血红素氧合酶-1在眼组织的表达和作用

血红素氧合酶-1(heme oxygenase-1,HO-1)是一种应激反应蛋白,能被多种氧化应激分子诱导.HO-1及其产物胆红素和一氧化碳具有抗炎、抗氧化及抗内皮细胞凋亡等作用.本文就HO-1在角膜、小梁网、晶状体、葡萄膜及视网膜等眼部组织的表达及作用作一综述.     

血红素加氧酶-1在眼科疾病中的应用

血红素加氧酶-1(heme oxygenase-1,HO-1)是体内分布最广泛的抗氧化酶之一,可催化血红素代谢为胆绿素、铁离子、一氧化碳,最终发挥抗炎、抗氧化等作用。在角膜病、白内障、青光眼、糖尿病视网膜病变等疾病中,HO-1表达上调保护组织免受氧化损伤,同时其过表达与眼部肿瘤的发生发展密切相关。本文就HO-1及其代谢终产物与眼科疾病的关系联合国内外最新研究进展作一综述。     

血红素氧合酶-1与糖尿病性视网膜病变:氧化应激的作用(英文)

糖尿病性视网膜病变(diabetic retinopathy,DR)是糖尿病最常见且严重的微血管并发症之一,其发病机制复杂,是多因素、多阶段作用的结果。近年研究表明,DR的发生与氧化应激密切相关,抗氧化治疗有助病情改善。血红素氧合酶-1(heme oxygenase-1,HO-1)是一种广泛存在的抗氧化防御酶,可对抗氧化应激造成的损伤,具有重要的生理作用。研究表明,在高血糖环境中,视网膜内HO-1的表达被诱导增高,且通过人为调节HO-1的表达水平可以加速或延缓病情的进展,提示将HO-1应用于DR的诊治有良好的应用前景。本文从氧化应激的角度对二者加以概述。     

血红素氧合酶-1对视网膜光损伤的保护作用

血红素氧合酶-1（heine oxygenase-1，HO-1）又称热休克蛋白32，是体内的一种内源性的保护因子，具有抗氧化应激、抗炎、抗凋亡等多种细胞保护作用。视网膜是接受光刺激、形成视觉的重要组织，当光线强度或光照时间等超过了视网膜的承受能力时，将会刺激视网膜产生大量的自由基、脂质过氧化物等，其病理过程与年龄相关性黄斑变性相似，因此对防治视网膜光损伤药物的研究有重要的临床价值。     

血红素氧合酶1的抗凋亡机制及其在相关眼病中的保护作用

血红素氧合酶1(HO-1)作为应激反应蛋白,其抗凋亡的研究成为新热点,而近年来研究表明,HO-1在眼病中起着重要的保护作用,HO-1是血红素分解代谢的限速酶,能催化血红素转化为胆绿素、一氧化碳和游离铁,均为抗凋亡的主要效应分子。就HO-1的生物学特性及其抗凋亡机制进行阐述,并分别对HO-1在角膜病、青光眼、白内障、葡萄膜炎、Graves眼病、糖尿病视网膜病变、视网膜色素变性等眼病中所起保护作用及相关机制的研究进展进行综述。     

血红素加氧酶-1可能的视网膜视神经保护作用

血红素加氧酶(HO)是血红素代谢途径中的限速酶,在体内分布广泛。该代谢不仅清除机体内细胞毒性的血红素,其伴随产物一氧化碳、胆绿素、铁/铁蛋白均在机体发挥重要的抗氧化、抗凋亡、抗炎症等效应,因此HO系统作为机体重要的防御系统被人们认识。视网膜组织中HO含量丰富,可以减轻视网膜组织对光刺激、缺血缺氧、氧化应激等危险因素的损伤,本文主要探讨HO-1可能的视网膜视神经的保护作用。     

