is no specific treatment available for targeting such vascular leakage thus far. This is due in part to the incomplete knowledge of the mediators and mechanisms underlying sepsis-elicited disruption of the endothelial barrier integrity. At present, most prior work has focused on pulmonary vascular leakage that results in lung edema and acute respiratory distress syndrome. Few studies have investigated coronary vascular leakage, which is a major cause of heart failure and death in human patients with septic shock. We recently discovered that expression of lipocalin 10 (Lcn10), a poorly characterized member of the lipocalin superfamily, was significantly downregulated in the hearts of both endotoxin LPS- and cecal ligation- puncture (CLP)-treated mice, compared to their controls. Interestingly, further cell-type specific analysis showed that such reduction of Lcn10 did not occur in either cardiomyocytes or fibroblasts but only in cardiac endothelial cells (ECs). These compelling data implicate a potential role of Lcn10 in sepsis-induced cardiovascular leakage. Indeed, using a global knockout mouse model, we observed that deficiency of Lcn10 significantly augmented LPS- induced vascular leakage, leading to greater cardiac depression and higher mortality, compared to LPS-treated wild-type control mice. By contrast, in vitro forced overexpression of Lcn10 in ECs showed greater resistance to LPS-induced monolayer leak relative to control cells. An initial mechanistic analysis by RNA-sequencing and RT- qPCR showed that both endogenous and exogenous elevation of Lcn10 in ECs caused significant upregulation of slingshot homolog 1 (Ssh1). Ssh1 is a phosphatase known to dephosphorylate and thus activate Cofilin, a key actin-binding protein that plays an essential role in controlling actin filament dynamics. Most importantly, knockdown of Ssh1 in ECs offsets the Lcn10-induced reduction of monolayer leakage upon LPS exposure. Based on these preliminary data, we hypothesize that Lcn10 is critical for protecting against sepsis-induced vascular leak via the activation of the Ssh1-Cofilin pathway. This hypothesis will be tested by pursuing three specific aims: 1) Define the precise role of Lcn10 in vascular permeability during polymicrobial sepsis, using a global knockout and an EC-specific Lcn10-transgenic mouse model; 2) Identify the mechanism by which Lcn10- elicited reduction of cardiovascular leakage is dependent on Ssh1-mediated actin dynamics, using a cross mouse model by mating EC-specific Lcn10-transgenic mice with Ssh1-KO mice; and 3) Investigate the therapeutic potential of recombinant Lcn10 protein in treating sepsis. The proposed studies are expected to identify Lcn10 as a potent and novel regulator of vascular permeability and a new protector against sepsis-induced heart failure. If completed, the findings from this proposal are likely to provide new therapeutic options for reducing vascular leakage during sepsis, with the hope of improving the survival of septic patients.

血管高通透性被公认为是脓毒症引发的器官衰竭和患者 死亡率。尽管进行了数十年的密集研究，但目前还没有针对这种血管的特定治疗方法。 到目前为止的泄漏情况。这在一定程度上是由于对调解人和潜在机制的不完全了解。 脓毒症-引起内皮屏障完整性的破坏。目前，大部分前期工作都集中在 导致肺水肿和急性呼吸窘迫综合征的肺血管渗漏。很少有研究发现 调查冠状动脉血管渗漏，这是人类心力衰竭和死亡的主要原因 感染性休克。我们最近发现Lipocalin 10(Lcn10)的表达，这是一个特征不佳的成员 在内毒素内毒素和盲肠结扎的心脏中，Lipocalin超家族的表达都显著下调。 穿刺法(CLP)治疗的小鼠与其对照组进行比较。有趣的是，进一步的细胞类型特异性分析显示 Lcn10的这种减少并不发生在心肌细胞或成纤维细胞中，而只发生在心脏内皮细胞中 细胞(ECs)。这些令人信服的数据暗示了Lcn10在脓毒症引起的心血管渗漏中的潜在作用。 事实上，使用全局基因敲除小鼠模型，我们观察到Lcn10基因的缺失显著增加了内毒素- 与内毒素治疗相比，诱导血管渗漏导致更大的心脏抑制和更高的死亡率 野生型对照小鼠。相比之下，在体外强制过表达Lcn10的内皮细胞显示出更强的抵抗力 脂多糖诱导的相对于对照细胞的单层渗漏。RNA测序和RT-PCR的初步机理分析 定量聚合酶链式反应显示内皮细胞Lcn10的内源性和外源性上调均可引起内皮细胞Lcn10的显著上调 弹弓同源基因1(Ssh1)。Ssh1是一种已知的磷酸酶，可以去磷酸化，从而激活cofilin，这是一个关键 肌动蛋白结合蛋白，在控制肌动蛋白细丝动力学中发挥重要作用。最重要的是， 敲除内皮细胞中的Ssh1可抵消Lcn10诱导的脂多糖暴露时单层渗漏的减少。基座 根据这些初步数据，我们假设Lcn10在预防败血症引起的血管渗漏中起关键作用。 通过激活Ssh1-cofilin途径。这一假设将通过追求三个具体目标来检验：1) 确定Lcn10在多菌败血症时血管通透性中的确切作用 2)鉴定Lcn10基因敲除和EC特异性Lcn10转基因小鼠模型的作用机制。 心血管渗漏的减少依赖于Ssh1介导的肌动蛋白动力学，使用交叉 通过将EC特异性Lcn10转基因小鼠与Ssh1-KO小鼠配对建立小鼠模型；以及3)研究 重组Lcn10蛋白治疗脓毒症的潜力拟议的研究预计将 Lcn10是一种有效的新型血管通透性调节剂和新的脓毒症保护剂 心力衰竭。如果完成，这项提案的发现可能会为减少 脓毒症时血管渗漏，以期提高脓毒症患者的存活率。