Objective To establish a rat model of blood ocular barrier breakdown induced by anterior segment intraocular analogic surgery. Methods One hundred and fifty healthy adult male rats were randomly divided into control group and model group,75 rats in each group.The rats were anesthetized with 1 ml/kg ketamine hydrochloride/xylazine hydrochloride solution.Three way pipes were attached to a phosphate buffer infusion bag and two intravenous catheters. One catheter was inserted 30° obliquelythrough the transparent cornea anterior to the limbus into the rat's anterior chamber.Then the needle was withdrawn and the sheath was indwelling.Another catheter was connected with a manometer.Intraocular pressure was varied from 0 to 12 mm Hg (1 mm Hg=0.133 kPa) 60 times,30 times per min.The catheter was removed.The eyes were treated with ofloxacin ophthalmic solution after surgery.The 1st,2nd,3rd,5th and 7th day after surgery,the integrity of the blood ocular barrier was assessed by immunohistochemical staining for albumin and quantitative measurement using Evan's blue as a tracer. Results Albumin immunohistochemical staining of the control group was confined to the iris and retinal blood vessels.The choroid was stained at each time point after surgery.Albumin immunohistochemical staining of the model group was abundant around the iris and the retinal vasculature on the 1st day after surgery.The albumin diffused throughout the iris and the retina on the 2nd and the 3rd day after surgery.The albumin reached the retinal vessels on the 5th and 7th day after surgery.The aqueous humor Evans blue leakages of the model group were higher than those of the control group on the 1st,2nd,3rd and 5th day after surgery.The differences were statistically significant (t=25.781,37.433,25.150,19.171; P＜0.01).The Evans blue leakage of the model group was close to that of the control group on the 7th day after surgery. The difference was no statistical significant(t=1.303,P=0.209).The retinal Evans blue leakages of the model group were higher than those of the control group on the 1st,the 2nd and the 3rd day after surgery.The differences were statistically significant (t=11.997,14.622,23.014; P＜0.01).The Evans blue leakage of the model group was close to those of the control group on the 5th and 7th day after surgery. The differences were not statistically significant(t=2.027,0.756 ； P=0.058,0.459).Conclusion This study establishes a rat model of blood ocular barrier breakdown induced by imitating the injury to the anterior segment during intraocular surgery.

Background Age-related cataract is a common cause of blindness.However,its cause and pathogenic mechanism have not been fully understood.Recent studies revealed that aquaporin 1 (AQP1) and AQP0 are closely related to the pathogenesis of cataract.Objective This study was to investigate the differential distribution and expression of AQP0 and AQP1 in lenses with age-related cataract and explore its effect on pathogenesis of age-related cataract.Methods Seventeen anterior capsular membrane samples and nucleus samples of lenses were collected from age-related cataract patients during the small incision nonphacoemulsification cataract extraction,and 6 normal lens samples were obtained from health donors in the First Affiliated Hospital of Fujian Medical University.The expression and distribution of AQP1 and AQP0 in the lenses were detected by immunohistochemistry,and the relative expression levels of AQP1 and AQP0 proteins in the lenses were assayed by using Western blot assay.This study protocol was approved by Ethic Committee of this hospital,and written informed consent was obtained from each patient.Results Immunohistochemistry showed that in the normal lenses,AQP1 expressed mainly in LECs;while AQP0 primarily expressed in fiber cells of the lens cortex and nucleus.The relative expression levels of AQP1 and AQP0 in the lenses with age-related cataract (absorbance) were 0.223±0.008 and 0.118±0.015,which were significantly lower than 0.246±0.007 and 0.149±0.007 in the normal lenses (t =-4.508,-3.291,both at P＜0.01).Western blot revealed that the relative expression levels of AQP1 and AQP0 in the lenses with age-related cataract (absorbance) were 0.663 ± 0.012 and 0.599 ± 0.015,which were significantly reduced in comparison with 0.844±0.041 and 0.955 ±0.064 in the normal lenses (t =-7.492,P＜0.05;t =-9.570,P＜0.01).Conclusions AQP1 and AQP0 distribute in different sites of lenses.The expressions of AQP1 and AQP0 are obviously down-regulated in lenses with age-related cataract,suggesting that AQP1 and AQP0 probably play different roles in the pathogenesis of age-related cataract.

