PURPOSE: To assess the reproducibility of circumpapillary retinal nerve fiber layer (cpRNFL) thickness measurement (measurement agreement) and its color-coded classification (classification agreement) by Cirrus spectral domain optical coherence tomography (OCT) in pseudophakic eyes. METHODS: Two-hundred five participants having glaucoma or glaucoma suspected eyes underwent two repeated Cirrus OCT scans to measure cpRNFL thickness (optic disc cube 200 x 200). After classifying participants into three different groups according to their lens status (clear media, cataract, and pseudophakic), values of intra-class coefficient (ICC), coefficient of variance, and test-retest variability were compared between groups for average retinal nerve fiber layer (RNFL) thicknesses and that corresponding to four quadrant maps. Linear weighted kappa coefficients were calculated as indicators of agreement of color code classification in each group. RESULTS: ICC values were all excellent (generally defined as 0.75 to 1.00) for the average and quadrant RNFL thicknesses in all three groups. ICC values of the clear media group tended to be higher than those in the cataract and pseudophakic groups for all quadrants and average thickness. Especially in the superior and nasal quadrants, the ICC value of the cataract group was significantly lower than that of the clear media and pseudophakic groups. For average RNFL thickness, classification agreement (kappa) in three groups did not show a statistically significant difference. For quadrant maps, classification agreement (kappa) in the clear media group was higher than those in the other two groups. CONCLUSIONS: Agreement of cpRNFL measurement and its color code classification between two repeated Cirrus OCT scans in pseudophakic eyes was as good as that in eyes with clear crystalline lens. More studies are required to ascertain the effect of lens status on the reproducibility of Cirrus OCT according to different stages of glaucoma patients.
Aged ; Cataract/complications ; Cataract Extraction ; Female ; Glaucoma/complications/*pathology ; Humans ; Lens, Crystalline/cytology/pathology ; Male ; Middle Aged ; Nerve Fibers/pathology ; Optic Disk/pathology ; Pseudophakia/complications ; Reproducibility of Results ; Retinal Ganglion Cells/*pathology ; Tomography, Optical Coherence/*methods/*standards

OBJECTIVETo investigate the effect of long-term power frequency electromagnetic field (50 Hz) exposure on the proliferation and apoptosis of human lens epithelial cells (SRA01/04 cells).

METHODSSRA01/04 cells in the exponential growth phase were exposed or sham-exposed to power frequency electromagnetic field (50 Hz, 2.3 mT) for 2 hours per day, 5 days every week. After 11 weeks of exposure, the cells were collected; the cell morphology was observed under a microscope, the cell viability was measured by MTT assay, the cell cycle and apoptosis were examined by flow cytometry, and the protein expression levels of cyclin D and proliferating cell nuclear antigen (PCNA) were determined by western blot.

RESULTSCompared with the sham-exposed SRA01/04 cells, most exposed cells became rounded and more stereoscopic, and heterochromatin gathered near the nuclear membrane in some exposed cells. The MTT assay showed that the viability of exposed cells was significantly increased compared with that of the sham-exposed cells (P < 0.05). Long-term power frequency electromagnetic field exposure led to significantly increased number of cells in S phase (P < 0.05), and the proliferation index was significantly higher in the exposed cells than in the sham-exposed cells (P < 0.05). There was no significant difference in apoptotic rate between the exposed cells and sham-exposed cells (P > 0.05). The exposed cells had significantly higher protein expression levels of cyclin D and PCNA than the sham-exposed cells (P < 0.05).

CONCLUSIONLong-term power frequency electromagnetic field exposure can promote cellular proliferation and change cell cycle in SRA01/04 cells, but it has no marked effect on the apoptosis of SRA01/04 cells.

