Retinal Factor Research Articles

Benzodiazepine receptor agonist carbacetam modulates the level of vascular endothelial growth factor in the retina of rats with streptozotocin-induced diabetes

One of the primary mechanisms of retinal neurodegeneration in diabetes mellitus is gamma-aminobutyric acid (GABA) deficiency that makes the use of GABA-benzodiazepine receptor modulators a promising option for the correction of this diabetic complication. The aim of this study was to determine the effect of the benzodiazepine receptor agonist carbacetam on the expression of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α) in retina of rats with hyperglycemia. Experimental diabetes was modeled by a single administration of streptozotocin (50 mg/kg) to three-month-old male Wistar rats. Immunoblotting and immunohistochemical studies were performed using monoclonal antibodies against VEGF and HIF-1α. It was shown that the development of diabetic retinopathy (DR) at the early stages was accompanied by a progressive multifold increase in the retina content of VEGF on 7-28 days and HIF-1α on 28th day. Insulin and insulin+carbacetam treatment significantly alleviated diabetes-induced overexpression of both HIF-1α and VEGF. Carbacetam was shown to block the diabetogenic increase in VEGF content in retina. The introduction of insulin with carbacetam significantly reduced the expression of VEGF and the development of specific morphological manifestations of DR. Thus, restoration of GABA-ergic signaling can be used as a promising therapeutic option for the correction of DR disorders. Keywords: carbacetam, GABA-benzodia­zepine receptors, HIF-1α hyperglycemia, retinopathy, streptozotocin-induced diabetes, VEGF

Novel Function of Nogo-A as Negative Regulator of Endothelial Progenitor Cell Angiogenic Activity: Impact in Oxygen-Induced Retinopathy.

Endothelial Progenitor Cells (EPCs) can actively participate in revascularization in oxygen-induced retinopathy (OIR). Yet the mechanisms responsible for their dysfunction is unclear. Nogo-A, whose function is traditionally related to the inhibition of neurite function in the central nervous system, has recently been documented to display anti-angiogenic pro-repellent properties. Based on the significant impact of EPCs in retinal vascularization, we surmised that Nogo-A affects EPC function, and proceeded to investigate the role of Nogo-A on EPC function in OIR. The expression of Nogo-A and its specific receptor NgR1 was significantly increased in isolated EPCs exposed to hyperoxia, as well as in EPCs isolated from rats subjected to OIR compared with respective controls (EPCs exposed to normoxia). EPCs exposed to hyperoxia displayed reduced migratory and tubulogenic activity, associated with the suppressed expression of prominent EPC-recruitment factors SDF-1/CXCR4. The inhibition of Nogo-A (using a Nogo-66 neutralizing antagonist peptide) or siRNA-NGR1 in hyperoxia-exposed EPCs restored SDF-1/CXCR4 expression and, in turn, rescued the curtailed neovascular functions of EPCs in hyperoxia. The in vivo intraperitoneal injection of engineered EPCs (Nogo-A-inhibited or NgR1-suppressed) in OIR rats at P5 (prior to exposure to hyperoxia) prevented retinal and choroidal vaso-obliteration upon localization adjacent to vasculature; coherently, the inhibition of Nogo-A/NgR1 in EPCs enhanced the expression of key angiogenic factors VEGF, SDF-1, PDGF, and EPO in retina; CXCR4 knock-down abrogated suppressed NgR1 pro-angiogenic effects. The findings revealed that hyperoxia-induced EPC malfunction is mediated to a significant extent by Nogo-A/NgR1 signaling via CXCR4 suppression; the inhibition of Nogo-A in EPCs restores specific angiogenic growth factors in retina and the ensuing vascularization of the retina in an OIR model.

