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.
ObjectiveTo observe the clinical characterisitics of choroidal excavation in the macula. MethodsA total of 22 patients (22 eyes) with choroidal excavation diagnosed by spectral domain high definition optical coherence tomography (HD-OCT) were enrolled in this study. The patients included 12 males (54.50%) and 57 females (45.50%). The age was ranged from 21 to 82 years old, with an average of (41.44±13.17) years. All the patients were affected unilaterally, including 9 right eyes and 13 left eyes. The corrected vision, slit lamp microscope with preset lens, fundus photography, HD-OCT and fluorescence fundus angiography (FFA)were measured for all patients. The clinical characterisitics and concomitant diseases were observed. Seventeen eyes were followed for a period between 3 to 12 months. The lesions change were evaluated by HD-OCT. ResultsThere were 18 eyes (81.8%) with symptoms of micropsia and metamorphopsia, 4 eyes (18.2%) without symptoms. The corrected vision was ranged from 0.3 to 1.2, 12 eyes (54.54%) with moderate or high myopia. Fundus examination presents yellowish-white exudation in 12 eyes (54.54%), yellowish-white exudation accompanied with hemorrhage in 9 eyes (40.91%), grayish yellow reflex halo in 1 eye (4.55%). HD-OCT showed that the retinal pigment epithelium (RPE) layer was involved in the excavation, and the photoreceptor outer segment and pigment junction (OPR) layer was disappeared in all eyes. The external limiting membrane and the photoreceptor inner segment/outer segment junction layer were preserved in 13 eyes (59.09%) and disappeared in 9 eyes (40.91%). There were 10 eyes (18.18%) with a single lesion, 4 eyes (18.18%) with idiopathic choroidal neovascularization, 4 eyes (18.18%) with punctate inner choroidopathy, 1 eye (4.55%) with polypoidal choroidal vasculopathy, 1 eye (4.55%) with macular preretinal menbrance, 1 eye (4.55%) with central serous chorioretinopathy. FFA showed hypofluorescence in early phase, hyperfluorescence in late phase, without obvious leakage. There was no noticeable changes in size and morphological changes in the follow-up period. ConclusionsChoroidal excavation in the macula occurs mostly in middle-aged people with myopia. It can be associated with many fundus diseases. The excavation is located in RPE layer, and OPR layer disappeared. Choroidal excavation in the macula develops slowly.
Objective To observe the inhibition of SARS-CoV-2 spike protein (S-protein) on the proliferation of human retinal pigment epithelium (RPE) cells. MethodsSARS-CoV-2 S-protein gene fragment expression plasmid (p3xflag-S) was constructed and transfected into human RPE, HEK293 cells. DNA sequencing was used for identification, and the expression of Flag-S was detected by Western blot. HEK293 cells were divided into the cells 1, 2, 3 and 4 and transfected with GFP11 plasmid and vector, GFP1-10plasmid and vector, transfected with GFP11 and pCMV-HA-ACE2 plasmid, GFP1-10 and p3xflag-S plasmid. Cell 1 was co-cultured with cell 2 (control group 1), cell 2 with cell 3 (control group 2), cell 3 with cell 4 (observation group), and cell 1 mixed with cells 2, 3 and 4 (control group 3). Bright-field microscopy and fluorescence microscopy were used to observe cell fusion. RPE cells were divided into control group and overexpression S-protein group. The cell cycle was detected by flow cytometry; the cell proliferation level was detected by Counting Kit 8 (CCK-8); and the S-protein expression level in RPE cells was detected by Western blot. The Student’s t-test was performed for comparison between groups. ResultsDNA sequence assay showed that S-protein cDNA was fused with flag-tagged protein. Western blot assay showed thatS-protein-related expression was elevated in transfected HEK293 cells compared with untransfected p3xflag-S cells. Large, multinucleated fused cell clusters were visible under bright-field microscopy; multiple nuclear with distinct green fluorescence were visible in the fused cells under fluorescence microscopy. Western blot assay showed elevated S-protein-related expression in transfected p3xflag-S plasmid RPE cells compared to untransfected p3xflag-S plasmid RPE cells. CCK-8 results showed that the proliferative capacity of RPE cells in the S-protein overexpression group was significantly reduced compared with the control group, with statistically significant differences (t=22.70, 16.75, 23.38; P<0.000 1). The results of flow cytometry showed that the G1 phase cells in the control and overexpression S-protein groups were 41.1% and 67.0%, respectively; compared with the control group, the G1 phase cells in the overexpression S-protein group were significantly higher, and the difference was statistically significant (t=4.76, P=0.018). The apoptosis rate was significantly increased in the S-protein overexpression group compared with the control group, and the difference was statistically significant (t=4.91, P=0.008). ConclusionOverexpression of the SARS-CoV-2 spike protein reduced the proliferation of human RPE cells.
