视网膜色素上皮、Bruch膜及脉络膜老年性改变的定量分析

目的 研究及定量分析视网膜色素上皮（retinal pigment epithelium,RPE)、Bruch膜及脉络膜的老年性改变。方法 应用改良Masson三色组织染色法及电脑图像分析软件对36例不同年龄(3～93岁)正常眼RPE数量和高度、Bruch膜厚度、脉络膜毛细血管的密度及脉络膜厚度的老年性改变进行定量分析。 结果 与年龄有显著相关性的老年性改变包括RPE细胞数量的减少(8个/ mm²·年)、RPE细胞高度的增加(0.01μm/年) 以及Bruch膜增厚(0.02μm/年);脉络膜毛细血管密度及脉络膜厚度的改变与年龄之间无显著的相关关系。 结论 RPE细胞数量减少、高度增加以及Bruch膜的增厚可能是黄斑区最早的老年性改变。(中华眼底病杂志,2000,16:236-239)     

自体带Bruch膜视网膜色素上皮复合体移植治疗渗出性老年黄斑变性近期疗效观察

目的 探讨自体带Bruch膜视网膜色素上皮复合体移植术治疗渗出性老年性黄斑变性近期疗效.方法 回顾性分析30例确诊为渗出性老年性黄斑变性的患者,采用脉络膜新生血管膜切除联合自体带Bruch膜视网膜色素上皮复合体移植手术方法.手术前后行视力、荧光素眼底血管造影、吲哚氰绿眼底血管造影、多焦视网膜电图、光学相干断层扫描检查评价疗效.结果 移植术后随访6个月时,25例(83.33%)患者视力获得提高,2例(6.66%)患者视力不变,3例(10%)患者视力下降;所有患者黄斑区RPE植片平铺在位,未见CNV复发.结论 自体带Bruch膜视网膜色素上皮复合体移植治疗渗出性老年性黄斑变性近期疗效确切,是治疗渗出性老年性黄斑变性安全有效的手段之一.     

自体带Bruch膜视网膜色素上皮复合体移植治疗渗出性老年黄斑变性近期疗效观察

目的 探讨自体带Bruch膜视网膜色素上皮复合体移植术治疗渗出性老年性黄斑变性近期疗效.方法 回顾性分析30例确诊为渗出性老年性黄斑变性的患者,采用脉络膜新生血管膜切除联合自体带Bruch膜视网膜色素上皮复合体移植手术方法.手术前后行视力、荧光素眼底血管造影、吲哚氰绿眼底血管造影、多焦视网膜电图、光学相干断层扫描检查评价疗效.结果 移植术后随访6个月时,25例(83.33%)患者视力获得提高,2例(6.66%)患者视力不变,3例(10%)患者视力下降;所有患者黄斑区RPE植片平铺在位,未见CNV复发.结论 自体带Bruch膜视网膜色素上皮复合体移植治疗渗出性老年性黄斑变性近期疗效确切,是治疗渗出性老年性黄斑变性安全有效的手段之一.     

自体带Bruch膜视网膜色素上皮复合体移植治疗渗出性老年黄斑变性近期疗效观察

目的 探讨自体带Bruch膜视网膜色素上皮复合体移植术治疗渗出性老年性黄斑变性近期疗效.方法 回顾性分析30例确诊为渗出性老年性黄斑变性的患者,采用脉络膜新生血管膜切除联合自体带Bruch膜视网膜色素上皮复合体移植手术方法.手术前后行视力、荧光素眼底血管造影、吲哚氰绿眼底血管造影、多焦视网膜电图、光学相干断层扫描检查评价疗效.结果 移植术后随访6个月时,25例(83.33%)患者视力获得提高,2例(6.66%)患者视力不变,3例(10%)患者视力下降;所有患者黄斑区RPE植片平铺在位,未见CNV复发.结论 自体带Bruch膜视网膜色素上皮复合体移植治疗渗出性老年性黄斑变性近期疗效确切,是治疗渗出性老年性黄斑变性安全有效的手段之一.     

