The expression of cytokines in aqueous humor of high myopic patients with cataracts.

Purpose: To analyze the expression of 440 human cytokines in aqueous humor of high myopic patients with cataracts. Methods: Eighty-five patients with cataracts were recruited in this study. In the screening stage, the RayBio G-Series Human Cytokine Antibody Array 440 was used to assay the aqueous humor samples collected from nine high myopic patients with cataracts and eight non-myopic patients with cataracts right before the surgery. The array was further used for verification of the screened cytokines, with aqueous humor samples obtained from 34 eyes of high myopic patients with cataracts and 34 eyes of non-myopic patients with cataracts. Results: Compared with the non-myopic patients with cataracts, the expression levels of decorin, receptor activator of NF-kB (RANK), angiopoietin-1 (ANG-1), C-X-C motif ligand 16 (CXCL16), ß-inducible gene-h3 (bIG-H3), insulin-like growth factor-binding protein 2 (IGFBP-2), and interleukin-17B (IL-17B) were statistically significantly higher in high myopic patients with cataracts (all p<0.000114). The matrix metalloproteinase-2 (MMP-2) level also increased in the aqueous humor of high myopic patients with cataracts (p = 0.0034). The concentrations of ANG-1 and MMP-2 were also increased in the aqueous humor of the confirmatory stage (all p<0.05). Conclusions: In this study, numerous cytokines in aqueous humor were detected in high myopic patients with cataracts and non-myopic patients with cataracts, and it was confirmed that the MMP-2 level in the aqueous humor of patients with high myopia was statistically significantly increased. Further verification also revealed the elevation of ANG-1 in the aqueous humor of high myopic patients with cataracts, which suggests that ANG-1 may be related to the pathogenesis of high myopia.

Effect of Aß protein on inhibiting proliferation and promoting apoptosis of retinal pigment epithelial cells.

AIM: To identify the effect and regulatory mechanism of amyloid ß (Aß) protein on retinal pigment epithelial (RPE) cells in cell proliferation and apoptosis, and clarify Aß role in the pathogenesis of age-related macular degeneration (AMD). METHODS: The model of Aß25-35 protein cytotoxicity in RPE cell was successfully established to investigate the effect of Aß protein on RPE cells in vitro. Based on Aß protein, the specific inhibitors (HY-50682 or BAY11-7082) or activating agent (lipopolysaccharide) was used to analyze the regulatory mechanism of Aß protein to RPE cells on cell proliferation and apoptosis by flow cytometry, real-time polymerase chain reaction, Western blotting, enzyme-linked immunosorbent assay and dual-luciferase reporter gene assay. RESULTS: The number of RPE cells, treated with Aß25-35 from 0.3 to 60 µmol/L, significantly reduce (P<0.01), and had the dose-dependent effect. Aß protein 60 µmol/L inhibits the G1/S phase transition (P<0.01) and down-regulated cyclin E mRNA level (P<0.01). Similarly, Aß25-35 induced a significant increase of cell apoptosis, accompanied by the significantly higher level of activated caspase 3 protein. Furthermore, nuclear factor-kappaB (NF-κB) activity and phosphorylated Iκ-Ba level would significantly lower in treated RPE cells. Using specific inhibitors or activating agent based on the Aß, the cell numbers, NF-κB activity, phosphorylated Iκ-Ba level, receptor for advanced glycation endproducts (RAGE) gene expression levels, cyclin E mRNA level and activated caspase 3 level had accordingly changed by different methods, confirming that RAGE/NF-κB signaling pathway involved in the regulation of Aß protein on RPE cell apoptosis and proliferation. CONCLUSION: Aß protein inhibits cell proliferation and activates apoptosis via inactivation of the RAGE/NF-κB signaling pathway in RPE cell.

Nrf2 protects human lens epithelial cells against H2O2-induced oxidative and ER stress: The ATF4 may be involved.

