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The prevention effect of aspirin on galactose cataract

Chinese Ophthalmic Research.2009;27(11):1015-1018.

Objective Experimental research demonstrated that oxidative damage leads to formation of cataract in rats and its machanism is the decline of activities of superoxide dismutase(SOD), glutathione peroxidase(GSH-PX) and catalase(CAT) . Aaspirin can improve the antioxidative ability of lens. The purpose of this study was to observe the inhibition of aspirin on D-Galactose-induced cataractous lenses of rats. Methods Galactose cataract model was established in 40 cleaning Wistar rals by intraperitoneal injection of 20 mL/kg 80% D-Galactose for 10 days. The models were divided into model group (20 rats) and aspirin group(20 rats). 150 mg/kg of aspirin was administered immediately by gastrogavaging in aspirin group for 20 days. Other 20 normal Wistar rats were as control group. At day 3, 6, 10, 14, 20, the transparency of rat lenses was observed under the slit lamp microscopy. At day 5 after experiment, the ultrastructure of the lenses was examined and evaluated under the scanning electron microscopy. The activities of SOD, GSH-PX and CAT were detected by Coomassie Brilliant Blue color comparator, respectively. The use of experimental animal followed the Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission. Results All lenses were transparent in the rats of control group. The degree of lens opacity was more mild in asprin group compared with model group. 25. 00%, 41. 67%, 58. 33%, 83. 33% of lenses in aspirin group showed swelling at day 6, 10, 14, 20, respectively, but 65% lenses were opacity in model group on day 3 and 100% lenses were nuclear cataracts in 6 days. The structure of lenses was normal in control group, but the process number, fiber thickness and fiber density of lens were significantly increased in model group compared with control group (P ＜0. 05), and only process number was increased in asprin group. The activities of SOD, GSH-PX and CAT in lens of model group were obviously lower than in normal control group(P＜0. 05), but those in asprin group were significantly increased in comparison with model group(P ＜0. 05). Conclusion Aspirin could protect lenses of rats against oxidative damage by elevating activities of SOD, GSH-PX and CAT in lens and inhibiting the generation and development of galactose-induced cataract at early stage of cataract.

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Expression of heranase and perlecan in the retina of mouse with oxygen-induced retinopathy

Lei, WAN ; Xiaoguang DONG ; Ting, LIU ; Jun, CHENG ; Lixin, XIE

Chinese Ophthalmic Research.2010;28(3):210-215. doi:10.3969/j.issn.1003-0808.2010.03.006

Background Heparanase degrade heparan sulfate side chains of heparan sulfate proteoglycans in the extracellular matrix.Heparanase induces angiogenesis and likely promotes the vascularization of tumor.ObjectiveThe present study is to investigate the expression of heparanase and perlecan in retinas with oxygen-induced retinopathy.Methods Sixty-five clean neonatal C57BL/6J mice were raised in a hyperbaric oxygen box with a volume percentage of 75%±2% for 5 days and then returned to the normal air room.Another 65 matched mice were raised in the normal environment as controls.Evans blue was infused by the superior vena cava in all the mice on postnatal days 12,13,17,21 and 30,afterwards fluorescein angiography was performed and then the mice were sacrificed.The retinas of mice were isolated and prepared and the retinal vessels were examined under a fluorescent microscope and optical microscope.Heranase and perlecan mRNA was detected using reverse transcription PCR (RT-PCR).Heranase and perlecan proteins were detected by Western blot.The analysis of variance was used to compare the mRNA and the protein levels of heranase and perlecan between the experimental and control groups.Results The expression of heparanase mRNA in the retinas of different ages of mice and the different groups showed significant differences (F_(group)=16.303,P=0.000;F_(time)=18.614,P=0.000;F_(interaction)=11.299,P=0.000),and the expression of heparanase mRNA was significantly enhanced in mice from postnatal days 12,13,17 and 21 compared with normal control mice (P=0.001,0.000,0.000,0.001,respectively).The expression of heparanase protein in the retinas of different ages of mice and the different groups followed the same tendency(F_(group)=458.134,P=0.000;F_(time)=78.466,P=0.000;F_(interaction)=71.398,P=0.000).The expression of perlecan mRNA in the retinas of different ages of mice and the different groups showed significant differences (F_(group)=7.351,P=0.013;F_(time)=9.098,P=0.000;F_(interaction)=3.349,P=0.000),and increase in differences also were clearly seen in mice from postnatal days 13,17 and 21 compared with normal control mice (P=0.048,0.000,0.003,respectively).Conclusion The expression of heparanase and perlecan is associated with the development and progression of retinal neovascularization,and perlecan and heparanase together produce a synergistic effect.Heparanase and perlecan may participate in the angiogenesis of oxygen-induced retinopathy.

