ObjectiveTo study the inhibitory effects of pigment epithelium derived factor (PEDF) on oxygen-induced retinal neovascularization in mice, and to investigate the possible involvement of interleukin-1β (IL-1β) in the neovascular-inhibitory function of PEDF. Methods A total of 140 postnatal day (P)7 C57BL/6 mice were randomly divided into normal control group, oxygen-induced retinopathy (OIR) model group, PEDF treatment group and PBS treatment control group. All mice except normal control group with their mothers were exposed to (75±2)% oxygen environment for 5 days and then kept in room air for another 5 days to establish the OIR model. Mice in normal control group were kept in room air only. At P12 and P14, respectively, mice in PEDF treatment group received intravitreous injections of 1 μl PEDF (2 μg/μl), while PBS treatment control group received the same volume of PBS (10 mmol/L, pH7.4).All mice were euthanized at P17 and eyes were isolated. The changes of retinal vessels were observed on retinal flat mounts and cryosections by fluorescence microscopy. Retinal specimens were prepared for IL-1β protein and mRNA analysis by Western blot and real time fluorescence quantitative reverse transcription-polymerase chain reaction (Real-time RT-PCR). ResultsChanges of retinal vessels had been viewed by fluorescence microscopy on flat-mounted retina, the relative retinal neovascularization areas were significantly increased in OIR model group compared with normal control group (t=15.02, P < 0.01), and the relative retinal neovascularization areas were obviously smaller in PEDF treatment group than those in PBS treatment control group (t=5.96, P < 0.01). Fluorescence staining revealed that retinal vascular tufts were extending from outer plexiform layer (OPL) to ganglion cell layer (GCL) of the retina along with multiple interconnections; Neovascular tufts in OIR model group and PBS treatment control group were presenting distinctly more than those of normal control group and PEDF treatment group. The specific expression levels of IL-1β protein in retinas of OIR mice by Western-blot analysis were higher than those of normal control group(t=3.35, P < 0.05), While these of PEDF treatment group showed a considerable decline in comparison with PBS treatment control group (P < 0.01), and there were no difference in normal control group and PEDF-treated group (F=11.764, P > 0.05). Similarly, expression levels of IL-1β mRNA tested by Real-time RT-PCR were obviously increased in the OIR model group when compared to normal control group(t=4.43, P < 0.01). After treated with PEDF, expression levels of IL-1β mRNA showed a considerable decrease when compared to PBS treatment control group (P < 0.01), and there were no difference in normal control group and PEDF-treated group (F=11.15, P > 0.05). ConclusionsPEDF can inhibit oxygen-induced retinal neovascularization. The mechanism may be related to that PEDF can downregulate the expression of IL-1β in retina.
Objective To observe the the inhibitory effect of recombined adenovirus mediated delivery of p21 (rAd-p21) on oxygen-induced retinal neovascularization in mice. Methods A total of 56 C57BL/6 mice at the age of seven days were divided into control group, phosphate buffer solution (PBS) group, rAd-p21 group and rAdno purpose gene control (rAd-NC) group, 14 mice in each group. The retinal neovascularization of PBS, rAd-p21and rAd-NC group were induced by oxygen, and received an intravitreal injection 1 mu;l PBS, rAd-p21 and rAd-NC at postnatal day 11, respectively.The rats of control group were not intervened. At postnatal day 17,RNV was determined by retinal flat mounts and retinal section; non-perfusion areas of retina were analyzed by Image-Pro plus 6.0 software; reverse transcription-polymerase chain reaction (RT-PCR) and Western blot was used to measure the mRNA and protein expression of p21 and CDK2. Results Compared with PBS and rAdNC groups, the retinal nonperfusion areas, neovascularization and the numbers of endothelial cell nuclei breaking through the internal limiting membrane in rAd-p21 group were reduced significantly. Nonperfusion areas of retina in rAd-p21 group was less than that in PBS and rAd-NC groups, the difference among these three groups was significantly (F=101.634，P＜0.05). Compared with the other three groups, the level of p21 mRNA and protein in rAd-p21 group increased significantly (F=839.664, 509.817;P＜0.05); the level of CDK2 mRNA and protein in rAd-p21 group decreased significantly (F=301.858, 592.882;P＜0.05). Conclusion rAd-p21can inhibit oxygen-induced retinal neovascularization, up-regulated p21 expression and down-regulated CDK2 expression may be the mechanism.
