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.

疱疹基质角膜炎（HSK）是美国感染引起的视力丧失的主要原因。 HSK 响应角膜疱疹单纯疱疹病毒-1（HSV-1）感染的发展。复发性角膜HSV-1感染 可能导致先天免疫细胞的持久性，例如单核细胞，中性粒细胞和效应子CD4 T细胞 感染角膜的角膜基质。作为中性粒细胞和效应子CD4 T细胞的积累 HSK角膜在策划组织损伤方面起着关键作用，操纵停留的策略 预计HSK角膜中的免疫细胞在降低HSK的严重程度上会产生影响。 趋化因子受体，例如CXCR4，可以在保留炎症细胞中发挥重要作用 受伤的组织。据报道，CXCR4信号传导可增强缺血组织中的血管生成。我们的初步 结果表明，在HSV-1感染的角膜中，CXCR4和CXCL12的表达增加。我们最近 标准化的协议，以将完整的角膜上皮（CE）和角膜基质（CS）与未感染 和HSV-1感染的角膜。由单个CE和CS制备的单细胞悬架可用于流量 细胞仪。使用此策略，我们在未感染的CE中检测到CXCR4表达细胞的存在 C57BL/6鼠。我们的结果表明，幼稚角膜CE中的大多数表达CXCR4的细胞显示 Langerhans细胞（LCS）或浆细胞类动物树突状细胞（PDC）的表型。最近，单胺是 证明可以在PDC上吸引CXCR4并抑制IFN-A产生。组胺是一种单胺，已释放 角膜HSV-1感染后。 AIM 1中提出的实验将检验组胺释放的假设 HSV-1感染的角膜在PDC上与CXCR4接合，而CE PDC的IFN-A产生降低，增加了 病毒负荷。我们的结果还表明，CE中LC的数量减少，但CS的LC数量从4--增加。 感染后的日至9天（P.I.），表明这些细胞可能从HSV-1感染中从CE到CS迁移 角膜。在CS中检测到CXCL12（CXCR4配体）的表达增加，而不是感染的角膜 在9天P.I.此外，受感染角膜CS中的大多数CD4+ T细胞显示出效应子（Ly6chi）表型 在9天P.I. AIM 1中提出的实验将检验以下假设：LCS介导的效应子的重新启用 感染角膜CS中的CD4 T细胞对于其保留和效应函数至关重要。目标1 实验还将测试LCS是否从CE迁移到HSV-1感染CE中的中性粒细胞。 角膜损害角膜上皮愈合。除了角膜居民先天免疫细胞外，我们的结果也 从新形成 HSK角膜CS中的血管。 AIM 2中提出的实验将测试阻止的假设 HSK角膜中的CXCR4信号传导将导致中性粒细胞的反向迁移，并抑制 血管生成。在本申请中提出的研究的成功完成将强调 CXCR4信号传导在HSK角膜中的多效作用，并将其记录为HSK中潜在的治疗靶标。