PURPOSE:In search of the mechanism for photic retinal injury. METHODS:A visible light damage model was established in the primary cultured healthy,adult human RPE cells by using intense fluorescence light (2 400 Lx). RESULTS:Electron microscopy revealed swelling of the mitochondria and obscurity of nuclear membranous structure in the light damaged cells. The decrease or dissolution of organelle,vacuolization of cytoplasm and myelinic degeneration were found in some severely damaged cells. The level of intracellular SOD was decreased to 41% of that of the controls (P＜0.05). CONCLUSION:The structure of the RPE was damaged by the light radiation and the level of intracellular SOD was decreased. These suggested the light damage might be associated with the production of free radicals and the lipid perioxide reaction in membranous structure of cell. （Chin J Ocul Fundus Dis,1996,12: 174-175 ）
Purpose To investigate retinoic acid (RA) induced apoptosis in retinal pigment epithelial (RPE) cells. Methods 10-5、10-6、10-7 mol/L were added to cultured PRE cells.Aridine orange fluorescence and TdT-mediated dUTP nick end labelling(TUNEL) techniques were used to observe apoptotic changes. Resultss 10-5、10-6、10-7 mol/L RA induced apoptosis in RPE cells.Cell shringkage,chromatin condensation and nuclear DNA fragmentation of RPE cells were observed by TUNEL technique.When 10-7、10-6、10-5mol/L RA treated RPE cells for 5 days,apoptotic index(AI)was 36.9%、4409% and 61.4% respectively,and 48.0%、59.9%、74.2% for 6 days.At the same concentration of RA,AI increased when time prolonged．At the same day,AI increased when the concentration of RA rose.There was significant difference in the results(Plt;0.05). Conclusion Our results showed that RA-induced apoptosis in RPE cells was detected with a good dose and time response. (Chin J Ocul Fundus Dis,1998,14:153-155)
Objective To investingate the ultrastructural changes of retinal pigment epithelium(RPE) and its permeability in spontaneously hypertensive rats(SHR)and explore the relation between these changes and hypertensive retinopathy.MethodsThe ultrastructure of RPE cells in the SHR aged five,six,seven months wasobserved with transmission electronmicroscope and compared to its normotensive control strain(WKY) with the same age.Then,lanthanum tracer procedures were carried out to investigate pathological changes of the blood-retinal barrier.Results (1)In SHR the main pathological changes involved swelling of mitochondria,enlargement of endoplasmic reticula,decrease of RPE cell infolding,and sparseness of microvilli.These degenerations were more serious in older rats with higher blood pressure.(2)The breakdown of outer blood-retinal barrier with permeation of lanthanum tracers were evident in SHR aged six or seven month,however,in WKY and five-month SHR the traces were prevented from passing by tight junctions.ConclusionThe degeneration of RPE owing to ischemia and anoxia arises in early periosd of hypertensive retinopathy.The pathological changes of ultrastructure and permeability might interact with the damage of visual cells and play a main role in the hypertensive retinopathy.
Purpose To investigate the effects of human vitreous fluid on proliferation of cultured human retinal pigment epithelial (RPE) cells and vascular endothelial cell lines(VEC304). Methods Human RPE cells and VEC304 were cultured and treated in different human vitreous-conditioned medium (VCM) with or without serum, vitreous volume concentrations of VCM were 1∶8, 1∶4 and 1∶2. Cells proliferation was assayed by tetrazolium (MTT) colorimetry at the 2nd, 4th and 6th day respectively. Results In the presence of serum, 1∶4, 1∶2 VCM had a significantly stimulative effect on RPE cells proliferation compared with control group at the 2nd, 4th, and 6th day retrospectively (P＜0.01), but exerted a bly inhibitory effect on VEC304 proliferation compared with control group at the 2nd, 4th, and 6th day retrospectively (Plt;0.05). In the absence of serum, only 1∶2 VCM had a stimulative effect on RPE cells growth compared with control group at the 2nd day (P＜0.05) and obviously at the 4th and 6th day respectively (P＜0.01). Conclusion Human vitreous fluid influences human RPE cells and VEC304 growth in vitro. This result suggests that vitreous may play different role in proliferative vitreoretinopathy and intraocular neovascular disease. (Chin J Ocul Fundus Dis, 2002, 18: 140-142)
Porpose To investigate the optimal concentration of epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) on DNA synthesis and their synergism indensity arrested human retinal pigment epithelial (RPE) cells. Methods Growth factor effects in cultured human RPE of the 6th generation were assessed by [3 H]-thymidine incorporation and radioautography. Results EGF and bFGF were potent stimulators when used alone,and their optimal concentrations were 10ng/ml in DMEM and 1ng/ml in 2% serum DMEM.When used in combination (10ng/ml EGF and 10ng/ml bFGF),they caused a significant enhancement of [3 H]-thymidine incorporation about 2.96 times. Conclusion EGF and bFGF were potent stimulators in RPE cells,and demonstrated synergism in their action. (Chin J Ocul Fundus Dis,1998,14:98-100)
