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Objective:To analyze the characteristics of eyes with congenital optic disc pits (ODPs) through multimodal imaging.Methods:A cross-sectional study was conducted.Thirty-eight patients (38 eyes) diagnosed with congenital ODPs in the Second Hospital of Hebei Medical University from January 2009 to January 2020 were enrolled.A comprehensive summary analysis of the imaging results including fundus photography, spectral domain-optical coherence tomography (SD-OCT), infrared imaging, fundus autofluorescence (FAF), fluorescein fundus angiography (FFA) and indocyanine green angiography (ICGA) was performed.This study protocol adhered to the Declaration of Helsinki and was approved by an Ethics Committee of The Second Hospital of Hebei Medical University (No.2021-P011). Written informed consent was obtained from each subject prior to any medical examination.Results:Among the 38 eyes, there were 32 eyes with ODPs located in or below the temporal side of optic disc, 4 eyes with ODPs located above the temporal side of optic disc, and 2 eyes with ODPs located at the center of optic disc, which were round or quasi-circular pale depression, and dark red eminences with clear or unclear boundaries between milk spots were found in 29 eyes with optical-disc macular degeneration (ODP-M) by fundus photography.SD-OCT examination showed that the structure of lamina cribrosa in the lesion area in all ODPs patients was incomplete, which presented a dark area with no tissue reflection, and the fissure led to the deep optic nerve.Fluid was found in the outer nuclear layer in all ODP-M patients, and there were 27 eyes with fluid in the inner nuclear layer, 13 eyes in the ganglion cell layer, and 4 eyes under the inner limiting membrane.Among the 29 eyes with ODP-M, there were 21 eyes with retinoschisis in outer layer, 27 eyes with neuroepithelial detachment.In the 27 eyes with neuroepithelial detachment, spot-like high reflection and reduced or disappeared ellipsoid band reflectance were seen above the neuroepithelium in 18 eyes.In infrared images, there were circular or quasi-circular low-reflection areas in the temporal side of the optic disc, and the lesion of ODP-M eyes presented low-reflection areas.FAF examination showed that in 27 eyes with ODP-M, there was a hypofluorescent region at the posterior pole consistent with the lesion range, among which, there was a granular or sheet-like hyperfluorescence at the center of the hypofluorescent region in 18 eyes.FFA showed that the optic disc depression in the arterial phase of patients was in a localized hypofluorescence state.During the venous phase, fluorescein dye extravasation along the temporal side of the optic disc could be found.A strong fluorescent arc with unclear boundaries at the temporal edge of the optic disc was formed in the late stage of angiography.Among the 29 eyes with ODP-M, the area of the macular lesion showed hyperfluorescence during the late stage of angiography in 27 eyes with neuroepithelial detachment, and no extension of dye toward the macula was found.ICGA showed that the optic disc depression of ODPs patients presented a localized hypofluorescence, and the lesion showed hyperfluorescence in 27 of the 29 ODP-M eyes with neuroepithelial detachment.Conclusions:Multimodal imaging can be helpful to realize the early diagnosis, etiology analysis of ODPs and make treatment plan.
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Objective To observe the expression ofRapl,guanosine triphosphate-Rapl (GTP-Rapl),vascular endothelial growth factor (VEGF) and β-catenin in experimental choroidal neovascularization (CNV).Methods Forty-two brown Norwegian rats were randomly divided into a blank control group (7 rats) and a model group (35 rats).Both eyes were enrolled.The CNV model was established by holmium ion laser photocoagulation in the model group.At 3,7,14,21,and 28 days after photocoagulation,fluorescein fundus angiography (FFA) and choroidal vascular smear were performed to observe the degree of fluorescein leakage and CNV area in rats;Western blot and real-time quantitative polymerase chain reaction (RT-PCR) were used to detect the expression ofRap1,GTP-Rap1,VEGF,β-catenin and mRNA in CNV.Results The results of FFA examination showed that a large disc-shaped fluorescein leaked in the photo-condensation spot 14 days after photocoagulation.Laser confocal microscopy showed that compared with 7 days after photocoagulation,CNV area increased at 14,21,28 days after photocoagulation,and the difference were statistically significant (t=3.725,5.532,3.605;P＜0.05).Western blot showed that there was no significant difference in the relative expression of Rap1 protein in CNV at different time points after photocoagulation between the two groups (P=0.156).Compared with the blank control group,the relative expression of GTP-Rap1 protein was significantlydecreased,the relative expression of VEGF and β-catenin protein were significantly increased in the model group (P=0.000).The results of RT-PCR showed that there was no significant difference in the relative expression of Rap 1 mRNA at different time points after photocoagulation between the two groups (P=0.645),but there were significant difference in the relative expression of β-catenin mRNA (P=0.000).At 7,14,21 and 28 days after photocoagulation,there were significant difference in the relative expression of GTP-Rap 1 and VEGF mRNA between the two groups (P=0.000).Conclusions The expression of GTP-Rap1 in experimental CNV is significantly lower than that in normal rats.