抗氧化剂防治噪声性聋的研究进展

氧化应激产生的活性氧自由基(ROS)参与了耳蜗毛细胞的坏死、凋亡过程,是噪声性聋（NIHL发生的重要机制。已有诸多针对性的抗氧化应激药物应用于NIHL的防治研究中,通过上调内源性抗氧化系统和补充外源性抗氧化剂维持体内氧化-抗氧化平衡。本文对内耳生理状态的氧化-抗氧化机制及抗氧化相关药物在NIHL防治中的研究进行总结。     

血红素氧合酶-CO-cGMP通路在体外培养人眼小梁细胞中的表达

目的 探讨体外培养人眼小梁细胞是否存在血红素氧合酶（HO）-一氧化碳（CO）-环磷酸鸟苷（cGMP）通路以及氯化血红素（hemin）对其的诱导作崩。方法 应用RT-PCR和免疫组织化学方法检测体外培养人眼小梁细胞中HO-1和HO-2 mRNA及蛋白的表达。向细胞培养液中加入氯化血红素，RT-PCR检测小梁细胞HO-1 mRNA，分光光度法检测培养上清液中碳氧血红蛋白（HbCO）浓度，并用放射免疫法检测细胞中cGMP浓度。结果 体外培养人眼小梁细胞存在HO-1和HO-2 mRNA及蛋白。氯化IffL红素对小梁细胞HO-1 mRNA、HbCO和cGMP浓度的诱导呈浓度依赖性。结论 人眼小梁细胞存在HO-CO-cGMP通路，并受氯化血红素诱导。药理诱导这一通路，可能为青光眼的治疗提供一个有效的新方法。     

基于Nrf2通道研究柚皮素磷脂复合物对氧化损伤ARPE-19细胞的保护作用

目的：研究柚皮素(Nar)及其磷脂复合物(NPC)对叔丁基氢过氧化物(t-BHP)诱导产生的氧化损伤人视网膜色素上皮细胞(ARPE-19细胞)的保护作用及其作用机制。

方法：根据文献最佳制作工艺制备NPC。将体外培养的ARPE-19细胞分为溶媒组(用DMSO培养)、模型组(用200μmol/L t-BHP干预)、Nrf2-siRNA组(针对Nrf2基因进行细胞转染)、柚皮素组(200μmol/L柚皮素培养基预处理后加入200μmol/L t-BHP)、NPC组(200μmol/L NPC培养基预处理后加入200μmol/L t-BHP)、Nrf2-siRNA+柚皮素组(200μmol/L柚皮素预处理后Nrf2基因干扰,再加入200μmol/L t-BHP)、Nrf2-siRNA+NPC组(200μmol/L NPC预处理后Nrf2基因干扰,再加入200μmol/L t-BHP)。检测各组细胞内活性氧、丙二醛、超氧化物歧化酶、总抗氧化能力水平,分别采用RT-PCR和Western blot法检测各组细胞内血红素加氧酶-1(HO-1)、醌氧化还原酶-1(NQO-1)、谷氨酸半胱氨酸连接酶(GCL)和核因子E2相关因子2(Nrf2)的表达水平。

结果：NPC较柚皮素能明显降低ARPE-19细胞内丙二醛、活性氧的含量,提高细胞内超氧化物歧化酶、总抗氧化能力水平。柚皮素和NPC预保护ARPE-19 细胞后,Nrf2、HO-1、NQO-1和GCL mRNA的相对表达量和蛋白表达水平均较模型组和Nrf2-siRNA组升高。柚皮素组与NPC组,Nrf2-siRNA+柚皮素组与Nrf2-siRNA+NPC组细胞内4种基因和蛋白相对表达水平均有差异。Nrf2-siRNA+柚皮素组与Nrf2-siRNA组Nrf2、HO-1、NQO-1蛋白表达无显著差异。Nrf2-siRNA+NPC组与Nrf2-siRNA组4种蛋白表达均有差异,NPC作用明显强于柚皮素。

结论：柚皮素磷脂复合物能明显提高细胞内抗氧化能力,降低氧化水平,其通过激活Nrf2/ARE抗氧化应激通路上调Nrf2及其下游抗氧化酶和Ⅱ相解毒酶的表达对氧化损伤的ARPE-19细胞起到更好的保护作用。     