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The clinical study on intravitreous injection of avastin for macular edema induced by branch retinal vein occlusion

Tong, QIAN ; Xiaoxin, LI ; Hong, YIN ; Jianhong LIANG ; Huijun, QI ; Wenzhen, YU

Chinese Ophthalmic Research.2010;28(1):79-82. doi:10.3969/j.issn.1003-0808.2010.01.021

Background Branch retinal vein occlusion (BRVO)often leads to macular edema as the result of elevating intravitreal VEGF level,and avastin,a anti-VEGF drug,show a good effectiveness on macular edema secondary to BRVO.ObjectiveThis study attempts to evaluate the clinical efficacy of avastin on macular edema induced by BRVO.MethodsThis is a retrospective case-observation study.The clinical data of 39 eyes from 39 patients with macular edema induced by BRVO were included in this analysis.All of the patients received intravitreal injection of 1.25mg (0.05mL)avastin without other any therapy prior to the injection of avastin.This procedure followed the Declaration of Helsinki,and written informed consent was obtained from all the patients before and initial of any management.Clinical indexes included best-corrected visual acuity(BCVA),slit-lamp examination,intraocular pressure and stereoscopic biomicroscopy examination before injection and 3,6 and 12 weeks after initial injection.Optical coherence tomography (OCT),fundus photography,fluorescein fundus angiography(FFA)were performed prior to injection and 6,12 weeks after initial injection.The follow-up period was 3-20 months.ResultsThe mean BCVA was significantly improved at 3,6,12 weeks after injection in comparison with before injection (t=-6.039,-6.182,-4.189,all P=0.000).The mean CMT showed a statistically significantly decline at 6,12 weeks after injection in comparison with before injection(t=8.684,5.019,all P=0.000).No ocular or systemic adverse events were found after intravitreal injection of avastin during the follow-up duration.The numbers of visual acuity-improved eyes were significantly increased in the patients with disease course ≤1 month duration in comparison to ones with the course ≥1 month (P<0.05)in 3 weeks after injection.CMT was obviously decreased in 12 weeks after injection in comparison to before injection between with and without macular perfusion eyes (P<0.05).ConclusionIntravitreal injection of avastin is safe and effective for macular edema induced by BRVO,especially the patients with shorter course of disease.

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The relationship between the genotype and clinical factors of filamentous fungus causing-keratitis

Xueqing, BAI ; Zhiqun, WANG ; Ran, LI ; Shiyun, LUO ; Shijing, DENG ; Qingfeng LIANG ; Xuguang, SUN

Chinese Ophthalmic Research.2010;28(1):71-74. doi:10.3969/j.issn.1003-0808.2010.01.019

Background The study on the classification of fungi is very important for the diagnosis and treatment of fungal keratitis.Identifying the different species of filamentous fungi is a critical factor for the application of anti-fungal drug in treating keratitis.ObjectiveThis report studies the relationship between the genotype of filamentous fungi and the clinical factors.MethodsFifty-two patients with filamentous fungal keratitis determined by clinical and laboratory examination were recruited in Tongren Hospital from January 2006-December 2006.The lesions were graded on the severity of the corneal ulcer and the presence of hypopyon.The filamentous fungal keratitis was treated with topical and systemic administration of anti-fungal drugs or corneal transplantation.The isolates were cultured in potato culture and identified by morphological characteristics based on the Nelson criterion and genotyped by the rDNA ITS method.The clinical data was retrospectively analyzed.ResultsForty-eight species (eubacteria are bacteria,not fungi)of fungus were identified by morphological characteristics,and the filamentous fungi were divided into 4 types based on the phylogenetic relationships within the rDNA ITS of the 52 filamentous fungi.The morphological characteristics and genotype were confirmed in 48 strains of eubacteria and 31 strains of 52 filamentous fungi (90.3%).The 4 groups of fungi were classified by genotype as follows:group 1 represents 22 strains including 20 strains of Fusarium solani and 2 strains of Fusarium oxysporum;group 2 represents 12 strains including 8 strains of Fusarium moniliformis,3 strains of Fusarium proliferatum and 1 strain of Fusarium incarnatum;group 3 represents 5 strains including 1 strain of Fusarium moniliformis and 4 unknown strains;group 4 represents 13 strains including 10 strains of Aspergillus spp.and 3 strains of Alternaria spp.Significant differences were found in the disease duration (P=0.00),inducing cause (P=0.03),ulcer grade (P=0.01)and outcome of the anti-fungal treatment (P=0.035)when compared between group 1 and 2 with group 3 and 4.Conclusion Filamentous fungi that cause keratitis could be correctly identified by sequencing the internal tanscribed spacer of rDNA.There are significant clinical differences among the groups classified by genotype.

