水通道蛋白1在老年性白内障及透明晶状体前囊膜上皮细胞的表达

目的明确水通道蛋白1(aquaporin1,AQP1)在透明晶状体及老年性白内障晶状体上皮细胞及前囊膜的表达及分布,并初步探讨AQP1在老年性白内障发病中的重要作用。方法收集52例老年性白内障及10例正常透明晶状体前囊膜,通过HE染色和免疫组化方法检测AQP1的表达。结果老年性白内障及正常透明晶状体上皮细胞中均可见AQP1的表达,透明晶状体组中AQP1的阳性细胞表达率(100%)显著高于老年性白内障组(53.8%)(P<0.05)。而在晶状体纤维中未见表达。结论晶状体上皮细胞是晶状体代谢最活跃的部位,终身进行有丝分裂,并移行至赤道部分化成晶状体纤维。晶状体上皮细胞有AQP1的活性表达,这对于维持晶状体上皮细胞一定的增殖潜能,保证晶状体的生理动态平衡和晶状体的透明性起到了重要的作用。提示AQP1在老年性白内障晶状体前囊膜及上皮细胞中的异常表达与老年性白内障的发生密切相关,具体调控机制有待进一步研究。     

色素上皮衍生因子在人晶状体上皮细胞表达的意义

目的:观察色素上皮衍生因子(PEDF)在人晶状体内的分布,探讨人晶状体上皮细胞(LEC)中PEDF表达水平与年龄及白内障发生发展的关系。方法:收集老年性和先天性白内障患者术中所取前囊膜、新鲜眼库眼透明晶状体样品,冰冻切片标本用间接免疫荧光组织化学法检测PEDF蛋白在人晶状体中的分布,前囊膜标本分别用蛋白质免疫印迹(Western-blot)及逆转录聚合酶链反应(RT-PCR)技术检测LEC中PEDF蛋白和基因的表达水平。结果:人晶状体中存在PEDF蛋白,主要分布于前囊下LEC胞浆中。按透明晶状体、轻度白内障、重度白内障及年龄分组分析,人LEC内PEDF基因及蛋白的表达水平在晶状体由透明变混浊、混浊由轻变重的过程中呈下调趋势(P<0.01),且随着机体的衰老显著下降(P<0.01)。结论:人LEC中PEDF表达随晶状体衰老和白内障发生发展显著下调。     

热休克蛋白27在老年性白内障晶状体上皮细胞中的表达及意义

目的　探讨热休克蛋白(HSP) 27在老年性白内障晶状体上皮细胞中的表达及意义。 方法　采用Westernblot蛋白印迹法,检测老年性白内障 50例(其中皮质型 17例、核型 18例、后囊下型 15例 )、正常人晶状体 4例上皮细胞中HSP27的表达。 结果　HSP27在老年性白内障(0 411±0 188)和正常人 (0 147±0 006)晶状体上皮细胞表达有显著性差异,老年性白内障皮质型、核型、后囊下型HSP27表达也不相同。 结论　HSP27可能在老年性白内障的发生、发展中起重要作用,HSP27在皮质型、核型、后囊下型白内障发生的作用机制可能不完全相同。     

波形纤维蛋白在老年性白内障晶状体上皮细胞的表达

目的观察波形纤维蛋白在老年性白内障晶状体上皮细胞的变化.方法用卵白素-生物素过氧化物酶法对22例老年性白内障患者晶状体前囊膜上皮细胞进行波形纤维蛋白染色,采用包埋前免疫酶电镜技术处理6个晶状体前囊膜标本,并观察其超微结构;利用十二烷基硫酸钠聚丙烯酰胺凝胶电泳及Westemnblot法分析4个晶状体表层组织(前囊膜、上皮细胞和表层皮质)中的波形纤维蛋白.结果老年性白内障晶状体上皮细胞的波形纤维蛋白表达减弱,与对照组比较差异有显著性(t=2.0948,P<     

