Expression of Robo4 in the fibrovascular membranes from patients with proliferative diabetic retinopathy and its role in RF/6A and RPE cells

Purpose: Robo4, a member of the roundabout (Robo) family, acts as a neuronal guidance receptor and plays some role in vasculogenesis and angiogenesis. This study investigated the effect of Robo4 on the formation of fibrovascular membranes (FVMs) from patients with proliferative diabetic retinopathy and its roles in choroid-retina endothelial (RF/6A) and human retinal pigment epithelial (RPE) cells. Methods: RT-PCR and immunohistochemistry were used to determine the levels of mRNA and the presence and distribution of Robo4 in FVMs. Small interfering RNA (siRNA) technology was used to knock down Robo4 expression and to study its effects on RF/6A and RPE cells in vitro. Cell proliferation, migration, spreading, cycling, and apoptosis were assessed with MTT assay, Boyden chamber assay, immunocytochemistry, and flow cytometry. Tube formation by RF/6A on Matrigel was also analyzed. Results: The level of Robo4 mRNA was high in FVMs. Robo4 was expressed in the vessels and fibrous-like tissue co-immunostained for CD31 and GFAP, respectively. Robo4 siRNA knockdown inhibited cell proliferation and migration. Tube formation by RF/6A cells was also disturbed. Under hypoxic conditions, more apoptotic cells were evident among the knockdown cells than among the control cells (p < 0.01). Conclusions: Robo4 may play a role in the formation of FVMs. Silencing the expression of Robo4 in RF/6A and RPE cells inhibited their proliferation and reduced their tolerance of hypoxic conditions, suggesting physiologic functions of Robo4 in the cells of the retina.http://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000267136400001&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=8e1609b174ce4e31116a60747a720701Biochemistry & Molecular BiologyOphthalmologySCI(E)PubMed15ARTICLE112-131057-10691

Protective effect of paeoniflorin against oxidative stress in human retinal pigment epithelium in vitro

Purpose: This study was conducted to determine whether paeoniflorin (PF) could prevent H(2)O(2)-induced oxidative stress in ARPE-19 cells and to elucidate the molecular pathways involved in this protection. Methods: Cultured ARPE-19 cells were subjected to oxidative stress with H(2)O(2) in the presence and absence of PF. The preventive effective of PF on reactive oxygen species (ROS) production and retinal pigment epithelium (RPE) cell death induced by H(2)O(2) was determined by 2', 7'-dichlorodihydrofluorescein diacetate (H(2)DCFDA) fluorescence and 3-(4, 5dimethylthiazol-2-yl)-2, 5 diphenyl tetrazolium bromide (MTT) assay. The ability of PF to protect RPE cells against ROS-mediated apoptosis was assessed by caspase-3 activity and 4', 6-diamidino-2-phenylindole (DAPI) staining. Furthermore, the protective effect of PF via the mitogen-activated protein kinase (MAPK) pathway was determined by western blot analysis. Results: PF protected ARPE-19 cells from H(2)O(2)-induced cell death with low toxicity. H(2)O(2)-induced oxidative stress increased ROS production and caspase-3 activity, which was significantly inhibited by PF in a dose-dependent manner. Pretreatment with PF attenuated H(2)O(2)-induced p38MAPK and extracellular signal regulated kinase (ERK) phosphorylation in human RPE cells, which contributed to cell viability in ARPE-19 cells. Conclusions: This is the first report to show that PF can protect ARPE-19 cells from the cellular apoptosis induced by oxidative stress. The results of this study open new avenues for the use of PF in treatment of ocular diseases, such as age-related macular degeneration (AMD), where oxidative stress plays a major role in disease pathogenesis.Biochemistry & Molecular BiologyOphthalmologySCI(E)PubMed1ARTICLE373-783512-35221

Is Asthma Related to Choroidal Neovascularization?

