Objective To observe the protective effects of Na2SeO3 on the damage of retinal neuron induced by microwave. Methods Cultured fluids of retinal neuron were divided into 4 groups,including 1 group of control, according to the concentration of Na2SeO3 in cultured fluid and then exposed to 30 mW/cm2 microwave for 1 hour.The targets of lipid peroxidation and the concentration of selenium in cells were measured.Apoptosis detection was taken by TUNEL detection kit. Results The activity of SOD and GSH-Px rised,meanwhile the content of MDA and the amount of apoptosis cells decreased in 1times;107 mol/L group compared with the group without Na2SeO3.The other groups was superior in antioxdant capacity to 1times;107 mol/L group. Conclusion Na2SeO3 might be possessed of the effect of protecting the damage of retinal neuron induced by microwave. (Chin J Ocul Fundus Dis,2000,16:97-99)
Objective To observe the effect of blue light on apoptosis of cultured human retinal pigment epithelial (RPE) cells in vitro. Methods Human RPE cells were exposed to blue light, and the cells were divided into 3 groups: group A, with various intensity of illumination; group B: with same intensity but different time of illumination; group C: with same intensity and time of illumination but different finish time of the culture. The apoptosis of RPE cells was observed by TdT-dUTP terminal nick-end labeling (TUNEL) and annexin V-fluoresein isothiocyanate (FITC)/propidium iodide (PI) flow cytometry, and transmission electron microscopy. Results The positive cells stained by TUNEL shrinked and turned round, whose nuclei concentrated and congregated like the crescent or hat. Cracked nuclei and membrane bleb were found. Swollen mitochondrial, disappeared inner limiting membrane of mitochondria, and dilation of the rough endoplasmic reticulum with metabolite were observed by transmission electronmicroscopy. In group A, mild damage of RPE cells was found when the threshold value of the intensity of illumination was less than（500±100）lx, and the apoptosis and necrosis of RPE cells aggravated as the intensity of illumination increased; in group B, as the time of illumination extended, the number of apoptotic RPE cells didn′t increase while the necrosis increased; in group C, 6 and 12 hours after illumination, apoptosis of cells was the main injury, while apoptosis with necrosis was found and necrotic cells increased as the time of illumination was prolonged. Conclusions Illumination with blue light may cause damages of human RPE cells in vitro, with the modalities of apoptosis, apoptotic necrosis and necrosis. The extent of injury is dependent on intensity and duration of the illumination. (Chin J Ocul Fundus Dis, 2005, 21: 384-387)
Objective To observe whether apoptosis was involved in cells of aspiration fluid from vitrectomy for proliferative vitreoretinopathy(PVR),and whether there was an association with expression of Fas antigen(Fas )and Fas ligand (FasL). Methods Cytocentrifuge slides of 11 fresh vitreous specimenｓ of PVR were prepared to be stained by TUNEL met hod for detection of apoptosis and by immunohistochemical technique for detection of Fas,FasL，and cytokeratin (CK),a cell-type specific antigen. Results Fas and FasL were expressed in normal human retina.Fas,FasL,CK，and apoptosis were found in all preparations.TUNEL-positive cells were 20.53% in total cells.70.35%,51.58%，and 82.97% of cells highly expressed Fas,FasL，and CK,respectively.The linear correlation coefficient of Fas and apoptosis was 0.99(Plt;0．001). Conclusion Vitrectomy specimens of PVR showed expression of Fas，FasL，and apoptosis.Prominent Fas and FasL expressions may be associated with apoptosis of proliferating retinal pigment epithelial cells in the vitreous of PVR. (Chin J Ocul Fundus Dis,1999,15:78-80)
