Geldanamycin treatment reduces neovascularization in a mouse model of retinopathy of prematurity

Background The benzoquinoid antibiotic 17-allylaminogeldanamycin (17-AAG) inhibits the Ras/Raf/MEK and PI3-Kinase signaling pathways and down-regulates vascular endothelial factor expression. Here we use a mouse model of oxygen-induced retinopathy to investigate the effect of 17-AAG on retinal neovascularization and vascular recovery. Material and methods C57BL/6 mice were exposed to 75% oxygen from postnatal day 7 (P7) to P12 and recovered in room air thereafter. Beginning with P12 mice were treated for 5 days by daily IP injection of 17-AAG (12.5 mg/kg body weight) micro dispersed in an emulsion of 4% Lipoid EPC, 5% sucrose, and 0.9% NaCl or Wortmannin (100 μg/kg body weight). On P17, the retinal vascular and avascular area, neovascular blood vessel tufts, and main vessel tortuosity were quantified after perfusion of the mice with FITC-Concanavalin A. The mRNA levels of VEGF, angiopoietin 1 and 2 were quantified by real-time RT-PCR. Results After 17-AAG treatment, a reduction of the vascular area was measured from 37.8±5.2% to 30.8±5.7% (P=0.005), and an increase of the avascular area from 10.8±5.6% to 20.3±6.6% (P=0.001). No alteration of the vascular pattern, the number of blood vessel tufts and the main vessel tortuosity was achieved by treatment with the PI-3 kinase inhibitor Wortmannin. After treatment with 17-AAG, the numbers of tufts (127.9±33.2) were different from the controls (173.7±55.2, P=0.035), but not the main vessel tortuosity. No significant change in VEGF and angiopoietin 1 mRNA expression could be achieved with either of the treatments. Wortmannin treatment also did not change the angiopoietin 2 mRNA level, whereas the level was reduced in 17-AAG treated mice retina from 436-fold (± 64) to 200-fold (±55) (P=0.035). Conclusion An IP injection of 17-AAG is able to reduce angioproliferative retinopathy in a mouse model for oxygen-induced retinopathy. Our data indicate that the mechanism does not involve a direct or indirect reduction of the VEGF mRNA level, but acts downstream of the VEGF pathway. Thus, 17-AAG probably does not work by PI-3 kinase inhibition but via the Ras/Raf/MEK pathway. These data underline the potential utility of tyrosine kinase inhibitors in hypoxia induced neovascularization.     

The effect of thalidomide on vascular endothelial growth factor and tumor necrosis factor-α levels in retinal ischemia/reperfusion injury

Background To evaluate the effects of thalidomide treatment on the temporal course of TNF-α, VEGF production and the histopathological changes in ischemia/reperfusion (I/R) injured guinea pigs retina. Methods Control, ischemia, and thalidomide/ischemia groups including seven animals each were formed. Retinal ischemia was induced in male guinea pigs by cannulating anterior chambers and lifting the bottle to a height of 205 cm for 90 min in the ischemia and thalidomide/ischemia groups. The thalidomide/ischemia group received thalidomide (300 mg/kg/day) via nasogastric tube 24 h before ischemia and during 7 days of reperfusion. Guinea pigs were sacrificed for histopathological examination to evaluate the mean thickness of the inner plexiform layer (IPL), polymorphonuclear leukocyte (PMNL) infiltration, and biochemical analysis of retinal VEGF and TNF-α levels by ELISA. Results The mean retinal VEGF and TNF-α levels of the control, ischemia, and thalidomide/ischemia groups were 10.22 ± 2.58 and 270.41 ± 69.77 pg/ml; 35.80 ± 5.97 and 629.93 ± 146.41 pg/ml; 19.01 ± 3.01 and 340.93 ± 158.26 pg/ml, respectively. The retinal VEGF levels were significantly higher in I/R injured groups. The thalidomide/ischemia group retinal VEGF level was significantly lower versus the ischemia group. The retinal TNF-α levels were significantly elevated in the ischemia group, but no difference was observed between the thalidomide/ischemia and control groups. Also, the retinal TNF-α level was significantly lower in the thalidomide/ischemia group versus the ischemia group. The mean thickness of IPL and PMNL infiltration showed no difference between the control and thalidomide/ischemia groups. However, there was a significant difference between the control and ischemia groups. Conclusion Thalidomide treatment decreases PMNL infiltration, retinal edema, VEGF, and TNF-α synthesis following I/R injury to the guinea pig retina. This study supported by Firot University Research Fund.     