Apoptosis ; Cell Line ; Cell Proliferation ; Cyclin D1 ; metabolism ; Electromagnetic Fields ; adverse effects ; Environmental Exposure ; adverse effects ; Epithelial Cells ; cytology ; Humans ; Lens, Crystalline ; cytology ; Proliferating Cell Nuclear Antigen ; metabolism

The formation of advanced glycation end products (AGEs) has been considered to be a potential causative factor of injury to lens epithelial cells (LECs). Damage of LECs is believed to contribute to cataract formation. The purpose of this study was to investigate the cytotoxic effect of AGEs on LECs both in vitro and in vivo. We examined the accumulation of argpyrimidine, a methylglyoxal-derived AGE, and the expression of apoptosis-related molecules including nuclear factor-kappaB (NF-kappaB), Bax, and Bcl-2 in the human LEC line HLE-B3 and in cataractous lenses of Zucker diabetic fatty (ZDF) rats, an animal model of type 2 diabetes. In cataractous lenses from twenty-one-week-old ZDF rats, LEC apoptosis was markedly increased, and the accumulation of argpyrimidine as well as subsequent activation of NF-kappaB in LECs were significantly enhanced. The ratio of Bax to Bcl-2 protein levels was also increased. In addition, the accumulation of argpyrimidine triggered apoptosis in methylglyoxal-treated HLE-B3 cells. However, the presence of pyridoxamine (an AGEs inhibitor) and pyrrolidine dithiocarbamate (a NF-kappaB inhibitor) prevented apoptosis in HLE-B3 cells through the inhibition of argpyrimidine formation and the blockage of NF-kappaB nuclear translocalization, respectively. These results suggest that the cellular accumulation of argpyrimidine in LECs is NF-kappaB-dependent and pro-apoptotic.
Animals ; Apoptosis/*drug effects ; Cell Line ; Epithelial Cells/*cytology/*drug effects ; Glycosylation End Products, Advanced/*pharmacology ; Lens, Crystalline/*cytology ; Male ; Ornithine/*analogs & derivatives/pharmacology ; Pyrimidines/*pharmacology ; Pyruvaldehyde/*chemistry ; Rats

OBJECTIVETo study the effects of ecdysterone (ECR) on the expression of nuclear factor (NF)-kappaB in H2O2 induced oxidative damage of human lens epithelial cells (HLECs).

METHODSThe cultured HLECs were divided into 5 groups, i.e., the control group, the H2O2 group, the beta-estradiol (E2) group, the ECR group, and the pyrrolidine dithiocarbamate group (PDTC) group. The expression rate of NF-kappaB p65 in the HLECs were detected by flow cytometer (FCM).

RESULTSThe expression of NF-kappaB p65 occurred in normal HLECs (9. 53%). The expression rate of NF-kappaB p65 in the H2O2 group obviously increased (39.87%, P < 0.01). The expression rate of NF-kappaB p65 in the PDTC group obviously decreased (5.90%, P < 0.01). The expression rates of NF-kappaB p65 in the ECR group (13.99%) and the E2 group (25.18%) ranged between the control group and the H2O2 group, but still lower than that of the H2O2 group (P < 0.01).

CONCLUSIONSThe activation of NF-kappaB in the HLECs could be induced by H2O2 ECR with the estrogenic activity could effectively inhibit the activation of NF-kappaB.

Cells, Cultured ; Ecdysterone ; pharmacology ; Epithelial Cells ; drug effects ; metabolism ; Humans ; Hydrogen Peroxide ; adverse effects ; Lens, Crystalline ; cytology ; Oxidative Stress ; drug effects ; Transcription Factor RelA ; metabolism

OBJECTIVE: To determine the apoptosis-inducing effect of ultraviolet light (UV) on human lens epithelial cell (HLEC) and to explore the involvement of changes in ALDH1 folowing UV radiation. METHODS: HLEC was exposed to the same UV light source and was subsequently divided into 6 groups according to UV radiation time of 0 (control group), 5, 10, 15, and 30 min. Apoptosis was detected by AO/EB staining. Changes of ALDH1 in HLEC were detected by immunohistochemical staining and Western blot. RESULTS: The intensity of immunohistochemical staining and the rate of positive cells decreased with increase of UV time (P<0.05). The rate of positive ALDH1 cells was negatively correlated with the rate of apoptosis (r= -0.92, P<0.05). Western blot showed the integrated absorbance values significantly decreased with the increase of UV time (P<0.05). CONCLUSION ALDH1 in HLEC decreases with an increase of UV exposure, which may be related to UV induced apoptosis of HLEC.
Aldehyde Dehydrogenase 1 ; Apoptosis ; radiation effects ; Cells, Cultured ; Epithelial Cells ; cytology ; metabolism ; radiation effects ; Humans ; Isoenzymes ; genetics ; metabolism ; Lens, Crystalline ; cytology ; Retinal Dehydrogenase ; genetics ; metabolism ; Ultraviolet Rays ; adverse effects

