ObjectiveTo observe the effect of complement receptor 1 (CR1) on barrier of cultured human retinal epithelial cells (hRPE) under complement-activated oxidative stress. MethodsThe third to fifth passage of hRPE cultured on Transwell insert were used to establish a stable hRPE monolayer barrier. The hRPE monolayer barrier was exposed to 500 μmol/L ten-butyl hydroperoxide and 10% normal human serum to establish the hRPE monolayer barrier model of complement-activated oxidative stress in vitro. hRPE monolayer barriers under complement-activated oxidative stress were divided into two groups including model group and CR1 treatment (1 μg/ml) group. Model group and CR1 treatment group were treated with 1 μl phosphate buffer solution (PBS) or CR1 for 4 hours. Normal hRPE monolayer barrier were used as control in transepithelial resistance (TER) measurement experiment. TER was measured to evaluate the barrier function of hRPE. The hRPE-secreted vascular endothelial growth factor (VEGF) and chemokine (C-C Motif) Ligand 2 (CCL2), together with complement bioactive fragments (C3a, C5a) and membrane-attack complex (MAC) in the supernatant were detected by enzyme-linked immune sorbent assay. ResultsStable hRPE monolayer barrier was established 3 weeks after hRPE seeded on Transwell insert. Complement-activated oxidative stress resulted in a sharp decrease of TER to 54.51% compared with normal hRPE barrier. CR1 treatment could significantly improve TER of barrier under complement-activated oxidative stress to 63.48% compared with normal hRPE barrier(t=21.60, P < 0.05). Compared with model group, CR1 treatment could significantly decrease the concentration of VEGF and CCL2 by 11.48% and 23.47% secreted by hRPE under complement-activated oxidative stress (t=3.26, 2.43; P < 0.05). Compared with model group, CR1 treatment could also decreased the concentration of C3a, C5a and MAC by 24.00%, 27.87%, 22.44%.The difference were statistically significant (t=9.86, 2.63, 6.94; P < 0.05). ConclusionsCR1 could protect the barrier function of hRPE cells against complement-activated oxidative stress. The underlying mechanism may involve inhibiting complement activation and down-regulating the expression of VEGF and CCL2.
The ultra.structural changes of photic injury to the retina of two patients caused by indirect ophthalmoscope were studied.The duration of exposure was 20 minutes. Under transmission electron microscope, dilation of the choroidal vessels,swelling and vacule formation at all retinal layers, disparsed pyknotic nuclei among the outer nuclear layer and swelling of the nerve fibers were found,The retina of the pregnant woman was injured more severely than that of the male patient. （Chin J Ocul Fundus Dis,1994,10:77-79）
ObjectiveTo observe the regulation of PTB-associated splicing factor (PSF) exerts on phosphatidylinositol 3 kinase (PI3K)/Akt signaling pathway in cultured retinal pigment epithelial (RPE) cells. MethodsARPE-19 RPE cells were divided into five groups including PSF overexpression (0.25, 0.50, 1.00 μg of pEGFP-C2-PSF plasmid DNA), PSF overexpression control (pEGFP-C2 empty vector DNA), PSF inhibition (0.25, 0.50, 1.00 μg of pGenesil-PSF-RNAi plasmid DNA), PSF inhibition control (pGenesil-scramble-siRNA empty vector) and sham transfected group (treated with lipofactamine 2000 reagent, but without adding plasmid DNA) groups. After transfecting with plasmid DNA, the cells were stimulated with insulin-like growth factor-1 (IGF-1). IGF-1-stimulated ARPE-19 cells were also treated with Wortmannin and /or PSF over-expression. WST-1 assay was used to detect the proliferation rates, the VEGF mRNA levels were analyzed using real time polymerase chain reaction (PCR), the levels of phosphorylation Akt and total Akt expression were measured by western blotting. ResultsAfter IGF-1 stimulation, the difference of the cell proliferation rates between PSF overexpression group, PSF overexpression control group and sham transfected group was statistically significant (F=29.728, P<0.05). The difference of the cell proliferation rates between PSF inhibition group, PSF inhibition control group and sham transfected was also statistically significant (F=14.121, P<0.05). Compared with control group, the VEGF mRNA levels was decreased in PSF overexpression group (P=0.000 3), but increased in PSF low expression group (P=0.030 9). Furthermore, overexpression of PSF could down-regulate the activation of pAkt after IGF-1 stimulation. When combined with Wortmannin treatment, the VEGF mRNA levels in PSF overexpression group was significantly lower than the control group (P<0.05). ConclusionsAfter IGF-1 treatment, PSF plays a role in suppressing the proliferation and VEGF expression in RPE cells by inactivating PI3K/Akt signaling pathway.
