Parthenolide protects human lens epithelial cells from oxidative stress-induced apoptosis via inhibition of activation of caspase-3 and caspase-9

The apoptosis of lens epithehal cells has been proposed as the common basis of cataract formation, with oxidative stress as the major cause. This study was performed to investigate the protective effect of the herbal constituent parthenolide against oxidative stress-induced apoptosis of human lens epithelial （HLE） cells and the possible molecular mechanisms involved. HLE cells （SRA01-04） were incubated with 50 μM H2O2 in the absence or presence of different doses of parthenolide （10, 20 and 50 μM）. To study apoptosis, the cells were assessed by morphologic examination and Annexin V-propidium iodide double staining flow cytometry; to investigate the underlying molecular mechanisms, the expression of caspase-3 and caspase-9 were assayed by Western blot and quantitative RT-PCR, and the activities of caspase-3 and caspase-9 were measured by a Chemicon caspase colorimetric activity assay kit. Stimulated with H202 for 18 h, a high fraction of riLE cells underwent apoptosis, while in the presence ofparthenolide of different concentrations, dose-dependent blocking of HLE cell apoptosis was observed. The expression of caspase-3 and caspase-9 induced by H202 in HLE cells was significantly reduced by parthenolide both at the protein and mRNA levels, and the activation ofcaspase-3 and caspase-9 was also suppressed by parthenolide in a dose-dependent manner. In conclusion, parthenolide prevents HLE cells from oxidative stress-induced apoptosis through inhibition of the activation ofcaspase-3 and caspase-9, suggesting a potential protective effect against cataract formation.

EDIL3 depletion suppress epithelial-mesenchymal transition of lens epithelial cells via transforming growth factor β pathway

AIM： To study the effect of discoidin I-like domaincontaining protein 3（EDIL3） depletion on the proliferation and epithelial-mesenchymal transition（EMT） in human lens epithelial cells（LECs）. METHODS： RNA interference was used to inhibit the expression of EDIL3 in human LECs in vitro. The morphology of cells was observed using an inverted microscope. Cell proliferation was assessed using Ed U kit. Cell migration was investigated using Transwell chamber and EMT of LECs was assessed using confocal microscope and Western blotting. The transforming growth factor β（TGFβ） pathway was investigated using Western blotting. RESULTS： The data showed that silencing EDIL3 expression changed LECs morphology and suppressed LECs proliferation（P〈0.05） and migration（P〈0.01）. Furthermore, the result of Western blotting showed that EDIL3 depletion reduced the expression of α-smooth muscle actin（α-SMA）（P〈0.001） and vimentin（P〈0.01）, while increased the expression of E-cadherin（P〈0.001）. EDIL3 depletion could suppress the phosphorylation of Smad2（P〈0.01） and Smad3（P〈0.01） and the activation of exracellular signal regulated kinase（ERK）（P〈0.05）. CONCLUSION： The findings indicate that EDIL3 might participate in the proliferation and EMT in LECs via TGFβ pathway and may be a potential therapeutic target for the treatment of posterior capsule opacification.

Effect of Pyruvate on Polyol Pathway and Lens Epithelial Cells Apoptosis in Diabetic Rats

Purpose: To investigate the effect of polyol pathway on lens epithelial cells apoptosis and the activity of caspase-3 and its reversal by pyruvate in diabetic rats. Methods: 220 Wister rats were divided into 3 groups: control group, model group and treatment group. After streptozotocin (STZ) induced cataract, the treatment group received 2% pyruvate in the diet and drinking. The opacification of lens was detected by microscope every 2 weeks. On 4W, 8W and 12W of the experiment, glucose and sorbitol in the lens were quantified by high-performance liquid chromatography. The percentage of lens epithelial cells undergoing apoptosis was measured by Annexin Ⅴ/PI staining. The activity of caspase-3 was analyzed by Western-blot. Results: Studies show that there was significant increase of glucose, sorbitol in lens of model group, the apoptosis rate and caspase-3 activity of lens epithelial cells were also gradually increase. Pyruvate treatment decreased the levels of sotbitol, glucose, lens epithelial cells apoptosis and caspase-3 activity. The progress of cataract was also significantly delayed. Conclusions: Polyol pathway, possibly through regulation of the activity of caspase-3, can induce apoptosis of lens epithelial cell. Pyruvate ingested orally can effective inhibit diabetic cataractogenesis in rats through inhibit polyol pathway.

Biosafety of a 3D-printed intraocular lens made of a poly (acrylamide-co-sodium acrylate) hydrogel in vitro and in vivo

AIM:To assess the biosafety of a poly(acrylamide-cosodium acrylate)hydrogel(PAH)as a 3D-printed intraocular lens(IOL)material.METHODS:The biosafety of PAH was first evaluated in vitro using human lens epithelial cells(LECs)and the ARPE19 cell line,and a cell counting kit-8(CCK-8)assay was performed to investigate alterations in cell proliferation.A thin film of PAH and a conventional IOL were intraocularly implanted into the eyes of New Zealand white rabbits respectively,and a sham surgery served as control group.The anterior segment photographs,intraocular pressure(IOP),blood parameters and electroretinograms(ERG)were recorded.Inflammatory cytokines in the aqueous humor,such as TNFαand IL-8,were examined by ELISA.Cell apoptosis of the retina was investigated by TUNEL assay,and macro PAHge activation was detected by immunostaining.RESULTS:PAH did not slow cell proliferation when cocultured with human LECs or ARPE19 cells.The implantation of a thin film of a 3 D-printed IOL composed of PAH did not affect the IOP,blood parameters,ERG or optical structure in any of the three experimental groups(n=3 for each).Both TNFαand IL-8 in the aqueous humor of PAH group were transiently elevated 1 wk post-operation and recovered to normal levels at 1 and 3 mo post-operation.Iba1+macroPAHges in the anterior chamber angle in PAH group were increased markedly compared to those of the control group;however,there was no significant difference compared to those in the IOL group.CONCLUSION:PAH is a safe material for 3D printing of personal IOLs that hold great potential for future clinical applications.