Background The imbalance of cell cycle regulation results in proliferative vitreoretinopathy (PVR).Studing the effects of cyclin dependent kinase (CDK) and CDK inhibitor (CKI) on the cell cycle regulation of retinal pigment epithelial (RPE) cells and fibroblasts in PVR formation is of important significance.Objective This study was to investigate the expressing trend of p21 ,p27 and CDK inhibitors in retinas of different ages of rabbits and explore the relationship between p21 or p27 and cell growth.Methods Nine clean New Zealand rabbits were assigned to 10-week group,20-week group and 30-week group according to the age and 3 rabbits for each.The eyeballs were enucleated binocularly after the animals were sacrificed and retinas were isolated.Real-time PCR and Western blot were employed to detect the expressions of p21 and p27 mRNA and their proteins in retinas of the rabbits.The use and care of the animals followed the Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Committee.Results The relative expressing levels of p21 mRNA were 1.631±0.063,1.506±0.012 and 1.585 ±0.015, and those of p27 m RNA were 1.581 ± 0.048,1.470 ± 0.012 and 1.490 ±0.013 in the 10-week group,20-week group and 30-week group, respectively, showing significant differences among the groups (p21 mRNA: F=9.311,P=0.014;p27 mRNA: F=12.360, P=0.007) , and the p21 and p27 mRNA expressing levels were significantly higher in the 10-week group and 30-week group than those in the 20-week group (all at P＜ 0.05).The expressing levels of p21 protein were 0.675 ± 0.061,0.089 ±0.001 and 0.200 ± 0.007, and those of p27 protein were 0.928±0.019,0.183±0.005 and 0.576±0.089 in the 10-week group,20-week group and 30-week group, respectively, with remarkable differences among the groups (p21 : F =228.905, P＜0.001;p27 : F =148.957,P＜0.001), and the expressions were significantly raised in the 10-week group and 30-week group in comparison with the 20-week group (all at P＜0.01).Significantly positive correlations were found in the expressing levels of p21 and p27 both in transcriptional and protein levels (mRNA : r =0.906, P＜0.01;protein : r =0.913, P＜0.01).Conclusions The expressions of p21 and p27 up-regulate in the retinas of developing stage of rabbits but gradually reduce with adultness.However, p21 and p27 levels appear to be increasingly raised with aging of the rabbits.It is implied that p21 and p27 play a balancing role in the process of cycle regulation in retina cells.

Background Retinal microglia (RMG) plays an important role in the pathogenesis of retinal degenerative diseases,while chemokine CX3CL1 participates in the regulation of steady-state of microglia.It has been determined that bone marrow-derived mesenchymal stem cells (BMSCs) have a remarkable role to modulate the immune response and protect the central nervous system through the release of soluble factors in a paracrine fashion and further affect the functional behavior of cells.However,whether BMSCs are able to interact with RMG and activate related signaling pathway for the maintaining of homeostasis in the retina is still unclear.Objective The aim of this study was to investigate the interaction between BMSCs and lipopolysaccharide (LPS)-activated RMG in vitro,and dissect the effects of CX3CL1/CX3CR1 signaling pathway on the biological behavior of BMSCs and RMG.Methods RMG was isolated from SD rats,cultured with mixed culture of retinal glial cells and purified by shaking.The cells were identified by detecting the expression of CD111b,Iba1 and glutamamine synthetase (GS) with indirect immunofluorescence assay.LPS (1 mg/ml,2 μl) was added in the medium for 24 hours to stimulate RMG,and then the cells were divided into LPS control group,BMSCs group (cocultured with BMSCs for 24 hours) and CB-BMSCs group (cocultured with CX3CL1-blocking-BMSCs for 24 hours).The cells without LPS stimulation served as the blank control group.The functions of RMG,including the release content of tumor necrosis factor-α (TNF-α) and interleukin-1 β (IL-1β),the proliferation,phagocytosis,and migration of RMG were examined.Results RMG was successfully isolated and harvested from SD rats by using mixed culture of retinal glial cells and purified by shaking.CD11b and Iba1 showed the positive expression with the green fluorescence in the cells and GS was absent.The contents of TNF-αt in the cell supernatant were (2.55 ±0.97) ng/ml,(24.91 ±3.07) ng/ml,(20.38 ±2.97) ng/ml and (24.90 ± 1.88) ng/ml in the blank control group,LPS control group,BMSCs group and CB-BMSCs group,respectively,showing a significant difference among the