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Background The residual astigmatism following Toric intraocular lens (IOL) rotation have received much attention.However,the variation of the optical performance and the wavefront abrrveation with Toric IOL rotation are unclear.Objective The aim of this study was to evaluate the optical performance,wavefront abrrveation and residual diopter spherical and cylinder lens with Toric IOL rotation.Methods T3,T4 and T5 Toric IOLs of +22.0 D were placed in Hwey-Lan Liou model eye respectively,with the posterior surface flat on the X axis and steep on the Y axis.Corneal astigmatism model was established by mimicing the model eye with Toric IOL using Zemax optical software.Then the Toric IOLs were rotated 5° to 10° individually under the 4 mm pupil diameter and 550 nm monochromatic light,and the image performance and wavefront abrrveation were recorded with all conditions,including modulation transfer function (MTF),out-of-focus aberration,astigmatism aberration,coma,trefoil aberration and spherical aberration.The refractive error of spherical power and cylinder power were calculated.Results Corneal astigmatism was fully corrected when Toric IOLs in the middle,and the MTF curves were near in T3,T4,T5 Toric IOLs.The image performance was worse under the high spatial frequency with the increase of rotation degrees of Toric IOLs,showing the gradually low of MTF curves,especially T5 Toric IOL.No obvious changes was seen in coma,trefoil aberration and spherical aberration after rotation of Toric IOLs,while out-of-focus aberration,astigmatism aberration were obviously increased.In addition,residual astigmatism and spherical error increase with the rotation of Toric IOLs.Conclusions Toricl IOL rotation leads to increase of astigmatism and spherical refractive error but not high order aberration.
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Objective To investigate the risk factors associated with neovascular glaucoma (NVG) after pars plana vitrectomy (PPV) in eyes with proliferative diabetic retinopathy (PDR).Methods Retrospective study.One hundred and thirty-seven patients (137 eyes) with PDR who underwent PPV were recruited.There were 85 males and 52 females.The average age was (60.1 ± 8.8) years old.The duration of diabetes was (10.2 ± 3.6) years.There were 49 patients with ipsilateral carotid artery stenosis.Fifty-three eyes underwent intravitreal ranibizumab or conbercept injection before PPV.All eyes were treated with 23G standard three-port PPV.The average follow-up time after PPV was 11.5 months.Fundus fluorescein angiography (FFA) was conducted in postoperative 4-6 weeks to observe non-perfused retinal areas.Risk factors,such as ipsilateral carotid artery stenosis,the presence of non-perfusion in retina after PPV and the application of anti-vascular endothelial growth factor (VEGF) drugs before PPV,were identified by logistic regression.Results Twenty of 137 patients (14.6％) developed postoperative NVG after PPV.Ipsilateral carotid artery stenosis [odds ratio (OR) =5.048,95％ confidence interval (CI) 2.057-12.389,P=0.000] and the presence of non-perfusion in retina after PPV (OR=4.274,95％CI 1.426-12.809,P=0.009) were significant risk factors for postoperative NVG,while the application of anti-VEGF drugs was not (OR=1.426,95％CI 0.463-4.395,P=0.536).But the time from PPV to the onset of NVG varies significantly between the two groups of injection of anti-VEGF drugs or not (t=-4.370,P=0.000).Conclusions Risk factors associated with NVG after PPV in eyes with PDR included ipsilateral carotid artery stenosis and the presence of non-perfusion in retina after PPV.The application of anti-VEGF drugs before PPV can delay the onset of NVG in PDR eyes after vitrectomy.
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Background Central retinal vein occlusion (CRVO) is a common retinal vascular disease.Macular edema is a common complication and can lead to the decrease of visual acuity.Intravitreal injection of antivascular endothelial growth factor (VEGF) drugs and triamcinolone acetonide has become the important treatment on macular edema.Objective This study was to systematically evaluate the clinical effects of anti-VEGF drugs and triamcinolone in patients with macular edema caused by CRVO.Methods The Databases including PubMed,Cochrane Library (Issue 11,2012),EMbase,CBM,CNKI,VIP and WanFang Database were electronically searched for the trials about the effects of anti-VEGF drugs and triamcinolone in patients with macular edema caused by CRVO from the date of establishment of the databases to September 2015.The combined effect was analyzed by using Review Manager 5.3 software.Results A total 7 trials involving 345 patients and 348 eyes were included.Meta-analysis showed that there was no statistical difference in best corrected visual acuity (BCVA) and macular central thickness between anti-VEGF drugs and triamcinolone in the 6-month follow-up (mean difference [MD] =-0.03,95％ confidence interval[CI]:-0.11-0.05,P =0.52;MD =-15.37,95％ CI:-36.29-5.55,P =0.15),but there was statistical difference in intraocular pressure (MD =-2.73,95％ CI:-3.59--1.86,P＜0.000 01).Twenty-two cases of lens opacity and 8 cases elevated intraocular pressure were observed in the triamcinolone group.Only 2 cases of lens opacity were observed in the anti-VEGF drugs group.Conclusions Anti-VEGF drugs and triamcinolone have similar improvement of BCVA and decrease of macular central thickness in CRVO patients,while the triamcinolone is accompanied with more side effects such as high intraocular pressure and progressing cataract.