BACKGROUND: Age-related degeneration (AMD) and asthma are both diseases that are related to the activation of the complement system. The association between AMD and asthma has been debated in previous studies. The authors investigated the relationship between AMD and asthma systemically. PRINCIPAL FINDINGS: The epidemiological study showed that asthma was related to choroidal neovascularization (CNV) subtype (OR = 1.721, P = 0.023). However, the meta-analysis showed there was no association between AMD and asthma. In an animal model, we found more fluoresce in leakage of CNV lesions by FA analysis and more angiogenesis by histological analysis in rats with asthma. Western blot demonstrated an elevated level of C3α-chain, C3α'-chain and VEGF. After compstatin was intravitreally injected, CNV leakage decreased according to FA analysis, with the level of C3 and VEGF protein decreasing at the same time. SIGNIFICANCE: This study first investigated the relationship between AMD and asthma systematically, and it was found that asthma could be a risk factor for the development of AMD. The study may provide a better understanding of the disease, which may advance the potential for screening asthma patients in clinical practice

Genetic and functional dissection of ARMS2 in age-related macular degeneration and polypoidal choroidal vasculopathy.

Age-related maculopathy susceptibility 2(ARMS2) was suggested to be associated with neovascular age-related macular degeneration (nAMD) and polypoidal choroidal vasculopathy (PCV) in multiple genetic studies in Caucasians and Japanese. To date, no biological properties have been attributed to the putative protein in nAMD and PCV. The complete genes of ARMS2 and HTRA1 including all exons and the promoter region were assessed using direct sequencing technology in 284 unrelated mainland northern Chinese individuals: 96 nAMD patients, 92 PCV patients and 96 controls. Significant associations with both nAMD and PCV were observed in 2 polymorphisms of ARMS2 and HTRA1 rs11200638, with different genotypic distributions between nAMD and PCV (p<0.001). After adjusting for rs11200638, ARMS2 rs10490924 remained significantly associated with nAMD and PCV (p<0.001). Then we overexpressed wild-type ARMS2 and ARMS2 A69S mutation (rs10490924) in RF/6A cells and RPE cells as in vitro study model. Cell proliferation, attachment, migration and tube formation were analyzed for the first time. Compare with wild-type ARMS2, A69S mutation resulted in a significant increase in proliferation and attachment but inhibited cell migration. Moreover, neither wild-type ARMS2 nor A69S mutation affected tube formation of RF/6A cells. There is a strong and consistent association of the ARMS2/HTRA1 locus with both nAMD and PCV, suggesting the two disorders share, at least partially, similar molecular mechanisms. Neither wild-type ARMS2 nor A69S mutation had direct association with neovascularisation in the pathogenesis of AMD

Recurrence of Retinopathy of Prematurity in Zone II Stage 3+ after Ranibizumab Treatment: A Retrospective Study

Objective. To determine the prevalence and risk factors for the recurrence of retinopathy of prematurity (ROP) in Zone II Stage 3+ after ranibizumab treatment. Methods. This was a retrospective, nonrandomized, noncontrolled study that excluded Zone I and aggressive posterior ROP (APROP) cases. Infants who developed Zone II Stage 3 ROP with plus disease and underwent initial intravitreal injection of ranibizumab (IVR) were recruited. Patients were divided into 2 groups based on the outcome after initial ranibizumab treatment: recurrence of ROP or favorable outcome. Data was collected and analyzed by SPSS 16.0. Results. Forty-two patients were included, and 80 eyes with Zone II Stage 3+ were subjected to IVR treatment. Eleven of 42 patients (26.2%, 18 eyes) had a recurrence of ROP after the initial treatment. On univariate analysis, preretinal hemorrhage before treatment was significantly different between the two groups (P = 0 000). Multivariate analysis found that preretinal hemorrhage before treatment was the only factor associated with the recurrence of ROP in our study (P = 0 004). Conclusions. The recurrence rate of ROP in Zone II Stage 3+ after initial ranibizumab treatment was notable and preretinal hemorrhage before treatment was associated with the recurrence of ROP in our study.National Natural Science Foundation of China Grant [81470649]SCI(E)ARTICL

Factors related to retinal nerve fiber layer thickness in bipolar disorder patients and major depression patients

Abstract Background We analyzed the correlation of the clinical data with retinal nerve fiber layer (RNFL) thickness and macular thickness in bipolar disorder patients and major depression patients. The aim of this study is to explore factors that affect RNFL thickness in bipolar disorder patients and major depression patients, with a view to providing a new diagnostic strategy. Methods Eighty-two bipolar disorder patients, 35 major depression patients and 274 people who were age and gender matched with the patients were enrolled. Demographic information and metabolic profile of all participants were collected. Best-corrected visual acuity of each eye, intraocular pressure (IOP), fundus examination was performed. RNFL and macular thickness were measured by optical coherence tomography (OCT). Correlations between RNFL and macular thickness and other data were analyzed. Results RNFL and macula lutea in bipolar dipolar patients and major depression patients are thinner than normal people. Triglyceride and UA levels are the highest in the bipolar disorder group, while alanine aminotransferase (ALT) and glutamic oxalacetic transaminase (AST) levels in the depression group are the highest. Age onset and ALT are positively while uric acid (UA) is negatively correlated with RNFL thickness in bipolar dipolar patients. Cholesterol level is positively correlated with RNFL thickness while the duration of illness is correlated with RNFL thickness of left eye in major depression patients. Conclusions RNFL and macula lutea in bipolar dipolar patients and major depression patients are thinner than normal people. In bipolar disorder patients, age-onset and ALT are potential protective factors in the progress of RNFL thinning, while UA is the pathological factor