Our previous study has shown heme oxygenase-1 (HO-1) protects human lens epithelial cells (LECs) against H2O2-induced oxidative stress and apoptosis. Nrf2, the major regulator of HO-1, is triggered during the mutual induction of oxidative stress and ER stress. In response to ER stress, unfolded protein response (UPR) serves as a program of transcriptional and translational regulation mechanism with PERK involved. Both Nrf2 and ATF4 are activated as the downstream effect of PERK signaling coordinating the convergence of dual stresses. However, the ways in which Nrf2 interacting with ATF4 regulates deteriorated redox state have not yet been fully explored. Here, the transfected LECs with Nrf2 overexpression illustrated enhanced resistance in morphology and viability upon H2O2 treatment condition. Intracellular ROS accumulation arouses ER stress, initiating PERK dependent UPR and inducing the downstream signal Nrf2 and ATF4 auto-phosphorylation. Further, converging at target promoters, ATF4 facilitates Nrf2 with the expression of ARE-dependent phase II antioxidant and detoxification enzymes. According to either Nrf2 or ATF4 gene modification, our data suggests a novel interaction between Nrf2 and ATF4 under oxidative and ER stress, thus drives specific enzymatic and non-enzymatic reactions of antioxidant mechanisms maintaining redox homeostasis. Therapies that restoring Nrf2 or ATF4 expression might help to postpone LECs aging and age-related cataract formation.

miRNA-1273g-3p Involvement in Development of Diabetic Retinopathy by Modulating the Autophagy-Lysosome Pathway.

BACKGROUND Diabetic retinopathy (DR) is one of the most common and serious complications of diabetes mellitus (DM). The autophagy-lysosome pathway (ALP) is one of the main intracellular self-digestive degradation systems. Lysosomal impairment and autophagic dysfunction are early events in the pathogenesis of DR, suggesting autophagy might be a novel therapeutic strategy for DR treatment. MATERIAL AND METHODS In our study, we screened a differentially expressed miRNA, miR-1273g-3p, in streptozotocin (STZ)-injected DR rat retinal pigment epithelial (RPE) cells. miR-1273g-3p inhibitor and mimic were employed to treat RPE cells to assess the role of miR-1273g-3p. QRT-PCR and Western blot analysis were performed to examine the function of miR-1273g-3p on ALP-related and DR-related proteins. RESULTS miR-1273g-3p was highly expressed in STZ-induced DM RPE cells. miR-1273g-3p mimic promoted the expression of DR-related MMP-2, MMP-9, and TNF-α proteins, and ALP-related LC3, cathepsin B, and cathepsin L factors, but miR-1273g-3p inhibitor suppressed the levels of these factors. CONCLUSIONS miR-1273g-3p is involved in the progression of DR by modulating the autophagy-lysosome pathway. These findings provided new evidence of the close relationship between DR and ALP, and reveal a new target for DR therapy.

Efficacy comparison between manual small incision cataract surgery and phacoemulsification in cataract patients: a meta-analysis.

OBJECTIVE: Systematic review of manual small incision cataract surgery (MSICS) and phacoemulsification (PHACO) on the postoperative visual quality and surgical complications. METHODS: Relevant literatures on clinical efficacy of PHACO and MSICS were included by retrieving in Medline, PubMed, Chinese Biomedical Literature Database (CBM) and Chinese Academic Journal (CNKI) databases. Meta-analysis was conducted by RevMan5.0 software with OR and its 95% CI for the effect size. RESULTS: A total of ten documents were included in the study. Uncorrected visual acuity 1 week after surgery (OR = 0.84, 95% CI: 0.67 ~ 1.06, P=0.15), post-operative capsular rupture (OR = 1.07, 95% CI: 0.73~1.58, P=0.72), and corneal edema (OR = 0.90, 95% CI: 0.70~1.16, P=0.42) between MSICS and PHACO showed no statistical difference (P > 0.05). CONCLUSION: Clinical efficacy and complications of MSICS was similar tothat of PHACO.

[Effects of matrix metalloproteinases inhibitor GM6001 on choroidal neovascularization].

