CXCR4: A potential therapeutic target in HSK

CXCR4：HSK 的潜在治疗靶点

• 批准号: 10752865
• 负责人: Susmit Suvas
• 金额: $ 38.5万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10752865
• 关键词: Affect; Antigen-Presenting Cells; Blindness; Blood Vessels; C57BL/6 Mouse; CD4 Positive T Lymphocytes; CXCR4 gene; Cell Degranulation; Cells; Cornea; Corneal Stroma; Data; Dendritic Cells; Deposition; Development; Endothelium; Epithelium; Flow Cytometry; Growth; Herpes Simplex Infections; Herpes stromal keratitis; Herpesvirus 1; Histamine; Histamine Release; Homing; Human; Immune; Impairment; Individual; Infection; Inflammation; Inflammatory; Interferon Type II; Interferon alpha; Ischemia; Langerhans cell; Lesion; Ligands; Lipids; Mediating; Methods; Mus; Neutrophil Infiltration; PECAM1 gene; PTPRC gene; Pathogenicity; Phenotype; Play; Production; Protocols documentation; RNA Viruses; Recurrence; Reporting; Role; Severities; Signal Transduction; Standardization; Stromal Cell-Derived Factor 1; Suspensions; T-Lymphocyte; Testing; Tissues; United States; Vascular Endothelial Cell; Viral Load result; Visual; Visual impairment; cell motility; chemokine receptor; corneal epithelium; effector T cell; experimental study; functional outcomes; healing; immune cell infiltrate; injured; langerin; mast cell; migration; monoamine; monocyte; mouse model; neovascularization; neutrophil; pleiotropism; response; therapeutic target

Herpes stromal keratitis (HSK) is a leading cause of infection-induced vision loss in the United States. The HSK develops in response to corneal herpes simplex virus-1 (HSV-1) infection. Recurrent corneal HSV-1 infection can cause the persistence of innate immune cells, such as monocytes, neutrophils, and effector CD4 T cells in the corneal stroma of the infected cornea. As the accumulation of neutrophils and effector CD4 T cells in the HSK cornea plays a pivotal role in orchestrating the tissue damage, the strategies to manipulate the stay of infiltrating immune cells in the HSK cornea are anticipated to have an effect in reducing the HSK severity. Chemokine receptors, such as CXCR4, can play an important role in the retention of inflammatory cells in the injured tissue. CXCR4 signaling is also reported to enhance hemangiogenesis in ischemic tissue. Our preliminary results showed an increased expression of CXCR4 and CXCL12 in HSV-1 infected cornea. We recently standardized a protocol to separate an intact corneal epithelium (CE) and corneal stroma (CS) from uninfected and HSV-1 infected cornea. Single-cell suspension prepared from individual CE and CS can be used for flow cytometry. Using this strategy, we detected the presence of CXCR4-expressing cells in uninfected CE of C57BL/6 mice. Our results showed that most CXCR4-expressing cells in CE of naïve cornea display the phenotype of Langerhans cells (LCs) or plasmacytoid dendritic cells (pDCs). Recently, monoamines have been shown to engage CXCR4 on pDCs and inhibit IFN-a production. Histamine is a monoamine that is released upon corneal HSV-1 infection. Experiments proposed in aim 1 will test the hypothesis that histamine released in HSV-1 infected cornea engages CXCR4 on pDCs, and downplays IFN-a production from CE pDCs, increasing viral load. Our results also showed that the number of LCs decreases in the CE but increases in the CS from 4- day to 9-day post-infection (p.i.), suggesting a possible migration of these cells from CE to CS in HSV-1 infected corneas. An increased expression of CXCL12 (CXCR4 ligand) was detected in CS than CE of the infected cornea at 9-day p.i. Furthermore, most CD4+ T cells in the CS of infected cornea displayed effector (Ly6Chi) phenotype at 9-day p.i. Experiments proposed in aim 1 will test the hypothesis that LCs-mediated restimulation of effector CD4 T cells in the CS of the infected cornea is essential for their retention and effector function. Aim 1 experiments will also test whether LCs migrating out of CE and neutrophils infiltrating in the CE of HSV-1 infected cornea impair corneal epithelial healing. In addition to corneal resident innate immune cells, our results also detected the expression of CXCR4 on infiltrated neutrophils and vascular endothelial cells from newly formed blood vessels in the CS of HSK cornea. Experiments proposed in aim 2 will test the hypotheses that blocking CXCR4 signaling in HSK cornea will cause reverse migration of neutrophils and inhibit the development of hemangiogenesis. The successful completion of the studies proposed in this application will underscore the pleiotropic effects of CXCR4 signaling in HSK cornea and document it as a potential therapeutic target in HSK.