Preliminary study of retinal pathological features in preterm birth pups exposed to an animal model of oxygen-induced retinopathy in mice

Background

The main risk factors of retinopathy of prematurity (ROP) are low gestational age and low birth weight, which are mainly caused by preterm birth. Currently, the animal model of oxygen-induced retinopathy (OIR) in mice is the most widely used model in ROP-associated studies. However, the experimental mice are normal-term pups, and may not mimic the pathogenic status of human ROP patients. In this study, we investigated the retinal pathological features in preterm birth pups exposed to an animal model of oxygen-induced retinopathy in mice.

Methods

Preterm-birth mice were obtained from pregnant C57BL/6J mice that were induced by an intraperitoneal injection of lipopolysaccharide (LPS). The preterm and control mice were treated with high oxygen (75 %) from postnatal day 7 (P7) to P12. The mice were perfused with high-molecular-weight FITC-dextran on P12, P15 and P17, and the retinas were whole-mounted and imaged. Vascular endothelial growth factor (VEGF) mRNA was also detected. Cross-sections of the retina were stained with hematoxylin and eosin (H&E) to identify preretinal neovascular tufts. For general observation, whole retinal images were also obtained using a microscope.

Results

Leakage of the retinal blood vessels was aggravated in the preterm mice, particularly on P12 and P15. The non-perfused areas of the retina (pixel value, 183,673?±?28,148 vs 132,110?±?23,732, P?=?0.009) and the number of preretinal endothelial cell nuclei were smaller (30.17?±?8.33 vs 22.17?±?6.74, P?<?0.0001) on P17. The VEGF mRNA levels in the retinas were higher on P12 and P15 but lower on P17, compared with the control mice. Retinal hemorrhage was observed in the preterm mouse group (five out of six examined eyes).

Conclusions

Preterm-birth mice that were subject to OIR exhibited several pathological features, such as retinal hemorrhage, severe retinal leakage and moderate retinal neovascularization, which were similar to the clinical manifestations in ROP patients.     

氧诱导的视网膜病变小鼠模型建立方法的改良

目的:建立简单、稳定、成模率高的氧诱导的视网膜病变小鼠模型。方法:用改良法(A组)和传统方法(B组)分别建立氧诱导的视网膜病变的C57BL/6小鼠模型,比较两组的母鼠死亡率、乳鼠成活率和成模率,视网膜冰冻切片GSL染色免疫组化观察两组的视网膜新生血管,荧光素灌注视网膜铺片检测视网膜无灌注区和新生血管的面积。结果:改良法(A组)与传统法(B组)相比较,乳鼠成活率未降低,且具有母鼠死亡率低,成模率高,动物消耗量少的特点。改良法制作的小鼠模型视网膜无灌注区和新生血管面积较传统法多而典型。结论:改良法操作简单,能构建出高效、稳定、典型的视网膜新生血管模型。该方法值得推广和借鉴。     

Peeling of internal limiting membrane during vitrectomy for complicated retinal detachment prevents epimacular membrane formation

