中性粒细胞促进实验性脉络膜新生血管形成。

Neutrophils promote experimental choroidal neovascularization.

作者信息

Zhou Jiehao, Pham Lucia, Zhang Ning, He Shikun, Gamulescu Maria-Andreea, Spee Christine, Ryan Stephen J, Hinton David R

机构信息

The Arnold and Mabel Beckman Macular Research Center, Doheny Eye Institute, Keck School of Medicine, University of Southern California, Los Angeles, CA 90033, USA.

出版信息

Mol Vis. 2005 Jun 16;11:414-24.

PMID:15988410

Abstract

PURPOSE

To investigate the role of neutrophils in the development of laser induced experimental choroidal neovascularization (CNV).

METHODS

CNV was induced by laser photocoagulation in adult male C57BL/6J mice. Neutrophil infiltration was evaluated by histology and confocal immunohistology. The expression of neutrophil chemotactic chemokines in the regions of laser injury was determined by quantitative real-time PCR. Animals were treated with NIMP-R14, an anti-murine neutrophil monoclonal antibody (mAb), intraperitoneally to deplete neutrophils. The specific neutrophil depletion was confirmed by flow cytometry. The CNV responses were compared between neutropenic and untreated control mice on the basis of fluorescein angiography (FA), CNV lesion volume and lesion histology, and vascular endothelial growth factor (VEGF) expression by ELISA. Expression of VEGF and Angiopoietin-1 and Angiopoietin-2 protein by murine neutrophils was evaluated by confocal immunohistochemistry.

RESULTS

Neutrophils infiltrated the sites of laser injury as early as day 1 after laser treatment and peaked at day 3. The neutrophil infiltration correlated with enhanced mRNA expression of neutrophil chemotactic chemokines MIP-2 and KC in the lesions. Administration of NIMP-R14 mAb specifically depleted neutrophils. Analysis of FA, CNV volume, and lesion histology, all demonstrated a moderate decrease in the CNV response in neutropenic mice compared to control mice (p<0.01). The reduction in the CNV response in neutropenic mice was associated with decreased VEGF protein levels in the ocular posterior segment. Murine neutrophils contained VEGF and Angiopoietin-1 and Angiopoietin-2 proteins.

CONCLUSIONS

Neutrophil invasion was part of early inflammatory responses during laser induced CNV. Neutrophil depletion correlated with reduced CNV responses and decreased VEGF protein expression. These data suggest that neutrophils promoted the early development of CNV possibly via secretion of angiogenic growth factors.

摘要

目的

研究中性粒细胞在激光诱导的实验性脉络膜新生血管形成（CNV）过程中的作用。

方法

通过激光光凝在成年雄性C57BL/6J小鼠中诱导CNV形成。通过组织学和共聚焦免疫组织化学评估中性粒细胞浸润情况。采用定量实时PCR测定激光损伤区域中性粒细胞趋化性细胞因子的表达。给动物腹腔注射抗小鼠中性粒细胞单克隆抗体（mAb）NIMP-R14以清除中性粒细胞。通过流式细胞术确认中性粒细胞的特异性清除。基于荧光素血管造影（FA）、CNV病变体积和病变组织学以及酶联免疫吸附测定法（ELISA）检测的血管内皮生长因子（VEGF）表达，比较中性粒细胞减少的小鼠和未处理的对照小鼠的CNV反应。通过共聚焦免疫组织化学评估小鼠中性粒细胞中VEGF、血管生成素-1和血管生成素-2蛋白的表达。

结果

中性粒细胞在激光治疗后第1天就开始浸润激光损伤部位，并在第3天达到峰值。中性粒细胞浸润与病变中中性粒细胞趋化性细胞因子MIP-2和KC的mRNA表达增强相关。给予NIMP-R14 mAb可特异性清除中性粒细胞。FA、CNV体积和病变组织学分析均显示，与对照小鼠相比，中性粒细胞减少的小鼠的CNV反应中度降低（p<0.01）。中性粒细胞减少的小鼠中CNV反应的降低与眼后段VEGF蛋白水平降低有关。小鼠中性粒细胞含有VEGF、血管生成素-1和血管生成素-2蛋白。