Intravitreal injection of anti-vascular endothelial growth factor (VEGF) agents is a first line therapy for neovascular age-related macular degeneration (nAMD).However,how to predict the respond for antiVEGF treatment is still a challenge in clinic practice.Optical coherence tomography angiography (OCTA) can offer dynamic following-up for choroidal neovascularization (CNV) in nAMD after antiangiogenic therapy.The change of vascular morphology based on OCTA enriches the novel theory of prognosis for tveatment of nAMD and can be considered as a biomarker of neovascularization in vivo,which can help us to evaluate the activity of CNV and understand the mechanism of anti-VEGF resistance.So,OCTA should be a standard strategy during the diagnosis,treatment and follow-up of nAMD.We should pay more attention to the guiding significance in the prognosis evaluation of nAMD basis on character of vascular morphology by OCTA following antiangiogenic therapy.

Objective To investigate the inhibiting effect of CGP77675 (CGP),a Src inhibitor,on epithelial-mesenchymal transition (EMT) of human retinal pigment epithelial (RPE) cells induced by transformation growth factor-β1 (TGF-β1).Methods Human RPE cell line (ARPE19 cells) was cultured in vitro and divided into control group,TGF-β1 group and TGF-β1 +CGP group.Corresponding agent was added into culture medium based on grouping.The morphology of the cells were examined under the optical microscope 3 days after culture.The expressions of EMT-related genes and proteins in the cells were detected by real-time quantitative PCR and Western blot,respectively,including fibronectin 1 (FN 1),and plasminogen activation inhibitor 1 (PAI1),and the expressions of zonula occludens protein 1 (ZO1) and cytoskeleton protein filamentous actin (F-actin) were detected by immunofluorescence staining.MTT assay was employed to evaluate the cell proliferation rate.The migration distance of the cells was measured by scratch test.Results The ARPE19 cells in the control group showed an epithelial-like morphology and F-actin and ZO-1 were expressed along cell membrane.In the TGF-β1 group,the cells appeared to be fibrous-like,and the fluorescence staining of F-actin was disordered and ZO-1 was discontinuous on the cell membrane.The cells in the TGF-β1 +CGP group remained to be an epithelial-like in shape with clear and complete expressions of F-actin and ZO-1.The relative expressions of FN1 mRNA and PAI1 mRNA in the cells were 0.211 ± 0.080 and 0.116±0.073,1.000±0.001 and 1.000±0.001,0.368±0.097 and 0.362±0.048 in the control group,TGF-β1 group and TGF-β1 +CGP groups,showing significant differences among the groups (F=33.14,82.92;both at P＜0.01),with the highest expressions ofFN1 mRNA and PAI1 mRNA in the TGF-β1 group (all at P＜0.05).The relative expressions of FN1 and PAI1 proteins were 0.166±0.055 and 0.327±0.066,1.000±0.001 and 1.000± 0.001,0.143 ± 0.030 and 0.260 ± 0.077 in the control group,TGF-β1 group and TGF-β1 + CGP group,with significant differences among three groups (F=181.90,48.85;both at P＜0.01),and the expressions FN1 and PAI1 proteins were significantly higher in the TGF-β1 than those in the control group and TGF-β1 +CGP group (all at P＜0.05).The cell proliferative rate in the TGF-β1+CGP group was (79.30±3.44) ％ and (54.80±7.39) ％ at the third day and seventh day after culture,which were significantly reduced in comparison with (99.50 ± 1.00)％ and (99.10±0.50)％ in the control group as well as (95.10±4.20)％ and (92.10±4.50)％ in the TGF-β1 group (all at P＜0.05).The migration distance was disappeared in the TGF-β1 group,and the scratch width was not obviously changed in the TGF-β1 +CGP group.Conclusions Src inhibitor can inhibit EMT process of ARPE19 cells induced by TGF-β1,indicating that Src signaling pathway may play a critical role in EMT of RPE cells.