老年性白内障合并2型糖尿病患者晶状体上皮细胞转化生长因子β表达及其与晚期糖基化终末产物的相关性

目的研究转化生长因子p（transforminggrowthfactor—β,TGF—β）在老年性白内障合并2型糖尿病患者中的表达变化及其可能的作用机制,为进一步阐明老年性白内障合并2型糖尿病患者发病的分子机制提供实验数据。方法分别采用点杂交和实时荧光聚合酶链反应方法定量检测和分析老年性白内障患者及老年性白内障合并2型糖尿病患者晶状体前囊膜上皮细胞中TGF-β蛋白、mRNA表达水平。免疫荧光方法观察晚期糖基化终末产物（advanced glycation endoproducts,AGEs）对体外培养人晶状体上皮细胞TGF-β表达的影响。结果与老年性白内障患者相比,老年性白内障合并2型糖尿病患者晶状体前囊膜上皮细胞中TGF-β（TGF-β／2／3）蛋白表达增加（P〈0．05）,mRNA表达变化分别为TGF—β1升高（P〈0．05）,TGF—β2降低（P〈0．01）,TGF-β3未见显著改变（P〉0．05）。AGEs与晶状体上皮细胞共培养后,细胞表面TGF—β（TGF-β1／2／3）表达上调。结论TGF—β在老年性白内障合并2型糖尿病患者晶状体上皮细胞中的表达上调。AEGs可诱导晶状体上皮细胞TGF—β表达上调。（中国眼耳鼻喉科杂志,2010,10：212514）     

老年性和糖尿病性白内障晶状体上皮细胞的超微结构改变

目的 观察正常人，老年性白内障、糖尿病性白内障晶状体上皮细胞的超微结构，探讨细胞凋亡在白内障发生中的作用。方法 收集正常人、老年性白内障、糖尿病性白内障晶状体前囊膜送透射电镜扫描，取老年性白内障和糖尿病性白内障前囊膜各15例，用脱氧核苷酸末端转移酶缺口标记原位细胞检测法，检测晶状体上皮细胞的凋亡细胞。结果 老年性和糖尿病性白内障晶状体上细胞的超微结构在形态上为扁平状，胞浆出现空泡变性，胞核出现固缩，染色质边聚，浓缩等改变。糖尿病性白内障晶状体上皮细胞的胞浆空泡变性更加严重。在光镜下老年性和糖尿病性白内障的晶状体上皮均有凋亡细胞。结论 老年性白内障和糖尿病性白内障的发生与晶状体上皮的细胞凋亡密切相关。     

老年性和糖尿病性白内障晶状体上皮细胞的超微结构改变

目的 观察正常人,老年性白内障、糖尿病性白内障晶状体上皮细胞的超微结构,探讨细胞凋亡在白内障发生中的作用。方法 收集正常人、老年性白内障、糖尿病性白内障晶状体前囊膜送透射电镜扫描,取老年性白内障和糖尿病性白内障前囊膜各15例,用脱氧核苷酸末端转移酶缺口标记原位细胞检测法,检测晶状体上皮的凋亡细胞。结果 老年性和糖尿病性白内障晶状体上皮细胞的超微结构在形态上为扁平状,胞浆出现空泡变性,胞核出现固缩,染色质边聚,浓缩等改变。糖尿病性白内障晶状体上皮细胞的胞浆空泡变性更加严重。在光镜下老年性和糖尿病性白内障的晶状体上皮均有凋亡细胞。结论 老年性白内障和糖尿病性白内障的发生与晶状体上皮的细胞凋亡密切相关。     

骨桥蛋白在白内障晶状体上皮细胞中的表达

目的：探讨骨桥蛋白（osteopontin，OPN）在白内障晶状体上皮细胞中的表达水平及作用。方法：采用免疫组织化学法检测22例前囊下性白内障、17例核性白内障、14例皮质性白内障患者和11例正常晶状体上皮细胞中OPN的表达，并进行阳性细胞计数和各组间比较。结果：OPN在前囊下白内障患者晶状体上皮细胞中的表达明显增多，其阳性表达率与核性白内障、皮质性白内障及正常人晶状体比较，差异有显著意义（P〈0．01）。结论：OPN在前囊下性白内障晶状体上皮细胞中表达阳性，在前囊下白内障的形成过程中可能起重要作用。     