groups (F=119.90,P＜0.05).The contents of IL-1 β in the cell supernatant were (1.12±0.36) ng/ml,(10.40±2.76) ng/ml,(7.00± 1.75) ng/ml and (9.55 ± 1.11) ng/ml in the blank control group,LPS control group,BMSCs group and CB-BMSCs group,respectively,showing a significant difference among the groups(F =34.96,P＜0.05).The secretory volume of TNF-α and IL-1 β were evidently lower in the BMSCs group than those in the LPS control group (both at P＜0.05),and no significant differences were found in the secretory volume of TNF-α and IL-1β between CB-BMSCs group and LPS control group (both at P＞0.05).The proliferative rate of RMG was lower in the BMSCs group than that in the LPS control group (P＜0.05),while there was no statistical difference between BMSCs group and CB-BMSCs group (P＞0.05).The mean fluorescence intensity (MFI) and the number of migrated RMG were considerably different among the four groups (F=70.55,15.49,both at P＜0.05),and those in the BMSCs group were significantly increased in comparison with the LPS control group (both at P＜0.05),while there was no significant difference between CB-BMSCs group and LPS control group (both at P＞0.05).Conclusions BMSCs could suppress the proliferation of LPS-activated RMG.Moreover,BMSCs might inhibit proinflammatory cytokines releasing,enhance phagocytosis and migration capabilities of RMG via CX3CL1/CX3CR1 signaling pathway.

Background Laser peripheral iridoplasty (LPI) is widely used in the treatment of glaucoma by flattening the iris and widening angle of anterior chamber (AA).However,no evidence suggests the optimal site of LPI in iris.Objective This study was to compare the therapeutic effects of LPI at different sites of iris for glaucoma.Methods Glaucoma models were established in the right eyes of 40 healthy adult male pigment rabbits by intrachamber injection of 0.1 ml compound carbomer solution with 0.3％ carbomer and 0.025％ dexamethasone.The models were randomly divided into model control group,corneoscleral limbus group,one spot from corneoscleral limbus group and two spots from corneoscleral limbus group.LPI was performed at corresponding site of iris by 532 nm argon laser with the spot diameter 500 μm,energy 300 mW,exposure time 0.3 seconds and laser number 24 spots,and the rabbits in the model control group did not receive LPI.Intraocular pressure (IOP),coefficient of outflow facility (C value) were measured and calculated with Schi(o)tz tonometer before LPI and 2,4,7,14 and 30 days after LPI,and anterior chamber depth (ACD),AA,anterior chamber angle opening distance within 500 μm radius from scleral spur (AOD500) were measured with ultrasound biomicroscope (UBM).The eyeballs were extracted 30 days after LPI,and the chamber angle were observed under the optical microscope after hematoxylin and eosin staining.The use and care of the animals complied with the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health.Results UBM showed that compared with the model control group,the anterior chamber angle was evidently widened in all the LPI groups,with the best effectiveness in the one spot from corneoscleral limbus group and the worst one in the two spots from corneoscleral limbus group.Compared with the model control group,the IOP was evidently reduced,and C values,AA and AOD500 were significantly increased in the corneoscleral limbus group,one spot from corneoscleral limbus group and two spots from corneoscleral limbus group after LPI,showing significant differences among the four groups (IOP:Fgroup =16.848,P ＜ 0.01;C value:Fgroup =9.629,P ＜ 0.01;AA:Fgroup =62.336,P＜0.01;AOD500:Fgroup =77.779,P ＜ 0.01).IOP was reduced and C value,AA and AOD500 were increased in 2,4,7,14 and 30 days after LPI as compared with before LPI,with significant differences over time (IOP:Ftime =3.041,P =0.011;C value:Ftime =4.311,P＜0.01;AA:Ftime =14.627,P＜0.01;AOD500:Ftime =20.378,P＜0.01).Compared with the model control group,the ACD was significantly increased in the corneoscleral limbus group and one spot from corneoscleral limbus group,and that in the two spots from corneoscleral limbus group was significantly reduced,and the ACD was insignificantly increased over time after LPI (Fgroup =18.017,P＜0.01;Ftime =0.022,P =1.000).Hematoxylin and eosin staining showed that the trabecular meshwork and adhesion of tissure were reopened and the anterior chamber angle was widened after LPI.Conclusions LPI can widen anterior chamber angle and lower the IOP.The best therapeutic outcome for glaucoma is displayed when LPI is performed at the iris site corresponding to one spot from the corneoscleral limbus.