α-硫辛酸对蓝光诱导损伤的ARPE-19细胞的保护作用

目的 研究α-硫辛酸(ALA)对蓝光诱导损伤的ARPE-19细胞的保护作用。方法 体外培养ARPE-19细胞，随机分为正常组(N组)、药物组(D组)、光照组(L组)、药物+光照组(D+L组)。倒置显微镜下观察ARPE-19细胞形态结构。CCK-8法检测各组ARPE-19细胞活性；流式细胞仪(Annexin V/PI双染色法)检测ARPE-19细胞凋亡率；倒置荧光显微镜检测ARPE-19细胞ROS水平；实时荧光定量PCR(RT-qPCR)检测ARPE-19细胞核因子E2相关因子2(Nrf2)、谷胱甘肽巯基转移酶(GST)、NAD(P)H醌氧化还原酶1(NQO1)、血红素氧合酶-1(HO-1) mRNA表达水平；Western blot检测ARPE-19细胞天冬氨酸特异性半胱氨酸蛋白酶-3(Caspase-3)、Nrf2、NQO1蛋白表达水平。结果 N组及D组细胞均呈贴壁生长，分布均匀，形态清晰，细胞存活率差异无统计学意义(P>0.05)。与N组相比，L组细胞皱缩、变小、变圆，失去正常形态，部分细胞不能贴壁生长，细胞存活率降低，细胞凋亡率升高，ROS水平升高(均为P<0.05)...     

Oxidative stress is an early event in hydrostatic pressure induced retinal ganglion cell damage

PURPOSE: To determine whether oxidative adduct formation or heme oxygenase-1 (HO-1) expression are altered in retinal ganglion cell (RGC) cultures exposed to elevated hydrostatic pressure and in a mouse model of glaucoma. METHODS: Cultured RGC-5 cells were subjected to 0, 30, 60, or 100 mm Hg hydrostatic pressure for 2 hours, and the cells were harvested. Parallel experiments examined the recovery from this stress, the effect of direct 4-hydroxy-2-nonenal (HNE) treatment, and the effect of pretreatment with resveratrol or quercetin. Mice were anesthetized and intraocular pressure was increased to 30, 60, or 100 mm Hg for 1 hour; then the retinas were harvested. HNE adduct formation and HO-1 expression were assessed by immunocytochemistry and immunoblotting. RESULTS: Increases of HNE-protein adducts (up to 5-fold) and HO-1 expression (up to 2.5 fold) in pressure-treated RGC-5 cells were dose dependent. During recovery experiments, HNE-protein adducts continued to increase for up to 10 hours; in contrast, HO-1 expression decreased immediately. HNE, at a concentration as low as 5 muM, led to neurotoxicity in RGC-5 cells. HNE adducts and HO-1 expression increased in the mouse retina and optic nerve after acute IOP elevation up to 5.5-fold and 2-fold, respectively. Antioxidant treatment reduced the oxidative stress level in pressure-treated RGC-5 cells. CONCLUSIONS: This study demonstrates that oxidative stress is an early event in hydrostatic pressure/IOP-induced neuronal damage. These findings support the view that oxidative damage contributes early to glaucomatous optic neuropathy.     

