老年白内障晶状体上皮细胞培养与表皮生长因子受体表达

目的观察老年白内障(≥60岁)晶状体上皮细胞培养与表皮生长因子受体(EGFR)表达。方法用消化分离法及组织块贴片法培养老年白内障、先天性白内障、胎儿晶状体及正常成人晶状体上皮细胞;应用免疫组织化学方法检测老年白内障晶状体上皮细胞、正常成人的晶状体上皮细胞及培养的晶状体上皮细胞EGFR蛋白表达,RT-PCR方法检测EGFRmRNA表达。结果老年白内障晶状体上皮细胞传代培养基本不能增殖,先天性白内障、胎儿可传6~7代,成人晶状体上皮细胞可传4~5代;老年白内障、培养的晶状体上皮细胞及正常成人晶状体上皮细胞EGFRmR-NA及蛋白均呈阳性表达。结论晶状体上皮细胞的培养对年龄有较高的敏感性,年龄愈大,细胞越易老化,老年白内障晶状体上皮细胞传代培养基本不能增殖;人眼的晶状体上皮细胞EGFR呈阳性表达。     

年龄相关性白内障晶状体上皮细胞中miR-181a和SIRT1的表达关系及意义

目的探讨白内障晶状体上皮细胞微小RNA-181a(miR-181a)与沉默信息调节因子(SIRT)1的表达关系,研究其在年龄相关性白内障患者的诊治及病情评估方面的临床意义。方法选取诊断为年龄相关性白内障患者62例为白内障组,另选取同期近视矫正手术患者42例为对照组,利用RT q-PCR法,检测两组晶状体上皮细胞miR-181a与SIRT1表达水平;Pearson法测定miR-181a与SIRT1相关性;利用Lipofectamine 2000转染miR-181a模拟物和抑制剂,调节人晶状体上皮细胞miR-181a表达水平,RT q-PCR法检测SIRT1表达水平;酶联免疫吸附试验(ELISA)检测晶状体上皮细胞中超氧化物歧化酶(SOD)、丙二醛(MDA)、谷胱氨酸(GSH)、谷胱氨酸过氧化物酶(GSH-Px)水平。结果与对照组相比,白内障组miR-181a表达水平显著升高,SIRT1表达水平显著降低(P<0.05);Pearson相关分析显示,miR-181a和SIRT1呈负相关关系(r=-0.719,P<0.05);miR-181a模拟物组SIRT1表达水平显著低于模拟物阳性对照组(P<0.05),miR-181a抑制剂组,SIRT1表达水平显著高于抑制剂阴性对照组(P<0.05);白内障组SOD、GSH、GSH-Px水平明显低于正常组(P<0.05),MDA水平明显高于正常组(P<0.05);miR-181a表达水平与SOD、GSH、GSH-Px因子呈负相关关系(r=-0.702,-0.739,-0.776),与MDA水平呈正相关关系(r=0.679);SIRT1表达水平与SOD、GSH、GSH-Px呈正相关关系(r=0.699,0.743,0.763),与MDA水平呈负相关关系(r=-0.684)。结论白内障晶状体上皮细胞中miR-181a高表达,SIRT1低表达,miR-181a可调控人晶状体上皮细胞中SIRT1表达;SIRT1低表达可促使晶状体上皮细胞凋亡,从而影响或导致了年龄相关性白内障的发病。     

老年白内障晶状体上皮细胞凋亡及凋亡基因的表达

目的探讨老年白内障与晶状体上皮细胞凋亡的关系。方法TUNEL法检测凋亡细胞百分率；免疫组化法检测P53、bax在老年白内障晶状体上皮细胞中的蛋白表达；RT-PCR检测p53、bcl-2在老年白内障晶状体上皮细胞中的mRNA的表达水平。结果凋亡细胞百分率为5%～47.4%，P53蛋白在老年白内障晶状体上皮细胞中的表达率为15%～28%，bax蛋白的表达率为5%～16%，p53mRNA的表达率为0.48～0.78，baxmRNA的表达率为0.34～0.67。结论老年白内障的发生与晶状体上皮细胞凋亡存在一定的相关性。     