Loranthus regularis Ameliorates Neurodegenerative Factors in the Diabetic Rat Retina

Diabetic retinopathy remains a primary source of blindness with the growing pandemic of diabetes. Numerous studies have shown that early neurodegeneration caused by elevated oxidative stress may initiate microvascular damage in the diabetic retina during the last few decades. A variety of preventive and treatment strategies using phytochemicals that possess high antioxidants have shown great promise in reducing diabetes-induced neurodegeneration retinal damage. In this investigation, we employed an extract of Loranthus regularis, a traditional medicinal herb which is found to improve diabetes and associated complications in experimental studies. We orally treated STZ-induced diabetic rats with L. regularis and analyzed the neurodegenerative factors in the retina. After treatments, we used Western blotting techniques to analyze the protein content of neurotrophic factors (NGF, BDNF, TrkB), apoptotic factors (cytochrome c, Bcl-2, Bax), and phosphorylation of AKT in the diabetic retina. Additionally, we used ELISA methods to measure the contents of BDNF and the activity of Caspase-3 and biochemical procedures to determine the levels of glutathione and lipid peroxidation (TBARS). Our findings show that L. regularis treatments resulted in a considerable increase in neurotrophic factors and a decrease in apoptotic factors in the diabetic retina. Furthermore, in diabetic retina treated with L. regularis, the level of Bcl-2 protein increased, while the phosphor-AKT signaling improved. As a result, L. regularis may protect against diabetic-induced retinal neuronal damage by increasing neurotrophic support and reducing oxidative stress and apoptosis. Therefore, this study suggests that in diabetic retinopathy, L. regularis could be a potential therapy option for preventing neuronal cell death.

Combination therapy with semaglutide and rosiglitazone as a synergistic treatment for diabetic retinopathy in rodent animals

ObjectiveTo investigate the protective efficacies and potent mechanisms of combination therapy with semaglutide and rosiglitazone (RSG) on the high-glucose incubated human ARPE-19 cells and diabetic retinopathy (DR) model rats. Main methodsThe CCK-8 methods were used to evaluate the protective effects of semaglutide and RSG alone or combination on the cell viability of high-glucose treated ARPE-19 cells. After the DR rat model was established, the effects of combined treatment on general indexes, retinal morphological changes, retinal Müller cells as well as PI3K/Akt/MTOR related factors of DR model rats were investigated. ResultsThe CCK-8 assay showed obviously enhanced protective efficacies of combination therapy with semaglutide and RSG on the ARPE-19 with oxidative stress induced by high-glucose with combination index all below 1.5 demonstrating obvious synergistic effects. Combined incubation could also effectively decrease the expression of inflammatory factors, including TNF-α, IL-1β, IL-6, and the increase of ROS content in ARPE cell culture supernatant induced by high-glucose. Combined use of the antioxidant, PI3K/Akt and mTOR inhibitors, we further demonstrated that combined incubation of semaglutide and RSG could effectively by reduce high glucose-induced inflammatory injury inhibiting ROS/PI3K/Akt/mTOR signaling. Furthermore, chronic combination treatment effectively improved the histopathological characteristics and down-regulated the GFAP expression in Müller cells as well as PI3K/Akt/MTOR signaling pathway-related factors in retina which was better than any monomer treatment group. ConclusionsCombined semaglutide with RSG exhibited synergistically protective efficacies on retinal cells by decreasing the GFAP expression, inhibiting oxidative stress and PI3K/Akt/MTOR signaling-transduction in DR model rats.

Dual-Function Analysis of Astaxanthin on Golden Pompano (Trachinotus ovatus) and Its Role in the Regulation of Gastrointestinal Immunity and Retinal Mitochondrial Dysfunction Under Hypoxia Conditions.