Objective To further investigate pathologic mechanism of retinal phototrauma. Methods Twenty Wistar rats were divided into control and experimental groups.Their eyes were extracted in 12，24 and 36 hours after light exposure.HE stained retina samples were examined and TDT-mediated dUTP nick end labelling(TUNEL)method was employed to distinguish apoptotic cells. Results After 12-hour light exposure,slight vesiculation was observed in the rod outer segment of the retinas.After 24-hour light exposure,the outer nuclear layer showed predominant fractured and condensed nuclei and fragmented DNA.After 36-hour light exposure,the rod outer and inner segments were lysed and most of the nuclei in the outer nuclear layer were disappeared. Conclusions Apoptosis of photoreceptor cell is one of the important mechanisms which cause experimental retinal photoinjury of rats. (Chin J Ocul Fundus Dis, 1999, 15: 167-169)
Objective To observe the effect of exogenous basic fibrob last growth factor (bFGF) on apoptosis of cultured human retinal pigment epithelial (RPE) cells exposed to visible light,and determine the role of bFGF, fibroblast growth factor receptor 1 (FGFR1),bcl-2 and caspase-3. Methods 2000±500) lx cold white light was used. Exogenous bFGF was utilized during culture. Annexin annexin V-fluoresce in isothiocyanate/propidium iodium (V-FITC/PI) labeling,flow cytometry, Immunocytochemical staining, enzyme associated absorb examing and reverse transcriptional polymerase chain reaction (RT-PCR) were used to determine the apoptosis, the expression levels of bFGF, FGFR1, bcl-2, as well as the activity of caspase-3. Results No protective effect of bFGF was observed under the concentration 5 ng/ml.A significant inhibition of apoptosis was found in 10 ng/ml and 20 ng/ml groups （P＜0.05）. The upregulation of bcl-2 was observed in bFGF (10 ng/ml, 20 ng/ml) protreated groups（P＜0.01）.Compared to no light exposure group,all light exposure groups (including bFGF pro-treated) had higher endogenous bFGF and FGFR1 levels （P ＜0.05）, and the increase was concentration dependent.The bFGF and FGFR1 levels were higher in exogenous bFGF applied (gt;5 ng/ml) groups than light exposure groups（P＜0.05）. The caspase-3 activity was significantly inhibited in bFGF (10 ng/ml) pro-treated groups. Conclusions Human RPE cells exposed to visible light were rescued by application of exogenous bFGF in vitro.The probable protective mechanism of bFGF partly is directly binding to FGFR1 or potentiating endogenous bFGF autocrine loop,to upregulate bcl-2 and to inhibit caspase-3 activation. (Chin J Ocul Fundus Dis，2003，19：24-28)
Objective To study the effect of hypoxia on proliferation of cultured bovine retinal pigment epithelium (RPE) cells and expression of the antiapoptotic protein bcl-2. Methods The bovine RPE cells were cultured under normal and hypoxic chamber respectively. After 24 hours, the proliferation of RPE cells was evaluated by［3-(4,5-dimethylthiazole-2yl)-2,5-diphenyl tetrazolium bromide, MTT］test. At the same time, anti-bcl-2 protein antibody was examined by immuno-histochemistry method. Results The A value in the hypoxia group was higher than that in the normal group after 24 hours (P＜0.05 )in MTT-test. Positive staining for anti-bcl-2 protein antibody was seen in 72.6% cells in hypoxia group and 38.64% in normal group. The positive staining was more obvious near the nucleus, and fine granules scattered in cytoplasm of some cells. Conclusion Hypoxia can stimulate the proliferation of RPE cells and expression of antiapoptotic protein bcl-2. The results indicate that bcl-2 may play an important role in mediating the proliferation activity of RPE cells. (Chin J Ocul Fundus Dis, 2002, 18: 293-295)
Purpose To investigate bax expression and induction of apoptosis in normal cultured retinal pigment epithelium (RPE) cells ． Methods Cultured human RPE cells were transfected by PMDNA3-hbax，which incoded the whole bax gene and may be induced by Zn2+ under the MTII promoter， through lepofectin mediated protocol．The tested RPE cells were divided into three groups of A，PMDNA3-hbax transfected ;B，PMDNA3 (nude vector) transfected and C ，normal RPE cells．After transfection， DNA gel electrophoreses were perform ed ，the tested RPE cell cycles were analyzed with flow cytometry (FCM)． Results The gel electrophoretogram showed DNA ladder phenomenon，FCM confirmed the apoptosis of RPE cells PMDNA3-hbaxtransfected ， consisting of significant apoptotic peak sited before the G 1 phase and the apoptotic rate was 36%． Conclusion The foreign bax gene can be effectively conducted in to the RPE cell through lepofectinmediated protocol and induced expression ． The foreign bax overexpression may induce the cultured human RPE cell susceptibi lity to apoptosis． (Chin J Ocul Fundus Dis, 2001,17:132-134)