Background  To investigate the clinical benefit of internal limiting membrane (ILM) peeling at the macula for the prevention of epimacular membrane formation following vitreous surgery using silicone oil for the treatment of complicated retinal detachment. Methods  This was a non-randomized, retrospective, interventional study of a case series. Patient charts were reviewed retrospectively for 20 consecutively recruited patients who underwent successful primary vitrectomy with ILM peeling at the macula using silicone oil (group 1) and 22 consecutively recruited patients who underwent successful primary vitrectomy using silicone oil without ILM peeling at the macula for complicated rhegmatogenous retinal detachment (group 2). The main outcome measures were distant visual acuity and epimacular membrane formation. The data were analyzed and compared using Fisher’s Exact test, Pearson Chi-square test, independent t-test, Mann–Whitney U-test, and a repeated ANOVA. Results  The mean age of patients was 52.7 ± 12.6 years in group 1 and 53.2 ± 13.3 years in group 2 (p = 0.89). The mean follow-up time was 24.6 ± 7.6 weeks in group 1 and 34.1 ± 12.6 weeks in group 2 (p = 0.01). Preoperatively, ten eyes in group 1 and 10 eyes in group 2 were pseudophakic; the macula was detached in all cases. Silicone oil had been removed from all eyes of both groups at least 3 months before the final examination. There were no significant differences between the two groups with regard to sex (p = 0.44), mean duration of retinal detachment (p = 0.12), mean preoperative visual acuity (logMAR), mean number of retinal breaks (p = 0.43), and grade of proliferative vitreoretinopathy (p = 0.35). The final visual acuity (logMAR) was 0.60 ± 0.30 in group 1 and 0.72 ± 0.35 in group 2 (p = 0.49). Four eyes in group 1 and two eyes in group 2 underwent cataract surgery during silicone oil removal. Epimacular membrane formation was observed in two eyes before silicone oil removal and in four eyes within 8 weeks after silicone oil removal in group 2. No epimacular membrane formation was seen in group 1 (p = 0.02). Conclusion  ILM peeling at the macula during vitreous surgery with silicone oil for the treatment of complicated retinal detachment may prevent epimacular membrane formation without negatively affecting distant visual acuity. The results of this study were presented at the 8th Euretina Congress 2008, Vienna, Austria. The authors have no conflicting interests in the subject matter presented.     

Circulatory parameters in the retrobulbar central retinal artery and vein of patients with diabetes and medically treated systemic hypertension

Background  We aim to study the circulatory parameters in the retrobulbar central retinal artery and vein in diabetic patients with and without medically treated systemic hypertension. Methods  The study included 108 patients with diabetes that were allocated in four different groups according to the presence of diabetic retinopathy (DR) and hypertension: group 1—patients without DR and without hypertension (n = 23), group 2—patients without DR and with hypertension (n = 21), group 3—patients with nonproliferative DR and without hypertension (n = 36), group 4—patients with nonproliferative DR and with hypertension (n = 28). The circulatory parameters that were evaluated were: peak systolic blood velocity (PSV), end-diastolic blood velocity (EDV), maximum venous velocity (Vmax), minimum venous velocity (Vmin) and the Pourcelot index which were measured using color Doppler imaging. Non-parametric tests were used to test inter-group differences. Spearman’s coefficient of correlation was tested between ocular perfusion pressure and the circulatory parameters in each of the patient groups. Contingency table was performed to test the relation of diabetic retinopathy and hypertension to the PSV in the central retinal artery. Results  The PSV and EDV in the central retinal artery was significantly higher in group 1 (p = 0.02, p = 0.04) and group 2 (p = 0.02, p = 0.02) than in group 3. The Pourcelot index in the central retinal vein was significantly lower in group 1 than in group 4 (p = 0.02), and in group 2 than in groups 3 and 4 (p = 0.02, p < 0.01). A significant relationship was detected between the presence of hypertension, the stage of diabetic retinopathy and the PSV in the central retinal artery of our patients (χ² = 8.29; p = 0.04). Conclusion  Medically treated hypertension affects the retrobulbar circulatory parameters in the central retinal artery and vein in diabetes.     