老年性白内障晶状体上皮细胞凋亡及caspase-3表达

目的 探讨晶状体上皮细胞(LECs)凋亡及凋亡相关基因caspase-3表达与老年性白内障发生的关系.方法 选择老年性白内障患者25例为研究组,收集患者行白内障手术时环形撕囊后获取的晶状体前囊膜标本.同时收集成人尸体解剖非白内障眼球所获取的晶状体前囊膜标本10例为对照组.采用免疫组化法检测两组的晶状体前囊膜组织中凋亡相关基因caspase-3的表达,并应用TUNEL方法检测两组LECs的凋亡情况.结果 研究组LECs凋亡细胞百分率为(30.4±3.7)％,对照组LECs凋亡细胞百分率为(0.27±0.12)％.研究组显著高于对照组(t=11.59,P＜0.01).研究组LECs caspase-3蛋白的阳性表达率为84.0％(21/25),而对照组为10.0％(1/10),研究组显著高于对照组(t=12.05,P＜0.01).25例老年性白内障患者的LECs中caspase-3蛋白的表达与细胞凋亡数间呈正相关(r=0.621,P＜0.01).结论 LECs凋亡及caspase-3表达在老年性白内障的发生发展过程中起重要作用.     

衰老与年龄相关性白内障的关系

年龄相关性白内障（age-related cataract，ARC）又称为老年性白内障，已成为致盲的主要原因。随着年龄的增长，晶状体的有关化学成分、含量和代谢都发生了改变，使晶状体逐渐老化，透明度降低，最终导致本病的发生。笔者就新近资料对衰老与年龄相关性白内障的关系作一综述。     

Egr-1特异脱氧核酶抑制血管平滑肌细胞增殖的研究

采用MTT比色法观察空白对照组、等浓度的早期生长反应因子-1(Egr-1)特异脱氧核酶(ED5)、杂码寡核苷酸(EDSSCR)转染后对血管平滑肌细胞(VSMC)增殖活性的影响.应用RT-PCR、Western blot方法观察Egr-1、转化生长因子-β1>(TGF-β1>)的表达.结果 显示,与对照组及EDSSCR相比,ED5对VSMC的增殖有明显的抑制作用;ED5转染后Egr-1、TGF-β1> mRNA及蛋白水平均明显下降.认为Egr-1 mRNA的脱氧核酶可抑制培养的VSMCEgr-1和TGF-β1>的表达,有效阻止VSMC增殖.     

Egr-1与肺纤维化

异常的细胞外基质的沉积破坏了正常的肺结构是肺纤维化的特征。Egr-1被发现与肺纤维化有关，据研究其作用大多是通过TGF-β1实现的。Egr-1能诱导TGF-β1的表达，而TGF-β1又能诱导Egr-1表达。Egr-1影响成纤维细胞增殖、向肌成纤维细胞分化，并且与肺泡上皮细胞、内皮细胞凋亡，肺泡、气道炎症以及细胞外基质形成有关。血管紧张素转换酶系统对肺纤维化的作用亦与Egr-1有关。但Egr-1对肺纤维化的具体作用还不完全明了，需要作深入研究。     

年龄相关性白内障患者非球面与球面人工晶状体植入术后对比敏感度的差异

目的 比较年龄相关性白内障患者植入非球面与球面人工晶状体(IOL)术后对比敏感度的差异.方法 对116例年龄相关性白内障患者实施白内障超声乳化吸除术,随机植入疏水性丙烯酸酯非球面(Alcon SN60 WF)或球面(Alcon SN60 AT)IOL,分别于术后1、3、6和12个月进行最佳矫正视力和夜、夜+周边眩光、昼及昼+周边眩光4种条件下功能性视力对比敏感度测试.结果 术后不同时间植入非球面IOL眼与球面IOL眼最佳矫正视力及昼条件下对比敏感度无显著性差异(P＞0.05),而在昼+周边眩光、夜及夜+周边眩光条件下植入非球面IOL眼的对比敏感度明显高于球面IOL眼(P＜0.05).结论 植入非球面IOL眼获得良好的视功能.     

早期生长反应因子1过表达促进肾小管上皮细胞转分化

目的:探讨早期生长反应因子1(Egr-1)在肾小管上皮细胞转分化中的作用. 方法:(1)体外实验:用pcDNA 3.1,pcDNA 3.1/Egr-1转染肾小管上皮细胞株HK-2,qRT-PCR法、Westem免疫印迹法检测Egr-1及转分化标志物E-cadherin、成纤维细胞特异性蛋白(Fsp-1)mRNA及蛋白水平的表达变化;用pcDNA 3.1/Egr-1和siEgr-1或psilencer 3.1共转染HK-2,Western免疫印迹法检测Egr-1、E-cadherin、Fsp-1蛋白水平的表达变化.(2)体内实验:建立大鼠慢性肾纤维化模型--5/6肾脏次全切除,造模成功后免疫组化法检测大鼠肾组织中Egr-1、E-cadherin、Fap-1的表达. 结果:(1)与转染空载体相比,HK-2细胞过表达Egr-1后,上皮细胞标志性蛋白E-cad-herin表达减弱,成纤维细胞标志性蛋白Fsp-1表达增强(P<0.05),提示出现肾小管上皮细胞转分化表型.而用小干扰RNA干扰掉Egr-1后,与空载体组及亲本细胞组相比,转分化表型得到逆转.(2)体内实验证实,与假手术组(SOR)相比,5/6肾脏次全切除肾组织中,转分化标志蛋白E-cadherin表达减弱、Fsp-1表达增加,同时肾小管上皮细胞Egr-1的表达明显升高,提示肾小管上皮细胞发生转分化,且其与Egr-1过表达密切相关. 结论:Egr-1的过表达促进了肾小管上皮细胞转分化,参与肾间质纤维化的发生发展.     