ObjectiveTo observe confocal scanning laser ophthalmoscope (cSLO) based retinal imaging and color fundus camera in pigment epithelial detachment (PED) of polypoidal choroidal vasculopathy (PCV).MethodsPED of 30 patients (32 eyes) were recruited from June 2016 to June 2017 in the Beijing Tongren Hospital who were detected in high-definition OCT (HD-OCT) and diagnosed as PCV by FFA and ICGA. There were 16 males (17 eyes) and 14 females (15 eyes); aged from 50-83 years, with the mean age of 66.59 years. The photographs of ocular fundus including color fundus camera, cSLO imaging, HD-OCT, FFA and ICGA were analyzed. Multimodal imaging results were regarded as gold standard. Sensitivity and specificity were calculated in serous and hemorrhagic PED diagnosis using color fundus camera and cSLO imaging. The positive number of PED was used to compare between two modes fundus imaging by using χ2 test.ResultsTwenty serous PED eyes, 3 hemorrhagic PED eyes and 9 serous/hemorrhagic PED eyes were determined using multimodal imaging. The sensitivity and specificity of color fundus camera were 45% and 100% in detecting serous PED and 100% and 91% in detecting hemorrhagic PED. The sensitivity and specificity of cSLO imaging were 83% and 100% in detecting serous PED and 50% and 86% in detecting hemorrhagic PED. The positive number of serous PED in cSLO imaging was significantly higher than color fundus camera (χ2=7.752, P=0.011). The positive number of hemorrhagic PED in cSLO imaging shows no obvious difference compared with color fundus camera (χ2=1.164, P=0.419).ConclusionThe sensitivity and positive number of detecting serous PED with PCV in cSLO fundus imaging were higher than the color fundus camera technology.
Replacement of diseased retinal pigment epithelium (RPE) cells with healthy RPE cells by transplantation is one option to treat several retinal degenerative diseases including age-related macular degeneration, which are caused by RPE loss and dysfunction. A cellular scaffold as a carrier for transplanted cells, may hold immense promise for facilitating cell migration and promoting the integration of RPE cells into the host environment. Scaffolds can be prepared from a variety of natural and synthetic materials. Strategies, such as surface modification and structure adjustment, can improve the biomimetic properties of the scaffolds, optimize cell attachment and cellular function following transplantation and lay a foundation of clinical application in the future.
Fluorescein angiography(FA)was performed in 31 pigmented rebbits.The angiograms were evaluated as prints and as negative film under a light microscope.The patterns of retinal pigment epithelial(RPE)cells were studied by scaning electron microscopy and fluorescein light one,compared with other rabbits belonging to the same species.In 58 eyes,we observed the hexagonal pattern of RPE cell.It showed central hypofluorescent area surrounded by hyperfluorescent rim,which was easily seen away from the medullary rays by three or more disc diameters and became larger in the periphery than that in the posterior pole.There were no finding in four lightly pigmented eyes. （Chin J Ocul Fundus Dis,1994,10:226-228）
ObjectiveTo assess the occurrence of CNV in patients presenting with flat irregular pigment epithelial detachments (FIPED). MethodsForty-five patients (49 eyes) with FIPED on OCT were enrolled in this retrospective study. There were 25 males (28 eyes) and 20 females (21 eyes). The mean age was 61.022±9.292 years. FFA, ICGA, spectral domain OCT and OCT angiography (OCTA) were performed in all patients during the same period. The FIPED was defined as an irregular elevation of the RPE allowing distinct visualization of Bruch’s membrane on OCT B-scan. The abnormal vascular signals from the deep retinal layer to the choroid layer on OCTA was defined as CNV. The CNV was classified into a type 1 CNV and a type 2 CNV according to the OCT characteristics. The CNV was classified into a typical and occult CNV according to the characteristics of the