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The effects of mitogen-activated protein kinase on EGF-induced expression of integrin α_5 in human RPE cells

Zhen, CHEN ; Yiqiao, XING ; Changzheng CHEN

Chinese Ophthalmic Research.2010;28(1):62-65. doi:10.3969/j.issn.1003-0808.2010.01.017

Background Human retinal pigment epithelial(RPE) cells play a critical role in the pathogenesis of proliferative vitreoretinopathy(PVR) and other related ocular diseases.Research demonstrated that epidermal growth factor(EGF) stimulates activation of RPE cells and therefore causes PVR,and integrin α_5 mediates the adhesion of cells and EGF.Objective This study aims to investigate the role of mitogen-activated protein kinase(MAPK) in regulation of EGF on integrin α_5 expression in human RPE cells.MethodsHuman RPE cells strain(CRL2302) was cultured in DMEM/F12 with 10% calf serum and passaged in 0.25% trypsin.Cultured cells were divided into DMEM/F12control group,20μmol/L PD98059 +10 ng/mL EGF＋DMEM/F12(PD98059) group and 10 ng/mL EGF＋DMEM/F12(EGF) group.The expression of integrin α_5 protein in CRL2302 cells was detected by RT-PCR and calculated as α_5 mRNA/β-actin mRNA,and the expression of integrin α_5 mRNA in CRL2302 cells was detected evaluated by flow cytometry.The MAPK phosphorylation level in each group of human RPE cells was tested by Western blot.ResultsThe expression value of integrin α_5 mRNA was 0.93±0.06 in the control group,1.06±0.07 in PD98059 group and 1.97±0.09 in EGF group,showing a significant difference among the three groups(F=458.896,P<0.01).The fluorescence intensity of integrin α5 protein in CRL2302 cells after 24 hours was 1.94±0.22,4.56±0.25,2.39± 0.14 in three groups respectively with a significant difference(F=21.05,P<0.05).After 30 minutes of culture,Western blot result showed that the strongest phosphorylation levels of ERK1/2 activation in EGF group and the weakest phosphorylation levels of ERK1/2 activation in the control group;While that in PD98059 group was significantly stronger than control group and weaker than EGF group(F=143.14,P<0.01).ConclusionEGF stimulates activation of ERK1/2 pathway in human RPE cells and the expressions of integrin α_5 mRNA and protein in human RPE cells in vitro.

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Effect of tTG inhibitor on the expression of FN and Col-Ⅳ induced by TGF-β_2 in human lens epithelial cells

Xing, XING ; Hu, YIZHEN ; Chen, BO

Chinese Ophthalmic Research.2010;28(1):58-61. doi:10.3969/j.issn.1003-0808.2010.01.016