年龄相关性白内障患者小切口术后前房深度与晶状体厚度、视力水平相关性

目的分析年龄相关性白内障患者小切口术后前房深度与晶状体厚度、视力水平的相关性。方法年龄相关性白内障超声乳化吸除术联合人工晶状体植入术患者根据手术切口大小,将行1.8 mm透明角膜切口的病例设为观察组,行2.2 mm透明角膜切口的病例为对照组。结果观察组视力水平、前房深度和晶状体厚度明显高于对照组(P0.05)。对照组散光、眼压水平高于观察组(P0.05);对照组白细胞介素(IL)-2水平、泪膜破裂时间、基础泪液分泌实验结果显著高于观察组(P0.05);观察组术后前房深度与视力及晶状体厚度呈现显著正相关。结论年龄相关性白内障微小切口患者术后前房深度与视力及晶状体厚度呈现显著正相关,短时间视力可恢复,机体炎症反应较小,手术安全效果较好。     

老年核性和皮质性白内障患者转化生长因子β2的表达及晶状体上皮细胞的变化

目的探讨老年人核性及皮质性白内障的发病机制.方法比较老年核性、皮质性白内障患者和健康人(对照组)晶状体上皮细胞(LECs)中转化生长因子β2(TGF-β2)mRNA、增殖细胞核抗原(PCNA)、Bcl-2/Bax、纤维连接蛋白(FN)及波形蛋白的表达,并对晶状体上皮细胞的密度进行比较. 结果老年核性和皮质性白内障患者LECs中TGF-β2mRNA的表达量与β-actin的比值分别为0.16±0.02和0.27±0.01,与对照组0.15±0.02比较,差异有统计学意义(均为P＜0.05);两组PCNA蛋白的表达与内参的比值分别为16.01、1.98,与对照组2.43比较,差异有统计学意义(均为P＜0.05);两组Bax蛋白的表达与内参的比值分别为15.97、16.93,对照组为16.97;两组Bcl-2蛋白表达与内参的比值分别为6.97、1.44,与对照组的6.12比较,差异有统计学意义(均为P＜0.05);免疫组化结果显示,20张切片中,核性白内障组LECs中FN阳性为1张,波形蛋白阳性为2张;皮质性白内障组分别为19张、17张,两组比较差异有统计学意义(均为P＜0.05);核性和皮质性白内障组LECs密度分别为(5438.40±262.03)个/mm2和(4250.63±275.05)个/mm2,与对照组(5368.63±211.07)个/mm2比较,差异有统计学意义(均为P＜0.05).结论TGF-β2在老年人白内障发生的过程中起重要作用.老年人皮质性白内障的发生与LECs的凋亡、转化有关,而老年核性白内障患者LECs的增殖较活跃.     

Outcome of extracapsular cataract surgery plus posterior chamber intraocular lens insertion for age-related cataract in eastern Nepal

As age-related cataract is the leading cause of blindness in Nepal, much more attention should be given to improving surgical outcomes.The aim of our study was to determine the effectiveness and risks of extracapsular cataract surgery (ECCE) for age-related cataract in a tertiary hospital of eastern Nepal. ECCE with posterior chamber intraocular lens (PC IOL) insertion on 797 eyes (754 patients) revealed improved visual acuity in 97.9%. In 595 (74.6%) the best corrected visual acuity was 6/18 or better, and 167 (20.9%) had 6/24 to 6/60 with negligible operative/postoperative complications.Thus it is concluded that ECCE with PC IOL could be performed safely and effectively even in developing countries where people are unaware that surgery is available and of the good surgical outcome.     