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Background Interleukin-1β (IL-1β) is an important inflammation-related factor in the initial stage of proliferative vitreoretinopathy (PVR).The previous research showed that curcumin can inhibit IL-1 β-induced proliferation of rabbit retinal pigment epithelium (RPE) cells,but the anti-inflammatory mechanism and effect of curcumin are still undefined.Objective This study was to observe the migration of IL-1β-induced rabbit RPE cells,and evaluate the function and mechanism of inhibition of curcumin on IL-1β-induced inflammation of RPE cells.Methods Cultured rabbit RPE cells of generation 4 were used in this experiment.The cells were cultured in serum-free DMEM and 0,0.1,1.0 and 10.0 μg/L IL-1β were separately added in the medium for 24 hours.The expressions of cyclooxygenase-2 (COX-2) protein and mRNA in the cells were detected by Western blot and reverse transcription PCR to determine the optimal concentration of IL-1β.The cells were divided into IL-1β group and curcumin+IL-1β group,and 1.0 μg/L IL-1 or 1.0 μμg/L IL-1 β combined with 10 μg/ml curcumin was respectively added into the medium for 24,48 and 72 hours.The cells cultured by only serum-free medium served as the control group.Hematoxylin and eosin staining was conducted for the cells to count the number of cells migrating into the injured area under the optical microscope.The relative expression levels of COX-2 protein and mRNA in the cells were detected by Western blot and reverse transcription PCR,and the relative expression levels of nuclear factor (NF)-κBp65 and inhibitor of NF-κB-α (IκB-α) protein were also detected by Western blot assay.The expression intensity and location of NF-κBp65,IκB-α and COX-2 in the cells were detected by immunochemistry.Results RPE cells just isolated from the rabbit eyes were in round shape and abundant in melanin.The melanin significantly decreased in the fourth generations of RPE cells.The shape of cells became long and narrow,and net shaped distribution.Immunochemistry demonstrated the strong positive response of RPE cells for keratin (AE1/AE3).There were (31.93 ±1.21),(36.27±2.50) and (38.33±2.40) migratory cells in the control group after 24,48 and 72 hours respectively.The number of migratory cells increased to 45.73 ± 2.30,71.13 ± 1.92 and 80.60 ± 1.71 in the IL-13 group,but obviously decreased to 13.13 ± 2.20,14.93 ± 1.10 and 12.60 ± 1.51 in the curcumin + IL-1β group.A Significant increase in the migrating cell number was found in the IL-1 β group compared with the control group and the curcumin+IL-1β group in various time points (all at P＜0.05).The relative expression levels of COX-2 protein and mRNA peaked in the 1.0 μg/L IL-1β group,so 1.0 μg/L of IL-1β was determined as the optimal concentration in the experiment.In 24,48 and 72 hours after culture,the expression levels of COX-2 protein and mRNA in the cells were significantly lower in the curcumin + IL-1β group than those in the control group (all at P＜0.05).The relative expression level reached peak in NF-κBp65 protein and lowed bottom in IκB-α proteins at 48 hours after cultured in the IL-1β group,and the reverse trend was seen in the curcumin+IL-1β group,with the significant differences between the two groups (both at P＜0.05).Immunochemistry showed that NF-κBp65 was expressed strongly in the cell nuclei and cytoplasm in the IL-1 β group and presented the weaker expression in the control group and the curcumin+IL-1 β group.Compared with the control group,the expression was weaker in IκB-α and stronger in COX-2 in the IL-1β group.In addition,the expression of IκB-α was enhanced and that of COX-2 was attenuated in the curcumin+IL-1β group in comparison with the IL-1β group.Conclusions Curcumin inhibits the movement of rabbit RPE cells induced by IL-1β.IL-1β up-regulates the expression of COX-2 by activating NF-κB signal pathway,and curcumin plays an anti-inflammatory role by blocking this pathway.