OBJECTIVE: To explore the apoptosis-inducing effect of ultraviolet(UV) radiation on human lens epithelial cells (HLEC), with particular focus on changes in Bcl-2 or Bax expression as possible mechanisms. METHODS: All experimental groups were exposed to the same UV light source. HLEC were divided into 6 groups according to duration of UV radiation : 0 min group (control group), 5 min group, 10 min group,15 min group, and 30 min group. Analysis on apoptosis of HLEC was performed by flow cytometry analysis (FCA, Annexin V + PI staining). Changes of Bax and Bcl-2 expression in HLEC were detected by hybridization in situ. RESULTS: Apoptosis in HLEC increased with UV exposure time. The expression level of Bax mRNA was increased with the increase of UV exposure time, whereas the expression level of Bcl-2 mRNA decreased with the increase of UV exposure time. The proportion of apoptotic cells was negatively correlated with ratio of Bcl-2/Bax (r=-0.874, P<0.05). CONCLUSION UA radiation can induce apoptosis of HLEC in vitro. Bcl-2 and Bax genes may play an important role in regulating this apoptotic process.
Apoptosis ; radiation effects ; Cells, Cultured ; Epithelial Cells ; cytology ; metabolism ; radiation effects ; Humans ; Lens, Crystalline ; cytology ; radiation effects ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Time Factors ; Ultraviolet Rays ; adverse effects ; bcl-2-Associated X Protein ; genetics ; metabolism

PURPOSE: The purpose of this study is to understand the mechanism of apoptosis occurring on a cultured human lens epithelial cell line after exposure to ultraviolet (UV) light. We intended to confirm the presence of cellular toxicity and apoptosis and to reveal the roles of p53, caspase 3 and NOXA in these processes. METHODS: Cells were irradiated with an ultraviolet lamp. Cellular toxicity was measured by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Hoechst staining and fluorescent anti-caspase 3 antibodies were used for apoptosis investigation. The quantities of p53, caspase 3, and NOXA were measured by Western blotting for to investigate the apoptosis pathway. RESULTS: Cellular toxicity on the human lens epithelium markedly increased with time after UV exposure. On Hoechst staining, we found that apoptosis also remarkably increased after exposure to ultraviolet light, compared with a control group. In the immunochemical study using anti-caspase 3 antibodies, active caspase 3 significantly increased after exposure to ultraviolet light. On Western blotting, p53 decreased, while caspase 3 and NOXA increased. CONCLUSIONS: Exposure of cultured human lens epithelial cell lines to ultraviolet light induces apoptosis, which promotes the expression of NOXA and caspase 3 increases without increasing p53. This may suggest that UV induced apoptosis is caused by a p53-independent pathway in human lens epithelial cells.
Apoptosis/*physiology ; Caspase 3/metabolism ; Cell Line ; Cell Survival/radiation effects ; Epithelial Cells/radiation effects ; Humans ; Lens, Crystalline/cytology/*physiology/*radiation effects ; Proto-Oncogene Proteins c-bcl-2/metabolism ; Tumor Suppressor Protein p53/metabolism ; *Ultraviolet Rays