老年性白内障晶状体上皮细胞超微结构的观察

目的 :观察老年性白内障晶状体上皮细胞的超微改变 ,探讨细胞凋亡在老年性白内障形成中的作用。方法 :老年性白内障患者 2 4例 ,行囊外或超声乳化联合后房型人工晶状体植入术 ,术中取中央部晶状体前囊膜 ,分别行光镜 ,扫描电镜 ,透射电镜观察。结果 :老年性白内障晶状体上皮细胞的超微改变有细胞形态大小不一 ,扁平细胞多无正常细胞结构 ,低柱状及柱状细胞 ,细胞结构大体正常 ,但细胞间隙增大 ,胞浆内可见空泡变性 ,部分细胞溶解坏死 ,部分细胞皱缩 ,呈早期细胞调亡改变。结论 :晶状体上皮细胞坏死与凋亡均与老年性白内障的发生有关。     

晶状体上皮细胞转分化对老年性白内障形成的影响研究

目的 :为研究晶状体上皮细胞转分化对老年性白内障形成的影响。方法 :对 14例老年性白内障晶状体前囊上皮细胞进行角质蛋白 -8、波形纤维蛋白和纤维粘连蛋白表达的免疫组化研究。结果 :皮质性白内障晶状体上皮的波形纤维蛋白表达明显强于核性白内障 (P <0 0 5 ) ;角质蛋白 -8在皮质性白内障晶状体上皮的表达比核性白内障弱 ((P <0 0 1) ;纤维粘连蛋白在皮质性白内障晶状体上皮的表达明显强于核性白内障 ,而在正常晶状体上皮不表达。结论 :晶状体上皮细胞具有转分化为成纤维样细胞的双向化潜能。晶状体上皮获得转分化能力后而失去原来的细胞学特性。在老年皮质性白内障中 ,晶状体上皮细胞转分化成为成纤维样细胞 ,伴随波形纤维蛋白的过度表达和角质蛋白表达下降。同时合成包括纤维粘连蛋白在内的细胞外基质增多 ,而纤维粘连蛋白能促进晶状体上皮的增殖、迁移及粘附 ,因而晶状体上皮细胞转分化对老年性白内障形成及后囊混浊的形成发挥重要作用。     

2型糖尿病合并白内障患者晶状体上皮细胞中PEDF和VEGF的表达及意义

目的：观察糖尿病合并年龄相关性白内障患者晶状体上皮细胞(LECs)中色素上皮衍生因子(PEDF)、血管内皮细胞生长因子(VEGF)的表达水平,探讨糖尿病合并年龄相关性白内障的发病机制。

方法：回顾性研究。收集2020-08/2021-04在蚌埠医学院第一附属医院眼科就诊的年龄相关性白内障和2型糖尿病合并年龄相关性白内障患者各30例。所有患者白内障超声乳化术中收取术眼晶状体中央区5.5～6.0mm直径的前囊膜标本,采用蛋白免疫印迹法(Western-blot)检测LECs中PEDF、VEGF蛋白表达水平。实时荧光定量PCR(qRT-PCR)方法检测PEDF、VEGF mRNA的相对表达量。

结果：两组患者LECs中均存在PEDF、VEGF的表达,2型糖尿病合并年龄相关性白内障组患者VEGF mRNA相对表达量为1.364±0.062,高于年龄相关性白内障组的1.000±0.0(P<0.01),PEDF mRNA的相对表达量为0.398±0.053,明显低于年龄相关性白内障组的1.000±0.0(P<0.001)。2型糖尿病合并年龄相关性白内障组患者LECs中VEGF和PEDF蛋白表达量为2.053±0.026、0.579±0.045,年龄相关性白内障组为1.680±0.064、1.058±0.007(均P<0.01)。

结论：2型糖尿病合并年龄相关性白内障患者LECs中PEDF和VEGF表达水平发生变化,可能与糖尿病患者白内障的发生和发展有关。     

Pigment Epithelium-derived Factor in Cataractous Aqueous Humor and Lens Epithelial Cells