Objective To explore whether brimonidine has a protective effect on retinal ganglion cells (RGCs) through improving mitochondrial function under the oxidative stress.Methods Mouse RGC-5 cells were cultured in DMEM medium containing low concentration of glucose (1 g/L),10％ fetal bovine serum and 100 U/ml penicillin-streptomycin solution.The cells were divided into normal control group,H2O2-treated group and brimonidine+ H2O2 group.H2O2 at the concentration of 800 μmol/L was added into the medium in the H2O2-treated group,and 1 μmol/L brimonidine was added into the medium for 2 hours prior to the addition of H2O2 in the brimonidine+H2O2 group.The cells were sequently cultured for 24 hours.The morphology of the cell nucleus was examined by Hoechst fluorscence staining.The expressions of apoptosis-related protein in the cells were detected by Western blot assay.Mitochondrial membrane potential was assessed by JC-1 staining.Results The cell nuclei showed round or oval in shape with consistent size in the normal control group.The pycnosis and karyorrhexis of the cell nuclei were seen in the H2O2-treated group,and less abnormal nuclei were found in the brimonidine+H2O2 group.The relative expression level of bcl-2 protein in the cells was 0.76±0.15,0.50±0.13 and 0.75±0.17 in the normal control group,H2O2-treated group and brimonidine + H2O2 group,respectively,and the expression of bcl-2 protein in the H2O2-treated group was significantly lower than that in the normal control group and brimonidine+H2O2 group (both at P＜0.05).The relative expression level of bax protein in the cells was 0.65±0.13,0.83±0.07 and 0.70±0.10 in the normal control group,H2O2-treated group and brimonidine+H2O2 group,respectively,and the expression of bax protein in the H2O2-treated group was significantly higher than that in the normal control group and brimonidine+H2O2 group (both at P＜0.05).A strong orange fluorescence was seen in the mitochondrial membrane of RGC-5 in the normal control group with a coexpression with the green fluorescence of cell membrane.In the H2O2-treated group,the orange fluorescence intensity in the cells was evidently weakened,and the number of JC-1 responsed cells was considerably increased and the orange fluorescence intensity was enhanced in the brimonidine + H2O2 group.Conclusions Brimonidine can prevent RGCs from oxidative-stress damage by improving the mitochondrial function and therefore play a potential neuroprotective effect on optic nerve.

Objective To investigate whether vascular endothelial cells in choroidal neovascularization whether hypoxia condition can up-regulate SNAI1 and activate matrix metalloproteinase (MMP)2 and MMP9 therefore to participate in choroidal neovascularization(CNV).Methods Sixteen SPF male C57 mice aged 6-8 weeks were divided into control group and model group.CNV models were induced by retinal laser photocoagulation,and flatmount and frozen sections of retinal pigment epithelium (RPE)-choroid-sclera compound were prepared at 7 days after modeling.The CNV in flat-mount was examined by Isolectin B4 staining,and the location of SNAI1,MMP2 and MMP9 in frozen sections was determined by immunofluorescence technology.The expression of SNAI1,MMP2 and MMP9 at mRNA level in CNV was detected by real-time fluorescence quantitative PCR (real-time PCR).The use and care of experimental animals complied with Statement for the Use of Animals in Ophthalmic and Visual Research.The RF/6A cells were divided into normoxia group and hypoxia group and cultured for 24 hours in 5％ CO2condition and mix condition of 94％ N2,5％ CO2 and 1％ O2,respectively.The expression of SNAI1,MMP2 and MMP9 in the cells at mRNA and protein levels was detected by real-time PCR and Western blot assay,respectively.Small interfering RNA of SNAI1 (siSNAI1) was transfected into the cells,and then the expression of MMP2 in the cells at mRNA and protein levels was detected by real-time PCR and Western blot assay,respectively,and the migrating number of the cells was assayed by Transwell chamber assay.Results CD31 and SNAI1 positive-response cells were seen in RPE-choroid-sclera flat-mounts under the laser scanning confocal microscope.The relative expression levels of SNAI1mRNA and MMP2 mRNA in RPE-choroid-sclera tissues were higher in the model group than those in the control group (SNAI1 mRNA:1.291 ±0.060 vs.0.759±0.074,P =0.001;MMP2 mRNA:1.610±0.424 vs.0.772 ±0.080,P =0.044).The expression of MMP9 mRNA was not significantly elevated between model group and control group (P＞0.05).The relative expression level of MMP2 mRNA was higher in comparison with MMP9 mRNA in the model group (P＜0.01).The relative expressions of hypoxic induced factor 1α (HIF-1α),SNAI1 and MMP2 at mRNA level and protein level in RF/6A cells were significantly higher in the hypoxia group than those in the normoxia group (all at P＜0.05) and no considerable difference was seen in MMP9 mRNA expression between the two groups (P＞0.05).The relative expressions of MMP2 mRNA in the cells were 0.217±0.036 and 0.818±0.105,and those of MMP2 protein in the cells were 0.236±0.009 and 1.043±0.120 in the hypoxia+siSNAI1 group and only hypoxia group,respectively,with significant differences between them (P =0.002,0.003).The migrating number of the cells was (254.60 ±71.31)/field in the hypoxia+siSNAI1 group,which was significantly less than (534.10±96.21) /field in the control group (P =0.029).Conclusions The hypoxic environment at CNV can activate MMP2 by up-regulating the expression of SNAI1,which promotes the migration of vascular endothelial cells and therefore participates in CNV formation,and the intervention of SNAI1 activation under the hypoxic condition can inhibit this process.