An increase in superoxide dismutase ameliorates oxygen-induced retinopathy in transgenic mice

PURPOSE: Oxygen therapy is a well-recognized risk factor for retinopathy of prematurity. We examined whether an increase in the naturally occurring enzyme copper-zinc superoxide dismutase (CuZnSOD), which controls oxygen, can reduce the severity of oxygen-induced retinopathy in a mouse model. METHODS: Seven transgenic mice overexpressing CuZnSOD and six wild-type mice were exposed to 75% oxygen from postnatal day 7 to 12. Seven transgenic mice and five mice of the wild type were kept in room air and served as controls. Fluorescein-conjugated dextran angiography of retinal vasculature was performed and flat-mounted preparations were evaluated by scoring blood vessel growth, blood vessel tuft formation, extraretinal neovascularization, degree of central constriction, and tortuosity of vessels. In addition, quantification of the number of blood vessel tufts was performed in a masked fashion with haematoxylin and eosin staining of paraffin-embedded eye sections. RESULTS: The mean retinal score+/-SD obtained by the wild-type mice was 9.4+/-2.0, whereas the transgenic mice overexpressing CuZnSOD obtained a value of 2.4+/-1.6 (P=0). The two control groups (wild type and transgenic) that were kept in room air, each obtained a score of 0. Significantly fewer extraretinal vascular tufts were seen in the transgenic mice (0.26+/-0.34) than in the wild-type mice (4.27+/-1.6) after both groups were exposed to oxygen (P<0.001). CONCLUSIONS: The results suggest that high SOD activity protects neonatal mice against oxygen-induced retinopathy, and support the assumption that oxygen radicals are a major causative factor in oxygen-induced retinopathy.     

Involvement of purinergic P2 receptors in experimental retinal neovascularization

PURPOSE: The present study was aimed to investigate the expression of purinergic P2 receptors in oxygen-induced retinal neovascularization. METHODS: Immunohistochemistry was used to study the expression of purinergic P2Y2 and P2X2 receptors in the neonatal mouse retina during normal vascular development and after oxygen-induced retinopathy (OIR). The effect of the P2 antagonists, suramin and PPADS, on the extent of oxygen-induced retinal neovascularization was analyzed. RESULTS: In normal mice, the expression of P2Y2 receptors was weak throughout the retina, whereas P2X2 receptor expression was detected in the outer plexiform layer. In mice treated with oxygen, P2Y2 expression was detected in the ganglion and in the nerve fiber layers, whereas P2X2 expression was found in the inner and outer plexiform layers. Oxygen-induced preretinal neovascularization was strongly inhibited by the P2 antagonists, suramin (p<0.05) and PPADS (p<0.05), and this was accompanied by a down-regulation of P2X2 receptor expression in the inner plexiform layer in suramin-treated mice. CONCLUSIONS: The data suggest that purinergic P2 receptors are involved in neovascularization associated with OIR.     

Attenuation of EphrinB2 Reverse Signaling Decreases Vascularized Area and Preretinal Vascular Tuft Formation in the Murine Model of Oxygen-Induced Retinopathy

Purpose. EphB4 and ephrinB2 are known key regulators of retinal vascular development, but due to their capacity for bidirectional signaling, delineation of their individual roles in this process remains unclear. To better dissect out individual contributions, a model of proliferative retinopathy in mice with attenuated ephrinB2 reverse signaling was studied. It was hypothesized that endothelial ephrinB2 reverse signaling regulates hypoxia-induced capillary sprouting, as well as the pathologic formation of neovascular tufts in postnatal retinal microvascular networks. Methods. Genetically manipulated mice with attenuated ephrinB2 reverse signaling (ephrinB2(lacZ/+)), along with wild-type (WT) controls, were exposed to oxygen-induced retinopathy (OIR), a postnatal model of proliferative retinopathy. At peak disease (postnatal day 18), microvascular networks were analyzed to examine intraretinal revascularization, capillary sprouting, and pathologic neovascularization responses. EphB4 and phosphorylated ephrinB protein expression patterns along retinal microvessels were also assessed. Results. EphrinB2(lacZ/+) mice exhibited reduced hypoxia-induced revascularization (P ≤ 0.04) and reduced formation of neovascular tufts (P < 0.001), as compared with WT controls. Corresponding to the observed inhibition of retinal angiogenesis, ephrinB2(lacZ/+) retinas displayed an increased number of blind-ended capillary sprout tips (P < 0.02) and endothelial filopodial processes (P = 0.001). In WT and ephrinB2(lacZ/+) OIR-exposed retinas, ephrinB was confined to endothelial cells, with expression detected along angiogenic vascular processes including neovascular tufts and blind-ended capillary sprouts. Conclusions. EphrinB2 reverse signaling is a regulator of key processes during retinal vascularization and controls pathologic retinal angiogenesis through direct effects on capillary sprouting and endothelial filopodia formation.     