老年性Bruch膜及视网膜色素上皮的脂类沉积

目的验证Bruch膜及视网膜色素上皮(retinal pigment epit helium,RPE)脂类沉积与年龄的关系,以及老年性黄斑变性(age-related macular degener ation,AMD)病眼的脂类沉积特点。方法应用组织化学染色的方法,对32例尸体正常眼球(20天～84岁)及2例尸体患AMD病眼进行研究,检测其Bruch膜及RPE 中的脂类沉积。结果正常眼中Bruch膜中脂类沉积随年龄增长而增加,以中性脂类为主。黄斑区较周边部显著。RPE染色的强度和面积也与年龄有关。AMD眼脂类染色较正常老年眼更强烈,亦以黄斑部为著。结论老年性Bruch膜与AMD眼有共同的区域性分布,可能在老年性改变及AMD的发展中有着重要作用。随年龄增强的RPE脂类染色可能反映了细胞内脂褐素的积聚。(中华眼底病杂志,1999,84-87)     

激光光凝后视网膜色素上皮、Bruch膜和脉络膜超微结构改变

目的观察激光光凝后视网膜色素上皮(RPE)、Bruch膜和脉络膜超微结构的改变和修复过程。方法收集眼眶恶性肿瘤患者7例,其眼球的视网膜及脉络膜结构基本正常。均经患者签署知情同意书后,于眶内容物剜除术前1、3、7d,用Ⅰ、Ⅱ、Ⅲ级光斑光凝视网膜。摘除眼球后,将其视网膜和脉络膜组织制成超薄切片、醋酸铀和枸橼酸铅染色,于透射电镜下观察其超微结构。结果激光光凝后1d,经Ⅰ级光斑光凝视网膜组织见部分RPE细胞肿胀、坏死、数量减少,微绒毛部分消失;Bruch膜完整;脉络膜毛细血管内皮肿胀,少数管腔闭塞。经Ⅱ级光斑光凝的视网膜组织见较多RPE细胞溶解破坏,Bruch膜空泡形成,脉络膜毛细血管广泛闭塞。经Ⅲ级光斑光凝的视网膜组织见大量RPE溶解消失,Bruch膜部分破坏,脉络膜血管受损。激光光凝后3d,RPE细胞和脉络膜水肿减退,破坏区内出现吞噬细胞,RPE细胞和脉络膜纤维组织开始增生。激光光凝后7d,增生的RPE细胞覆盖破坏区,脉络膜血管减少和基质纤维化。结论激光光凝可引起RPE细胞水肿、坏死,Bruch膜和脉络膜血管不同程度破坏,随后RPE细胞和纤维组织增生修复破坏区。不同参数的激光治疗可修复病变的RPE和脉络膜组织。（中华眼科杂志,2004,40：692-695)     

视网膜色素上皮的老年性改变

视网膜色素上皮对维持视网膜的正常功能具有重要意义,其老年性改变可能导致多种形式的视网膜病理变化,特别是老年性黄斑变性。     

老年黄斑变性的视网膜色素上皮及其视功能的改变

目的：应用黄斑区密集点矩阵视野并结合眼底荧光血管造影电脑眼底图像分析老年黄斑变性视网膜色素上皮损害与ＤＭＭＭ的视野光敏感度的改变，找出改变的规律有评价其应用价值。方法：用Ｈｕｍｐｈｒｅｙ－６４０视野计自行设计ＤＭＭＭ检测ＡＭＤ干性组５１人６４只眼湿性且２３人２７只眼，用Ｚｅｉｓｓ公司生产的ＫＯＮＴＲＯＮＥＬＥＫＴＲＯＮＩＫ Ｉｎａｇｅ Ａｎａｌｙｓｉｓ Ｄｉｖｉｓｉｏｎ－计算机图像分析仪总丢失量显     

高脂血症和视网膜色素上皮层-Bruch膜-脉络膜毛细血管复合体的相关性研究进展

高脂血症和许多眼底临床疾病相关,它是引起眼科疾病的一个危险因素,国内外学者对此进行了多方面的研究。近年来研究发现,血脂异常可以导致视网膜色素上皮细胞的形态损害、Bruch膜脂质的沉着且破坏其完整性和导致脉络膜毛细血管内皮细胞的病理改变。本文就高脂血症对视网膜色素上皮层-Bruch膜-脉络膜毛细血管复合体的影响及其机制的研究进展进行综述,为预防和治疗相关眼科疾病提供思路。     

自体视网膜色素上皮-脉络膜薄片移植术治疗年龄相关性黄斑变性

目前对于渗出性年龄相关性黄斑变性（AMD）的治疗主要有温热疗法、光动力疗法和玻璃体内注射抗血管内皮生长因子等疗法。手术彻底去除脉络膜新生血管（CNV）不可避免地会造成视网膜色素上皮（RPE）、Bruch膜和脉络膜毛细血管的损伤。黄斑转位术虽可使RPE维持黄斑中心凹光感受器的功能，但由于增生性玻璃体视网膜病变、黄斑水肿和复视等并发症的发生率较高而限制其应用。     

Clathrin and adaptin accumulation in drusen, Bruch's membrane and choroid in AMD and non-AMD donor eyes