The expression of the Slit-Robo signal in the retina of diabetic rats and the vitreous or fibrovascular retinal membranes of patients with proliferative diabetic retinopathy.

The Slit-Robo signal has an important role in vasculogenesis and angiogenesis. Our study examined the expression of Slit2 and its receptor, Robo1, in a rat model of streptozotocin-induced diabetes and in patients with proliferative diabetic retinopathy.Diabetes was induced in male Sprague-Dawley rats via a single, intraperitoneal injection of streptozotocin. The rats were sacrificed 1, 3 or 6 months after the injection. The expression of Slit2 and Robo1 in retinal tissue was measured by real-time reverse transcription polymerase chain reaction (RT-PCR), and protein levels were measured by western blotting and immunohistochemistry. Recombinant N-Slit2 protein was used to study the effects of Slit2 on the expression of VEGF in vivo. The concentration of Slit2 protein in human eyes was measured by enzyme-linked immunosorbent assay in 27 eyes with proliferative diabetic retinopathy and 28 eyes in control group. The expression of Slit2, Robo1 and VEGF in the excised human fibrovascular membranes was examined by fluorescence immunostaining and semi-quantitative RT-PCR.The expression of Slit2 and Robo1 in the retina was altered after STZ injection. Recombinant N-Slit2 protein did not increase the retinal VEGF expression. Vitreous concentrations of Slit2 were significantly higher in the study group than in the control group. In the human fibrovascular membranes of the study group, the co-localization of VEGF with the markers for Slit2 and Robo1was observed. The expression of Slit2 mRNA, Robo1 mRNA, and VEGF mRNA was significantly higher in human fibrovascular proliferative diabetic retinopathy membranes than in the control membranes.The alteration of Slit2 and Robo1 expression in the retinas of diabetic rats and patients with proliferative diabetic retinopathy suggests a role for the Slit-Robo signal in the various stages diabetic retinopathy. Further studies should address the possible involvement of the Slit-Robo signal in the pathophysiological progress of diabetic retinopathy

Clinical and Mutation Analysis of Patients with Best Vitelliform Macular Dystrophy or Autosomal Recessive Bestrophinopathy in Chinese Population

Mutations in the gene BEST1 usually cause bestrophinopathies, such as the rare progressive diseases Best vitelliform macular dystrophy (BVMD) and autosomal recessive bestrophinopathy (ARB). This study aimed to investigate the clinical characteristics of patients with BVMD or ARB carrying BEST1 mutations. A total of 12 probands including 9 patients with a clinical diagnosis of BVMD and 3 patients with a clinical diagnosis of ARB were recruited for genetics analysis. All patients underwent detailed ophthalmic examination. All coding exons of the BEST1 gene were screened by PCR-based DNA sequencing. Programs of PolyPhen-2, SIFT, and MutationTaster were used to analyze the potential pathogenicity of the mutations in BEST1. In the 9 unrelated patients with BVMD, one heterozygous BEST1 mutation was revealed in 8 patients and two compound heterozygous mutations in 1 patient. In the 3 unrelated patients with ARB, two compound heterozygous mutations were revealed in 2 patients and three compound heterozygous mutations in 1 patient. Molecular analyses identified a total of 15 mutations, including 3 novel mutations (c.424A>G p.S142G, c.436G>A p.A146T, and c.155T>C p.L52P). Antivascular endothelial growth factor (VEGF) drugs were given to two affected eyes, especially those also exhibiting choroidal neovascularization (CNV), and no serious adverse events occurred. Our study indicates that there is wide genotypic and phenotypic variability in patients with BVMD or ARB in China. The screening of BEST1 gene is significant for the precise diagnosis of BVMD and ARB