Heme oxygenase in the retina in diabetes

INTRODUCTION: Heme oxygenase (HO) isoforms, HO-1, and HO-2, are responsible for heme breakdown to iron and carbon monoxide (CO). HO may respond to oxidative stress and may modulate the expression of vasoactive factors like nitric oxide (NO). Since diabetes induced oxidative stress may change HO, the present study examined whether diabetes is associated with HO alterations, its relationship with NO, endothelin-1(ET-1) and the functional significance. MATERIALS AND METHODS: Male SD rats with Streptozotocin induced diabetes were investigated after six-weeks. Poorly controlled diabetic animals were randomized to one of three treatment groups (n = 6 each group); a) untreated, b) HO-1 inhibitor SnPP-IX (50 micromol/kgIP/day), c) NO donor molsidomine (120 mg/L PO/day) and were compared with age and sex matched non diabetic control animals with or without SnPP-IX treatment. Color Doppler ultrasound analysis was used to determine retinal resistivity index (RI). mRNA for HO-1, HO-2, ET-1, eNOS and iNOS were analyzed with competitive RT-PCR. HO distribution in the retina was investigated by immunocytochemistry. RESULTS: Diabetic animals expressed lower body weight, higher blood glucose and increased glycated hemoglobin levels. HO-1 and HO-2 immuno-reactivity were identified in the retina. Diabetes induced increased RI was associated with up-regulation of both ET-1 and HO-1 mRNA expression but not eNOS or iNOS mRNA. Both SnPP-IX and molsidomine treatments prevented a diabetes increase of RI, in spite of increased ET-1 expression and were associated with increased iNOS mRNA. CONCLUSIONS: The present data suggests that the HO system is up-regulated in short term diabetes leading to HO and NO interactions which may modulate vascular function in the retina.     

Protective effects of upregulated HO-1 gene against the apoptosis of human retinal pigment epithelial cells in vitro

AIM: To assess the correlation between disorganization of the retinal inner layers (DRIL) and best-corrected visual acuity (BCVA) in patients with uveitis and macular edema (UME) who underwent systemic treatment using optical coherence tomography (OCT). METHODS: A retrospective clinical study of 23 patients (30 eyes) with DRIL and 23 patients (31 eyes) without DRIL secondary to UME were included. All patients underwent comprehensive ophthalmic examinations at baseline, 3, 6, and 12mo after local and systemic treatment. The OCT-based parameters included: foveal center point thickness (FCPT), mean thickness (MT), and diameters of DRIL in horizontal and vertical directions. BCVA and OCT-based parameters were compared between the two groups. The relationship between each OCT parameter and BCVA was evaluated using linear correlation and regression analysis. RESULTS: At the initial visit, the mean baseline FCPT was 441.03±128.68 μm in the eyes with DRIL and 337.26±99.31 μm in the eyes without DRIL (P=0.001). No significant differences were observed in MT (P=0.357). The mean size of transverse and vertical diameters of DRIL was 684.07±267.51 μm, and 267.07±104.61 μm at baseline, respectively. There was significant improvement in BCVA and OCT-based parameters at baseline, 3, 6, and 12mo in all cases (P<0.001 for each timepoint). In addition, significant differences were detected in BCVA and OCT parameters between eyes with and without DRIL at each time point (P<0.01 for each timepoint). A greater DRIL range at baseline was associated with a worse baseline BCVA (transverse diameter of DRIL: r=0.875, P<0.001; vertical diameter of DRIL: r=0.622, P<0.001). The transverse diameter of baseline DRIL was found to be significantly correlated with the final BCVA (P=0.003). CONCLUSION: The improvement in BCVA is associated with DRIL in patients with UME. DRIL is an easy-to-determine and robust imaging biomarker that could help predict BCVA prognosis in eyes with UME.     

Biliverdin Rescues the HO-2 Null Mouse Phenotype of Unresolved Chronic Inflammation Following Corneal Epithelial Injury