The role of Fas-FasL in the development and treatment of ischemic retinopathy

PURPOSE: Define a role for Fas-FasL in oxygen-induced retinopathy and explore the mechanism of pigment-epithelium-derived growth factor (PEDF) inhibition in this model. METHODS: Seven-day-old mice C57BL/6J (B6), FasL-defective (B6-gld), or Fas-defective (B6-lpr) mice were exposed to 75% oxygen for 5 days (postnatal day [P]7-P12) and returned to room air. On day P17, vascular architecture was assessed microscopically after perfusion with FITC-dextran, and preretinal nuclei were quantified by PAS and hematoxylin staining. In some experiments, mice were treated intraperitoneally with PEDF. Vascular architecture and preretinal nuclei counts were compared with those in PBS-treated control animals. RESULTS: Oxygen-induced retinopathy was significantly increased in FasL-defective gld mice compared with wild-type B6 animals. This was manifested by an increase in the number of microaneurysms, neovascular tufts, and preretinal nuclei. PEDF treatment prevented retinopathy in B6, B6-gld, and B6-lpr mice. CONCLUSIONS: Fas-FasL interactions regulate the extent of oxygen-induced retinal neovascularization. The inhibition of neovascularization in B6 gld, and B6-lpr mice by PEDF suggests that Fas-FasL interactions are probably not the mechanism for inhibition in this model.     

Intravitreal injection of the heparin analog 5-amino-2-naphthalenesulfonate reduces retinal neovascularization in mice

The effect of the heparin analog 5-amino-2-naphthalenesulfonate (5-amino-2-NMS) on retinal neovascularization was investigated in the mouse model for oxygen-induced retinopathy (OIR). From postnatal day 7 (P7) until P12, mice were kept in a 75% oxygen environment. On P12, they received an intravitreal injection of 10mM 5-amino-2-NMS in one eye and PBS as control substance in the fellow eye. The animals were intracardially perfused with fluorescein-dextran solution on P17. Retinal whole mounts were prepared and ischemic retinopathy was evaluated in 30 animals using a standardized retinopathy score. A single intravitreal injection of 5-amino-2-NMS reduces significantly angioproliferative changes (blood vessel tufts, extra-retinal neovascularization, and blood vessel tortuosity) compared to the contralateral control eye (p=0.025). The median retinopathy score (maximal 13) for the 5-amino-2-NMS treated eyes was 6 versus 8 for the control eyes. 5-Amino-2-NMS binds to the heparin-binding site of FGF1 and FGF2 and thus may be a promising substance for the local treatment of retinal neovascularization.     

内皮抑素抑制氧致视网膜病变小鼠 新生血管形成的实验研究

目的观察血管生成抑制因子内皮抑素（ES）对视网膜新生血管形成的抑制作用。方法将鼠龄为7 d的32只C57BL/6J小鼠随机分为给氧组（12只）、ES治疗组（12只）和对照组（8只）。将给氧组和ES治疗组小鼠置于浓度为（75±5）％高氧环境中生活5 d，然后回到正常氧环境中。ES治疗组小鼠在出氧箱后12、36 h，一只眼玻璃体腔内注射1μgES，另一只眼注射1 μl的磷酸盐缓冲溶液（PBS）作为对照。对照组小鼠生活在正常氧环境中。右旋糖苷-异硫氰酸荧光素（FITC-dextran）视网膜造影整装铺片了解视网膜新生血管改变；计数突破内界膜的内皮细胞数反映视网膜血管增生情况，观察ES对视网膜新生血管形成的抑制作用。结果与给氧组相比，ES治疗组视网膜铺片见视网膜血管分支走行正常，未见明显的无灌注区；ES治疗眼平均每个视网膜切面可见突破内界膜的内皮细胞核数减少，为（5.39±1.52）个，与给氧组［（22.56±2.13）个］比较，差异有统计学意义（P<0.001）。结论ES可有效抑制视网膜新生血管的形成，有望成为治疗血管增生性视网膜病变的新途径。(中华眼底病杂志,2005,21:314-317)     