Purpose: To study the characteristics of PEDF in cataractous aqueous humor and its expression in human lens epithelium. Methods: The PEDF concentration in the aqueous humor was measured by enzyme -linked immunosorbent assay in senile (130cases) and congenital (18cases) cataract patients who underwent cataract phacoemulsification extraction surgery. Anterior lens capsular specimens were obtained from these patients to count lens epithelial cells (LEC) density. The Lens Opacities Classification System Ⅲ was used to classify the senile cataracts as cortical, nuclear, posterior subcapsular and mixed types of opacity, and quantitative analysis of the nuclear opacities was performed by Pentacam Scheimpflug imaging system. Anterior lens capsular specimens from another senile (10cases) and congenital (10cases) cataract were collected for immunofluorescence with polyclonal antibodies specific to human pigment epithelium -derived factor (PEDF). Results:The mean aqueous level of PEDF was(178. 9±87. 5)ng/ml, and there was negative linear correlation of PEDF level and age (r=0. 811, P<0. 001). In senile cases, the aqueous PEDF concentration decreased with increasing nuclear opacities (r=0. 447, P < 0.01) , and the mean PEDF level in nuclear cataract was significantly lower than that in posterior subcapsular opacity (P < 0.01) . PEDF immunostaining was detected in LEC of all capsular specimens. Conclusion : The PEDF level in human aqueous humor is related to age, types of cataracts and lens opacity. PEDF also express in human LEC. The study results suggest PEDF may regulate and/or protect LEC by paracrine and autocrine, and lack of PEDF may play a role in cataractogenesis.     

Downregulation of gene expression in the ageing lens: a possible contributory factor in senile cataract

PURPOSE: To study the molecular characteristics of lens epithelial cells from patients with senile cataract by cDNA microarray technique. METHODS: Lens epithelial cells adhering to anterior capsules taken during cataract surgery collected from 108 patients, aged 56-92 years (senile cataract group), were pooled. Pooled epithelial cells of normal, noncataractous lenses from one patient with ocular trauma, one patient with lens subluxation, and 25 cadaveric eyes, all under the age of 55 years, served as a control. Total RNA was extracted by conventional methods from the two groups of cells, and a fluorescent probe was prepared for each group. The probes were hybridized on 9700 known human cDNA clones. Hybridized clones were analysed using a scanning laser and the results were processed by GEMTools (Incyte Genomics) software. RESULTS: A total of 1827 clones hybridized with the two probes. Of these, 400 showed differences of more than two-fold in gene expression between the two probes. Relative to controls, gene expression in the senile cataract lenses was upregulated in 318 clones and downregulated in 82. Three genes-filensin, inwardly rectifying potassium channel (IRPC), and pigment epithelium-derived factor (PEDF) were strongly downregulated (by 41.3-, 6.8-, and 5.9-fold, respectively) in senile cataract. CONCLUSIONS: Cataractogenesis is associated with numerous changes in the genetic profile of the lens epithelial cells. Since filensin, IRPC, and PEDF genes are known to have important roles in the physiology and morphology of the transparent lens, substantial downregulation of their expression might contribute to the formation of senile cataract.     

老年性白内障晶状体上皮细胞凋亡及相关基因蛋白的表达

目的 探讨老年性白内障与晶状体上皮细胞凋亡的关系。方法透射电镜下观察老年性白内障晶状体上皮细胞的超微结构；Tunel法检测凋亡细胞百分率；并对其晶状体上皮细胞DNA进行琼脂糖凝胶电泳；免疫组化法检测P53、bcl-2在老年性白内障晶状体上皮细胞中的蛋白表达。结果 透射电镜下发现老年性白内障晶状体上皮细胞中有凋亡细胞；凋亡细胞百分率为8．4％～37．8％，琼脂糖凝胶电泳出现梯状条带；P53在老年白内障晶状体上皮细胞中蛋白表达率为16．9％～19．1％，bcl-2无蛋白表达。结论 老年性白内障的发生可能与其晶状体上皮细胞凋亡有关。     

