Doheny Eye Institute and Department of Ophthalmology, David Geffen School of Medicine at UCLA, Los Angeles, CA, USA.
Hamilton Glaucoma Center, Shiley Eye Institute, Viterbi Family Department of Ophthalmology University of California, San Diego, CA, USA.
Exp Eye Res. 2020 Jul;196:108064. doi: 10.1016/j.exer.2020.108064. Epub 2020 May 18.
This work sought to compare aqueous angiographic segmental patterns with bead-based methods which directly visualize segmental trabecular meshwork (TM) tracer trapping. Additionally, segmental protein expression differences between aqueous angiographic-derived low- and high-outflow human TM regions were evaluated. Post-mortem human eyes (One Legacy and San Diego eye banks; n = 15) were perfused with fluorescent tracers (fluorescein [2.5%], indocyanine green [0.4%], and/or fluorescent microspheres). After angiographic imaging (Spectralis HRA+OCT; Heidelberg Engineering), peri-limbal low- and high-angiographic flow regions were marked. Aqueous angiographic segmental outflow patterns were similar to fluorescent microsphere TM trapping segmental patterns. TM was dissected from low- and high-flow areas and processed for immunofluorescence or Western blot and compared. Versican expression was relatively elevated in low-flow regions while MMP3 and collagen VI were relatively elevated in high-flow regions. TGF-β2, thrombospondin-1, TGF-β receptor1, and TGF-β downstream proteins such as α-smooth muscle actin were relatively elevated in low-flow regions. Additionally, fibronectin (FN) levels were unchanged, but the EDA isoform (FN-EDA) that is associated with fibrosis was relatively elevated in low-flow regions. These results show that segmental aqueous angiographic patterns are reflective of underlying TM molecular characteristics and demonstrate increased pro-fibrotic activation in low-flow regions. Thus, we provide evidence that aqueous angiography outflow visualization, the only tracer outflow imaging method available to clinicians, is in part representative of TM biology.
本研究旨在比较水相血管造影的节段模式与直接可视化节段小梁网（TM）示踪剂滞留的珠基方法。此外，还评估了水相血管造影衍生的低和高流出人 TM 区之间的节段蛋白表达差异。死后人类眼球（One Legacy 和圣地亚哥眼库；n=15）用荧光示踪剂（荧光素[2.5%]、吲哚菁绿[0.4%]和/或荧光微球）灌注。血管造影成像（Spectralis HRA+OCT；Heidelberg Engineering）后，标记周边低和高血管造影血流区域。水相血管造影的节段流出模式与荧光微球 TM 滞留的节段模式相似。从低和高流量区分离 TM，并进行免疫荧光或 Western blot 处理并进行比较。在低流量区，蛋白聚糖表达相对升高，而在高流量区，MMP3 和胶原 VI 相对升高。TGF-β2、血小板反应蛋白-1、TGF-β受体 1 和 TGF-β下游蛋白如α-平滑肌肌动蛋白在低流量区相对升高。此外，纤连蛋白（FN）水平不变，但与纤维化相关的 EDA 同工型（FN-EDA）在低流量区相对升高。这些结果表明，节段水相血管造影模式反映了 TM 的分子特征，并显示低流量区中存在增加的促纤维化激活。因此，我们提供的证据表明，水相血管造影流出可视化是临床医生唯一可用的示踪剂流出成像方法，部分代表 TM 生物学。