建立可定量的视网膜新生血管小鼠模型的探索

目的:建立可对视网膜新生血管进行定量研究的动物模型,为研究视网膜新生血管发生机制及治疗提供实验基础。方法:将32只7d龄C57BL/6J幼新生鼠在浓度为(75±2)%的高氧状态下生活5d,然后回到正常氧环境下,作为氧诱导模型组;另32只同日龄新生鼠置于正常空气环境中生活,作为正常对照组。采用荧光素血管灌注法视网膜铺片及作视网膜冰冻切片GSA(griffoniasimplicifolialectinB4)染色,了解视网膜血管的改变,定量计算新生鼠视网膜新生血管面积。结果:实验组新生鼠的视网膜血管在高氧中生长5d后,视网膜血管收缩、闭塞,出现大片无灌注区;回到正常空气环境下2d后,开始出现新生血管;回到空气中5d后,新生血管达到高峰。结论:该动物模型可重复性高,并可进行定量研究,是进行视网膜新生血管发生机制和药物治疗的合适模型。     

The blood pressure-induced diameter response of retinal arterioles decreases with increasing diabetic maculopathy

Background The aim of the study was to compare the diameter response of retinal arterioles and retinal thickness in patients with different stages of diabetic maculopathy during an increase in the arterial blood pressure.Methods Four groups each consisting of 19 individuals were studied. Group A consisted of normal individuals and groups B–D consisted of type 2 diabetic patients matched for diabetes duration, age, and gender, and characterized by: Group B no retinopathy, Group C mild retinopathy, Group D maculopathy not requiring laser treatment. The diameter changes of a large retinal arteriole were measured using the Retinal Vessel Analyzer (RVA, Imedos, Germany) before, during, and after an increase in the blood pressure induced by isometric exercise. Additionally, the retinal thickness was measured using optical coherence tomography scanning.Results The arterioles contracted during isometric exercise in normal persons (diameter response: −0.70±0.48%) and in patients with no retinopathy (−1.15±0.44%), but dilated in patients with mild retinopathy (0.41±0.49%) and diabetic maculopathy (0.54±0.44%), p=0.01. Retinal thickness was normal in Group A (260±5.0 μm), Group B (257±4.5 μm), and Group C (253±4.4 μm), but was significantly (p=0.006) increased in Group D (279±5.3 μm).Conclusions The diameter response was reduced in type 2 diabetic patients with retinopathy, whereas retinal thickness was increased in patients with diabetic maculopathy. This suggests that impairment of diameter response in retinal arterioles precedes the development of diabetic macular edema.     

Ophthalmodynamometry and ischemic ophthalmopathy

Background To report on the clinical application of a modified ophthalmodynamometer for the detection of ischemic ophthalmopathy. Methods A 70-year-old patient showed unilateral loss of vision to 1/20, thin retinal arteries, tiny intraretinal hemorrhages, and iris neovascularization. We performed a modified ophthalmodynamometry using a Goldmann contact lens in the holding grip of which a pressure sensor was incorporated. Results Ophthalmodynamometry showed that the diastolic central retinal artery pressure was significantly (p < 0.001) lower in the affected eye than in the contralateral eye (14.6 ± 2.2 arbitrary units versus 45.5 ± 5.1 arbitrary units). These ophthalmodynamometric measurements of both eyes were significantly (p < 0.05) lower than in a control group (73.8  ± 6.2 arbitrary units) consisting of 149 normal eyes. Doppler sonography eventually revealed a marked stenosis of the right internal carotid artery, consistent with the diagnosis of a unilateral ischemic ophthalmopathy. Conclusions Ophthalmodynamometry is a helpful additional tool in the assessment of the oculo-afferent and cerebroafferent vessels in patients with symptomatic ocular ischemia.     