PURPOSE. The heme oxygenase system (HO-1 and HO-2) represents an intrinsic cytoprotective and anti-inflammatory pathway based on its ability to modulate leukocyte migration and to inhibit the expression of inflammatory cytokines and proteins by its products biliverdin/bilirubin and carbon monoxide. Corneal injury in HO-2 null mice leads to impaired healing and chronic inflammatory complications, including ulceration and neovascularization. The authors examined whether topically administered biliverdin can counteract the effects of HO deficiency in a corneal epithelial injury model. METHODS. HO-2 null mice were treated with biliverdin 1 hour before epithelial injury and twice a day thereafter. Reepithelialization and neovascularization were assessed by fluorescein staining and vital microscopy, respectively, and were quantified by image analysis. Inflammation was quantified by histology and Gr-1-specific immunofluorescence, and oxidative stress was assessed by DHE fluorescence. RESULTS. Treatment with biliverdin accelerated wound closure, inhibited neovascularization and reduced epithelial defects. It also reduced inflammation, as evidenced by a reduction in the appearance of inflammatory cells and the expression levels of inflammatory and oxidant proteins, including KC and NOXs. CONCLUSIONS. The results clearly show that biliverdin, directly or through its metabolism to bilirubin by biliverdin reductase-the expression of which is increased after injury-rescues the aberrant inflammatory phenotype, further underscoring the importance of the HO system in the cornea for the execution of an ordered inflammatory and reparative response.     

Oxidative stress--induced single-strand breaks in chromosomal telomeres of human retinal pigment epithelial cells in vitro

PURPOSE: To demonstrate that chronic hyperoxia induces single-stranded breaks in chromosomal telomeres as a measure of oxidative DNA damage in cultured RPE cells. METHODS: RPE340 cells were cultured in 40% and 20% (control) O(2). DNA damage was assessed by mean terminal restriction fragment (TRF) length, and the S1 nuclease assay was used to determine the frequency of single-strand breaks in telomeric DNA. The degree of oxidative stress in cells was estimated by flow cytometric analysis of reactive oxygen intermediate (ROI)-induced 2',7'-dichlorodihydrofluorescein diacetate fluorescence and Northern blot analysis of heme oxygenase-1 (HO-1) mRNA induction. RESULTS: The mean TRF length of cells grown in 40% O(2) shortened at a faster rate than those grown in 20% O(2). The S1 nuclease assay showed that the accelerated mean TRF length shortening was due to an increased accumulation of single-stranded breaks in telomeric DNA. The degree of ROI production and HO-1 mRNA induction was greater in cells treated with 40% than 20% O(2), an effect that was also larger in old than young passaged cells. CONCLUSIONS: RPE340 cells in vitro grown in chronic hyperoxia exhibited evidence of DNA damage with accelerated telomeric shortening via an increased accumulation of single-strand breaks in telomeric DNA. These changes could provide insight into aging of RPE cells by oxidative DNA damage.     

Effect of N-acetylcysteine on the early expression of inflammatory markers in the retina and plasma of diabetic rats

Purpose:  The aim of this study is to investigate markers of inflammation and oxidative stress in an early model of diabetic retinopathy, correlate retinal and plasma results and evaluate the influence of treatment by N -acetylcysteine (NAC), a free radical scavenger.
Methods:  Four groups were studied: control (C), streptozotocin (STZ)-induced diabetic rats (D), STZ rats following 8 weeks of NAC (DT), and control rats following 8 weeks of NAC (CT). Plasma levels of free 15-F2t-isoprostane (15-F-2t-IsoP), superoxide dismutase (SOD) and tumour necrosis factor-alpha (TNF-α) were obtained. Primary antibodies against macrophages (ED-1), microglia (Ox-42), pericytes (NG-2), endothelial and perivascular cells (IB-4), haem oxygenase 1 (HO-1) and vascular endothelial growth factor (VEGF) were used.
Results:  Expression of NG-2 was robust in C, CT, DT, and mild in D. The intensity of IB-4 was higher in D and DT compared with the C and CT. Ox-42 and ED-1 expression was higher in the D than in the DT, C or CT. Expression of VEGF and HO-1 was non-specific across the four groups. Plasma levels of 15-F-2t-IsoP and TNF-α were higher in the D as compared with the C, CT and DT. SOD levels were lower in the D when compared with the C, CT and D.
Conclusions:  Macrophage/microglia activation, pericyte loss and endothelial/perivascular cell changes occur early in the pathogenesis of DR. These changes are associated with an increase in plasma markers of oxidative stress and inflammation and are minimized by treatment with NAC. The results suggest that therapies that reduce free radicals will help minimize the early events in diabetic retinopathy in the STZ model.