Objective To establish a central serous chorioretinopathy (CSC) model on different strains of rabbits by intravenous injection of adrenaline,which may contribute to related researches of CSC.Methods This study was approved by Bioethics Committee of Fourth Military University and complied with Statement for the Use of Animals in Ophthalmic and Visual Research.Fundus fluorescein angiography (FFA) was initially performed on male New Zealand white rabbits (10),Belgium rabbits (5) and Chinchilla rabbits (10) to make sure that the retinas of subjects were normal.For the New Zealand white rabbits,adrenaline was injected via ear vein at a dose of 0.04 mg/kg once per day for the first 8 weeks and followed by a dose of 0.08 mg/kg for the next 4 weeks,while 0.04 mg/kg adrenaline was injected in the same way for 8 weeks in the Belgium rabbits and Chinchilla rabbits.FFA was performed every week after injection of adrenaline to evaluate the fluorescence leakage in ocular fundus.New Zealand white rabbits were sacrificed in 4 (3 rabbits),8 (3 rabbits) and 12 weeks (4 rabbits) after injection respectively,and Belgium rabbits and Chinchilla rabbits were sacrificed in the 8 weeks after injection.The eyeballs of the rabbits were enucleated to prepare the retinal sections for histopathological examination after hematoxylin-eosin staining.The results of FFA and retinal structure were compared among different strains of rabbits.Results No fluorescence leakage was found by FFA in ocular fundus,and the retinal structure was normal in all the 10 New Zealand white rabbits during the experiment.Fluorescence leakage was found by FFA in 2 Belgium rabbits at 1 week and 2 weeks after injection respectively,and retinal detachment and depigmentation of retinal pigment epithelium (RPE) with an enlarged intercellular space were shown by hematoxylin-eosin staining.For the Chinchilla rabbits,fluorescence leakages were found in 7 rabbits throughout the whole period of adrenaline administration.Circumscribed retinal detachment,depigmentation of RPE with enlarged intercellular space were also found in leakage lesions.Conclusions Repeated intravenous injection of adrenaline can induce CSC-like lesions in colored rabbits but not in albino rabbits.

Objective To investigate the effect of Genistein,a tyrosinase inhibitor,on retinal neovascularization in mice.Methods Thirty-six 7-day-old C57BL/6J mice were randomly assigned into hyperoxiainduced group,Genistein group,DMSO group and normal control group.The mice in the hyperoxia-induced group,Genistein group and DMSO group were fed in a static chamber with the oxygen volume fraction (75±2)％ for 5 days and then sent back to natural environment for 5 days to establish retinal neovascular models,and 1 μl Genistein diluted by 5％ dimethyl sulfoxide (DMSO) (400 mg/L) and 1 μl DMSO were intravitreally injected in the 12-dayold mice of Genistein group and DMSO group,respectively.The mice in the normal control group were bred in natural environment.The fluorescence angiography was carried out in 17-day-old mice (2 mice in each group) to prepare the whole retinal flatmounts,and the morphology of retinal vessels was observed under the fluorescence microscope.The other mice in various groups were sacrificed and the retinas were collected.The expressions of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) in mRNA and protein levels were detected by realtime fluorescence quantitative PCR and Western blot,respectively.The use and care of the mice complied with regulations for the management of laboratory animals.Results Retinal vessels were normal in the mice of normal control group.In the mice of the hyperoxia-induced group and DMSO group,retinal vessels were tortuous,and neovacularization and non-perfusion areas were visible.In the Genistein group,retinal vessels were clearly visible,but non-perfusion areas were exhibited.The relative expression levels of VEGF mRNA in retinas were 0.64±0.25,0.37±0.23,0.03±0.02 and 0.04±0.02,and the relative expression levels of bFGF mRNA in retinas were 21.40±3.07,17.22±2.63,0.52±0.25 and 0.67±0.23,in the hyperoxia-induced group,DMSO group,Genistein group and normal control group,and the expressions of VEGF and bFGF in mRNA level in the hyperoxia-induced group and DMSO group were significantly higher than those in the Genistein group and normal control group (all at P＜0.05).The protein expression levels of VEGF and bFGF were 1.01 ±0.05 and 0.97±0.06 in the hyperoxia-induced group,1.06±0.07 and 1.03±0.08 in the DMSO group,0.73±0.05 and 0.76±0.07 in the Genistein group,0.52±0.05 and 0.56± 0.05 in the normal control group.The expressions of VEGF and bFGF in protein level in the hyperoxia-induced group and DMSO group were significantly higher than those in the Genistein group and normal control group (all at P＜0.05).Conclusions Genistein can inhibit hyperoxia-induced retinal neovascularization may be by downregulating the expressions of VEGF and bFGF in retinas.