Clathrin was identified in a recent proteomic analysis of Bruch's membrane from age-related macular degeneration (AMD) donor eyes. The present study was conducted to determine the localization of clathrin in AMD tissues and to compare this distribution and relative content with that in non-AMD control tissues. The distribution of adaptin, which is functionally linked to clathrin, was also evaluated. Human eyes were from donors between 66 and 94 years of age; 13 eyes were from donors with AMD and 13 from non-AMD donors. Bruch's membrane and choroid from the macula of each donor eye were prepared for immunohistochemistry and Western blotting. Differences in immunoreactivity were quantitated. Drusen, Bruch's membrane and choroid from AMD tissues showed greater immunoreactivity for clathrin and adaptin than did non-AMD tissues. Western blots also showed more intense clathrin and adaptin immunoreactivity in AMD tissues than were present in non-AMD samples. This study suggests that accumulation of clathrin and adaptin in drusen, Bruch's membrane and choroid may reflect a higher rate of clathrin mediated endocytosis in AMD tissues. Alternatively, the accumulation of these proteins in these extracellular compartments may reflect a higher susceptibility to oxidative damage.     

脂肪干细胞体外诱导获得RPE样细胞及其治疗AMD相关研究进展

年龄相关性黄斑变性（age-related macular degeneration,AMD）是一种随年龄增长而发病率逐渐升高的黄斑部疾病.主要由视网膜色素上皮（retinal pigment epithelium,RPE）和视网膜退行性变引起的不可逆性中心视力下降.人类脂肪干细胞（adipose-derived stem cells,ADSC）具有多向分化潜能.近年研究发现,ADSC联合视网膜细胞诱导因子可体外诱导ADSC向RPE细胞分化,并表达RPE细胞的标志.相关动物模型显示,由ADSC分化而来的RPE样细胞移植后可参与视网膜重建及修复.ADSC移植为AMD的治疗提供了新思路.     

干性ARMD中视网膜色素上皮细胞的作用及损伤机制

年龄相关性黄斑变性(age-related macular degeneration,ARMD)是导致老年人中心视力不可逆丧失的主要原因。ARMD的典型特征是视网膜色素上皮(retinal pigment epithelium,RPE)和脉络膜毛细血管发生退行性改变及黄斑区出现玻璃膜疣。临床上ARMD分为两种亚型：非渗出型(干性或萎缩型)和渗出型(湿性或新生血管型)。该病的发生是年龄、环境、遗传、吸烟、氧化应激和心血管功能障碍等多种因素相互作用的结果。鉴于RPE细胞在ARMD发病中的重要作用,现以RPE细胞为重点,总结了蓝光、吸烟、氧化应激、脂褐素积累、慢性炎症和蛋白质稳态对干性ARMD发病的作用和可能机制,为认识和预防干性ARMD的发生提供新的帮助。     

Effect of Tetramethylpyrazine on RPE degeneration, choroidal blood flow and oxidative stress of RPE cells

AIM: To study the effects of Tetramethylpyrazine (TMP) on retinal pigment epithelium (RPE) degeneration, choroidal blood flow and oxidative stress of RPE cells. METHODS: The 35mg/kg NaIO3-induced RPE degeneration rat eyes was given 25μg 1% TMP eye drops 3 times a day for 7 days before NaIO3 injection, and then 2 to 4 weeks after NaIO3 injection. RPE function was measured with c-wave of electroretinogram (ERG). Colored microsphere technique was used for in vivo experiments to determine the choroidal blood flow in ocular hypertensive (40mmHg) rabbit eyes. Methylthiazoltetrazolium (MTT) assay was used to study in vitro effect of TMP on various oxidants induced injury in the hRPE (ARPE-19 (ATCC, Manassas, VA, USA)) . RESULTS: Two weeks after NaIO3 injection, the amplitude of ERG c-wave fell markedly in NaIO3 group to 36% of control group(P <0.01). No apparent difference was observed in TMP+NaIO3 group. Four weeks later, the NaIO3 group fell to 46% of control group (P<0.01), while the TMP+NaIO3 group fell to only 77% of control group (P<0.01). There was a 67% reversal of the ERG c-wave by TMP as compared to NaIO3 group(P<0.01). The choroidal blood flow was significantly increased at all time points (at 30, 60 and 120 minutes after TMP instillation) as compared with corresponding controls. TMP had no effect on hypoxia-(1%O2), t-BHP- and H2O2-induced damage in RPE cells. 10(g/mL TMP could reverse 1 and 3mM NaN3-induced loss of viability of RPE by 18.5% (P <0.01) and 23% (P<0.01), respectively. 30μg/mL TMP could reverse 30 and 100mM NaIO3 induced loss of viability of RPE by 18.1% (P <0.05) and 16.8% (P <0.01), respectively. CONCLUSION: TMP can significantly protect RPE from NaIO3 induced degeneration in vivo and oxidative stress in vitro and can increase choroidal blood flow markedly in vivo.     