人原代晶状体上皮细胞中PEDF基因下调引起波形蛋白和αB-晶状体蛋白含量的变化

目的　研究人原代晶状体上皮细胞中,下调色素上皮衍生因子（ｐｉｇｍｅｎｔｅｐｉｔｈｅｌｉｕｍ－ｄｅｒｉｖｅｄｆａｃｔｏｒ,ＰＥＤＦ）的表达对波形蛋白和αＢ－晶状体蛋白表达的影响。方法　取２０～３５岁青年人晶状体上皮细胞进行原代培养。构建三种短发卡ＲＮＡ（ｓｈｏｒｔｈａｉｒｐｉｎＲＮＡ,ｓｈＲＮＡ）以特异性下调原代人晶状体上皮细胞中的ＰＥＤＦ基因ＲＮＡ表达水平。感染细胞后,用Ｒｅａｌ－ｔｉｍｅＰＣＲ检测ＰＥＤＦ基因ｍＲＮＡ的表达水平,确定ＲＮＡ干扰效率。人原代晶状体上皮细胞ＰＥＤＦ基因ＲＮＡ干扰４８ｈ后,用ＷｅｓｔｅｒｎＢｌｏｔ检测其中波形蛋白以及αＢ－晶状体蛋白表达水平的变化,以ｓｈＲＮＡ－ｓｃｒａｍｂｌｅ组为阴性对照组。结果　人原代晶状体上皮细胞被成功分离培养,带有绿色荧光蛋白的表达ＰＥＤＦｓｈＲＮＡ的病毒载体对人原代晶状体上皮细胞的感染效率都在８０％左右。Ｒｅａｌ－ｔｉｍｅＰＣＲ结果显示ＰＥＤＦ基因ｍＲＮＡ的表达明显被三种ｓｈＲＮＡ下调。ＷｅｓｔｅｒｎＢｌｏｔ结果显示ＰＥＤＦ基因的表达下调引起了波形蛋白的表达下降（在三个实验组中分别下降了７２％、７９％和９３％）以及αＢ－晶状体蛋白的表达增加（在三个实验组中分别增加了２１９％、２０４％、９３％）。结论　人原代晶状体上皮细胞中ＰＥＤＦ基因下调可引起维持晶状体透明性的两种重要蛋白的改变,提示ＰＥＤＦ与白内障形成有一定的联系。     

Expression of pigment epithelium-derived factor in normal adult rat eye and experimental choroidal neovascularization

PURPOSE: Pigment epithelium-derived factor (PEDF) is a protein produced by the retinal pigment epithelial (RPE) cells. Recent studies have implicated PEDF in activities that are inhibitory to angiogenesis. In this study, the expression of PEDF was investigated in normal rat eyes and in eyes with experimentally induced choroidal neovascularization and compared with the expression of vascular endothelial growth factor (VEGF). METHODS: Choroidal neovascularization was induced by laser photocoagulation in rat eyes. At intervals of up to 2 weeks after photocoagulation, the eyes were removed and prepared for in situ hybridization and immunohistochemical study. In situ hybridization was performed with digoxigenin-labeled PEDF riboprobes. Protein expression of PEDF and VEGF was studied immunohistochemically. RESULTS: In normal adult rat eyes, PEDF mRNA was observed mainly in the corneal epithelial and endothelial cells, lens epithelial cells, ciliary epithelial cells, retinal ganglion cells, and the RPE cells. During the development of choroidal neovascularization, PEDF mRNA, PEDF protein, and VEGF protein were strongly detected in many cells within the laser lesions at 3 days after photocoagulation, after which levels gradually declined. However, PEDF was still expressed in the RPE cells that proliferated and covered the neovascular tissues at 2 weeks, whereas VEGF protein was weakly expressed in endothelial cells in choroidal neovascularization. CONCLUSIONS: PEDF is expressed in different cell types of normal rat eyes. The expression of PEDF was detected in the choroidal neovascular tissues induced by photocoagulation, and these findings suggest that PEDF may modulate the process of choroidal neovascularization.     

正常幼儿及不同年龄段白内障患者晶状体中常见miRNA的表达

目的：检测并分析miRNA在正常幼儿及不同年龄段白内障患者晶状体中的表达差异,初步探讨其在维持晶状体正常功能和不同年龄段白内障形成中的可能作用。

方法：使用stem-loop RT-PCR法检测正常幼儿和幼儿(先天性白内障)、中青年及老年(年龄相关性白内障)白内障患者晶状体中miRNA的表达情况,比较各组间miRNA表达差异。

结果：正常幼儿晶状体中miR-184表达量最高。与正常幼儿相比,白内障幼儿晶状体中miR-184、miR-182表达升高,miR-124、miR-204表达降低。与幼儿患者相比,中青年患者晶状体中miR-204、miR-124和let-7d表达升高,miR-184、miR-183和let-7a表达降低,而老年患者晶状体中所有被检测miRNA表达均有改变,其中miR-182、miR-204、miR-124表达均升高,miR-184、miR-181b、miR-183、miR-125b、let-7a/b/d表达均降低。

结论：正常幼儿及不同年龄段白内障患者晶状体中miRNA表达差异显著,部分miRNA与晶状体的正常形态、功能及某些病理状态相关,这为进一步研究miRNA在维持幼儿晶状体正常功能及不同年龄段白内障形成过程中的可能作用机制提供了理论依据。