OBJECTIVE: To explore the efficiency of GM6001 for the inhibition of choroidal neovascularization. METHODS: Experimental study. Twenty-four Brown Norvy (BN) rats after photocoagulation were randomly divided into 3 groups as GM6001 group, DMSO group and CONT group. GM6001 (0.2 ml of 0.1% suspension) was injected retrobulbar for GM6001 group and 0.2 ml DMSO was injected for DMSO group on days 1, 3, 6, 9, and 12 after photocoagulation. No injection was performed in the CONT group. Fundus fluorescence angiography, histopathology, immunohistochemistry and quantitative analysis of choroidal neovascularization (CNV) were performed 3 weeks after photocoagulation. One-way ANOVA was used in conjunction with SNK-t test to assess statistical significance within groups. RESULTS: The fluorescein leakage appeared in all three groups; but fluorescein leakage of GM6001 group (74.56 ± 2.33) was less than that of DMSO group (119.57 ± 1.15)and CONT group (122.36 ± 2.38) (F = 403.23, P = 0.001; LSD-t test, all P value < 0.01), whereas fluorescein leakage of DMSO group was similar to that in the CONT group. The retinal and choroidal capillaries in the CONT group were damaged and disordered; a great deal of CNV and migration and proliferation of retinal pigment epithelium cells, fibrocytes and collagen fibers were discovered. Pathological changes in DMSO group were similar to those in the CONT group. There were a small quantity of retinal pigment epithelium cells, fibrocytes and CNV in GM6001 group. Although the immunohistochemical staining for CD105 displayed positive results in all three groups, positive staining of GM6001 group (19.85 ± 1.59) was significantly less than that of the CONT group (38.02 ± 2.57) and DMSO group (39.02 ± 3.12) (F = 55.57, P = 0.001; LSD-t test, all P value < 0.01). Positive staining of CD105 in the CONT group was similar to that of DMSO group (P > 0.05). The size of CNV in GM6001 group (15.35 ± 0.77) was significantly less than that of CONT group (28.38 ± 1.60) and DMSO group (28.74 ± 1.19) (F = 114.85, P = 0.001; LSD-t test, all P value < 0.01). There was no statistical difference for the size of CNV between CONT group and DMSO group (P > 0.05). CONCLUSION: GM6001 effectively inhibits CVS induced by krypton laser photocoagulation.

IL-33 is induced by amyloid-ß stimulation and regulates inflammatory cytokine production in retinal pigment epithelium cells.

Age-related macular degeneration (AMD) is the predominant cause of irreversible blindness in the elderly population. Despite intensive basic and clinical research, its pathogenesis remains unclear. However, evidence suggests that immunological and inflammatory factors contribute to the pathogenesis of AMD. A newly identified cytokine, IL-33, appears to be an important pro-inflammatory cytokine promoting tissue inflammation. In this study, IL-33 was increased through amyloid-beta(1-40) (Aß(1-40)) stimulation and regulated inflammatory cytokines including IL-6, IL-8, IL-1ß, and TNF-α secretion using different signaling pathways in retinal pigment epithelium (RPE) cells. Furthermore, ST2L, the important component of the IL-33 receptor, was significantly increased following recombinant human IL-33 stimulation in RPE cells. These findings suggest that IL-33-mediated inflammatory responses in RPE cells are involved in the pathogenesis of AMD. Greater understanding of the inflammatory effect of IL-33 and its role in RPE cells should aid the development of future clinical therapeutics and enable novel pharmacological approaches towards the prevention of AMD.

[Some attentive issues in the selection of intraocular lenses].

Different kinds of new intraocular lenses (IOL) provide more choices for different clinical demands. However, there are many misunderstandings about how to select the proper IOL. These misunderstandings arise from the believes that include "the expensive IOL is good IOL", "new product is absolutely perfect", etc. Furthermore, we should pay attention to avoid the "confusion concept", "comprehend error of monovision and matching different types of IOL", "ignore the results from evidence-based medicine", etc. When we select the IOL, we should concern about how to meet the patients' demands and how to provide the best benefits to the patients.

[Protective effects of puerarin on lens epithelial cells in rat diabetic cataract].