BackgroundOur previous research and other reports disclosed that the expression of tissue transglutaminase(tTG)in lens epithelial cells(LECs) of patients with cataract is enhanced,indicating tTG is related to formation of posterior capsule opacification(PCO).ObjectivePresent study is to observe the effect of tTG specific inhibitor monodansyl-cadaverineon(MDC) on the expression of fibronectin(FN) and collagen Ⅳ(Col-Ⅳ) induced by TGF-β_2 in human LECs.MethodsHLE-B3 was cultured in vitro in DMEM containing 10% fetal bovine serum and then were divided into 5 groups.The free-serum culture was used as normal control group.Free-serum culture containing 10μg/L TGF-β_2 was utilized as treatment group.10μg/L TGF-β_2 plused 100μmol/L,200μmol/L and 400μmol/L MDC respectively in different concentrations as MDC-treatment group.Semiquantitative RT-PCR was used to assay the expression of tTG,FN and Col-Ⅳ in HLE-B3.A(tTG/β-actin),A(FN/β-actin) and A(Col-Ⅳ/β-actin) was calculated separately as the detecting indexes.ResultstTG,FN and Col-Ⅳ were positively expressed in cultured HLE-B3.The expression levels of tTG,FN and Col-Ⅳ in HLE-B3 were remarkably increased in the group with 10μg/L TGF-β_2 compared with normal control group(t=33.95,P<0.01;t=38.24,P<0.01;t=13.48,P<0.01).The expression levels of FN and Col-Ⅳ were gradually declined in 100,200 and 400μmol/L MDC groups in comparison with TGF-β_2 treatment(P<0.01).The significant differences were also found in the expressions of FN and Col-Ⅳ in HLE-B3 among 100,200 and 400μmol/L MDC groups(P<0.01).ConclusionMDC inhibits the expression of FN and Col-Ⅳ induced by TGF-β_2 in human LECs at a concentration-dependent manner.tTG may be involved in the formation of posterior capsule opacification through up-regulating the expressions of FN and Col-Ⅳ in human LECs.

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Sustained release ability and safety of matrine polyactic acid microsphere intravitreal injection

Danyan, LIU ; Jingxue, MA ; Deying, CAO ; Jianxin, WANG ; Jianzong LIU ; Lancun, LÜ

Chinese Ophthalmic Research.2010;28(1):34-38. doi:10.3969/j.issn.1003-0808.2010.01.010

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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Inhibition of corneal neovascularization by liposome mediated plasmid encoding endostatin with RGD sequence

Ping, LIU ; Hua, LI ; Hong, ZHANG ; Lijuan, ZHANG ; Zhen, SONG ; Hongyan, GE

Chinese Ophthalmic Research.2010;28(1):19-22. doi:10.3969/j.issn.1003-0808.2010.01.006

Background It has been demonstrated that αvβ3/αvβ5 and α5β1 integrins are overexpressed in neovascular tissue.Consequently,peptide containing the RGD (Arg-Gly-Asp) sequence,which exists in ligands of integrins,is effective in targeting therapeutic reagents to neovascular endothelium.ObjectivePresent study aims to investigate the antiangiogenetive effects of liposome mediated plasmid encoding endostatin with RGD sequence on alkali burn-induced corneal neovascularization (CNV) in rabbits.MethodsCNV models induced by alkali burn were established in 72 eyes of 36 New Zealand white rabbits by putting the filter paper with 1 mol/L NaOH at the central cornea for 20 seconds.The animal models were divided into four groups randomly.0.2mL of liposome and plasmid encoding RGD-ES complex (liposome mediated pCI-RGD-ES injection group),liposome and plasmid encoding ES complex (pCI-ES injection group),liposome and carrier plasmid (pCI) complex (pCI-ES injection group),and PBS were subconjunctivally injected respectively in the models from four groups twice a week for two weeks.The growth status of CNV was observed on day 1,3,7,14 after alkali burn under the slim lamp microscope.Experimental animals were sacrificed on the 3rd,7th and 14th day and the expression of VEGF in CNV was detected by immunohistochemistry.The number of corneal microvessels was counted based on the number of CNV cross-section under the light microscope.ResultsCNV area was significantly smaller in liposome mediated pCI-RGD-ES injection group and pCI-ES injection group compared with PBS group at different time points (P＜0.01),but no significant difference was seen in CNV area between pCI-ES injection group and PBS group at different time points (P＞0.01).The changes of number of corneal microvessels was followed a similar fashion as the change of CNV area.Expression of VEGF in cornea was obviously stronger in pCI-ES injection group and PBS group than liposome mediated pCI-RGD-ES injection group and pCI-ES injection group.ConclusionEndostatin with RGD sequence could effectively inhibit corneal neovascularization,and liposome is proved to be a potent carrier in gene transfer.