Objective To investigate the role of anti apoptosis gene bcl-2 in the survival of cultured uveal melanoma UM cells. Methods Antisense oligodeoxynucleotides (AS-ODN) bcl-2 were delivered with cationic lipid to primary cultured UM cells. The inhibitory effect of AS-ODN bcl-2 on proliferation of UM cells was examined by 3,-4,5 Dimethyliazol-2,5 diphenyl tetrazolium bromide (MTT) method. Using DNA ladder to determine if the UM cells had been apoptotic. Bcl-2 expression was detected by RT-PCR and Westernblot technics. Results The effect of AS-ODN bcl-2 in inhibiting the proliferation of cultured UM cells had opposite relation to dosage. It down regulated the mRNA and protein level of bcl-2 gene, and the sense ODN didn′t have this effect. Conclusion AS-ODN bcl-2 can down regulate bcl-2 expression, inhibits UM cells proliferation and induces apoptosis. (Chin J Ocul Fundus Dis, 2002, 18: 38-41)
Objective To investigate the correlation of ascorbic acid distribution and retinal susceptibility to iron toxicity of the retina.Methods Autoclaved iron particles of 5 mg and 15 mg were implanted into the vitreous cavities of 32 Spragu-Dawley (SD) rats and 9 rabbits, respectively. The retinal sections of rats and rabbits were examined after hemotoxylin-eosin (HE) staining. Apoptos is of rabbits′retinal neurons was investigated by TdT-mediated dUTP-biotin nick-end labeling (TUNEL). Chinoy′s method was used to observe the distribution of as corbic acid in the retinae of the 2 kinds of animals.Results In rats, histological and structural densification was observed only in the photoreceptor cells after implantation of the iron particles. In rabbits, however, histological and structural destruction as well as TUNEL-positive nuclei were observed in all neuronal layers of the retina 3 days after the implantation of the iron particles. Silver granules reduced by ascorbic acid from silver nitrate were observed only in the outer nuclear layer in normal rats retinae, while they were observed evenly throu ghout all layers of rabbits′retinae. Conclusions The suscept ibility of retina to iron toxicity is correlated to the distribution of ascorbic acid in retina. (Chin J Ocul Fundus Dis，2003，19：269-332)
Purpose To study the proteins expression of genes related to apoptosis of retinal cells in development of human fetus． Methods Fifty cases of retinas of human fetus aged from 12 to 38 we eks were collected and paraffin embedded sections were made． Immunohistochemical method was used． Results Fas protein was expressed by cells of ganglion cell layer, inner and outer nuclear later, which were just formed on 16th week． It was not expressed until 38th week, Fas(+) staining appeared in layers of retina． Fas-L(+) staining was detected in cells of layers of retina on 26th week and the positive staining located in ganglion cell layer on 32th week． Neuronal fiber layer was Fas-L positive． Bax positive staini ng was detected on 8th week． Bax positive nucleus were observed mainly in GCL and ONL on 16th week． It was in INL on 24th week and in Muuml;ller cells inner terminates on 26th week． After this time, all cells of retina were bax immune ne gative staining． Bcl-2(+) staining appeared in differentiating neuroblastic layer on 16th week． Beginning on 24th week, bcl-2 (+) staining was observed in glial cells of GCL and inner terminates of Muuml;ller cell． Conclusion Apoptosis of developing retinal cell may be Fas/Fas-L independent and bax may be involved in apoptosis of the cells． (Chin J Ocul Fundus Dis, 2001,17:55-57)