Objective To investigate the effect of suppression of ischemia-induced retinal neovascularization by VEGF antisense oligodeoxyribonucleotides. Methods Mouse models of hyperoxia-induced ischemic retinopathy were established. Retrobulbar injections were performed with VEGF antisense oligodeoxyribonucleotides or NS in 4 groups:normal control and various doses respectively. The nuclei of new vessel buds extending from the retina into the vitreous in differ ent groups were counted and compared under the light microscope. Results There were plenty of new vessel buds in the eyes of mice in hyperoxic condition., while the number of the nuclei of new vessel buds is less in the murine eyes with retrobulbar injection of VEGF antisense oligodeoxyribonucleotides,especially the nuclei were redused with 59.3% in eyes with large dose. Conclusion The proliferation of retinal new vessel may be suppressed by using the retrobulbar injection of VEGF antisense oligodeoxyribonucleotides. (Chin J Ocul Fundus Dis, 2001,17:141-143)
Since anti-vascular endothelial growth factor (VEGF) therapy has recently become the first-line treatment of wet age related macular degeneration in China, as well as retinopathy of prematurity, neovascular glaucoma and macular edema secondary to diabetic retinopathy or retinal vein occlusion in other countries. It is worth thinking about that how to perform anti-VEGF treatment properly to benefit more patients. We reviewed the fields of clinical researches to explore the best role of anti-VEGF treatment in prevention and treatment of retinal disease in future.
Objective To explore the inhibitory effects of r-k4k5 on retinal neovascularization. Methods Eighty-eight one-week-old C57BL/6J mice were put into the environment with 75% oxygen for 5 days to establish models of vascular proliferation retinopathy. One eye of each mouse received an intravitreal injection of 500 ng of r-k4k5 (large-dosage group) and of 250 ng of r-k4k5(small-dosage group), and the same volume of BSS was injected into the other eye of the mice both in these two groups as a control. The ADPase histochemical staining was used for retinal flatmount to observe changes of retinal vessels. The inhibitory effects of r-k4k5 on retinal neovascularization were evaluated by counting the endotheliocyte nuclei of new vessels extending from retina to vitreous in the tissue-slice. Results Regular distributions and reduced density of retinal blood vessels in eyes in the treatment group were found in retinal flatmount. The number of the endotheliocyte nuclei of new vessels extending from retina to vitreous was less in the eyes in the treatment group than which in control group (Plt;0.001). The nuclei of new blood vessels in the large-dosage group were less than which in small-dosage group (Plt;0.001). No histologic evidence of retinal toxicity or inflammatory response was found in the tissue-slice after the injection of r-k4k5. Conclusions Retinal neovascularization can be inhibited by intravitreal injection of r-k4k5,which suggests that intravitreal injection of r-k4k5 may have potential therapeutic benifits in retinal vascular disease. (Chin J Ocul Fundus Dis，2003，19：121-124)
Objective To observe the inhibitory effects of gene transfer of canstatin on retinal neovascularization in mice. Methods Fifty-six 7-day-old C57BL/6J mice were randomly divided into control group,oxygen-induced retinopathy (OIR) group, empty vector group and treated group,14 mices in each group. Except for the control group,the mice in the other groups were exposed to (75plusmn;2)% oxygen for 5 days and then back to the normal air to establish the model of OIR. On postnatal 12 day, the treated group was received intravitreal injection of canstatin pCMV-HA, while the empty vector group was received the same volume of empty plasmid.The changes of retinal vessels were observed by Evans blue angiography on postnatal 17 day. With parafin section which stained by hematoxylin and eosin, then the number of endotheliocyte nuclei breaking throuhgh the internal limiting membrane（ILM） was observed and counted by optical microscope.Results Retinal blood vessels distributed regularly in treated group compared with OIR group and empty vector group.The differences of the number of endotheliocyte nuclei breaking throuhgh ILM in treated group was significant compared with the other two groups（F=39.006，Plt;0.001).Conclusion The canstatin pCMV-HA can effectively inhibit the retinalneovascularization in OIR.