Objective This study was to investigate the role of recombinant human platelet derived growth factor-BB(rhPDGF-BB) in the proliferation and migration of human retinal vascular endothelial cells (hRVECs).Methods hRVECs were cultured in DMEM with 10％ fetal bovine serum.The rhPDGF-BB at the concentrations of 10,50 and 200 ng/ml were added into the medium of exponential phase-growth cells for 24 and 48 hours,respectively,and no rhPDGF-BB was added in the normal control group.The proliferation of the cells (absorbancy) was assayed by cell counting kit 8 (CCK8) method.Cell scratch test was employed to evaluate the relative migration area of cells (migrated acellular area/initial acellular area).The relative expression of rhPDGF-BB recepter (rhPDGF-BBR) mRNA in the cells was detected by reverse transcription PCR.The relative expression of VEGF mRNA and integrin mRNA in the cells was detected using real-time fluorescence quantitative PCR.Results hRVECs grew well and a expressing band according with rhPDGF-BBR prime was displayed.The absorbancy values of thecells were 1.01±0.05,1.09±0.04,1.10±0.02 and 1.13±0.05 in the normal control group and 10,50,200 ng/ml rhPDGF-BB groups at 24 hours after culture,and those in the 10,50 and 200 ng/ml rhPDGF-BB groups were significantly increased in comparison with the normal control group (t =2.504,3.430,3.483,all at P＜0.05).The relative migrated areas of the cells in the normal control group and 10,50,200 ng/ml rhPDGF-BB groups were 0.42±0.10,0.38±0.09,0.55±0.06 and 0.61±0.05 at 24 hours after culture,and those at 48 hours were 0.75±0.06,0.81 ±0.02,0.87±0.02 and 0.98±0.02,showing significant differences among the groups (Fgroup =16.283,P =0.000;Ftime =209.129,P=0.000),and the relative migrated areas was depended upon the rhPDGF-BB dose and time.The relative expressions of integrin mRNA were 1.06 ± 0.02,1.30 ±0.10,1.20 ± 0.16 and 1.27 ± 0.08,and those of VEGF mRNA were 0.97±0.05,1.06±0.16,1.58 ±0.18 and 1.66 ±0.21 in the normal control group and 10,50 ng/ml,200 ng/ml rhPDGF-BB groups,respectively,and increased expressions of integrin mRNA and VEGF mRNA were found in the 50 and 200 ng/ml rhPDGF-BB groups compared with the normal control group (integrin mRNA:t =3.900,4.014,both at P ＜ 0.05;VEGF mRNA:t =6.940,7.210,both at P ＜ 0.05).Conclusions rhPDGF-BB/rhPDGF-BBR signal promotes the proliferation and migration of hRVECs probably by up-regulating the expressions of integrin and VEGF.

Objective To develop a remote diabetic retinopathy (DR) screening system and to evaluate the effectiveness of the screening system in community.Methods A cross-sectional study was carried out under the informed consent of subjects in Peking University People's Hospital and Beijing Xicheng District Desheng Community Health Service Center from June 2015 to December 2016.A remote DR screening system was established in Peking University People's Hospital and Beijing Xicheng District Desheng Community Health Service Center during June 2015 to December 2016.Based on non-mydriatic digital eye fundus camera photography and the internet transmission technology,anterior ocular segment and fundus images of 2 473 eyes from 1 355 community subjects with type 2 diabetes mellitus were transmitted from Beijing Xicheng District Desheng Community Health Service Center to the reading center of Peking University People's Hospital,and the results were provided to the subjects after analysis,including visual examination,diagnosis and follow-up rate of the subjects,the agreement between remote screening system and conventional screening method was analyzed and compared.Results The visual acuities of the 2 473 eyes of 1 355 subjects were obtained by trained community physician,and the visual acuity was ≤0.05 in 103 eyes (4.2％),＞0.05-0.3 in 780 eyes (31.5％),＞0.3 in 1 590 eyes (64.3％).A good consistency was found in the diagnosis and grading of DR (Kappa value =0.895) and in diagnosis of macular disorder (Kappa value =0.763)between the remote screening system and conventional screening method.In addition,the diagnosis results of retinal photocoagulation were consistent between the two methods (Kappa value =1.000).The mean duration of the remote screening system for one subject was 10 minutes,which was shorter than 23 minutes of conventional screening method.The follow-up rate of remote screening system was 75.2％.Conclusions There is a high consistency in the DR diagnosis and evaluation between the remote non-mydriatic screening system and conventional screening method.The screening program with follow-up requests has a satisfying follow-up rate,which could meet the demand of DR screening.