Tetramethylpyrazine在视网膜色素上皮细胞变性及脉络膜血流和RPE细胞氧化应激中的作用

目的：研究1% Tetramethylpyrazine （TMP）在视网膜色素上皮细胞（RPE）变性,脉络膜血流和RPE细胞氧化应激中的作用.方法：在碘酸钠诱导的大鼠RPE变性研究中,1%TMP滴眼液预先处理1wk,3次/d,1wk后予碘酸钠舌下静脉注射,在2wk和4wk末,视网膜电图（ERG）测量c波.色素微球体技术分析高眼压状态下TMP对脉络膜血流的影响.Methylthiazoltetrazolium （MTT）分析TMP在各种氧化应激中对RPE的保护作用.结果：碘酸钠注射后2wk,碘酸钠组ERG的c波下降至对照组的36%（P〈0.01）.4wk后,碘酸钠组下降至对照组的46%（P〈0.01）,而1% TMP ＋碘酸钠组下降至对照组的77%（P〈0.01）.与碘酸钠组比较,1%TMP ＋碘酸钠组控制了67%的c波下降（P〈0.05）.在脉络膜血流的测量中,30,60,和120min的结果显示,TMP显著增加脉络膜血流.在氧化应激部分,不同浓度的TMP在各种氧化应激损伤中,对RPE都有各种程度的保护作用.结论：浓度为1% Tetramethylpyrazine可以显著保护碘酸钠和氧化应激诱导的RPE变性,增加脉络膜血流,并可能在AMD的治疗中发挥作用.     

Biological effects of native and oxidized low-density lipoproteins in cultured human retinal pigment epithelial cells

Age-related macular degeneration (AMD) and artherosclerosis share common characteristics in their pathogenesis. In this study, we investigated the effects of lipoproteins like native (n)-LDL, oxidized (ox)-LDL and high-density lipoprotein (HDL) on advanced senescence, extracellular matrix accumulation, cell loss, and transforming growth factor-beta2 (TGF-β2) expression in cultured human retinal pigment epithelial (RPE) cells. Primary human RPE cells were incubated with 10-100 μg/ml n-LDL, ox-LDL, and HDL for 24 h. For determination of advanced senescence, beta-galactosidase staining was used. The induction of fibronectin (Fn), laminin alpha 1 (Laa1), and collagen type IV alpha 2 (Col4a2) mRNA was quantified by real-time PCR. Cell loss was investigated by live dead assay. Expression of TGF-β2 was analyzed by real-time PCR and ELISA assays. Ox-LDL accelerated dose-dependently the onset of RPE senescence, whereas LDL and HDL had no effect. LDL and ox-LDL led to induced expression of Fn, Laa1 and Col4a2, whereas HDL had no influence. Incubation of RPE cells with 100 μg/ml ox-LDL induced marked cell death compared to untreated control cells. Expression of TGF-β2 was dose-dependently increased by LDL and ox-LDL.LDL and ox-LDL induced cellular changes in RPE cells in vitro, which may resemble pathogenic events of AMD. These results may provide further information about the effects of LDL and ox-LDL in the human RPE and their potential role in the pathogenesis of AMD.     

Tetramethylpyrazine在视网膜色素上皮细胞变性及脉络膜血流和RPE细胞氧化应激中的作用

目的:研究1%Tetramethylpyrazine(TMP)在视网膜色素上皮细胞(RPE)变性,脉络膜血流和RPE细胞氧化应激中的作用。方法:在碘酸钠诱导的大鼠RPE变性研究中,1%TMP滴眼液预先处理1wk,3次/d,1wk后予碘酸钠舌下静脉注射,在2wk和4wk末,视网膜电图(ERG)测量c波。色素微球体技术分析高眼压状态下TMP对脉络膜血流的影响。Methylthiazoltetrazolium(MTT)分析TMP在各种氧化应激中对RPE的保护作用。结果:碘酸钠注射后2wk,碘酸钠组ERG的c波下降至对照组的36%(P<0.01)。4wk后,碘酸钠组下降至对照组的46%(P<0.01),而1%TMP+碘酸钠组下降至对照组的77%(P<0.01)。与碘酸钠组比较,1%TMP+碘酸钠组控制了67%的c波下降(P<0.05)。在脉络膜血流的测量中,30,60,和120min的结果显示,TMP显著增加脉络膜血流。在氧化应激部分,不同浓度的TMP在各种氧化应激损伤中,对RPE都有各种程度的保护作用。结论:浓度为1%Tetramethylpyrazine可以显著保护碘酸钠和氧化应激诱导的RPE变性,增加脉络膜血流,并可能在AMD的治疗中发挥作用。     