Protective effect of resveratrol on lens epithelial cell apoptosis in diabetic cataract rat

Objective:To study the protective effect of resveratrol on lens epithelial cell apoptosis in diabetic cataract rat.Methods:A total of 84 Wistar rats were divided into 4 groups:12 in Group A(control group).24 in Group B(diabetic cataract group),24 in Group C(therapeutic-dose of resveratrol group) and 24 in Group D(low-dose of resveratrol group).Rats in Group B-D were given with60 mg/kg streptozotocin through intraperitoneal injection.Rats in Group C were given with 100mg/kg resveratrol and rats in Group D were given with 20 mg/kg resveratrol.The caspase-3expression levels and apoptosis ratios of LEC among each group were observed:the degrees of lens opacity in Group B-D after 12 weeks were compared.Results:There were significant differences in caspase-3 expression levels,apoptosis ratios of T.F.C among groups at 4 w,8 w and12 w(P0.05).After 12 weeks,in Group B the degree of lens opacity was as follow:0(0.00%) in grade Ⅰ,3(37.50%) in grade Ⅱ,2(25.00%)in grade Ⅲ,2(25.00%)grade Ⅳ,and 1(12.50%) in gradeⅤ:in Group C:2(25.00%)in grade Ⅰ,4(50.00%) in grade Ⅱ.2(25.00%)in grade Ⅲ,0(0.00%)gradeⅣ,and 0(0.00%) in grade Ⅴ;in Group D:1(12.50%)in grade Ⅰ,4(50.00%) in grade Ⅱ,2(25.00%)in grade Ⅲ,1(12.50%) grade Ⅳ,and 0(0.00%) in grade Ⅴ.The.difference among Group B-D was statistically significant(P0.05).Conclusions:Resveratrol has protective effect on lens epithelial cell apoptosis in diabetic cataract rat,and the effect is relative to its dose.     

Alpha lipoic acid protects lens from H2O2-induced cataract by inhibiting apoptosis of lens epithelial cells and inducing activation of anti-oxidative enzymes

ObjectiveTo determine whether alpha lipoic acid (LA) can effectively protect lenses from hydrogen peroxide (H₂O₂)-induced cataract.MethodsLens from adult Sprague-Dawley rats were cultured in 24-well plates and treated without or with 0.2 mM of H₂O₂, 0.2 mM of H₂O₂ plus 0.5 mM, 1.0 mM, or 2.0 mM of LA for 24 h. Cataract was assessed using cross line grey scale measurement. Superoxide dismutase (SOD), glutathione (GSH-Px), lactate dehydrogenase (LDH), and malondialdehyde (MDA) activity or level in lens homogenates was measured. Apoptosis of lens epithelial cells in each group were detected by Terminal Deoxynucleotidyl Transferase dUTP Nick End Labeling (TUNEL) Assay.ResultsA total of 0.2 mM of H₂O₂ induced obvious cataract formation and apoptosis in lens' epithelial cells, but 0.5-2.0 mM of LA could block the effect of 0.2 mM H₂O₂ in inducing cataract and apoptosis. Furthermore, 0.2 mM of H₂O₂ significantly decreased SOD, GSH-Px, and LDH activity and significant increased MDA level in the lens, but 0.5-2.0 mM of LA blocked the effect of 0.2 mM H₂O₂. One mM of LA was found to be the most effective.ConclusionsLA can protect lens from H₂O₂-induced cataract. LA exerts protective effects through inhibition of lens' epithelial cell apoptosis and activation of anti-oxidative enzymes.     

Decreases in Raf-1 levels in galactosaemic lens epithelial cells are partially reversed by myo-inositol

Changes in the amounts and localization of protein kinase C (PKC) isoforms occur in galactosaemic lens epithelial cells. A link between PKC changes and myo-inositol depletion has been suggested. Raf-1, a component of a Ras pathway, is a substrate for PKC. Raf-1 levels were measured in galactosaemic lens epithelial cells grown with or without myoinositol. Raf-1 levels were measured by densitometric scanning of Western blots from cells grown with or without 40 mmol/l galactose or 40 mmol/l galactose plus 1.0 μmol/l myoinositol for 1, 3, 5 or 7 days. Scans were compared to those for PKCα, an isoform of PKC and to 14-3-3, a protein which binds to Raf-1. Cell growth was quantitated by thymidine incorporation. Raf-1 levels were decreased in bovine lens epithelial cells after 3, 5 or 7 days (33% of control) of growth in 40 mmol/l galactose. Addition of 1 μmol/l myoinositol reversed this decrease at day 3, but not after 5 or 7 days of growth in 40 mmol/l galactose. PKCα and 14-3-3 levels were not affected by galactose. The decrease in Raf-1 was not a result of cell growth as measured by thymidine incorporation. These results suggest that Raf-1 levels are decreased during galactosaemia. This was only partially reversed by the addition of myoinositol. Received: 14 November 1997 / Accepted in revised form: 24 June 1998