The present study investigated the potential mechanisms of astaxanthin in the regulation of gastrointestinal immunity and retinal mitochondrial function of golden pompano (Trachinotus ovatus). Triplicate groups of juvenile T. ovatus (mean initial weight: 6.03 ± 0.01 g) were fed one of six diets (D1, D2, D3, D4, D5, and D6) for 8 weeks, with each diet containing various concentrations of astaxanthin (0, 0.0005, 0.001, 0.005, 0.01, or 0.1%, respectively). Growth performance of fish fed the D2–D5 diets was higher than that of fish fed the D1 diet; however, growth performance and survival of fish deteriorated sharply in fish fed the D6 diet. Gut villus in fish fed the D2–D5 diets were significantly longer and wider than that of fish fed the D6 diet. Feeding with D2–D5 diets led to increased abundance of Bacillus, Pseudomonas, Oceanobacillus, Lactococcus, Halomonas, Lactobacillus, and Psychrobacter while abundance of Vibrio and Bacterium decreased. Additionally, feeding with the D6 diet resulted in a sharp decline in Pseudomonas and Lactobacillus abundance and a sharp increase in Vibrio abundance. A low dissolved oxygen environment (DO, 1.08 mg/L) was conducted for 10 h after the rearing trial. No fish mortality was observed for any of the diet treatments. Lysozyme (LZY) activity in fish fed the D6 diet decreased sharply and was significantly lower than that in other groups. ROS production also decreased sharply in fish fed the D6 diet. Moreover, the conjunctiva and sclera in the fish fed the D6 diet were indistinguishable. Suitable dietary astaxanthin supplementation levels (0.005–0.1%) exerting a neuroprotective effect from low dissolved oxygen environments is due to up-regulated expression of anti-apoptotic factors, such as phosphorylated Bcl-2-associated death promoter (pBAD), phosphorylated glycogen synthase kinase-3β (pGSK-3β), Bcl-2 extra large (Bcl-xL), and down-regulated expression of Bcl-2-associated X protein (Bax) pro-apoptotic factor in retinas. Furthermore, suitable dietary astaxanthin levels (0.0005–0.01%) suppressed up-regulation of critical mitochondrial components, such as peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α), mitochondrial transcription factor A (TFAM), and mitochondrial DNA (mtDNA), while excessive astaxanthin supplementation produces the opposite effect. In brief, high-dose astaxanthin arouses and aggravates low dissolved oxygen-induced inflammation, oxidative stress, intestinal disorder, retinal apoptosis, and retinal mitochondrial dysfunction in T. ovatus. Second-degree polynomial regression of WG indicated that the optimum dietary astaxanthin for juvenile T. ovatus is 0.049%.

Col4a1 mutations cause progressive retinal neovascular defects and retinopathy.

Mutations in collagen, type IV, alpha 1 (COL4A1), a major component of basement membranes, cause multisystem disorders in humans and mice. In the eye, these include anterior segment dysgenesis, optic nerve hypoplasia and retinal vascular tortuosity. Here we investigate the retinal pathology in mice carrying dominant-negative Col4a1 mutations. To this end, we examined retinas longitudinally in vivo using fluorescein angiography, funduscopy and optical coherence tomography. We assessed retinal function by electroretinography and studied the retinal ultrastructural pathology. Retinal examinations revealed serous chorioretinopathy, retinal hemorrhages, fibrosis or signs of pathogenic angiogenesis with chorioretinal anastomosis in up to approximately 90% of Col4a1 mutant eyes depending on age and the specific mutation. To identify the cell-type responsible for pathogenesis we generated a conditional Col4a1 mutation and determined that primary vascular defects underlie Col4a1-associated retinopathy. We also found focal activation of Müller cells and increased expression of pro-angiogenic factors in retinas from Col4a1+/Δex41mice. Together, our findings suggest that patients with COL4A1 and COL4A2 mutations may be at elevated risk of retinal hemorrhages and that retinal examinations may be useful for identifying patients with COL4A1 and COL4A2 mutations who are also at elevated risk of hemorrhagic strokes.

Expression changes of brain-derived neurotrophic factor in retina of oxygen-induced retinopathy

