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Objective To study the relationship between the expression of sonic hedgehog (Shh) and vascular endothelial growth factor (VEGF) in hypoxic human retinal pigment epithelial (hRPE) cells.Methods Cultured hRPE-19 cells (3rd-6th generations) were used in this experiment.hRPE-19 cells were divided into three groups including the control group,the hypoxia experimental group (100 μmol/L CoCl2) and the inhibition group (pretreatment with 20 μmol/L cyclopamine 1 hour before hypoxia).After culturing for 4,8,12 and 24 hours,the mRNA level of Shh and VEGF genes in these cells were measured by fluorescence quantitative polymerase chain reaction,and the protein level of Shh and VEGF in the supematants were measure by enzymelinked immunosorbent assay.The relationship between the expression of Shh and VEGF was analyzed by Pearson correlation analysis.Results The control group expressed low levels of Shh and VEGF mRNA/protein.The expression of Shh and VEGF mRNA/protein in the hypoxia experimental group was significantly higher than that in the control group (F=178.364,183.732,77.456,91.572;P＜0.01).The expression of Shh and VEGF mRNA in the inhibition group was significantly lower than that in the hypoxia experimental group (F=68.745,121.834;P＜0.01).In the hypoxia experimental group,the expression of VEGF protein was positively correlated with the expression of Shh protein (r=0.942,P＜ 0.05);and the expression of VEGF and Shh mRNA was positively correlated (r=0.970,P＜0.01).However,there was no significant correlation in the expression of VEGF and Shh mRNA in the inhibition group (r=0.915,P＞ 0.05).Conclusion There is a positive correlation between the expression of Shh and VEGF in hypoxic hRPE cells.
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Objective To investigate the role of sonic hedgehog (Shh) signal transduction pathway in the expression of vascular endothelial growth factor (VEGF) under hypoxia in cultured human retinal pigment epithelial (hRPE) cells.Methods ARPE-19 were cultured and divided into normal ARPE-19 (Cont) and hypoxia group (100 μmol/L CoCl2 Cobalt Chloride +-ARPE-19);hypoxia group was further divided into CoCl2 group,cyclopamine group (CYA) and dimethyl sulfoxide (DMSO) group.20 μmol/L cyclopamine was added to the CYA group 1 hour before hypoxia,1‰ DMSO was added into DMSO group at the same time.The hRPE cells were cultured under hypoxia for 4,8,12,24 hours.The expression of Shh and VEGF were determined by Real-time fluorescent quantitate PCR (RT-PCR).The amount of VEGF in the hRPE-conditioned supernatant was measured using enzyme linked immunosorbent assay (ELISA) at 4,8,12,24 hours,respectively.Results RT-PCR tests showed that the level of Shh and VEGF of hRPE was time dependently increased (Shh:F=45.260,P=0.001;VEGF:F=264.938,P=0.001).The level of Shh and VEGF of hRPE in the group treated with cyclopamine was decreased (P＜0.01).ELISA tests showed that the amount of VEGF in hRPE supernatant was significantly increased in time-dependent manner (F =3 156.676,P =0.001),and it was down-regulated by cyclopamine under hypoxia (P＜0.01).Conclusion Shh signal transduction pathway could play a role in the VEGF expression induced by hypoxia in hRPE cells.
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From August 1996 to October 2004,totally 22 patients(22 eyes) of entire blepharocoloboma after malignant palpebrae superior tumorectomy,admitted to Department of Ophthalmology in the Affiliated Hospital of Binzhou Medical College,were repaired with sclera transplantation for the eyelid reconstruction.The alcohol-preserved scleral allograft was utilized for a substitute of tarsal plate.The internal surface of scleral graft was covered with remaining fornical conjunctiva and the external surface with musculus orbicularis oculi and skin.Bilateral sutures were fixed on the stump of tarsal plate or exterior and interior external palpebral ligament.Then eyelid margin was sutured and cut off after 6 months.All patient were followed up after operation for average 28.8 months.No reject reaction was found,catacleisis was fine and external appearance was satisfying.The eyelid reconstruction with scleral allograft is effective to keep eyelid normal function and external appearance.
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Objective To explore the effect of endothelin (ET)、nitric oxide (NO) in plasma on retinopathy in the pregnancy-induced hypertension (PIH). Methods The level of ET and NO in plasma of 75 cases of in-patient women with PIH and 20 cases of women with the full terms and normal pregnancy before and after delivery was determined by radioimmunoassay. The retinopathy of the patients with PIH before and after delivery was detected by appointed doctor. The levels of ET and NO in both groups were compared and the relationship between ET and NO in plasma and the retinopathy before and after the delivery was detected. Results The levels of ET [(145.00?54.41) ng/L] in serious PIH patients were much higher than that in the control [(81.50?43.80) ng/L], the minor [(85.30?33.33) ng/L] and middling PIH group [(90.20?39.25) ng/L]. The levels of ET in plasma before and after pregnancy were not changed in PIH patients [(118.70?33.44) ng/L], but were higher than that in the control group. The levels of plasma NO in serious [(87.56?35.58) ng/L] and middling [(78.11?28.96) ng/L] PIH group were both higher than that in the control group [(46.70?32.64) ng/L], and the levels in minor[(52.56?28.35) ng/L] and middling PIH group were lower than that in the serious PIH group. The level of NO in plasma of PIH patients after the delivery was much lower than that before the delivery, while higher than that in the control. The positive correlation between levels of ET and NO and retinopathy was found in PIH patients. Conclusions The levels of plasma ET and NO in PIH patients are related to the extent of the disease, and the level of ET in plasma is highly related to the retinopathy in PIH patients. ET and NO might be played an important role in pathogenesis of retinopathy and ET might be a good index in reflecting the rank of retinopathy in PIH.