OBJECTIVE: The therapeutic effects of general and local applications of puerarin in the treatment of streptozotocin (STZ)-induced rat diabetic model were compared. METHODS: Experimental research. We equally divided normal Sprague-Dawley (SD) rats into a STZ group, a peritoneal injection group, a peribulbar injection group and a control group. STZ, peritoneal injection and peribulbar injection groups were first treated with STZ. Subsequently, the STZ group was injected with normal saline intraperitoneally, while in the later two groups puerarin was injected through peritoneal and peribular routes, respectively. Control group only received peritoneal injection of saline. The morphology of lens epithelial cells (LEC) and their subcellular structure were examined by bright-field microscopy, transmission electron microscopy (TEM) and scanning electron microscopy (SEM) 20, 40 and 60 days after the injection. Nitrogen oxide (NO) and nitric oxide synthase (NOS) were measured by biochemistry methods. Finally, inducible nitric oxide synthase (iNOS) protein and mRNA levels were monitored by Western blot and RT-PCR, respectively. Data was processed with two factorial experiment analysis of variance. RESULTS: Twenty to sixty days after the injection, marked or complete lens opacities appeared in the STZ group, whereas only slight opacities appeared in the lens in peritoneal and peribulbar puerarin groups and the lens in the control group remained clear. At the 20th, 40th and 60th day after the injection, optical microscope detected pathological changes of LEC in the STZ group. The cell volume was decreased with a dense nucleus and many bubbles appeared around the equator area. Under TEM, enlargement of cell gap, vacuoles in the cytoplasm, swelling of mitochondria and unclear structure of rough endoplasmic reticulum appeared in the LEC of the STZ group. Part of the nucleus was in karyopyknosis and peripheral nucleus gap was enlargement. Under SEM, normal fiber conjunction structure of the lens disappeared, fibers were swelling, part of fiber membranes were discontinuous, detached, and accumulated in certain areas. Mild lens opacities detected by bright-field microscope were developed in peritoneal and peribulbar puerarin injection groups. Nucleus and fibers in the lens cells of both groups appeared to be normal, with minor swelling of mitochondria, minor enlargement of endoplasmic reticulum and slight increase of intracellular space. NO, NOS and iNOS protein and mRNA of the lens were increased and up-regulated in STZ group. In the other two groups only minor changes were present and the changes were significantly less than that of the STZ group but greater than that in the control group. CONCLUSION: Peritoneal and peribulbar injection of puerarin have similar therapeutic effects in the treatment of rat diabetic cataract.

[Alternative splicing of vascular endothelial growth factor A and ocular neovascularization].

Ocular neovascularization is the primary cause of blindness in a wide range of ocular diseases. The vascular endothelial growth factor A (VEGF-A) is the key factor involved in ocular angiogenesis, which can cause eye diseases through the development of pathological angiogenesis and increase of vascular permeability. There are two families of VEGF-A isoforms formed by alternative splicing, the angiogenic VEGF-A family (VEGF(xxx)), known to contribute to ocular neovascularization, and the anti-angiogenic VEGF-A family (VEGF(xxx)b), which is found in normal ocular tissues but downregulated in human diabetic retinopathy. The first member of the VEGF(xxx)b family to be isolated was VEGF(165)b. It can significantly reduce preretinal neovascularization without inhibition of physiological intraretinal angiogenesis. As the studies on the VEGF(xxx)b family proceed more deeply, controlling the balance of VEGF(xxx) to VEGF(xxx)b isoforms may be therapeutically valuable in the treatment of angiogenic eye diseases such as diabetic retinopathy and age-related macular degeneration.

Toxicity of endogenous peroxynitrite and effects of puerarin on transplanted retinal pigment epithelial sheets in the subretinal space in mice.

AIM: To evaluate the toxicity of endogeneous peroxynitrite on transplanted retinal pigment epithelial (RPE) sheets and the effect of puerarin on their survival in the C57BL/6 mice after RPE sheets have been transplanted into SD rats' subretinal space . METHODS: C57BL/6 mice eyes were used to culture RPE cells. Ninety-six SD rats were involved in the experiment. They were divided into control (block control), streptozotocin (STZ, negative control), untransplanted RPE (positive control) and transplanted RPE groups respectively. Diabetes was induced in SD rats by intra-peritoneal STZ injection in the latter three groups. Saline was injected into the subretinal space of 24 SD rats in the untransplanted RPE group and primary RPE sheets were injected into the subretinal space of 24 SD rats in the transplanted RPE group. Puerarin (45mg/kg) was administrated into both untransplanted RPE and transplanted RPE groups of diabetic rats through intra-peritoneal injection route after RPE sheets transplantation. At 20, 40, 60 days after surgery, Western blotting analysis, DNA ladder and RT-PCR were used for determining the differences in expression of nitrotyrosine (NT, the foot print of peroxynitrite ), apoptosis and iNOS mRNA in the control, STZ, untransplanted RPE and transplanted RPE groups respectively. HE staining was used for determining the RPE survival in the subretinal space of the transplanted RPE group. RESULTS: Apoptosis and expression of NT and iNOS mRNA were observed in STZ, untransplanted RPE and transplanted RPE groups, but were delayed in untransplanted RPE and transplanted RPE groups in a time-dependent manner compared with control and STZ groups (P<0.01). There were no differences between the two groups (P>0.01). NT, DNA ladder, iNOS mRNA were down-regulated, which were associated with the decrease of expression of peroxynitrite. Numerous pigmented cells emerged and increased in number in the subretinal space during the 60-day observation period after transplantation. On day 20, heavily pigmented cells were visible at the transplant site; On day 40, monolayer and multilayered transplant was visible in the subretinal space; On day 60, heavily pigmented monolayer and multilayered transplants with round apical profile were present along Bruch's membrane. CONCLUSION: Puerarin increased the 60-day survival of C57BL/6 mice RPE xenografts in the SD rats' subretinal space, which may be related to its direct inhibition of apoptosis of RPE cells and antagnism of damage of peroxynitrite to RPE cells.