在美国，疱疹性角膜基质炎（HSK）是感染引起视力丧失的主要原因。洪水桥 角膜单纯疱疹病毒-1（HSV-1）感染后发生。复发性角膜HSV-1感染 可导致先天性免疫细胞，如单核细胞、中性粒细胞和效应CD4 T细胞的持续存在， 感染角膜的角膜基质。随着中性粒细胞和效应性CD4 T细胞在淋巴细胞中的积累， HSK角膜在组织损伤中起着关键作用， HSK角膜中浸润的免疫细胞预期具有降低HSK严重性的作用。 趋化因子受体，如CXCR4，可以在炎症细胞滞留在炎症细胞中发挥重要作用。 受伤的组织CXCR4信号传导也被报道增强缺血组织中的血管生成。我们的初步 结果显示CXCR 4和CXCL 12在HSV-1感染的角膜中表达增加。我们最近 标准化了一种方案，将完整的角膜上皮（CE）和角膜基质（CS）与未感染的角膜上皮（CE）和角膜基质（CS）分离， 和HSV-1感染的角膜。由单独的CE和CS制备的单细胞悬液可用于流动 细胞仪使用这种策略，我们在未感染的CE中检测到表达CXCR 4的细胞的存在。 c57bl/6小鼠我们的结果表明，在未处理角膜的CE中，大多数CXCR4表达细胞显示出 表型的朗格汉斯细胞（LC）或浆细胞样树突状细胞（pDC）。最近，单胺已被 显示其接合pDC上的CXCR4并抑制IFN-α产生。组胺是一种单胺， 角膜HSV-1感染。在目标1中提出的实验将检验组胺释放的假设， HSV-1感染的角膜与pDC上的CXCR4结合，并降低CE pDC的IFN-α产生，从而增加CXCR4的表达。 病毒载量我们的研究结果还表明，LC的数量减少，但在CE中增加，从4- 感染后第1天至第9天（p.i.），提示这些细胞在HSV-1感染中可能从CE迁移到CS， 角膜在感染角膜的CS中检测到CXCL12（CXCR4配体）的表达高于CE 在9天p.i.此外，感染角膜CS中的大多数CD4 + T细胞显示效应（Ly6Chi）表型 在9天p.i.目的1中提出的实验将检验LC介导的效应子再刺激的假设。 感染角膜CS中的CD4 T细胞对于其保留和效应功能至关重要。要求1 实验还将测试是否LC迁移出CE和嗜中性粒细胞浸润在HSV-1感染的CE中 角膜损害角膜上皮愈合。除了角膜固有免疫细胞外，我们的结果还 检测新生血管内皮细胞浸润的中性粒细胞和血管内皮细胞CXCR4的表达 HSK角膜冠状窦内血管。在目标2中提出的实验将检验阻塞 HSK角膜中的CXCR4信号传导将引起中性粒细胞的反向迁移并抑制HSK角膜的发展。 血管生成成功完成本申请中建议的研究将强调 CXCR4信号转导在HSK角膜中的多效性作用，并证明其作为HSK的潜在治疗靶点。