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The effects of steriod on expression of TLR2 in rat corneas after penetrating keratoplasty

Lang, BAI ; Xiaohe, LU ; Yanyan, ZHONG ; Jing, ZHANG ; Jin, ZHOU ; Haijun, WU

Chinese Ophthalmic Research.2009;27(11):1011-1014.

Objective Our previous study demonstrated that toll-like receptor 2(TLR2) can distinguish extraneous antigen and prevent the immunological response. This study was designed to detect the expression of toll-like receptor 2 (TLR2) mRNA in cornea and investigate the effect of steroid on TLR2 expression in rats cornea following allograft penetrating keratoplasty. Methods The penetrating keratoplasty models were established in SPF rats with the 108 SD rats as receiptors and 36 SPF Wistar rats as donors, and other 6 SPF SD rats worked as normal controls. The receiptor rats were divided randomly into autograft group, allograft group and steriod group. The clarity and neovascularization of corneas of experiment rats were examined under the slit-lamp microscope and the rejection index was calculated based on Holland criteria. Corneal histopathological examination was carried out by hemotoxylin and eosin staining under the light microscope, and real time-PCR was employed for the detect of TLR2 mRNA in the corneas at the fifth, seventh and ninth day after operation. The experimental animals were obtained from the Animal Experimental Center of Southern Medical University and the procedure followed the Statement of Association for Research in Vision and Ophthalmology. Results The rejection occurred in 7 days after operation in allograft group, and only mild edema, opacity and neovascularization of corneas were found at different degrees in 9 days after operation in autograft group and steriod group. Severe corneal edema, a lots of inflammatory cells infiltration and new vessels in stroma were seen in allograft group, and mild inflammatory response was found in autograft group and steriod group. Normal comeal structure was exhibited in normal control group under the light microscope. The fold differences of TLR2 mRNA expression in cornea after amplification was significantly different among three groups and different time points (F_(group) = 39. 46, P = 0. 00; F_(time) =35. 38, P = 0. 00 ; F_(interaction) = 45. 66, P =0. 00), and the evident enhance of TLR2 mRNA expression was revealed in allograft group compared with autograft group (P ＜ 0. 05) and declined in steriod group (P ＜ 0. 05). Conclusion Steriod may restrain the acute allograft rejection by down-regulating the expression of TLR2 in corneas and its signals transaction. This result suggests that steriod offer a protection from rejection of cornea after penetrating keratoplasty.

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Acceleration of apoptosis by transfection of Apoptin gene in retinoblastoma cells

Jing, WANG ; Yuangui, WANG ; Man, LIANG ; Xuting, LI

Chinese Ophthalmic Research.2009;27(11):996-999.

Objective Present study aimed to observe the effects of Apoptin gene on killing retinoblastoma HXO-RB_(44) cells and illustrates its mechanisms. Methods Human retinoblastoma cells strain, HXO-RB_(44), was cultured and passaged in RPMI 1640 medium containing bovine serum. Apoptin gene was transfected into HXO-RB_(44) cells by liposome into HXO-RB_(44)/Apoptin, and pcDNA_3 was transfected in HXO-RB_(44)/peDNA_3 group. The expression of Apoptin mRNA was detected using Reverse Transcription Polymerase Chain Reaction (RT-PCR). The expression of protein of Apoptin and p53 were detected by SABC immunohistochemistry. The growth rate of HXO-RB_(44) cells was studied by constructing the growth curve and calculated as the formula: inhibitory rate = 1-cell number in experiment group/cell number in control group x 100%. Cellular apoptosis was determined by flow cytometry. Results The RT-PCR result showed the 450 kb specific band in UXO-RB_(44)/Apoptin group and absent amplification result in HXO-RB_(44) group and HXO-RB_(44)/pcDNA_3 group. The difference in SABC-positive cell number between HXO-RB_(44)/Apoptin group and control group was statistically significant (P ＜ 0. 05). The growth of HXO-RB_(44) cells was significantly inhibited in HXO-RB_(44)/Apoptin group compared with control group (P ＜ 0.05). Apoptosis cells increased significantly. The apoptosis rate was 38. 5% . Conclusion Apoptin gene could inhibit the growth of HXO-RB_(44) cells effectively. Up-regulation of expression of p53 gene might not be one of cell apoptosis mechanisms.

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