BackgroundThere is absence of specific biomarkers and an incomplete understanding of the pathophysiology of exudative age-related macular degeneration (AMD).Methods and FindingsEighty-eight vitreous samples (73 from patients with treatment naïve AMD and 15 control samples from patients with idiopathic floaters) were analyzed with capillary electrophoresis coupled to mass spectrometry in this retrospective case series to define potential candidate protein markers of AMD. Nineteen proteins were found to be upregulated in vitreous of AMD patients. Most of the proteins were plasma derived and involved in biological (ion) transport, acute phase inflammatory reaction, and blood coagulation. A number of proteins have not been previously associated to AMD including alpha-1-antitrypsin, fibrinogen alpha chain and prostaglandin H2-D isomerase. Alpha-1-antitrypsin was validated in vitreous of an independent set of AMD patients using Western blot analysis. Further systems biology analysis of the data indicated that the observed proteomic changes may reflect upregulation of immune response and complement activity.ConclusionsProteome analysis of vitreous samples from patients with AMD, which underwent an intravitreal combination therapy including a core vitrectomy, steroids and bevacizumab, revealed apparent AMD-specific proteomic changes. The identified AMD-associated proteins provide some insight into the pathophysiological changes associated with AMD.

Purpose: To detect intravitreal functional plasminogen in vitreous samples of patients with recent onset of central retinal vein occlusion (CRVO) and to demonstrate significantly higher intravitreal plasminogen in CRVO patients in comparison to controls. Methods: Prospective clinical case series of 13 consecutive patients with recent onset of CRVO scheduled for core pars plana vitrectomy and 10 consecutive patients undergoing standard pars plana vitrectomy for routine macular surgery or vitreal floater removal. In all 23 cases, vitreous taps were extracted from the central vitreous body, and plasminogen was functionally determined in a new ultrasensitive p-nitroanilide reaction after activation with streptokinase (100% of normal, %N = functional plasminogen in pooled normal citrated plasma). Results: Plasminogen was detected in all analyzed samples (n = 23), and mean plasminogen was revealed to be 1.33 ± 1.73% (mean ± SD), with a range of 0.03-7.8%N. Patients with recent onset of CRVO exhibited significantly higher intravitreal plasminogen (2.19 ± 1.89%N) in comparison to controls (0.20 ± 0.21%N; p < 0.001, Mann-Whitney U test). Conclusion: Due to significantly increased intravitreal plasminogen in patients with recent onset of CRVO, intravitreally administered tissue plasminogen activator might be an option to induce posterior vitreous detachment (enzymatic vitreolysis) in CRVO patients.

Purpose: To evaluate whether intravitreal functional plasminogen is elevated in eyes with branch retinal vein occlusion (BRVO) and to discover whether intravitreal plasminogen activities are correlated with the extent of blood-retina barrier (BRB) breakdown. Methods: Our study is a prospective case series of 20 consecutive patients with BRVO and 10 consecutive patients serving as controls. Vitreous taps were extracted from the central vitreous body and plasminogen was functionally determined in an innovative, ultrasensitive p-nitroanilide reaction after activation with streptokinase (100% of normal, %N = functional plasminogen in pooled normal citrated plasma). Intravitreal VEGF levels were assayed to estimate BRB breakdown. Results: Intravitreal functional plasminogen was detected in all analyzed samples (n = 30) and mean (±SD) plasminogen activities were found to be 0.97 ± 1.06%N (range: 0.03-3.9%N). Patients suffering from BRVO exhibited significantly higher intravitreal plasminogen (1.35 ± 1.11%N) in comparison with controls (0.20 ± 0.21%N, p < 0.001). Intravitreal VEGF concentrations in the BRVO group (576 ± 547 pg/ml) were significantly higher than these in controls (111 ± 120 pg/ml, p = 0.003). There was a significant correlation between intravitreal functional plasminogen and intravitreal VEGF levels (r = 0.519, p = 0.003). Conclusions: Intravitreal functional plasminogen is significantly elevated in eyes suffering from BRVO and correlates with the extent of BRB breakdown. The induction of posterior vitreous detachment by using intravitreally administered recombinant tissue plasminogen activator (enzymatic vitreolysis) should be explored in further investigations.

There were significant differences in intravitreal functional plasminogen and VEGF between eyes with CRVO, BRVO and controls. Intravitreal activity of plasminogen was significantly correlated with the severity of BRB breakdown in RVO affected eyes. The knowledge of intravitreal activities and concentrations of different components of the fibrinolytic cascade could offer new therapeutic strategies in RVO-affected eyes in the future.

To evaluate whether intravitreal thrombin activity is elevated in eyes with branch retinal vein occlusion (BRVO) and central retinal vein occlusion (CRVO) in comparison to healthy controls. Prospective clinical case series of 19 patients with BRVO, 13 patients suffering from CRVO and nine participants serving as controls. Vitreous taps were extracted from the central vitreous body, 200 μl frozen/thawed sample was immediately stabilized with 200 μl 5% human albumin, and 200 μl mixture thereof was stabilized with 200 μl 2.5 mol/l arginine, pH 8.6. Thrombin activity was determined chromogenically. Intravitreal levels of vascular endothelial growth factor (VEGF) as a marker for blood-retina barrier (BRB) breakdown were measured by a commercial chemiluminescent enzyme immuno assay (R&D). Intravitreal thrombin activity and VEGF levels were 1.6 ± 1.2 mIU/ml (mean value ± SD; range: 0.2-4.2 mIU/ml) and 554 ± 568 pg/ml (range: 20-2005 pg/ml) in BRVO-affected eyes, 2.6 ± 1.2 mIU/ml (range: 0.8-5.2 mIU/ml) and 1332 ± 1350 pg/ml (range: 58-3943 pg/ml) in eyes suffering from CRVO as well as 0.8 ± 0.8 mIU/ml (range: 0.2-2.7 mIU/ml) and 115 ± 120 pg/ml (range: 32-431 pg/ml) in controls. There are significant differences of intravitreal thrombin activity and intravitreal VEGF levels between eyes with BRVO, CRVO, and controls (P = 0.007 and P = 0.003, Kruskal-Wallis test). Intravitreal thrombin activity is significantly correlated with intravitreal VEGF levels (r = 0656; P < 0.001, Pearson correlation). Intravitreal thrombin activity might serve as a new marker for BRB breakdown or macular fibrin deposition in ophthalmology. Significant differences of intravitreal thrombin activity between eyes with BRVO, CRVO, and healthy controls might offer new therapeutic strategies for RVO-affected eyes. The effect of oral and intravitrealy injected direct thrombin inhibitors needs to be evaluated in further investigations.