Torpedo maculopathy is a rare, congenital lesion of RPE, which locates temporal to the macula and along the horizontal raphe. The lesion is torpedo-shaped with its torpedo-like tip pointing towards the fovea. As an incidental finding, it often affects only one eye with no damage to central visual acuity. According to its characteristics on OCT, it is divided into 2 types: typeⅠ, attenuation of outer retinal structures without outer retinal cavitation; typeⅡ, those with both attenuation of outer retinal structures and outer retinal cavitation. Diseases with pigment changes in the RPE layer similar to torpedo maculopathy include congenital hypertrophy of the RPE, RPE lesions in Gardner syndrome, etc. The main point to distinguish the disease from other diseases is its unique location and shape. Most of the torpedo maculopathy lesions are stable and do not require special treatment, but the disease can be complicated by neurosensory retinal detachment, choroidal neovascularization and so on, and symptomatic treatment is needed if necessary.
ObjectiveTo observe the influence of down-regulation of HtrA1 expression by small interfering RNA on light-injured human retinal pigment epithelium (RPE) cells. MethodsCultured human RPE cells(8th-12th generations)were exposed to the blue light at the intensity of (2000±500) Lux for 6 hours to establish the light injured model. Light injured cells were divided into HtrA1 siRNA group, negative control group and blank control group. HtrA1 siRNA group and negative control group were transfected with HtrA1 siRNA and control siRNA respectively. The proliferation of cells was assayed by CCK-8 method. Transwell test was used to detect the invasion ability of these three groups. Flow cytometry was used to detect the cell cycle and apoptosis. The expression of HtrA1 and vascular endothelial growth factor (VEGF)-A was detected by real time-polymerase chain reaction and Western blot respectively. ResultsThe mRNA and protein level of HtrA1 in the light injured cells increased significantly compared to that in normal RPE cells (t=17.62, 15.09; P<0.05). Compared with negative control group and blank control group, the knockdown of HtrA1 in HtrA1 siRNA group was associated with reduced cellular proliferation (t=6.37, 4.52), migration (t=9.56, 12.13), apoptosis (t=23.37, 29.08) and decreased mRNA (t=17.36, 11.32, 7.29, 4.05) and protein levels (t=12.02, 15.28, 4.98, 6.24) of HtrA1 and VEGF-A. Cells of HtrA1 siRNA group mainly remained in G0/G1 phase, the difference was statistically significant (t=6.24, 4.93; P<0.05). ConclusionKnockdown of HtrA1 gene may reduce the proliferation, migration capability and apoptosis of light-injured RPE cells, and decrease the expression of VEGF-A.
Severely coagulated retinae by argon laser of 20 Chinese hamsters were investigated with transmission electron-microscopy. The results revealed destruction of retinal pigment epithelium-Bruch's membrane-choroid capillary complex at the coagulated foci, and leakage of fluid and blood cells through the choroidal vessels into the subretinal space. Several days after laser burn the subretinal fluid was found to subside and the RPE cells surrounding the burned lesions started to proliferate. The smaller lesions were covered by the proliferating RPE 10 days after coagulation, but poor regeneration of RPE in large necrotic areas. Neovascularization was usually associated with obvious defect of Bruch's membrane and restoration of RPE barrier was most likely impossible. （Chin J Ocul Fundus Dis,1992,8:14-16）
In thiis study,we show thai carbachol stimulates the accumulation of inositol phosphates(InsPs)in human rellnal pigment epithelium (RPE)cells and atropine blocks the carbachol-induced effect ,suggesting the existence of musearinie acelyleholine receptors in human RPE cells. In contrast,noradrenaline,serotonin, cpidermal growth factor (EGF),isoproterenol,and NECA (5'-[N-ethyl]-carboxamido-adenosine)do not influence the basal levels of InsPs.Moreover,isoprmerenol and NECA do not affect the carhaehol elevated levels of InsPs.EGF,howcvcr,does potentiate the carhaehol stimulated elevation of InsPs in a dose-dependent manner ,suggesting an interaction between EGF and musearinie receptors in cultured human RPE cells. （Chin J Ocul Fundus Dis,1994,10:220-222）