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Background Proliferative vitreoretinopathy (PVR) is a common cause of vision loss clinically,and retinal pigment epithelium (RPE) cells play a major part in this disease.Studying the effect of traditional Chinese medicine on RPE cells are of great importance to reveal the pathogenesis and prevention of PVR,which were rarely reported.Objective This study was to study and compare the inhibition effect among curcumin,salvia miltiorrhiza and matrine on IL-1β-induced proliferation of rabbit RPE cells.Methods RPE cells at passages 3-4 were enrolled for the research and identified by transmission electron microscope.The proliferation effect of IL-1 β (2.5,5.0,10.0,20.0 μg/L) and inhibitory effect of curcumin (5,10,20 μg/ml),salvia miltiorrhiza (5,10,20 μg/ml)or matrine (100,200,400 μg/ml) on RPE cells 24,48 and 72 hours after cultivation were studied by MTT assay.The 50％ inhibitory dose (IC50) of the three medicines were analyzed by regression analysis.The use and feeding of the experimental animals were followed by the ARVO Statement.Results RPE cells isolated from the rabbit eye were in round shape and abundant in melanin;The melanin significantly decreased in the fourth generations of RPE cells.Immunohistochemistry showed that the RPE cells was positive for keratin (AE1/AE3).The proliferation rates of RPE cells were statistically different among different concentrations of IL-1β 24,48 and 72 hours after cultivation (Ftime =30.33,P =0.00;Fconcentration =9.37,P =0.00);The proliferation rates of RPE were significantly different among different time points or different concentrations of IL-1β (all at P ＜ 0.05).And the proliferation rate run up to maximum at 10 μg/L after 72 hours of cultivation.The inhibitory rates of the three medicines were statistically different among different time points or different concentrations (curcumin:Ftime =128.75,P =0.00;Fconcentration =334.05,P=0.00.salvia miltiorrhiza:Ftime =39.32,P=0.00;Fconcentration =165.57,P=0.00.matrine:Ftime =267.76,P =0.00;Fconcentration =912.34,P =0.00).The three medicines dose-dependently and time-dependently inhibit IL-1β-induced proliferation of RPE cells,with significant differences between the adjacent time points and concentrations (all at P＜0.05).The IC50 were 26.77,19.01 and 9.45 μg/ml for curcumin;33.72,23.47 and 12.56 μg/ml for salvia miltiorrhiza,570.96,352.25 and 97.50μg/ml for matrine 24,48 and 72 hours after cultivation.Conclusions The proliferation of RPE cells can be stimulated by IL-1β,and the maximal proliferation occurred with a concentration of 10.0 μg/L IL-1β.Curcumin,salvia miltiorrhiza and matrine dose-dependently and time-dependently inhibit proliferation of RPE cells induced by IL-1β.Curcumin is the best medicine to inhibit the proliferation of RPE cells.
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Background Choroidal neovascularization (CNV) is a common pathological basis of many ocular fundus diseases.Some treating methods are proved to be effective on CNV but there exist their own shortages.Celecoxib can inhibit experimental neovescularization.Sustained release drug of celecoxib and application approach can offer a basis for the therapy of CNV.Objective This study was to evaluate the sustained release ability of celecoxib-poly lactide-co-glycolide microsphere (CEL-PLGA-MS) in vitro and its inhibitory ability on experimental CNV in vivo.Methods CEL-PLGA-MS was prepared by Hebei Medical University and examined under the scanning electron microscope.The size of CEL-PLGA-MS was measured by Laser Particle Size Analyzer.The drugloading in vitro releasing was monitored by high performance liquid chromatograph (HPLC).Experimental CNV was induced by laser photocoagulation of retina in the right eyes of 72 male brown Norway (BN) rats and then were randomized into the CEL-PLGA-MS group,celecoxib group,blank PLGA group and PBS group.CEL-PLGA-MS with 320 μmol/L celecoxib,80 μmol/L celecoxib,blank PLGA microspheres solution and 0.01 mol/L PBS was intravitreally injected separately according to the grouping.CNV was assessed by fundus fluorescein angiography (FFA) on the 14th day after injection.The fibrovascular proliferation (FVP) thickness at photocoagulation spots was measured by OCT.The retinal pigment epithelium (RPE)-choroid-sclera sections were prepared for the histopathologieal examination of FVP.On the 7th and 28th day after intravitreal injection,the relative expression levels of VEGF mRNA and COX-2 mRNA in the photocoagulation area were detected by reverse transcription PCR (RTPCR).The use and feeding of the experimental animals were followed by the ARVO statement.Results CELPLGA-MS showed the spherical shape with the mean size of 2 467.9 nm and the drug-loading of 7.77％ and the drugrelease rate of 80.91％ in vitro for 45 days.It presented the controllable release characteristics.CEL-PLGA-MS agglomerated in vitreous body after injection.On the 14th day after intravitreal injection,the mean FVP thicknesses were (94.67±4.64),(98.56±4.72),(71.00±4.77),(50.44±3.01) μm in the blank PLGA microspheres group,PBS group,celecoxib group and CEL-PLGA-MS group,respectively,showing significant increases in mean FVP thickness in the blank PLGA microspheres group and PBS group compared with the celecoxib group and CEL-PLGAMS group (all at P＜0.01),and the CEL-PLGA-MS group appeared a lower mean FVP thickness value than the celecoxib group (P＜0.01).FFA revealed a large number of strong hyperfluorescences at the photocoagulation area in the rat eyes of the blank PLGA microspheres group and PBS group;while only weak hyperfluorescences were seen in the eelecoxib group and CEL-PLGA-MS group.Histopathological examinations verified the same results in the FVP thickness to OCT image.The relative expression levels of COX-2 mRNA and VEGF mRNA in the RPE-choroid-sclera were all significantly elevated in the blank PLGA microspheres group compared with the celecoxib group and CELPLGA-MS group both on the 7th and 28th day after intravitreal injection (all at P＜0.01).On the 7th day after injection,the relative expression levels of COX-2 mRNA were lower on the 7th day and the relative expression levels of COX-2 mRNA and VEGF mRNA were higher on the 28th day in the celecoxib group in comparison with the CEL-PLGA-MS group (all at P＜0.01).Conclusions CEL-PLGA-MSs are even in size with the spherical shape and controllable release characteristics in vitro.CEL-PLGA-MS can inhibit experimental CNV and was more durable effective than celecoxib after intravitrea] injection.