Objective To examine the influence of retinal pigment epithelium(RPE) cells on antigen-specific activatedlymphocytes in vitro,and to explore the role of RPE cells in the immune privilege of the eye. Methods Co-culture systems of RPE cells with antigen-specific T lymphocyte lines and resting T lymphocytes were established in vitro．Induction of apoptosis was detected by genomic DNA electrophoresis,DNA in situ end-labelling and flow cytometry. Results RPE cells induced apoptosis in antigen-specific activated T lymphocytes. 24 hours after culture,the signs of apoptosis appeared in lymphocytes co-incubated with RPE cells．As time of co-culture went on,the number of apoptosic cells increased．Quantitative analysis of apoptosic cells showed that apoptosic cells accounted for 5.95% after 24 hours, 9.38% after 48 hours,and 17.95% after 72 hours．In contrast,RPE cells induced few apoptosis in resting T lymphocytes. Conclusions These results suggest that RPE cells possess the ability to induce the apoptosis of invading lymphocytes. This phenomenon serves as a restrain mechanism of immune response and may be of vital importance in the maintenance of immune privilege in posterior segment of eye and in the protection of eye from the damage of immunogenic inflammation. (Chin J Ocul Fundus Dis, 1999, 15: 241-244)
Objective To explore the expression and activation of transcription factor E2F1 in cultured human retinal pigment epithelium (RPE) cells. Methods Cultured human RPE cells were divided into two groups after synchronization: one was cultured in Dulbecco′s modified Eagle′s medium (DMEM) without serum; the other was cultured in DMEM supplemented with 20% serum of newborn calf. The expressions of E2F1 protein in two groups were detected by Western blot analysis. The E2F1-DNA binding activities were measured by gel mobility-shift assay(EMSA). Results E2F1 protein of 60 000 molecular weight was detected in the nuclear extract of human RPE cells, and serum stimulation could increase its expression（P＜0.001）. EMSA exhibited the increased binding activity of E2F1 in the serum-stimulated RPE cells with DNA. Conclusions E2F1 is expressed in the nuclei of human RPE cells. Serum stimulation can increase its protein expression as well as binding activity, so as to play a regulation role of gene transcription. (Chin J Ocul Fundus Dis, 2002, 18: 224-226)
To investigate the characteristics of fundus autofluorescence (AF) in the leakage site of central serous chorioretinopathy (CSC). Methods Sixty-seven CSC patients (67 eyes) underwent fundus fluorescein angiography (FFA) examination with a confocal scanning angiography (HRA2). Autofluorescence was elicited by the wavelength of 488 nm. The patterns of autofluorescence corresponding to the leakage site on FFA were observed. All the enrolled patients were grouped by age (agele;45 in 47 eyes and age ＞45 in 20 eyes) and courses (acute CSC in 25 eyes and chronic or recurrent CSC in 42 eyes), the patterns of autofluorescence were analyzed respectively. Results There are 4 patterns of AF in the leakage site on FFA of CSC patients: no AF changes, punctuate hypo-AF, expanded hypo-AF or speckled AF, hyper-AF. The percentages of those patterns in all 67 eyes are 52.2%, 23.9%, 14.9% and 9.0% respectively. The percentages of those patterns in the group of age le;45 (n=47) are 55.3%, 23.4%, 14.9% and 6.3% respectively. The percentages of those patterns in the group of age ＞45 (n=20) are 45.0%, 25.0%, 15.0% and 15.0% respectively. The percentages of those patterns in acute CSC (n=20) are 80.0%, 16.0%, 4.0% and 0% respectively. The percentages of those patterns in chronic or recurrent CSC (n=42) are 35.7%, 28.6%, 21.4% and 14.3% respectively. Conclusion There are different patterns of fundus autofluorescence in different age and courses of CSC patients.
Objective To observe the inhibition effect of curcumin on the proliferation of rabbit retinal pigment epithelial (RPE) cells and investigate its mechanism. Methods The 4th generation of RPE cells were selected and divided into curcumin group and blank control group. The concentration of curcumin included 10, 15, and 20 mu;g/ml. The MTT assay was used to evaluate the inhibition effect on the proliferation of RPE cells at the 24th, 48th, 72nd and 96th hour after cultured with curcumin (10, 15, and 20 mu;g/ml). The IC50 value of curcumin at different time points were calculated by Linear Regression. Flow cytometry was used to detect the effect on the cell cycle at the 72nd hour after cultured with curcumin (15 mu;g/ml); the expression and apoptosis of proliferating cell nuclear antigen (PCNA) were also determined at the 24th,48th, and 72nd hour after cultured with curcumin (15 mu;g/ml) respectively. The configuration of RPE cells were observed by transmission electron microscope. Results The IC50 value of curcumin at the 24th,48th, 72nd and 96th hour was 29.31, 17.50, 13.24, and 10.99 mu;g/ml respectively. Cell cycel analysis indicated that curcumin blocked cells in G0/G1 phase. At the 24th, 48th, and 72nd hour after cultured with curcumin (15 mu;g/ml), the expression of PCNA of RPE cells were 565.04plusmn;23.60, 473.61plusmn;36.88, and 396.15plusmn;32.45; the apoptosisrate were （12.83plusmn;0.13）%，（32.27plusmn;4.51）%，（56.81plusmn;8.67）%, respectively. The differeces of curcumin groups compared with the control group were significant (P＜0.05). Apoptosis of RPE cells was observed under transmission electron microscope. Conclusions Curcumin can inhibite the proliferation of RPE cells by inhibit the synthesization of PCNA and inducing the apoptosis of RPE cells. Curcumin may become a potential drug to prevent and treat PVR.