OBJECTIVE: To investigate the mechanism and effect of diallyl disulfide (DADS) on the bystander effect of herpes simplex virus kinase/ganciclovir (HSV-tk/GCV) suicide gene therapy system which was mediated by recombinant adeno-associated virus (rAAV) in lens epithelial cells. METHODS: Immunohistochemistry method was used to determine the distribution of connexin 43 (Cx43) protein in cultured rabbit lens epithelial cells and rabbit lens epithelial cells transfected by HSV-tk suicide gene. Cx43 protein was measured and analyzed in cultured rabbit lens epithelial cells and rabbit lens epithelial cells transfected by HSV-tk suicide gene that was induced by various DADS. Cell survival was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method. RESULTS: DADS increased the Cx43 protein expression in cultured rabbit lens epithelial cells and rabbit lens epithelial cells transfected by HSV-tk suicide gene, and especially in 50 μmol/L DADS. After combining with DADS, the bystander effect was significantly improved (P<0.05). CONCLUSION DADS can elevate the expression of Cx43 protein and enhance the bystander effect of HSV-tk/GCV suicide gene therapy system.
Adenoviridae ; genetics ; metabolism ; Allyl Compounds ; pharmacology ; Animals ; Antiviral Agents ; pharmacology ; Bystander Effect ; drug effects ; Cells, Cultured ; Connexin 43 ; metabolism ; Disulfides ; pharmacology ; Epithelial Cells ; metabolism ; Ganciclovir ; pharmacology ; Genes, Transgenic, Suicide ; Lens, Crystalline ; cytology ; drug effects ; metabolism ; Plant Oils ; Rabbits ; Simplexvirus ; enzymology ; Thymidine Kinase ; genetics ; metabolism

OBJECTIVETo investigate the effect of cinobufagin on the proliferation of lens epithelial cells (LECs) and the bcl-2 and bax mRNA expressions of rabbits.

METHODSCultured LECs were treated for 72 h with cinobufagin of different concentrations, the end titer was 0.1 mg/L for low, 0.2 mg/L for moderate and 0.3 mg/L for high concentration, respectively. The inhibitory rate of cinobufagin on LECs' proliferation was detected using MTT method; the mRNA expressions of bcl-2 and bax genes in LECs were examined using reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSProliferation of LECs was inhibited significantly by cinobufagin in a concentration dependent manner, the inhibitory rate was 24.65%, 30.13% and 36.98% respectively for low, moderate and high concentration. With the drug concentration increasing, the mRNA expression levels of the pro-apoptotic bax were increased, whereas those of the anti-apoptotic bcl-2 decreased.

CONCLUSIONCinobufagin can remarkably inhibit the proliferation and induce the apoptosis of LECs in vitro, it might be taken as one of the drugs for the prevention and treatment of after-cataract.

Animals ; Apoptosis ; drug effects ; Bufanolides ; pharmacology ; Cell Proliferation ; drug effects ; Epithelial Cells ; drug effects ; metabolism ; Lens, Crystalline ; cytology ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; RNA, Messenger ; genetics ; Rabbits ; bcl-2-Associated X Protein ; metabolism

PURPOSE: To evaluate the protective effects of epigallocatechin gallate (EGCG) against UV irradiation of cultured human lens epithelial cells. METHODS: We irradiated cultured human lens epithelial cells with a 30-second pulse from a UV lamp with an irradiance of 0.6 mW/cm2. Five minutes and 1 hour after UV irradiation, we administered 0, 5, 10, 15, 25, 50, or 100 uM EGCG. The cell number was measured with a microscopic counting chamber and cell viability was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. RESULTS: Compared to untreated cells, the total number of cultured human lens epithelial cells was markedly higher after UV irradiation. In a dose-dependent manner, viability was also higher in EGCG-treated cells. CONCLUSIONS: EGCG increased the cell count and cell viability after UV irradiation of cultured human lens epithelial cells, indicating that EGCG can protect lens epithelium against UV damage.
Antioxidants/*pharmacology ; Catechin/*analogs & derivatives/pharmacology ; Cell Count ; Cell Survival/drug effects ; Cells, Cultured ; Coloring Agents/diagnostic use ; Dose-Response Relationship, Drug ; Epithelial Cells/radiation effects ; Humans ; Lens, Crystalline/cytology/*radiation effects ; Radiation Injuries/*prevention & control ; Radiation-Protective Agents/*pharmacology ; Tetrazolium Salts/diagnostic use ; Thiazoles/diagnostic use ; *Ultraviolet Rays