Early attachment of uncultured retinal pigment epithelium from aged donors onto Bruch's membrane explants

Retinal pigment epithelium (RPE) transplantation might replace cells lost as a consequence of choroidal neovascular membrane excision in patients with age-related macular degeneration (AMD). Autologous transplantation of RPE cells harvested from a peripheral biopsy may overcome problems of immune rejection. To study the feasibility of autologous RPE cell transplantation, the authors examined the attachment of freshly harvested RPE cells from aged donors onto Bruch's membrane explants, debrided to (1) remove or (2) preserve the RPE basement membrane. Human retinal pigment epithelial sheets were harvested from adult donor eyes (N = 12, mean age 79.00 +/- 9.40 years) and, following incubation in collagenase, were mechanically fragmented into microaggregates. Microaggregates (approximately 120 000 cells) were seeded onto the paired explants (7 mm diameter) and incubated for 20 min, 1, 4, or 24 hr at 37 degrees C. The percent coverage of the debrided surface by microaggregates was determined by sampling the center of the explants with scanning electron microscopy. RPE microaggregate attachment to Bruch's membrane was significantly greater at all time points analysed in samples with intact basement membrane versus those with an exposed inner collagenous layer. Coverage of debridements retaining intact RPE basement membrane was 1.83 +/- 1.10% at 20 min, 3.54 +/- 2.14% at 1 hr, and 8.68 +/- 2.63% at 4 hr. Coverage of debridements lacking basement membrane was 0.10 +/- 0.04% at 20 min, 0.39 +/- 0.25% at 1 hr, and 0.63 +/- 0.42% at 4 hr. Based on their morphologic appearance, many cells were dying as early as 1 hr following seeding. To increase surface coverage, the authors seeded four times the above number of cells and incubated the specimens for 1 hr. Coverage on explants lacking RPE basement membrane showed no increase in the number of cells attached to the inner collagenous layer. There was a significant approximately three-fold increase in the number of cells attached in the presence of basement membrane. These results indicate that if RPE cells from aged human donors are used for transplantation, some modification of the Bruch's membrane surface or the cells must be considered for cell attachment and eventual cell survival.     

Growth-related effects of oxidant-induced stress on cultured RPE and choroidal endothelial cells

Mounting evidence suggests that oxidative stress caused by reactive oxygen intermediates is a significant mechanism in the pathogenesis of age-related macular degeneration (AMD). Although vascular endothelial growth factor (VEGF) and other cytokines are involved in choroidal neovascularization (CNV) it is largely unknown whether oxidative stress may predispose the eye to increased levels of proangiogenic factors. In an in vitro study we have determined viability and proliferation of both human retinal pigment epithelial (RPE) cells and bovine choroidal endothelial cells (CECs) and assessed the release of basic fibroblast growth factor (bFGF) and VEGF from RPE cells after exposing them to oxidative stress. Permanent presence of tert-butyl-hydroperoxide (tBH), a pro-oxidative stressor, in the cell cultures resulted in decreasing viability and proliferation of RPE cells and CECs. Loss of RPE cell viability was associated with activation of apoptosis by tBH in a dose-dependent manner. The antioxidant, N-acetyl-l-cysteine (NAC), and secreted soluble mediators of RPE cells were appropriate to attenuate the effects of tBH-mediated oxidative stress. RPE cells exposed to tBH were found to release increasing amounts of bFGF but not VEGF after 24 h of culture, thereby supporting proliferation of CECs. These findings suggest that oxidative stress compromises the viability of RPE cells and CECs. However, increased bFGF levels concomitantly released from RPE cells may attenuate the CEC-directed effect, protect CECs from oxidative insults, and are likely to promote CNV.     

干细胞来源的RPE细胞移植治疗视网膜退行性疾病的研究进展

在视网膜退行性疾病中,年龄相关性黄斑变性(age-related macular degeneration,ARMD)和Stargardt黄斑营养不良(SMD)分别是威胁老年人和青少年人群视力最常见的疾病.目前仍缺乏可改善干性ARMD和Stargardt黄斑营养不良患者视力的确切疗法.近年来,干细胞(stem cells)技术不断发展,干细胞来源的RPE细胞移植疗法成为视网膜退行性疾病治疗的热点及新方向,本文将对该研究领域的最新研究进展作一综述.