Cammalleri Maurizio, Dal Monte Massimo, Locri Filippo, Lista Liliana, Aronsson Monica, Kvanta Anders, Rusciano Dario, De Rosa Mario, Pavone Vincenzo, André Helder, Bagnoli Paola
Department of Biology University of Pisa, Pisa, Italy.
Department of Chemical Sciences, University of Napoli Federico II, Napoli, Italy.
Invest Ophthalmol Vis Sci. 2016 May 1;57(6):2600–2611. doi: 10.1167/iovs.15-18758.
A mouse model of age-related macular degeneration (AMD) was used to investigate the anti-angiogenic and anti-inflammatory role of UPARANT in laser-induced choroidal neovascularization (CNV).
Choroidal neovascularization was induced by laser photocoagulation, and UPARANT was intravitreally injected. Some experiments were also performed after either intravitreal injection of anti-VEGF drugs or systemic administration of UPARANT. Immunohistochemistry using CD31 antibodies was used to evaluate the area of CNV. Evans blue dye extravasation was quantitatively assessed. Transcripts of markers of outer blood retinal barrier were measured by quantitative RT-PCR, also used to evaluate angiogenesis and inflammation markers. Western blot was used to determine levels of transcription factors encoding genes involved in angiogenesis and inflammation. Levels of urokinase-type plasminogen activator (uPA), its receptor (uPAR), and formyl peptide receptors (FPRs) were determined at the transcript and the protein level.
Intravitreal UPARANT reduced the CNV area and the leakage from the choroid. The uPA/uPAR/FPR system was upregulated in CNV, but was not influenced by UPARANT. UPARANT recovered laser-induced upregulation of transcription factors encoding angiogenic and inflammatory markers. Accordingly, angiogenic and inflammatory factors were also reduced. UPARANT as compared to anti-VEGF drugs displayed similar effects on CNV area.
UPARANT mitigates laser-induced CNV by inhibiting angiogenesis and inflammation through an action on transcription factors encoding angiogenesis and inflammatory genes. The finding that UPARANT is effective against CNV may help to establish uPAR and its membrane partners as putative targets in the treatment of AMD.
利用年龄相关性黄斑变性（AMD）小鼠模型研究UPARANT在激光诱导脉络膜新生血管（CNV）中的抗血管生成和抗炎作用。
通过激光光凝诱导脉络膜新生血管形成，并玻璃体内注射UPARANT。在玻璃体内注射抗VEGF药物或全身给药UPARANT后也进行了一些实验。使用CD31抗体的免疫组织化学方法评估CNV的面积。对伊文思蓝染料外渗进行定量评估。通过定量RT-PCR测量视网膜外血视网膜屏障标志物的转录本，也用于评估血管生成和炎症标志物。蛋白质印迹法用于测定参与血管生成和炎症的编码基因的转录因子水平。在转录本和蛋白质水平上测定尿激酶型纤溶酶原激活剂（uPA）、其受体（uPAR）和甲酰肽受体（FPRs）的水平。
玻璃体内注射UPARANT可减少CNV面积和脉络膜渗漏。uPA/uPAR/FPR系统在CNV中上调，但不受UPARANT影响。UPARANT恢复了激光诱导的编码血管生成和炎症标志物的转录因子的上调。因此，血管生成和炎症因子也减少了。与抗VEGF药物相比，UPARANT对CNV面积显示出相似的作用。
UPARANT通过作用于编码血管生成和炎症基因的转录因子来抑制血管生成和炎症，从而减轻激光诱导的CNV。UPARANT对CNV有效的这一发现可能有助于将uPAR及其膜伴侣确立为AMD治疗中的假定靶点。
