Center for Excellence in Functional and Molecular Imaging, Brigham and Women's Hospital, Boston, MA 02115, USA.
Invest Ophthalmol Vis Sci. 2013 Jun 3;54(6):3830-6. doi: 10.1167/iovs.12-10341.
Inflammation and immune cells regulate choroidal neovascularization (CNV) and could become therapeutic targets in age-related macular degeneration (AMD). Lymphangiogenesis is a key component of various inflammatory diseases. Whether lymphangiogenesis and lymph node-mediated immunity are involved in the pathogenesis of AMD is not understood.
To investigate lymphangiogenesis in CNV, we generated CNV in animals by laser and studied surgically removed CNV membranes from uveitis and AMD patients. Immunohistochemistry was performed with lymphatic vessel endothelial hyaluronate receptor 1 (LYVE-1) and podoplanin antibodies. VEGF-C and VEGFR-3 expressions were examined with immunohistochemistry and Western blotting. To examine the role of lymph node in CNV, we lasered lymphotoxin alpha-deficient mice (LTα-/-) and measured the CNV volume.
Immunohistochemistry showed that LYVE-1(+) macrophages infiltrated in acutely induced CNV, although lymphatic tubes did not form. CNV membranes from patients did not show LYVE-1(+)podoplanin(+) vessels, suggesting the lack of lymphangiogenesis in AMD and uveitis. Western blots and immunostaining revealed VEGF-C and VEGFR-3 expression in CNV lesions, mainly in macrophages and angiogenic endothelial cells. Using fluorescent microsphere tracers, we show a path for cellular migration from the eye to the cervical lymph nodes (LNs) during CNV. However, CNV injury did not cause LN swelling. CNV volume did not differ between wild-type and LN-deficient mice, suggesting that LN is not a key component of early CNV formation.
Laser-induced CNV is not primarily dependent on acquired immunity, nor does the fundus injury affect peripheral LNs. Our results reveal a previously unknown cellular connection between the ocular fundus and the cervical LNs. This connection that in function resembles lymphatics is actively utilized in CNV.
炎症和免疫细胞调节脉络膜新生血管（CNV），并可能成为年龄相关性黄斑变性（AMD）的治疗靶点。淋巴管生成是各种炎症性疾病的关键组成部分。然而，目前尚不清楚淋巴管生成和淋巴结介导的免疫是否参与 AMD 的发病机制。
为了研究 CNV 中的淋巴管生成，我们通过激光在动物中诱导 CNV，并研究了从葡萄膜炎和 AMD 患者手术切除的 CNV 膜。采用淋巴管内皮透明质酸受体 1（LYVE-1）和 podoplanin 抗体进行免疫组织化学染色。采用免疫组织化学和 Western blot 检测 VEGF-C 和 VEGFR-3 的表达。为了研究淋巴结在 CNV 中的作用，我们对淋巴毒素 alpha 缺陷型小鼠（LTα-/-）进行了激光处理，并测量了 CNV 体积。
免疫组织化学显示，LYVE-1（+）巨噬细胞浸润在急性诱导的 CNV 中，尽管淋巴管并未形成。来自患者的 CNV 膜未显示 LYVE-1（+）podoplanin（+）血管，提示 AMD 和葡萄膜炎中缺乏淋巴管生成。Western blot 和免疫染色显示 VEGF-C 和 VEGFR-3 在 CNV 病变中表达，主要在巨噬细胞和血管生成内皮细胞中。我们使用荧光微球示踪剂显示，在 CNV 期间，细胞从眼睛迁移到颈淋巴结（LN）的途径。然而，CNV 损伤并未导致 LN 肿胀。野生型和 LN 缺陷型小鼠之间的 CNV 体积无差异，提示 LN 不是早期 CNV 形成的关键组成部分。
激光诱导的 CNV 主要不依赖于获得性免疫，眼底损伤也不会影响外周 LN。我们的研究结果揭示了眼睛和颈 LN 之间以前未知的细胞连接。这种功能上类似于淋巴管的连接在 CNV 中被积极利用。
