Department of Ophthalmology, University of Ulm Medical School, Prittwitzstrasse 43, D-89075 Ulm, Germany.
Br J Ophthalmol. 2011 Aug;95(8):1151-6. doi: 10.1136/bjo.2010.192229. Epub 2011 Jan 27.
Deregulated expression of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) or insulin-like growth factor-1 (IGF-1) is associated with the pathogenesis of diabetic retinopathy. The VEGF(165)-induced increase in permeability of retinal endothelial cells (REC), probably resulting in diabetic macular oedema (DME), could be completely restored by the VEGF-binding Fab fragment ranibizumab in vitro. We investigated whether bFGF and IGF-1 as single factors or in combination with VEGF(165) influence permeability and tight junctions in immortalised bovine REC (iBREC) and if these effects could be restored by inhibition of VEGF.
As a measure of changes in cellular permeability, transendothelial electrical resistance (TER) was monitored during long-term treatment of iBREC with growth factors in the absence or presence of ranibizumab or KRN951 (an inhibitor of VEGF receptors). Expression of claudin-1, as an indicator of functional tight junctions, was assessed by western blot analysis.
Whereas VEGF(165) decreased TER and expression of claudin-1 in a concentration-dependent manner, long-term treatment of iBREC with 10-100 ng/ml bFGF or/and IGF-1 did not. Changes in claudin-1 expression or TER, induced by 25 ng/ml VEGF(165), were slightly enhanced by bFGF and/or IGF-1 and were accompanied by a slightly increased secretion of VEGF. Complete reversion of these effects was achieved by prolonged treatment with ranibizumab and partly by exposure to KRN951.
Our findings indicate that VEGF(165), but not IGF-1 or bFGF, is mainly responsible for changes in cellular permeability observed in REC. This supports VEGF targeting as a therapeutic concept for DME.
血管内皮生长因子（VEGF）、碱性成纤维细胞生长因子（bFGF）或胰岛素样生长因子-1（IGF-1）的表达失调与糖尿病性视网膜病变的发病机制有关。VEGF（165）诱导的视网膜内皮细胞（REC）通透性增加，可能导致糖尿病性黄斑水肿（DME），这种增加可以在体外通过 VEGF 结合 Fab 片段 ranibizumab 完全恢复。我们研究了 bFGF 和 IGF-1 作为单一因素或与 VEGF（165）联合是否会影响永生化牛 REC（iBREC）的通透性和紧密连接，以及这些作用是否可以通过抑制 VEGF 来恢复。
作为细胞通透性变化的衡量标准，在缺乏或存在 ranibizumab 或 KRN951（VEGF 受体抑制剂）的情况下，长期用生长因子处理 iBREC 时，监测跨内皮电阻（TER）。通过 Western blot 分析评估紧密连接蛋白-1 的表达，作为功能性紧密连接的指标。
VEGF（165）以浓度依赖性方式降低 TER 和紧密连接蛋白-1 的表达，而长期用 10-100ng/ml bFGF 或/和 IGF-1 处理 iBREC 则不会。25ng/ml VEGF（165）诱导的紧密连接蛋白-1 表达或 TER 的变化，被 bFGF 和/或 IGF-1 轻微增强，并且伴随着 VEGF 的分泌略有增加。通过延长 ranibizumab 的治疗和部分暴露于 KRN951，可以完全逆转这些作用。
我们的发现表明，VEGF（165），而不是 IGF-1 或 bFGF，是 REC 中观察到的细胞通透性变化的主要原因。这支持将 VEGF 靶向作为 DME 的治疗概念。