Peroxynitrite-induced expression of inducible nitric oxide synthase and activated apoptosis via nuclear factor-kappa B pathway in retinal pigment epithelial cells and antagonism of cholecystokinin octapeptide-8 in vitro.

AIM: To explore that if peroxynitrite induced the expression of inducible nitric oxide synthase (iNOS)via nuclear factor-kappa B (NF-κB) pathway in retinal pigment epithelial (RPE) cells and the antagonism of cholecystokinin octapeptide-8 (Melatonin, CCK-8) in vitro. METHODS: RPE cells were obtained from eyes of C57BL/6 mouse and divided into control, peroxynitrite and CCK-8 groups. Control group was treated with saline, peroxynitrite group was treated with peroxynitrite, and CCK-8 group was treated with CCK-8 after added with peroxynitrite. All changes were observered at 6, 12 and 24 hours after treatment. Gene array analysis, Reverse Transcription Polymerase Chain Reaction (RT-PCR) were used to determine the expression of inducible nitric oxide synthase (iNOS) mRNA in RPE cells. Western blotting was used to test the apoptosis of RPE cells. Immunofluorescent staining was used to determine the NF-κB pathway signal transduction. RESULTS: Compared to the control group, the expression of iNOS mRNA was up-regulated in peroxynitrite group and down-regulated in CCK-8 group with gene array analysis. Apoptosis was increased in peroxynitrite group and decreased in CCK-8 group with western blotting. The NF-κB pathway signal transduction was more and more stronger in the peroxynitrite group. But in CCK-8 group, little stronger could be observed at 12 hours, then weak at 24 hours with immunofluorescent staining (P<0.001). CONCLUSION: This study suggested that apoptosis of RPE cells was partly induced by peroxynitrite, which may be the new way of oxidative damage to the RPE cells. The NF-κB signal transduction may affect and reinforce apoptosis mediated by peroxynitrite. CCK-8 decreased apoptosis of RPE cells induced by peroxynitrite and is a potential agent for therapy of retinopathy. The mechanism of CCK-8 dealing with RPE cells may be related to its direct inhibition of the formation of iNOS to produce peroxynitrite and antagnism of damage of peroxynitrite to the RPE cells.

Effect of puerarin on retinal pigment epithelial cells apoptosis induced partly by peroxynitrite via Fas/FasL pathway.

AIM: To evaluate the peroxynitrite (ONOO(-)) of puerarin on retinal pigment epithelial (RPE) cells apoptosis induced partly by peroxynitrite via Fas/FasL. METHODS: RPE cells from C57BL/6 mice eyes were cultured. Diabetes was induced in Sprague-Dawley (SD) rats by streptozotocin (STZ) intraperitoneal injection. Puerarin was administrated to cultured RPE cells and diabetic rats. Western blotting analysis, DNA ladder, RT-PCR, immunohistochemistry were used for determining the expression of nitrotyrosine (NT, the foot print of ONOO(-)), complement 3 (C3); apoptosis and inducible nitric oxide synthase (iNOS) mRNA as well as Fas/FasL signal transduction in RPE cells. RESULTS: Both RPE cells in ONOO(-) and puerarin group developed apoptosis and expressed NT, C3, iNOS mRNA and Fas/FasL. But latter delayed the all changes in a time-dependent manner compared with control and STZ group (P<0.001). iNOS, C3 and Fas/FasL were up-regulated and associated with an increase of expression of ONOO(-)in vivo and in vitro. CONCLUSION: Puerarin decreases RPE cells apoptosis partly induced by ONOO(-) for diabetic retinopathy.

[Several notable problems in current development of cataract surgery].