Objective To investigate the inhibitory effects and possible related mechanism of OTX008 [a selective inhibitor of galectin-1 (Galectin-1)] on retinal neovascularization (RNV) in mouse model of oxygen-induced retinopathy (OIR). Methods 7-day-old (P7) C57BL/6J mice were randomly (according to random number table) divided into 4 groups including normal group, OIR group, OIR-OTX008 group and OIR-phosphate buffered saline (PBS) group. To establish the OIR mouse model, mice from all groups except normal group were expose to (75±2)% oxygen for 5 days and then to room air. OIR-OTX008 group received an intravitreal injection of 1 μl (0.25 μg/μl) OTX008 at P12, OIR-PBS group received the equal volume (1 μl) of PBS injection. Mice from 4 groups were euthanized at P17, and retinas were collected for molecular biological analysis and morphological study. RNV was evaluated by counting the number of pre-retinal neovascular nuclei and the whole-mount immunofluorescent staining of mouse retina. Cyrosections of retinas were imaged via confocal microscopy to observe the enrichment of staining of Galectin-1. Protein levels of Galectin-1, Neuropilin-1 and phosphorylation of vascular endothelial growth factor receptor 2 (pVEGFR2) were determined with Western blot. Results At P17, Galectin-1 expressed higher in retinal ganglion cell layer, inner plexiform layer and inner nuclear layer from OIR group and OIR-PBS group than normal group. Galectin-1 expressed less in cryosection retinas from OIR-OTX008 group than OIR group and OIR-PBS group. The numbers of pre-retinal neovascular cell nuclei from OIR group and OIR-PBS group were obviously more than that from normal group (t=9.314,P＜0.05). The number of pre-retinal neovascular cell nuclei from OIR-OTX008 group were obviously lower than those from OIR group and OIR-PBS group (t=8.038, 7.774;P＜0.05). The RNV tufts area (t=13.250, 12.570), non-perfusion area (t=15.590, 12.430) and hypoxic area (t=9.542, 9.928) from OIR-OTX008 group were significantly smaller than those in OIR group and OIR-PBS group (P＜0.05). Protein levels of Galectin-1 (t=24.800, 23.060), Neuropilin-1 (t=4.120, 3.530) and pVEGFR2 (t=25.880, 15.480) in the OIR-OTX008 group were significantly down-regulated than those from OIR group and OIR-PBS group (P＜0.05). Conclusion Intravitreal injection of OTX008 inhibits RNV and ameliorates retinal hypoxia in mice model of OIR possibly through down-regulating Galectin-1, Neurolinpin-1 and pVEGFR2.
Purpose To evaluate the efficacy of vitreous surgery and endolaser in a series of patients with retinal vein occlusion(RVO)with vitreous hemorrhage,neovascular membranes(NVM) and/or traction retinal detachment(TRD). Methods Clinical records were reviewed on 37 consecutive patients(38 eyes)who underwent vitreous surgery and endolaser for RVO with persistent vitreous hemorrhage,NVM and/or TRD.There were 19 patients(20 eyes)with retinal branch vein occlusion (BRVO)and 18 patients(18 eyes)with central retinal vein occlusion(CRVO). Results NVM and TRD were confirmed during operation in 27 and 23 eyes,respectively.Visual acuity improved postoperatively in 34 eyes(89.5%)including 22 eyes with 0.1 or better vision,and 4 eyes remained unchanged.CRVO group had longer history and less visual improvement after surgery. Conclusions Vitreous surgery and endolaser photocoagulation can improve the outcome in the majority of patients with RVO with vitreous hemorrage,NVM and/or TRD. (Chin J Ocul Fundus Dis,1998,14:3-6)