精氨酸-谷氨酰胺在幼鼠早产儿视网膜病变模型中的应用(英文)

目的:观察精氨酸-谷氨酰胺(Arg-Gln)对早产儿视网膜病变动物模型视网膜新生血管的抑制作用。方法:48只7日龄的C57BL/6J新生鼠暴露在750mL/L高氧环境中5d,然后回到正常空气中建立早产儿视网膜病变的动物模型。在鼠龄12d时实验组(36只)新生鼠每天两次腹腔注射Arg-Gln(剂量分别为1.0,3.0,5.0g/kg,每组12只),连续注射5d;对照组(12只)每天两次腹腔注射PBS,连续5d。所有小鼠均于17d处死,视网膜铺片,ADP酶染色观察视网膜血管情况。HE染色,在光学显微镜下观察并计数突破视网膜内界膜的血管内皮细胞细胞核数目。Real-time RT-PCR方法测量每组视网膜VEGF mRNA水平。结果:与对照组相比,实验组以剂量依赖方式无灌注区面积和新生血管团逐渐减少;实验组中最大剂量组[5.0g/(kg·d)]突破内界膜的内皮细胞细胞核数目比对照组大约减少75%(P<0.01);实验组视网膜VEGF mRNA水平与对照组相比明显下降。结论:Arg-Gln能够有效抑制早产儿视网膜病变动物模型视网膜新生血管的生成,可能为临床提供一种预防和治疗早产儿视网膜病变安全有效的新方法。     

氧诱导视网膜病变鼠模型血管内皮 生长因子mRNA的表达

目的分析氧诱导视网膜病变动物模型血管内皮生长因子（VEGF）基因的调节规律，阐明早产儿视网膜病变（ROP）新生血管形成的可能机制。方法将36只7 d 龄C₅₇BL/6J幼鼠暴露在（75±2）% 浓度的高氧状态下5 d，随后在正常氧环境下5 d，作为氧诱导模型组；另24只同日龄幼鼠作为正常对照组。采用荧光素血管灌注及视网膜铺片法观察视网膜血管形态；半定量逆转录-聚合酶链反应(RP-PCR)观察各组VEGF mRNA的变化。结果氧诱导模型的视网膜血管形态特征为高氧状态下表层和深层血管的中心区出现无灌注，相对低氧状态下2 d后开始出现新生血管，其部位在中周部。RF-PCR结果显示，VEGF的表达与眼内新生血管的发生存在明确的时空对应关系，即高氧状态下，VEGF mRNA转录下降，相对低氧状态下，VEGF mRNA过度转录。结论缺氧是视网膜新生血管发生的主要原因；高氧之后的相对低氧使VEGF表达增加，可能会降低ROP新生血管的发生。(中华眼底病杂志,2005,21:292-295)     

Bevacizumab (Avastin) treatment in patients with retinal angiomatous proliferation