The technologies about cataract surgery and intraocular lens has been developing expeditiously. However, some problems have been occurring and should be corrected. Cataract blindness can not restore the sight after surgery has been performed still existed. Some new technologies blindly introduced and applied. The comprehension about individualized selection of intraocular lens still has some bias. To solve the above problems and to hold rhythm of popularizing new techniques and to stick the key points of the technologies may healthily promote and advance the cataract surgery in China.

[MicroRNA and progresses in the region of ophthalmology].

MicroRNAs (miRNAs) consist of a growing class of short non-coding RNAs (ncRNAs) that negatively regulate the expression of genes involved in development, differentiation, proliferation, apoptosis and other important cellular processes. They regulate gene expression at the post-transcriptional level by either degradation or translational repression of a target mRNA. Nowadays more than 400 miRNAs have been identified in the human genome. A growing number of reports have established a link between a specific group of microRNAs and tissues in eyes, including cornea, len, retina. Here, we describe our current understanding of the structure, biogenesis and function of MicroRNAs, as well as their potential as novel therapeutic approaches in eye disease.

[Screening and identification of abnormal expression proteins from hypoxia human retinal pigment epithelium cell in vitro].

OBJECTIVE: To screen and identify the abnormal proteins expressed by hypoxia human retinal pigment epithelium (RPE) in vitro. METHODS: It was a experimental study. Protein of normal/hypoxia human RPE cells in exponential phase of growth was extracted, and frozen by liquid nitrogen for proteome study. All samples were performed isoelectric focusing electrophoresis (IEF), separated by isoelectric point (pI); progressed sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE) and separated by protein molecular weight (Mr). 2D-gels revealed by coomassie brilliant blue, analyzed by Image Master 2D Elite. Differential proteins spots were screened in normal/hypoxia RPE cell. For identification of differential proteins, peptide masses were analyzed by matrix-assisted laser desorption ionization time-of flight mass spectrometry (MALDI-TOF/MS). RESULTS: Five hundred and seventy-eight protein spots were obtained in normal group (matching rate within group was 92.90%), and hypoxia group 559 (91.41%). The average matching rate between two groups was 85.47%. There were 32 differential protein spots, which volume value changed > or = 2.0 times. 7 spots were selected randomly for MS, and 5 proteins were identified successfully: HSP70 and HSP60 up-regulated; while beta-actin, beta-tubulin and peroxiredoxin 3 down-regulated. CONCLUSIONS: Findings of the in vitro study provide the evidence for expression changes of many proteins in RPE cell with hypoxia state; some of those regulated proteins can result in increasing stress capability obviously in cells, while the major cytoskeletal proteins down-regulated, and the ability of sustained-shapes, keeping order internal structure in cells decreasing. Proteome analysis provides a useful platform in systematic screening of various protein expressions in CNV related diseases.

[To guard against mistakes in selection of intraocular lenses].

Development of different kinds of new intraocular lenses (IOL) provide more selections to meet various clinical requirements, which may plays an active role in making cataract surgery more perfectible. But there are many misunderstandings about how to select the proper IOL. For example, "the expensive one is the best one", "new product is absolutely perfect", and so on. Some surgeons prefer more practice and ignore the summary of the experiences. Someone unilaterally exaggerates some special function of IOL When selecting the IOL, it is a common challenge for us to fully consider the patients needs, and properly manage the relationship between the basic need and special need, and to avoid fanaticism, and provide the best benefits to the patients.

[The effect of puerarin on prevention of oxidative damage of cultured lens capsule epithelial cells].