Background Neovascular diseases such as retinopathy of prematurity often leads to irreversible vision loss.The study on oxidative damage mechanism is becoming more and more important.Whether brain derived neurotrophic factor (BNDF) has protection to retinal ganglion cells(RGCs) has few research reports. Objective The study was to investigate the expression changes of BDNFin mouse retinas of oxygen-induced retinopathy (OIR). Methods Thirty SPF C57BL/6J immature rats were divided into OIR group and normal control group and to fifteen rats for each group.OIR models were established by raising 7-day-old (P7) mice together with maternal mouse in (75±3)% oxygen environment for 5 days and then returned to the normal air environment, and the mice of the normal control group were raised in the normal air environment.The P17 mice were sacrificed for retinal histopathological examination by hematoxylin and eosin staining, and the number of vascular endothelial cell nucleus extending the inner limiting membrane was counted.Whole retinal mounts were prepared after fluorescein isothiacyanate-dextran (FITC-dextran) (9 ml/kg) was retrosorbitally injected, and the distribution of retinal vessels was observed in P17 mice.The relative expression levels of BNDF in retinas were detected in P13, P15, P17 mice, and the results were compared between the normal control group and the OIR group. Results Histopathological examination showed that retinal inner limiting membrane was smooth in the normal control group, but a lots of vascular endothelial cell nucleus extending the inner limiting membrane were seen under the optical microscope in the OIR group.The number of the vascular endothelial cell nucleus extending the inner limiting membrane was 1.70±0.68 in the normal group and 45.3±3.13 in the OIR group, showing significant difference between them (t=86.5, P=0.00). Whole retinal mount revealed that normal retinal vessels and network-like capillaries were exhibited in the mice in the normal group, while tortuous vessels, capillary loss and non-perfusion areas were revealed in the OIR group on the whole retina mounts.The relative expressing levels of BDNF in retinas were 263.992±9.451 and 218.432±9.710 in P15 and P17 mice in the OIR group, which were significantly higher than 230.324±7.779 and 115.846±7.305 in the normal control group (t=14.2, 42.3, P<0.05). Conclusions OIR can be inhibited by increasing the expression of BNDF. Key words: Brain-derived neurotrophic factor; Hyperoxia/complications; Retinal neovascularization/pathology; Disease models, animal; Mice, inbred C57BL

Insulin-like growth factor-1 binding protein 3 (IGFBP-3) promotes recovery from trauma-induced expression of inflammatory and apoptotic factors in retina

Ocular trauma affects 20% of Americans in their lifetime and can cause permanent visual system damage. We have used a mouse model of ocular trauma (exposure to an air blast from a paintball gun) to examine pathways that trigger the resulting retinal damage and to develop treatment strategies that might ameliorate the deleterious effects of trauma on retinal tissue. Our previous studies have shown that ocular blast causes an increase in protein levels of inflammatory mediators and apoptotic factors, including tumor necrosis factor alpha (TNFα) and interleukin-1-beta (IL-1β), as well as the apoptotic markers, Bax, cytochrome C, and cleaved caspase 3. Furthermore, topical treatment by eye drop application of a β-adrenergic receptor agonist, Compound 49b, was shown to decrease these inflammation/apoptosis markers and thus ameliorate the effects of blast trauma. We postulate that the protective effect of Compound 49b may be linked to its demonstrated ability to activate the β-adrenergic receptor and in turn trigger production of insulin-like growth factor binding protein 3 (IGFBP-3). In the current study, we tested this hypothesis using mice with minimal IGFBP-3 activity (IGFBP-3 knockdown mouse) vs. wildtype mice. We found that ocular blast alone did not affect IGFBP-3 levels in retinas of wild type or knockdown mice and surprisingly, the lower levels of IGFBP-3 in knockdown animals did not exacerbate the blast-induced increase in protein levels of inflammation/apoptosis markers. Nevertheless, the levels of IGFBP-3 were significantly increased in knockdown mouse retina by treatment with Compound 49b 24h post-trauma and as expected, the increase in IGFBP-3 was linked to a decrease in inflammation/apoptosis markers. We conclude that while lowered IGFBP-3 may not make the retina more vulnerable to blast injury, an increase in IGFBP-3 post-trauma may play an important role in limiting trauma-induced inflammatory and apoptotic pathways leading to retinal damage. Eye drop application of the β-adrenergic receptor agonist, Compound 49b, provides a promising treatment strategy for increasing IGFBP-3 levels to promote recovery from retinal inflammation and apoptosis after ocular blast.

Expression of Toll-like receptor 4 and inflammatory factors in retina of diabetic rats

