Interplay between hypoxia and oxidative phosphorylation in herpes stromal keratitis

疱疹性基质角膜炎中缺氧与氧化磷酸化之间的相互作用

基本信息

批准号：
10586030
负责人：
Susmit Suvas
金额：
$ 42.25万
依托单位：
WAYNE STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-03-01 至 2025-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10586030
关键词：
Acetylcysteine Antioxidants Biological Assay Blindness Blood Vessels CD147 antigen CD4 Positive T Lymphocytes Cell Respiration Cell membrane Cell physiology Cells Chemicals Chronic Cicatrix Confocal Microscopy Cornea Corneal Neovascularization Corneal Opacity Development Dimerization Disease Management Disease model Enzymes Epithelial Cells Equilibrium Eye Infections Flow Cytometry GLUT-3 protein Gene Expression Genes Genetic Transcription Glucose Transporter Glycolysis Goals HK2 gene Herpes stromal keratitis Herpesvirus 1 Hypoxia Hypoxia Inducible Factor Immune Impairment Infection Inflammation Inflammatory Knockout Mice Knowledge Lactate Transporter Lesion Link Mediating Metabolic Metabolism Mitochondria Mus Myeloid Cells NADPH Oxidase Neutrophil Infiltration Nuclear Oxidative Phosphorylation Oxygen Pathogenesis Pathologic Pharmaceutical Preparations Phenformin Pimonidazole Prevalence Proteins Publishing Reactive Oxygen Species Recurrence Reporting Research Resolution Respiration Respiratory Burst Role SLC2A1 gene Severities Shapes Signal Pathway Signal Transduction Stains Tamoxifen Testing Tissues Up-Regulation Vascular Endothelial Cell Vascular blood supply angiogenesis chetomin conditional knockout diphenyleneiodonium ebselen efficacy evaluation experimental study functional outcomes immune cell infiltrate immune modulating agents inhibitor innovation neutrophil novel novel strategies novel therapeutic intervention pharmacologic prevent response transcription factor

项目摘要

Herpes stromal keratitis (HSK), a corneal chronic inflammatory condition that develops in response to recurrent corneal herpes simplex virus-1 infection, can cause permanent scarring and vision loss. Chronic inflammation often makes tissue hypoxic due to the lack of the blood vessel, reduced supply of blood, and a high metabolic demand of infiltrating immune cells. Development of hypoxia leads to the stabilization of hypoxia inducible factor (HIF), a transcription factor, which enhances the expression of glycolysis regulating genes and thereby promotes glycolytic metabolism. However, blocking of HIF signaling may promote oxidative phosphorylation to sustain the energy demand of inflammatory cells in inflamed tissue, suggesting an interplay between hypoxia and oxidative phosphorylation to prevent the resolution of an ongoing inflammation. Our preliminary results showed the development of hypoxia in corneas with HSK. Furthermore, the development of hypoxia was linked with the extent of neutrophils in HSV-1 infected corneas. When hypoxia associated signaling pathway PCR array and RT-qPCR were carried out on progressing HSK lesions, we detected an elevated expression of genes encoding key glycolytic enzymes, including PFKFB3 in infected corneas. PFKFB3 is a glycolytic activator, which is reported to enhance hemangiogenesis. We also found an increased level of expression of lactate transporters, MCT4 and MCT1, in corneas with HSK. RT-qPCR results were confirmed with confocal microscopy and flow cytometry. Our results also showed the nuclear localization of HIF-2 in epithelial cells, and HIF-1 in infiltrating neutrophils and CD4 T cells in HSK corneas. Interestingly, blocking of HIF dimerization, while using acriflavine, in HSV-1 infected mice exacerbated the corneal opacity, but decreased the hemangiogenesis. Our preliminary results and supportive published evidence together led us to hypothesize that neutrophils in HSK lesion shape the development of hypoxia resulting in the prevalence of HIF-regulated glycolytic metabolism that promotes HSK pathogenesis, and blockers of HIF, when given in association with inhibitors of mitochondrial respiration or reactive oxygen species (ROS) should reduce the severity of HSK. Three aims are proposed to test this hypothesis. Aim 1 will test the hypothesis that neutrophils in HSK lesion shape the development of hypoxia, and hypoxia enhances the severity of HSK. Aim 2 will test the hypothesis that inhibition of glycolytic activator PFKFB3, and the lactate transporter MCT4 and MCT1 protein reduces the severity of HSK. Aim 3 will test the hypothesis that blockers of HIF in association with an inhibitor of mitochondrial respiration or ROS will reduce the severity of HSK. Given the results from these studies, novel therapeutic approaches for treating HSK lesions may be developed. The knowledge gained from this application will provide innovative information that could be applicable to other ocular infection disease models.

疱疹基质角膜炎（HSK），一种角膜慢性炎症疾病，响应反复发作角膜疱疹单纯疱疹病毒1感染，可能导致永久性疤痕和视力丧失。慢性感染由于缺乏血管，血液供应减少和高代谢，通常会导致组织缺氧浸润免疫细胞的需求。缺氧的发展导致缺氧诱导因子的稳定（HIF），一种转录因子，可增强糖酵解调节基因的表达，从而促进糖酵解代谢。但是，HIF信号传导的阻塞可能促进氧化磷酸化以维持发炎组织中炎症细胞的能量需求，表明缺氧与氧化性之间存在相互作用磷酸化以防止持续的炎症解决。我们的初步结果表明使用HSK在角膜中的缺氧发展。此外，缺氧的发展与 HSV-1感染角膜中嗜中性粒细胞的程度。当缺氧相关的信号通路PCR阵列和 RT-QPCR是在进展的HSK病变上进行的，我们检测到编码的基因表达升高关键的糖酵解酶，包括感染角膜中的PFKFB3。 PFKFB3是一种糖酵解活化剂，报告增强血管生成。我们还发现乳酸转运蛋白，MCT4和 MCT1，与HSK的角膜。通过共聚焦显微镜和流式细胞术证实了RT-QPCR结果。我们的结果还表明，HIF-2在上皮细胞中的核定位和浸润性中性粒细胞和HIF-1的核位置和 HSK角膜中的CD4 T细胞。有趣的是，在HSV-1中使用acriflavine时，HSV-1感染了HIF二聚化的阻塞小鼠加剧了角膜不透明度，但减少了血管生成。我们的初步结果支持性发表的证据共同导致我们假设hsk病变中的中性粒细胞塑造缺氧的发展导致HIF调节的糖酵解代谢的流行，促进HSK 当与线粒体呼吸抑制剂相关时，发病机理和HIF的阻滞剂或活性氧（ROS）应降低HSK的严重程度。提出了三个目标来测试这一点假设。 AIM 1将检验以下假设：HSK病变中的中性粒细胞塑造缺氧的发展，并且缺氧增强了HSK的严重程度。 AIM 2将检验抑制糖酵解活化剂的假设 PFKFB3以及鞋底转运蛋白MCT4和MCT1蛋白降低了HSK的严重程度。 AIM 3将测试假设HIF的阻滞剂与线粒体呼吸或ROS的抑制剂相关的假设会降低 HSK的严重程度。鉴于这些研究的结果，治疗HSK腿的新型治疗方法可以开发。从本应用中获得的知识将提供可能是的创新信息适用于其他眼感染疾病模型。