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Background The optic quality of Toric intraocular lens (IOL)-implanted eye is affected by the residual astigmatism and individual difference of corneal spherical aberration and different magnification from steep and flat axis refraction.Whether correcting Toric IOL spherical aberration can effectively improve the image quality of individual patient is a question to be studied.Objective This study attempted to collect eye parameters of cataract patients to reconstruct the customized vision model by using Zemax optical software,and to evaluate the image performance with different Toric IOL spherical aberration.Methods A prospective study was performed.Forty-five eyes of 45 cataract patients were included in Second Hospital of Hebei Medical University from August 2012 to October 2013.Several relevant parameters were measured by Pentacam,including anterior and posterior surface height of cornea,corneal thickness,curvature radius of flat and steep meridians of anterior surface astigmatism,refractive diopter and curvature radius of posterior surface.The astigmatism of anterior and posterior corneal surface was described by Matlab 4.5 software.Corneal astigmatism model were set as aspheric state,and the effective position of Toric IOL was calculated using Holladay Ⅰ formula.Customized individual model eyes were constructed by Zemax software.The contrast sensitivity function (CSF) of different spherical Toric IOLs at different spatial frequencies were calculated and compared between 300 Td light environment with 3 mm pupil diameter (photopia light) and 0.3-1.0 Td light environment with 5 mm pupil diameter (mesopia light).This study was approved by Second Hospital of Hebei Medical University ethics committee,all the patients signed the informed consent.Results The mean astigmatism power was (1.51 ± 0.36) D and (1.49 ± 0.37) D,and the mean astigmatism meridian was (101.5 ± 59.8) ° and (101.9±58.5) ° in the model eyes and cataract eyes,respectively,without significant differences between them (t=0.886,0.652;both at P＞0.05).Bland-Altman test showed a good agreement in astigmatism power and astigmatism meridian between model eyes and cataract eyes.The LogCSF values at 1.5,3.0,6.0,12.0 and 18.0 c/d spatial frequencies were significantly higher in the aspherical Toric IOL model eyes than those in the spherical Toric IOL model eyes,and the LogCSF values at various spatial frequencies were significantly higher in the Toric IOLs with spherical aberrations of-0.13 μm and-0.26 μm than those in the zero spherical aberrations in both photopia light and mesopia light (all at P＜0.05).Conclusions A precise corneal astigmatism model based on cornea high data of cataract eyes was successfully constructed through special formulas with Zemax software.Aspherical Toric IOL can compensate for spherical aberration of cornea and enhance the optic quality in individual model eye.
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The hallmark of the recent latest advances in diagnostic fundus imaging technology is combination of complex hierarchical levels and depths,as well as wide-angle imaging,ultra-wide imaging.The clinical application of wide-angle and ultra-wide imaging,not only can reevaluate the role of the peripheral retina,the classification types and treatment modalities of central retinal vein occlusion,and enhance the reliability of diabetic retinopathy screening,improve the classification and therapeutic decision of diabetic retinopathy,and but also can help guide and improve laser photocoagulation.However we must clearly recognize that the dominant role of ophthalmologists in the diagnosis of ocular fundus diseases cannot be replaced by any advanced fundus imaging technology including wide-angle imaging.We emphasize to use the three factors of cognitive performance (technology,knowledge and thinking) to improve the diagnosis of ocular fundus diseases in China.