Objective To observe the expression of Toll-like receptor 4 (TLR4) and inflammatory cytokines,leucocytic density and permeability in retina of diabetic rat.Methods A total of 106 Brown Norway rats were randomly divided into experimental group and control group with 53 rats in each group.Diabetic model was established in experimental group by intraperitoneal injection of streptozotocin,and control rats received intraperitoneal injection of an equal volume of citric acid-sodium citrate buffer.Four weeks later,the retinas were collected for further analysis.TLR4 RNA and protein expression were measured by quantitative polymerase chain reaction and Western blot.Inflammatory cytokines,including tumor necrosis factor-α (TNF-α),interleukin-1β (IL-1β),monocyte chemo-attractant protein-1 (MCP-1),were measured by enzyme-linked immunosorbent assay in rat retina homogenate.Leukocyte density in the retina was measured by acridine orange fundus angiography.The retinal permeability was evaluated by Evans blue (EB) staining.Results TLR4 expression was significantly increased in diabetic rats of experimental group compared with non-diabetic rats of control group (F =1.606,0.789; P ＜ 0.05).Inflammatory cytokines (TNF-α,IL-1β and MCP-1) were significantly increased in retina of diabetic rats of experimental group versus non-diabetic rat of control group (F=24.622,5.758,4.829; P＜0.05).The retinal leukocyte density was (6.2±0.5) × 10-5,(2.2±0.3) × 10-5 cells / pixel2 in experimental and control group respectively,the difference was statistically significant (F=2.025,P＜0.05).The amount of retinal EB leakage was (23.41 ± 4.47),(13.22 ± 3.59) ng/mg in experimental and control group respectively,the difference was statistically significant (F=21.08,P＜0.05).Conclusion TLR4 and inflammatory cytokines expression,leucocytic density and permeability increased significantly in retina of diabetic rat. Key words: Diabetic retinopathy/pathophysiology; Toll-like receptor 4/immunology; Drosophila proteins/immunology

Effect of minocycline on the expression of glial cell-derived neurotrophic factor in retina of rats after optic nerve injury

Objective To explore the neuroprotective mechanism of minocycline on retinal ganglion cells (RGC) by investigating the effect of minocycline on the expression of glial cell-derived neurotrophic factor (GDNF) in retina of rats after optic nerve injury.Methods Fifty-three SD rats were randomly divided into three groups:control group (3 rats),saline group (25 rats) and experimental group (25 rats).The left eyes in the saline and experimental group were made into the model of optic nerve injury.45 mg/kg minocycline or the same dose of normal saline was injected intraperitoneally in the rats of experimental group or saline group,respectively.The rats were euthanized at 1 d,3 d,7 d,14 d and 28 d postoperatively and the retina was collected.The morphological changes and the counting results of RGC were shown by hematoxylineosin staining.The concentration of active GDNF in retina was tested by semi-quantitative immunohistochemistry.Results The RGCs in the experimental group were more tidy with less degree of cytoplasmic vacuole formation than that in the saline group.The average number of RGC was higher in the experimental group than the saline group at all time points except 1 d.The differences were statistically significant (P ＜ 0.05).The concentrations of GDNF were higher in the experimental group than the saline group at all time points (P＜ 0.05).Conclusion Minocycline protects RGC through enhancing the expression of GDNF in the early stage after injury. Key words: Minocycline; Injury, nerve optic; Factor, neurotrophic, glial cell-derived

Exposure to As, Cd and Pb-mixture impairs myelin and axon development in rat brain, optic nerve and retina

Arsenic (As), lead (Pb) and cadmium (Cd) are the major metal contaminants of ground water in India. We have reported the toxic effect of their mixture (metal mixture, MM), at human relevant doses, on developing rat astrocytes. Astrocyte damage has been shown to be associated with myelin disintegration in CNS. We, therefore, hypothesized that the MM would perturb myelinating white matter in cerebral cortex, optic nerve (O.N.) and retina. We observed modulation in the levels of myelin and axon proteins, such as myelin basic protein (MBP), proteolipid protein, 2′-, 3′-cyclic-nucleotide-3′-phosphodiesterase, myelin-associated glycoprotein and neurofilament (NF) in the brain of developing rats. Dose and time-dependent synergistic toxic effect was noted. The MBP- and NF-immunolabeling, as well as luxol-fast blue (LFB) staining demonstrated a reduction in the area of intact myelin-fiber, and an increase in vacuolated axons, especially in the corpus-callosum. Transmission electron microscopy (TEM) of O.N. revealed a reduction in myelin thickness and axon-density. The immunolabeling with MBP, NF, and LFB staining in O.N. supported the TEM data. The hematoxylin and eosin staining of retina displayed a decrease in the thickness of nerve-fiber, plexiform-layer, and retinal ganglion cell (RGC) count. Investigating the mechanism revealed a loss in glutamine synthetase activity in the cerebral cortex and O.N., and a fall in the brain derived neurotrophic factor in retina. An enhanced apoptosis in MBP, NF and Brn3b-containing cells justified the diminution in myelinating axons in CNS. Our findings for the first time indicate white matter damage by MM, which may have significance in neurodevelopmental-pediatrics, neurotoxicology and retinal-cell biology.