FFA image. Of all 49 eyes, fundus angiography revealed 18 eyes (36.7%) with CNV, and 31 eyes (63.3%) with no characteristic signs of CNV. FFA examination found that CNV in 8 eyes (classic CNV in 1 eyes, occult CNV in 7 eyes), which confirmed by OCT were type 1 CNV; transmitted fluorescence in 41 eyes. ICGA examination showed that CNV-like hyperfluorescence spots in 18 eyes, suspicious hyperfluorescence spots in late stage in 20 eyes, and choroidal high permeability in 11 eyes, respectively; and 18 CNV eyes were confirmed to be type 1 CNV by OCT. To compare the detection of CNV by OCTA and fundus angiography. ResultsOf the 49 eyes with FIPED, OCTA detected 36 eyes (73.5%) of type 1 CNV, and full or partial strong reflex signals were seen in FIPED; 13 eyes (26.5%) were not associated with CNV, and some strong reflection signals were found in FIPED in 9 eyes, 4 eyes with weak reflection signal. The FFA was examined for 1, 7 eyes of the classic and occult CNV, which confirmed to be type 1 CNV by OCTA. Among the 18 eyes with CNV which detected by ICGA, OCTA also found type 1 CNV. Among the 20 eyes with ICGA’s late suspicious strong fluorescent spots, OCTA showed 17 eyes of type 1 CNV; in 11 eyes with high choroidal permeability, OCTA showed type 1 CNV in 1 eye. Among the 36 eyes with CNV which detected by OCT, there were SRD in 32 eyes, no SRD in 2 eyes and retinal interlamellar cavities in 2 eyes. ConclusionOCTA can detect 73.5% of FIPED eyes with CNV. Compared with traditional fundus angiography, OCTA has a higher detection rate of CNV under FIPED. The FIPED of the internal strong reflection signal has a certain diagnostic value for the type 1 CNV.
ObjectiveTo observe the expression of hot shock protein 47 (HSP47) in pre-retinal membrane of proliferative vitreoretinopathy (PVR) and the influence of transforming growth factor-β2 (TGF-β2) on the expression of HSP47 in retinal pigment epithelial (RPE) cell. MethodsPre-retinal membranes were collected and observed by hematoxylin-eosin, Masson and immunohistochemical staining. Cultured ARPE-19 cells were treated with TGF-β2 at serial concentration (0, 1, 5, 10 ng/ml) and time (0, 12, 24, 48 hours), respectively. And then the mRNA and protein expressions of HSP47 and Col-Ⅰ were measured by fluorescence quantitative reverse transcription polymerase chain reaction and Western blot at the same time. ResultsA lot of epithelial cells with pigmental particles were observed in pre-retinal membranes of PVR, much accumulated collagen protein was observed in the specimens, and HSP47 positive expression was bserved in cytoplasm and stroma of most of the epithelioid cells. Compared with 0 ng/ml group, the expressions of HSP47 mRNA in ARPE-19 were up-regulated by 1.32, 2.35, 1.85 fold, significant differences were observed in all groups (F=27.21, P<0.05); the expressions of protein were up-regulated by 2.33, 2.89, 2.60 fold, significant differences were observed in all groups (F=39.78, P<0.05). The expressions of Col-Ⅰ mRNA were up-regulated by 1.29, 1.52, 2.11 fold, significant differences were observed in all groups (F=23.45, P<0.05); the expressions of protein were up-regulated by 1.18, 1.49, 2.11 fold and significant differences were observed in all groups (F=29.10, P<0.05). Compared with 0 hour group, the expressions of HSP47 mRNA were up-regulated by 1.56, 1.84, 2.86 fold in ARPE-19 cells stimulated by 5 ng/ml TGF-β2 for 12, 24 and 48 hours, and the differences were all significant (F=31.56, P<0.05); the expressions of protein were up-regulated by 2.08, 2.37, 2.80 fold, and the differences were all significant (F=49.18, P<0.05). The expressions of Col-Ⅰ mRNA were up-regulated by 1.57, 1.86, 2.78 fold and the differences were all significant (F=54.43, P<0.05), the expressions of protein were up-regulated by 1.38, 1.59, 2.16 fold and the differences were all significant (F=42.52, P<0.05). ConclusionTGF-β2 may play a role in the pathologic process of PVR by promoting the expression of HSP47 and then increasing the synthesis and accumulation of Col-Ⅰ.