Aim To determine the anatomical and functional outcome after injection of bevacizumab (Avastin, Genentech) in eyes with retinal angiomatous proliferation (RAP). Design Prospective interventional case series. Methods Sixteen eyes of 16 consecutive patients with visual loss due to RAP underwent intravitreal injections of 1.25 mg (0.05 ml) bevacizumab. Best corrected visual acuity testing, fluorescein and ICG-angiography as well as OCT imaging were performed at baseline and at each follow-up visit within a 3-month period. Results Mean visual acuity pre-injection was 0.68 ± 0.36 logMAR (n = 16), mean reading ability 0.58 ± 0.26 logRAD (n = 11). Far vision increased significantly by a mean of 1.7 ± 2 lines 4 weeks after the injection (p = 0.004), as did reading (0.6 ± 2.3 lines, p > 0.05). Both remained stable up to 3 months. Central retinal thickness decreased from 367 ± 112 μm (mean±SD) to 272 ± 123 μm 3 months after injection (p = 0.006). Leakage decreased angiographically in 12 eyes (75%) and remained stable in four eyes (25%). Re-injection of bevacizumab within the 3-month follow-up period was performed once in eight eyes, and twice in one eye. No adverse events were observed. Conclusion Intravitreal bevacizumab (Avastin) resulted in a reduction of leakage, intra- and subretinal fluid. An increase in visual acuity was seen already 4 weeks after first injection. However, a complete occlusion of feeder vessels could not be achieved within this 3-month period. Randomized clinical trials would be required to evaluate dose and frequency of injections and possible beneficial effects of combination therapies, as well as the long-term results.     

Diabetic patients with retinopathy show increased retinal venous oxygen saturation

Background  Longstanding diabetes mellitus results in a disturbed microcirculation. A new imaging oximeter was used to investigate the effect of this disturbance on retinal vessel oxygen saturation. Methods  The haemoglobin oxygen saturation was measured in the retinal arterioles and venules of 41 diabetic patients (65 ± 12.3 years) with mild non-proliferative through proliferative diabetic retinopathy (DR). Twelve individuals (61.3 ± 6.2 years, mean ± standard deviation) without systemic or ocular disease were investigated as controls. Measurements were taken by an imaging oximeter (oxygen module by Imedos GmbH, Jena). This technique is based on the proportionality of the oxygen saturation and ratio of the optical density of the vessel at two wavelengths (548 nm and 610 nm). Results  Whereas there were no significant differences in the arterial oxygen saturation between controls and diabetic retinopathy at any stage, the venous oxygen saturation increased in diabetic patients with the severity of the retinopathy: controls 63 ± 5%, mild non-proliferative DR 69 ± 7%, moderate non-proliferative DR 70 ± 5%, severe non-proliferative DR, 75 ± 5%, and proliferative DR 75 ± 8%. Conclusions  The increase of retinal vessel oxygen saturation in diabetic retinopathy points to a diabetic microvascular alteration. This may be due to occlusions and obliterations in the capillary bead and the formation of arterio-venous shunt vessels. On the other hand, hyperglycaemia-induced endothelial dysfunction, with subsequent suppression of the endothelial NO-synthase and disturbance of the vascular auto-regulation, may contribute to retinal tissue hypoxia. Walthard Vilser and Thomas Riemer have proprietary interests in the Vessel Map software. All primary data are under full control of the authors.     

Role of unc5b in retinal neovascularization in mice with oxygen-induced retinopathy

AIM: To explore the role of unc5b in retinal neovascularization in murine oxygen-induced retinopathy (OIR). METHODS: On postnatal 7(P7), C57BL/6J mice were exposed to 75%±2% oxygen for 5 days. On postnatal 12(P12), the mice were brought back to the room air (21% oxygen) to induce retinal neovascularization. Western blot analysis was performed to examine the temporal expression of unc5b in murine retinas. Double staining for unc5b and isolectin B4 were employed to determine the location of unc5b in murine retinas. The effect of unc5b on retinal neovascularization was evaluated by intravitreal injection of unc5b-FC in mice with OIR. Retinal neovascularization was measured by counting neovascular cell nuclei above the internal limiting membrane and by angiography of flat-mounted retinas perfused with fluorescein dextran. RESULTS: Compared to age-matched normal mice, the expression of unc5b was significantly increased in retinas of OIR mice on P17 and P21. Unc5b was apparently expressed in retinal vessels of OIR while being negative in normal retinal vessels. Retinal neovascularization in eyes injected with unc5b-FC was significantly reduced. CONCLUSION: Unc5b-FC can effectively inhibit retinal neovascularization induced by OIR. It may serve as a powerful and novel therapy for ischemia-induced retinal disease.