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Background Choroidal neovascularization (CNV) is one of the causes of blindness in multiple eye diseases.Researches showed that complement system participates in the pathogenesis of CNV.Objective This study was to construct the recombinant of complement factor B-small interference RNA (CFB-siRNA) expression vector and to observe its inhibitory effect on human umbilical vein endothelial cells (ECV-304).Methods CFB gene primers were designed based on human CFB gene,and an expression vector of CFB-siRNA was constructed by inserting CFB-siRNA into pRNAT-U6.1/Neo plasmid.Recombinant plasmids were confirmed by the digestion analysis of restriction endonuclease,and all inserted sequences were verified by DNA sequencing.The recombinant pRNAT-U6.1/CFB-siRNA plasmid and the blank plasmid were transfected into ECV-304 cells in the CFB-siRNA group and blank plasmid group by electroblot,respectively,and non-transfected cells served as the normal control group.The cells were observed under the fluorescence microscope 48 hours after transfection,and the transfective efficiency was calculated.The relative expression of CFB mRNA in the cells of different groups was detected by semi-quantitative reverse transcription PCR (RT-PCR).MTT was employed to calculated the growth inhibitory rates of the cells 24,48 and 72 hours after transfection.The percentages of the cells in different cell cycles were detected by flow cytometry.Results The sequence of the target vector was identical to the designed sequence.The green fluorescence protein (GFP) was seen in both the CFB-siRNA group and the blank plasmid group.The relative expression levels of CFB mRNA were 0.07 ±0.04,0.14 ±0.02 and 0.14 ±0.03 in the CFB-siRNA group,the blank plasmid group and the normal control group,respectively,a significant difference was obtained among the three groups (F=233.05,P =0.00);the expression level of CFB mRNA in the CFB-siRNA group was significantly declined in comparison with the blank plasmid group and the normal control group (both at P＜0.05).The growth inhibitory rates of the cells were (23.45 ±0.01) ％,(33.48 ±0.02) ％ and (45.49±0.01) ％ at 24,48 and 72 hours after transfection,respectively,a significant difference was obtained among the three groups (Fgroup =212.99,P =0.00);the growth inhibitory rates in CFB-siRNA group were significantly higher than that in the blank plasmid group and normal control group (all at P＜ 0.05).The percentages of G1 phase cells were (44.4 ±0.5) ％,(25.8 ±0.4) ％ and (27.9 ± 0.6) ％ in the CFB-siRNA group,the blank plasmid group and the normal control group respectively,a significant difference was obtained among the three groups (F=58.98,P=0.00).The percentages of G1 phase and G2 phase cells in the CFB-siRNA group were significantly higher than those in the blank plasmid group and the normal control group (all at P＜0.05).Conclusions Recombinant pRNAT-U6.1/CFB siRNA inhibits the proliferation of ECV-304 cells effectively by arresting the cells in G1 intermediate phase of the growth cycle.
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Background Culture of retinal pigment epithelium(RPE) cells is very important for establishment of proliferative vitreoretinopathy (PVR) model,prevention and treatment of PVR as well as RPE cell transplantation.Isolation of animal RPE cells by trypsinization is a critical step.ObjectiveThe present study is to establish the methods of isolation and culture of retinal pigment epithelium (RPE) cells in rabbit and comparied with that of pig RPE culture.MethodsRPE cells were isolated by trypsinization in pigmented rabbit and pig and cultured in DMEM containing 20% fetal bovine serum.Cultured RPE cells were identified by immunochemistry.The fourth generations of cells were used in this experiment.Morphology and characteristics of cultured RPE cells from rabbit and pig were examined and compared under the light microscope.ResultsIsolated RPE cells from pig were obtained by once trypsin digestion,but two times of trypsinization were needed in rabbit RPE cells isolation.The differentiation in response to trypsinization was related to anatomic difference between the two types of cells .The adherence time of pig RPE cells was 24 hours ,however,the rabbit RPE needed 48-72 hours after culture.Proliferation and vitality of cultured cells were gradually attenuated and melanin decreased after several times subculture.The morphology of culture RPE cells was obviously different between rabbit and pig because species difference.Immunohistochemistry demonstrated the positive response of RPE cells for keratin.ConclusionRPE cells can be acquired from both rabbit and pig by trypsinization and culture.The culture process of RPE cells of pig is simpler than that of rabbit.Cells within the fourth generations are suitable for experimental application.
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Background The anti-proliferative effect of matrine has been demonstrated and its relevance to prevention and treatment of proliferative retinovitreopathy is concerned.Howeverthe intravitreous injection of free-matrine reiteratively may raise the risk of ocular infection.ObjectiveThe goal of the present study is to investigate the sustained releasing ability and safety of matrine polyactic acid microsphere(MAT-PLA-MSintravitreal injection.MethodsMAT-PLA-MS was prepared by Hebei Medical University and examined under the transmission electron microscope.The release of MAT-PLA-MS was monitored by ultraviolet spectrophotometry.Free-matrine with the dose of 1,2,4mg was intravitreally injected respectively in 12 eyes of New Zealand albino rabbits in free-matrine group and MAT-PLA-MS with matrine(2,4,6mg respectively was administered in 16 eyes separately in matrine microsphere group.The blank microsphere was injected in 6 right eyes as blank control group and normal saline solution was injected in 6 fellow eyes as control group.The retinal function change was evaluated by electroretinogram(ERG),and the morphological and histological change of retina following drug injection were assessed under the slit lamp biomicroscope,indirect ophthalmoscope,light microscope and transmission electron microscope.The decomposed process of MAT-PLA-MS in vitreous was recorded with ocular anterior segment and fundus color camera.Results MAT-PLA-MS containing matrine showed the spherical shape with the mean diameter of 2.28±47μm under the transmission electron microscope and the drug-loading rate 6.17% and drug-release rate 87.93% in vitro for 672 hours,presenting the controllable release characteristics.After implantation into the vitreous,the MAT-PLA-MS containing matrine decomposed gradually with the prolong of time.The b amplitudes of ERG maximum response were significantly declined in 4mg free-matrine injection group in comparison with before injection in various time points(P<0.01).However,no considerably differences were found in MAT-PLA-MS with matrine groups and control groups in various time points following the intravitreal injection(P>0.05).No obvious abnormal was seen under the slim lamp and ophthalmoscope through the study period.The changes of retinal ultrastructure were found from 1 through 28 days after injection of 4mg free-matrine,and slight retinal structural damage was seen from 7 through 28 days after injection of 6mg MAT-PLA-MS containing matrine.ConclusionThese results suggest that MAT-PLA-MS possesses good sustained release feature.MAT-PLA-MS containing matrine has less toxicity to retina than free-matrine after intravitreal injection.MAT-PLA-MS is an excellent drug delivery system.
