Suppression of alkali burn-induced corneal neovascularization by dendritic cell vaccination targeting VEGF receptor 2
Suppression of alkali burn-induced corneal neovascularization by dendritic cell vaccination targeting VEGF receptor 2
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DOI：
10.1167/iovs.07-1396
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发表时间：
2008-05-01
影响因子：
4.4
通讯作者：
Ishida, Susumu
中科院分区：
文献类型：
--
作者：
Mochimaru, Hiroshi;Usui, Tomohiko;Ishida, Susumu
目的：探讨诱导靶向血管内皮生长因子受体2（VEGFR2）的细胞毒性T淋巴细胞（CTLs）是否能抑制碱烧伤所致的角膜新生血管形成。
方法：将对应于VEGFR2的400至408位氨基酸的H - 2Db限制性肽（VEGFR2(400 - 408)）用作表位肽。从C57BL/6小鼠的骨髓祖细胞中获取树突状细胞（DCs）。6周龄的C57BL/6小鼠每隔6天皮下注射VEGFR2(400 - 408)或gp70脉冲的成熟DCs共3次。第三次免疫后，通过碱烧伤诱导角膜新生血管形成。损伤2周后，通过凝集素血管造影评估角膜血管化区域。为了证实C57BL/6小鼠中肽特异性CTL活性，对免疫小鼠的CD8(+) T细胞进行干扰素（IFN）-γ和肿瘤坏死因子（TNF）-α产生的酶联免疫吸附测定（ELISA）以及(51)Cr释放细胞毒性试验。为了确定体内效应T细胞，给免疫小鼠腹腔注射抗CD4或抗CD8耗竭抗体。
结果：与未免疫或用gp70免疫的小鼠相比，用VEGFR2(400 - 408)免疫的小鼠角膜新生血管形成显著减轻。VEGFR2(400 - 408)或gp70（而非β - gal(96 - 103)）的应用导致与刺激细胞共培养的CD8(+) T细胞中IFN - γ和TNF - α呈剂量依赖性诱导。细胞毒性试验显示对表达VEGFR2的主要组织相容性复合体匹配细胞有特异性裂解，但对β - gal(96 - 103)无此作用。体内耗竭CD8(+)（而非CD4(+)）T细胞可使VEGFR2(400 - 408)免疫对角膜新生血管形成的抑制作用显著逆转至未免疫或用gp70免疫动物中所观察到的水平。
结论：这些结果表明以VEGFR2为靶点的DC疫苗接种作为一种治疗角膜化学损伤的新策略是有可能的。
PURPOSE. To investigate whether the induction of cytotoxic T lymphocytes (CTLs) targeting VEGF receptor 2 inhibits corneal neovascularization caused by alkali injury.METHODS. H-2Db-restricted peptide corresponding to amino acids 400 to 408 of VEGF receptor 2 (VEGFR2(4 00 -408)) was used as an epitope peptide. Dendritic cells (DCs) were harvested from bone marrow progenitors of C57BL/6 mice. Sixweekold C57BL/6 mice received subcutaneous injections of VEGFR2(400-408)- or gp70-pulsed mature DCs three times at 6-day intervals. After the third immunization, corneal neovascularization was induced by alkali injury. Two weeks after the injury, the corneal vascularized area was evaluated by lectin angiography. To confirm the peptide-specific CTL activities in C57BL/6 mice, CD8(+) T cells from immunized mice were subjected to ELISA for interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha production and (51)Cr-release cytotoxicity assay. To determine the in vivo effector T cells, the immunized mice were intraperitoneally injected with an anti-CD4 or -CD8 depletion antibody.RESULTS. Corneal neovascularization was significantly attenuated in mice immunized with VEGFR2(400-408) compared with those not immunized or immunized with gp70. VEGFR2(400-408) or gp70, but not beta-gal(96-103), application led to dose-dependent induction of IFN-gamma and TNF-alpha in the CD8(+) T cells cocultured with stimulator cells. Cytotoxicity assays showed the specific lysis of major histocompatibility complex-matched cells expressing VEGFR2, but not beta-gal (96-103). In vivo depletion of CD8(+), but not CD4(+), T cells significantly reversed the suppressive effect of VEGFR2(400-408) immunization on corneal neovascularization to the level observed in nonimmunized or gp70-immunized animals.CONCLUSIONS. These results indicate the possibility of DC vaccination targeting VEGFR2 as a novel therapeutic strategy for corneal chemical injury.