OBJECTIVE: To investigate the peroxynitrite damage to the lens epithelial cells (LEC) and the prevention of this damage by puerarin in vitro. METHODS: This paper was experimental study. Rabbit LEC were isolated and cultured and the third or forth passage LEC were used in this experiment The experiment groups included: (1) CONTROL GROUP: Heat-pathogen free saline (NS) 200 microl was added to the medium; (2) ONOO- group: ONOO- 200 microl was added to obtain the terminal concentration at 0. 5 mmol/L; (3) Puerarin group: 5 microg/ml ONOO- and 10 microg/ml puerarin were added simultaneously. Then, the cells were cultured and collected after 6,12 or 24 hours. The nitrotyrosine (NT), a symbol of the ONOO-, was tested with immunofluorescence technique. The expression of NT protein was examined with Western blot method. The cell morphology was observed with light microscope. Cell apoptosis was examined via DNA ladder, flow cytometry and Fas/FasL immunohistochemical staining. These datas were analyzed by one-way-ANOVA and q test. RESULTS: During the 6 to 24 hours of experiment period, green color could be observed in the cell nucleus and cytoplasm of control group. Staining ranged from yellow to brown-yellow, then to brown color were observed in STZ group. Staining ranged from faint green to yellow green or faint green color were observed in puerarin group. Slight expression of nitrotyrosine (NT) could be seen in the control group. A moderate to strong expression of NT was observed at different stages in the STZ group (A = 77.22 +/- 2.44, 145.00 +/- 3.94, 235. 8 +/- 5.97). At 6 hours, a slight expression of NT could be seen in the control group (A = 72.78 +/- 2.64), this increased at 12 hours (A =89. 94 +/- 3.01) and decreased at 24 hours (A = 74. 44 +/- 3.00). With computer photo-analysis, there were significant differences between the control, STZ and puerarin groups at different period during the experiment (q = 78.12, 82.76, 69.98, P <0. 01). In the control group, cell morphology and gene DNA ladder were normal, minor apoptosis could be observed but no expression of Fas/FasL in the membrane and cytoplasm of the cells. Distinctive cell morphology changes and the typical "ladder bands" as well as the expression of Fas/FasL could be observed in STZ group. All of these aspects were comparatively normal in puerarin group. CONCLUSIONS: The LEC apoptosis induced by ONOO- in vitro could be alleviated by puerarin. Fas/FasL cell signal transduction pathway may affect and strengthen the apoptosis process mediated by ONOO-.

[The preliminary study of Phosphomannopentaose sulfate (PI-88) on the experimental choroidal neovascularization].

OBJECTIVE: To probe the intervention and mechanism of the inhibitor of heparanase (PI-88) on the experimental CNV model . METHODS: Experimental CNV was induced by laser photocoagulation in 29 mail (Brown Norway) BN rats (647 nm wave length Krypton laser, 360 mW power, 50 microm spot size, 0.05 second duration), and randomly divided into vacant control group, physiologic saline control group, preventive group and treated group, another 3 rats were acted as normal group, 15 days continuous intraperitoneal injection of PI-88 (25 mg kg(-1) d(-1) was administrated in preventive group (PI-88 was administered the same day as photocoagulation) and treated group (PI-88 was administered 1 week after photocoagulation when CNV had been formed), for the physiologic saline control group, PI-88 was replaced by physiologic saline. To comprehensively evaluate the effect of PI-88 on the CNV by the quantitation of CNV area marked by FITC-dextran in choroid-sclera flat mounts, fluorescence fundus angiography and histopathology; the changes of HPA expression were surveyed by western blot and immunohistochemistry. RESULTS: CNV area of preventive group and treated group decreased 52.1% and 53.8% respectively at the time point of 3 weeks after photocoagulation; the relative thickness of CNV membrane in eyes of treated group had been decreased 46% as that of control group by histopathology; CNV occurrenced 1 week after photocoagulation both in the control group and preventive group, while the fluorescein leakage of the preventive group had been inhibited significantly; 3 weeks after photocoagulation, there had been 7 days discontinuation for the prevent group, the fluorescein leakage did not enhanced, while there had been 15 days continuous administration for the treated group, the fluorescein leakage had been decreased significantly compared to the time point of 2 week; western blot showed that the relative expressed levels of HPA protein in both preventive and treated group decreased; HPA displayed distribution at the leading edge of CNV membrane migrating toward inner retina and the vascular tissue by immunohistochemistry, the expression of HPA was inhibited significantly in treated group. CONCLUSION: PI-88 may not only prevent CNV in certain degree, but also make it partially subsidize for the existed CNV, and the effect is associated with the inhibition of HPA production.

[To recognize monovision correctly and matching different types of intraocular lenses in two eyes].

The production of different kinds of novel intraocular lens (IOLs) with various functions plays an active role in improving cataract surgery by providing more choices to meet different clinical demands. However, each type of IOL has its own advantages and disadvantages. Therefore, it is important to implant two different kinds of IOLs into two eyes of a patient, which can optimize the advantages of these two IOLs and can obtain a better mixing vision. But there are many misunderstandings about how to comprehend the principle of monovision and matching two different types of IOLs. When we consider monovision and matching two different types of IOLs, there are many challenges for us, e.g. to explain to the patients about the difference between expected and practical results, and the psychological adaptation requires a long period; and we should to avoid selection the IOLs blindly and to provide the best results and benefits for the patients.