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Objective To establish a matrine delivery system in vitreous is very important for the dynamic treatment of proliferative vitreoretinopathy(PVR) . Present study was to evaluate the efficacy of matrine polyactic acid microsphere(MAT-PLA-MS) in prevention of PVR. Methods The suspension of cultured fibroblasts was injected into vitreous cavity of 30 healthy adult New Zealand albino rabbits to induce PVR. Then the experimental rabbits were divided into 3 groups and 10 rabbits for each. The animals received intravitreal injection of 0.3 mL MAT-PLA-MS(4 mg) matrine in MAT-PLA-MS group. Free matrine normal sodium solution 0.3 mL(containing 2mg matrine) was injected in vitreous cavity in free matrine group. 0. 3 mL normal saline solution was injected into the vitreous of the left eyes and the equivalent volume of blank polyaetic acid microsphere(blank-PLA-MS) into the right eyes in control group. The changes of cornea, aqueous humor, lens, vitreous and fundus were examined and recorded by slit lamp biomicroscope, indirect ophthalmoscope, fundus color camera and B ultrasonogram on the 1st, 3rd, 7th, 14th, 21st, 28th and 35th day following injection of drug. The inhibition effect of matrine on PVR was evaluated according to Ryan' s grading criteria of PVR. Results On the 14th days after implantation of MAT-PLA-MS, the rate of retinal detachment was 60%, 10%, 5% and 60% in normal saline group, free matrine group, MAT-PLA-MS group and blank-PLA-MS group respectively. Statistically significant difference was found among normal saline group, blank-PLA-MS group, MAT-PLA-MS group and free matrine group(P ＜0. 05). On the 21st day after injection of fibroblasts, the morbidity of retinal detachment was 80%, 30%, 10% and 80% in normal saline group, free matrine group, MAT-PLA-MS group and blank-PLA-MS group respectively, showing a significant difference among different groups. On the 28th day, the incidence rate of retinal detachment was 90%, 50%, 15% and 90% respectively, presenting statistical difference among various groups (P ＜ 0. 05) as well as between free matrine group and MAT-PLA-MS group (P＜0. 05). On the 35th day, considerably difference also was seen in the morbidity of retinal detachment among various groups (90%, 60%, 15% and 90% respectively) (P＜0.05). Conclusion Implantation of MAT-PLA-M S into vitreous cavity can effectively inhibit the development of PVR induced by fibroblasts in rabbit model.
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Objective hole.Research demonstrated that amnion promotes the proliferation of and its mechanism.Methodswhite rabbit.The cells were primarily cultured and passaged in DM/F12 with 10% fetal bovine serum and identified with glial fibrillary acidic using 0.25% trypsin and 0.05%EDTA and suspended in 10% fetal bovine serum+DMEM/F12.0.5 mL of the amnion supernatant was added to the medium for 12 hours.The expression of EGFR in the cells was detected by immunohistochemistry.The difference of EGFR expression with or without the amnion was evaluated.ResultsCultured cells showed a positive response for GFAP and S-100.The intermediate filaments with a length of 8-10 nm were exhibited under the transmission electron microscope.EGFR was fainly expressed in cultured hours,the expression of EGFR was obviously increased with a significant difference in the gray scale between the two groups(571 588.80±67 862.68 vs.1 000 352.00±98 386.22)(t=4.035,P＜0.01).Conclusion
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Objective To observe the effect of amniotic homogenate on closing holes in experimental rhegmatogenous retinal detachment and investigate its mechanism.Methods Forty rabbits were randomly divided into group A,B,C and D with 10 rabbits in each group.Group A and C were the treatment groups,and group B and D were the control groups.All eyes of rabbits underwent pars plana vitrectomy,retinectomy,and fluid-air exchange.The surface of the breaks was treated with 0.1 ml amniotic homogenate in experimental groups and 0.1 ml PBS in control groups.At the end of operation,20% SF6 was tamponaded and the retina reattaced.The animals were executed 14 (group A and B) and 28 days (group C and D) after the surgery.The tissue sections were observed by light microscope,electron microscope and immunocytochemistry method.Results Fourteen days after the surgery,the retina reattached in 6 eyes in group A (60%) and 2 eyes in group B (20%) (P=0.021),Twenty-eight days after the surgery,the retina reattached in 8 eyes in group C (80%) and 3 eyes in group D (30%) (P=0.046).The difference of the rate of retinal reattachment among the 4 groups were statistical significant (P<0.05).Light postoperative inflammation of ocular anterior segment was observed,which was controlled 3-5 days after treated with topical steroids.The result of light microscopy showed that the eyes in treatment groups had multilayer of fibroblast like cells around the retinal breaks,adhering to the choroid and retinal pigment epithelial cells.The proliferative cells around the retinal breaks obvious less in control groups than that in the treatment groups,and the retina could not adhere to the choroid.The results of electron microscopy were the same as that of light microscopy.Immunohistochemistry staining of the fibroblast-like cells revealed positve glial fibrillary acidic protein,which suggested that the proliferative cells around the retinal breaks were retinal glial cells.Conclusions Amniotic homogenate helps to seal retinal breaks and promote retinal reattachment by stimulating the proliferation of retinal glial cells around the breaks.
