Quercetin protects retina external barrier from oxidative stress injury by promoting autophagy.

Purpose: This study aimed to investigate the protective effects of quercetin on the tight junction proteins of human retinal pigment epithelial cells (ARPE-19 cells) suffering from oxidative stress injury and explore the possible mechanism.Methods: H2O2 (300 µM) was used to establish an oxidative stress model of ARPE-19 cells. ARPE-19 cells were pretreated with different concentrations (0-80 µM) of quercetin before H2O2 exposure. The expression and distribution of tight junction proteins and autophagy-related proteins were detected by Western blot and immunostaining. ARPE-19 cells were pretreated with 5 mM 3-methyladenine (3-MA).Results: The cell viability weakened in the H2O2 group compared with the control group. However, it was preserved after pretreatment with quercetin. It was observed that the expression levels of occludin, claudin-1 were decreased in the H2O2 group. Quercetin treatment significantly enhanced the expression levels of them as compared to the H2O2 group. H2O2 alone strongly decreased the Zonula occludens protein 1 (ZO-1) expression in the cytomembrane. Quercetin supplementation enhanced the accumulation of ZO-1 in ARPE-19 cells. The expression levels of Beclin-1 and Microtubule associated protein light chain 3 II (LC-3II) increased, and that of P62 decreased in the quercetin protection group. The appearance of LC-3II, which examined by immunofluorescence experiments, enhanced in the quercetin protection group as compared with the control group. The expression levels of beclin-1 and LC-3II increased, and that of P62 increased in the autophagy-inhibited group compared with the quercetin protection group. The levels of occludin and claudin-1 also decreased.Conclusion: Quercetin prevents the loss of tight junction proteins by upregulating autophagy after oxidative stress in ARPE-19 cells.

[Chinese herbal medicine Lingqi Huangban Granule protects retinal pigment epithelial cells against oxidative stress-induced injury in vitro].

OBJECTIVE: To observe the protective effects of drug-contained serum of Lingqi Huangban Granule (LQHBG), a compound traditional Chinese herbal medicine, on oxidative stress-induced injury in rabbit retinal pigment epithelial (RPE) cells in vitro. METHODS: The oxidative stress of rabbit RPE cells in vitro was induced with hydrogen peroxide (500µmol/L) and different concentrations of LQHBG were administered to rats to prepare medicated serum. RPE cells were randomized into normal control group (no hydrogen peroxide), model group (hydrogen peroxide), model plus serum group (hydrogen peroxide and 10% control serum), model plus low-dose LQHBG group (hydrogen peroxide and low-dose LQHBG-medicated serum) and model plus high-dose LQHBG group (hydrogen peroxide and high-dose LQHBG-medicated serum). Teminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) assay and flow cytometry (FCM) were used to measure apoptosis of cultured rabbit RPE cells. Protein expressions of caspase-3 and Bcl-X(L) were observed by Western blot method. RESULTS: FCM results showed that the apoptotic rates of the normal control group, model group, control serum group and serum containing low- and high-dose LQHBG groups were (4.85±0.26)%, (20.02±1.37)%, (21.84±0.94)%, (13.56±0.55)%, and (8.58±0.39)%, respectively; compared with the model group, the apoptotic rates of RPE cells in the low- and high-dose LQHBG groups were obviously reduced in a dose-related manner (P<0.05). TUNEL results showed that nuclei of apoptotic cells were stained brown; the number of apoptotic cells in the low- and high-dose LQHBG groups was obviously less than that in the model group. The protein expression of caspase-3 was up-regulated in the model and control serum groups, which was higher than that in the high-dose LQHBG group (P<0.05). The protein expression of Bcl-X(L) was down-regulated in the model and control serum groups, which was lower than that in the low- and high-dose LQHBG groups (P<0.01). CONCLUSION: Drug-contained serum of LQHBG obviously reduces apoptosis and partly protects rabbit RPE cells from oxidative stress-induced injury. The protective function is due to an improvement in antioxidant abilities, down-regulation of the expression of caspase-3 and up-regulation of the expression of Bcl-X(L).

[Protective effect of Huangban Granule against light-induced retinal damage in rats].

OBJECTIVE: To observe the protective effect of Huangban Granule, a compound of traditional Chinese herbal medicine, on rats with retinal damage induced by light. METHODS: A total of 24 male Sprague-Dawley (SD) rats were randomly divided into normal control group, untreated group and Huangban Granule group. Retinal light damage was induced by exposure to constant white fluorescent light for 5 hours at an illumination of 2,800 Lux. The Huangban Granule was given 10 days before light exposure until the animals were sacrificed in Huangban Granule group, and an equal volume of distilled water for the rats in untreated group. Electroretinogram (ERG) was recorded in all animals 2 weeks after light exposure and the animals were sacrificed for histopathological examination of retina. The outer nuclear layers (ONLs) on the superior and inferior retina were counted. RESULTS: Fourteen days after light exposure, the ONLs on the superior retina were 3 to 6 in the untreated group and 7 to 9 in treatment group. There were 9 to 11 layers in normal group. The mean number of ONLs in the untreated group (4.68+/-1.64) was less than that in the treatment group (8.23+/-1.35) (P<0.01). B-wave amplitudes were (319.38+/-71.96) muV and (135.16+/-42.30) muV in Huangban Granule group and the untreated group respectively (P<0.01). A-wave amplitudes were (184.63+/-47.23) muV and (83.35+/-27.75) muV (P<0.01), and oscillatory potential amplitudes were (239.38+/-20.19) muV and (125.44+/-26.23) muV (P<0.01) respectively in the two group. There was no significant difference in implicit times of a-wave and b-wave among the three groups. CONCLUSION: Huangban Granule obviously protects both function and morphology of the retina from light-induced retinal damage in rats.