Expression of Endoplasmic Reticulum Stress-Related Factors in the Retinas of Diabetic Rats

Recent reports show that ER stress plays an important role in diabetic retinopathy (DR), but ER stress is a complicated process involving a network of signaling pathways and hundreds of factors, What factors involved in DR are not yet understood. We selected 89 ER stress factors from more than 200, A rat diabetes model was established by intraperitoneal injection of streptozotocin (STZ). The expression of 89 ER stress-related factors was found in the retinas of diabetic rats, at both 1- and 3-months after development of diabetes, by quantitative real-time polymerase chain reaction arrays. There were significant changes in expression levels of 13 and 12 ER stress-related factors in the diabetic rat retinas in the first and third month after the development of diabetes, Based on the array results, homocysteine- inducible, endoplasmic reticulum stress-inducible, ubiquitin-like domain member 1(HERP), and synoviolin(HRD1) were studied further by immunofluorescence and Western blot. Immunofluorescence and Western blot analyses showed that the expression of HERP was reduced in the retinas of diabetic rats in first and third month. The expression of Hrd1 did not change significantly in the retinas of diabetic rats in the first month but was reduced in the third month.

Effects of curcumin on expression of macrophage migration inhibitory factor in retina of diabetic rats

目的 检测巨噬细胞游走抑制因子(MIF)在糖尿病大鼠视网膜中的表达,探讨MIF在糖尿病视网膜病变(DR)中的作用及姜黄素早期干预对其表达的影响.方法 对30只清洁级SD大鼠行腹腔注射质量分数1%链脲佐菌素(STZ)诱导建立糖尿病动物模型,选取其中的20只随机分为糖尿病组和姜黄素治疗组,各10只,另取10只正常匹配的大鼠作为正常对照组.姜黄素治疗组自STZ注射后3d起给予含羧甲基纤维素的姜黄素200mg/(kg*d)灌胃,共8周,正常对照组和糖尿病组给予等量的羧甲基纤维素灌胃.动物处死后收集各组大鼠眼球,苏木精-伊红染色观察大鼠视网膜组织的病理变化;采用ELISA法检测各组大鼠视网膜组织中MIF的表达;应用免疫组织化学法观察各实验组大鼠视网膜中MIF的表达及定位;透射电镜下观察大鼠视网膜超微结构的改变.结果 与正常对照组比较,糖尿病组大鼠和姜黄素治疗组大鼠血糖明显升高,差异均有统计学意义(P＜0.01);姜黄素治疗组大鼠血糖水平明显低于糖尿病组,差异有统计学意义(P＜0.01).与正常对照组比较,糖尿病组和姜黄素治疗组大鼠体重明显下降,差异均有统计学意义(P＜0.01);姜黄素治疗组大鼠体重明显大于糖尿病组,差异有统计学意义(P＜0.01).糖尿病组大鼠视网膜变薄,但姜黄素治疗组大鼠视网膜接近正常.免疫组织化学检测表明,糖尿病组视网膜组织中含有大量MIF阳性细胞,姜黄素治疗组视网膜中MIF表达明显少于糖尿病组(P＜0.01).ELISA法检测结果显示,糖尿病组MIF的表达较正常对照组显著升高(P＜0.01);姜黄素治疗组MIF的表达较糖尿病组明显下降(P＜0.01).透射电镜下糖尿病组大鼠视网膜光感受器内外节水肿,毛细血管内皮细胞和周细胞染色体边集;姜黄素治疗组大鼠视网膜超微结构的改变轻微.结论 DR与视网膜的炎性改变有关,视网膜组织中MIF的高表达参与早期DR的发生发展.姜黄素可下调MIF的表达,其早期干预可延缓DR的发病进程。

Lens regeneration in adult newt eyes related to retina pigment cells and the neural retina factor

Lens regeneration in adult newt eyes related to retina pigment cells and the neural retina factor