Objective To observe the clinical characteristics of multi-mode images of retinal arterial macroaneurysms and provide reference for the accurate diagnostic.Methods The clinic data of 24 patients (25 eyes) with retinal arterial macroaneurysms who were diagnosed in the First Affiliated Hospital of Zhengzhou University from August 2012 to May 2016 were retrospectively analyzed.All patients received ophthalmologic examinations including visual acuity,fundus photography,and fundus fluorescein angiography (FFA).The patients who could not be diagnosed by fundus photography and FFA underwent indocyanine green angiography (ICGA) and spectral-domain optical coherence tomography (SD-OCT) examinations.The visual acuity of the three types of retinal arterial macroaneurysms and the diameters of retinal arterial macroaneurysms based on FFA images was analyzed.Results A single macroaneurysm appeared in all the 25 eyes.Retinal arterial macroaneurysms of 22 eyes were on temporal artery branches and those of 3 eyes were on the nasal artery branches.Sixteen retinal arterial macroaneurysms were determined as hemorrhagic type,2 were exudative type and the other 7 were quiescent type.The difference of vision acuity in the three types of retinal arterial macroaneurysms was significantly different (x2=15.117,P=0.001).Retinal arterial macroaneurysms of 20 eyes could be clearly exhibited by FFA,and the retinal arterial macroaneurysms in other 5 eyes which were concealed due to bleeding were displayed by ICGA and OCT.The average diameter of retinal arterial macroaneurysms and normal arteries were (330.65±43.09)μm and (134.70±10.74)μm,respectively,showing a significant difference (t =21.034,P =0.000) and a positive correlation between them (r =0.867,P=0.000).Conclusions Most retinal artcrial macroaneurysms can be diagnosed by FFA,moreover,both ICGA and OCT can provide necessary supplement for the concealed retinal arterial macroaneurysms.The larger the diameter of the normal artery is,the larger the diameter of the corresponding retinal arterial macroaneurysms is.The typing of retinal arterial macroaneurysms can offer basis for the evaluation of management.

Objective To evaluate the efficacy and safety of 27-gauge sutureless vitrectomy with air tamponade for rhegmatogenous retinal detachment (RRD).Methods The clinical data of 35 consecutive eyes with primary RRD from 35 patients who received 27-gauge vitrectomy with intraocular air tamponade in Zhongshan Eye Center from April 2016 to January 2017 were retrospectively analyzed.The mean follow-up duration was 8.6 months.Best corrected visual acuity (BCVA) (LogMAR) and intraocular pressure (IOP) were examined before surgery,1 week and 3 months after surgery.The operative duration,sclerotomy sites,retinal reattachment rate,intraoperative and postoperative complications were recorded.Results The mean duration of vitreous removal was (15.3 ± 3.6) minutes,and the mean duration of operation was (34.5 ± 4.8) minutes.No suturing process was performed at sclerotomy sites in all eyes.The retinal reattachment rate following a single procedure was 100％.The mean BCVA was significantly different among before surgery,1 week and 3 months after surgery (F =64.12,P＜0.01),and the BCVA at 1 week and 3 months after surgery was evidently improved in comparison with before surgery (0.82±0.31 vs.1.01 ±0.40;0.68±0.30 vs.1.01 ±0.40) (both at P＜0.05).The mean IOP was (14.69±3.66),(17.37±2.32) and (16.69±2.45) mmHg (1 mmHg =0.133 kPa) before surgery,1 week and 3 months after surgery,showing a significant difference among them (F=14.82,P＜0.01),and the IOP 1 week and 3 months after surgery was evidently higher than that before surgery (both at P＜0.05).The complications included intraoperative iatrogenic retinal breaks in 2 eyes,postoperative hypotony in 1 eye and hypertension in 5 eyes.These complications were curable.Conclusions 27-Gauge vitrectomy and air tamponade for RRD is an effective and safe approach.