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Objective To investigate the effects of liposome mediated intraocular gene transfection of endostatin on the inhibition of the development of choroidal neovascularization (CNV) in a rat model. Methods Experimental CNV model in Brown Norway rats was induced by laser photocagulation. The recombinant eukaryotic expression plasmid pSecTagA hEndostatin or control plasmid pSecTagA and liposome complexes were injected into the subretinal space of the model rats. In situ hybridization and immunohistochemical observation confirmed the presence of endostatin mRNA and protein expression two weeks after injection. Intraocular and serum levels of endostatin were measured by enzyme linked immunosorbent assay. The intensity of fluorescein leakage from the photocoagulated lesions was studied at 13 d after photocoagulation. The area of CNV was measured using high molecular weight FITC dextran (MW2?10 6) for high resolution angiography in RPE choroid sclera flat mounts. In addition, sections of CNV lesions were studied by light microscopy and endoglin (CD105) immunohistochemical evaluation. Results The retina, RPE, choroidal were infected by subretinal delivery of the pSecTag hEndostatin and expressed the endostatin. Two weeks after intraocular injection, the level of endostatin in the whole eye homogenates were (50 14?3 43) ng/eye and (31 5?2 21) ng/eye, respectively. Fluorescein leakage from the CNV lesions decreased significantly as compared with that in the control groups. The average area of CNV at the sites of the Bruch's membrane rupture showed significant difference in eyes injected with endostatin as compared with that in the control eyes. Endothelial cells demonstrated strong immunoreactivity of CD105 in CNV lesions in the control eyes. Conclusion Liposome mediated endostatin gene transfection can significantly inhibit the development of CNV.
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Objective To observe the clinical characteristics of patients with macular branch retinal vein occlusion (MBRVO) and the changes of the area of foveal avascular zone (FAZ). Methods The data of 69 eyes of 69 patients with MBRVO , who had been diagnosed by ophthalmoscopy, slit-lamp examination and fluorescein angiography, were retrospectively studied. The relationship of locations between artery and vein on the obstructive site, and the characteristics of fundus pictures, retinal vasculature changes and the complications were analyzed. In 69 patients with MBRVO, 36 had the course of disease for more than 3-6 months, of whom the area of FAZ was compared with that of 30 healthy people. Results In 69 patients, superior MBRVO occurred in 45 eyes (65.22%), and inferior MBRVO occurred in 24 eyes (34.78%). Most of the arteries were anterior to the veins at the obstructive site. Four clinical types of MBRVO were found, and the main complication was macular edema. There was a significant difference in area of FAZ between patients with MBRVO and healthy people (P
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ObjectiveTo detect the content of vascular endothelial growth factor（VEGF）in aqueous humour and in vitreous of patients with proliferative diabetic retinopathy（PDR）．MethodsEnzyme linked immunosorbent assay（ELISA）was used，which is one of the basic methods of protein quantitative analysis．ResultsThe VEGF levels in aqueous humour of PDR（mean level 398．6 pg/ml）were higher than those of the controls（mean level 106.5 pg/mlP＜0.05）．The VEGF levels in vitreous of the four PDR（mean level 1395.2 pg/ml)were higher than those in aqueous humour of the four PDR(P＜0.05，Primer t test）．The increased VEGF level in vitreous was positively correlated with the level in aqueous humour．In the normal group，the mean value of VEGF in the vitreous was 64.6 pg/ml．In comparison with that of 1140.4 pg/ml in the PDR group，there was a significant difference between the two groups （P＜0.05，Primer t test）．ConclusionVEGF might play an important role in the pathogenesis of neovascularization of PDR．
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0.05). But the mean light sensitivity at central 10? eccentricity were significantly decreased 3 months after photocoagulation (P