Caspase-3 and its inhibitor Ac-DEVD-CHO in rat lens epithelial cell apoptosis induced by hydrogen in vitro

Objective To investigate the role of caspase 3 and its inhibitor Ac DEVD CHO in rat lens epithelial cell apoptosis induced by hydrogen peroxide (H 2O 2) in vitro Methods Rat lenses were incubated in modified Eagle's medium containing 2 mmol/L H 2O 2 to induce apoptosis in vitro Apoptosis in lens epithelial cells was assessed by transmission electron microscopy and annexin V propidium iodide (PI) double staining flow cytometry after 12, 24 and 48 h of incubation The activity of caspase 3 was analyzed by western blotting Results Observations under transmission electron microscopy revealed that 2 mmol/L H 2O 2 could effectively induce lens epithelial cell apoptosis in vitro Caspase 3 activity increased during cell apoptosis and the peak measurement occurred at 24 h after treatment with H 2O 2 Cell apoptosis was blocked by caspase 3 inhibitor Ac DEVD CHO Conclusions The activation of caspase 3 plays an important role in executing apoptosis in H 2O 2 treated lens epithelial cells and in the formation of cataract The caspase 3 inhibitor Ac DEVD CHO may effectively prevent lens epithelial cell apoptosis caused by oxidative injury

人参皂苷Rg_(3)对人晶状体上皮细胞氧化应激损伤的影响

目的探讨人参皂苷Rg_(3)对过氧化氢诱导的人晶状体上皮细胞氧化损伤的改善作用及对核转录因子E2相关因子2(nuclear factor E2 related factor 2,Nrf2)/血红素加氧酶-1(heme oxygenase 1,HO-1)信号通路的调节作用。方法用不同浓度人参皂苷Rg_(3)处理过氧化氢诱导的SRA01/04细胞,用噻唑盐(methyl thiazolyl tetrazolium,MTT)法检测细胞存活率。将第3代对数生长期SRA01/04细胞随机分为正常组、氧化损伤组(用200μmol∙mL^(−1)过氧化氢处理)、人参皂苷Rg_(3)低剂量组和人参皂苷Rg_(3)高剂量组(分别用40、80μg∙mL^(−1)人参皂苷Rg_(3)处理6 h,更换培养基后用200μmol∙mL^(−1)过氧化氢处理12 h),用MTT法检测细胞存活率,用流式细胞仪检测细胞凋亡率,用试剂盒检测丙二醛(malondialdehyde,MDA)、超氧化物歧化酶(superoxidedismutase,SOD)和谷胱甘肽过氧化物酶(glutathioneperoxidase,GSH-Px)的含量,用蛋白印迹法检测Nrf2、Kelch样环氧氯丙烷相关蛋白1(Kelch like epichlorohydrin related protein 1,Keap1)和HO-1蛋白的相对表达量。结果与0μg∙mL^(−1)人参皂苷Rg_(3)组比较,10、20、40、80μg∙mL^(−1)人参皂苷Rg_(3)组的细胞存活率逐渐升高(P<0.05)。与正常组比较,氧化损伤组的细胞存活率、SOD和GSHPx含量以及Nrf2、Keap1和HO-1蛋白相对表达量降低,细胞凋亡率和MDA含量升高(P<0.05);与氧化损伤组比较,人参皂苷Rg_(3)低剂量和人参皂苷Rg_(3)高剂量组的细胞存活率、SOD和GSH-Px含量以及Nrf2、Keap1和HO-1蛋白的相对表达量升高,细胞凋亡率和MDA含量降低(P<0.05);人参皂苷Rg_(3)低剂量和人参皂苷Rg_(3)高剂量组各项指标水平变化规律相同,人参皂苷Rg_(3)高剂量组更显著(P<0.05)。结论人参皂苷Rg_(3)可抑制过氧化氢诱导的人晶状体上皮细胞凋亡,减轻氧化应激损伤,其可能是通过激活Nrf2/HO-1信号通路发挥调节作用的。

年龄相关性白内障晶状体上皮细胞的Smac、caspase-3表达及死亡方式

目的探讨年龄相关性白内障晶状体上皮细胞(LECs)中Smac、caspase-3的表达和LECs超微结构变化及死亡方式。方法收集皮质性(30例)、核性(24例)、后囊下性(28例)年龄相关性白内障患者晶状体前囊片,以12例高度近视摘出的透明晶状体的晶状体前囊片为对照,应用免疫组织化学链酶菌抗生素蛋白-过氧化物酶法检测Smac、caspase-3蛋白在LECs中的表达,并采用透射电镜观察LECs超微结构的改变及死亡方式。结果Smac、caspase-3蛋白在年龄相关性白内障组LECs中的阳性表达率分别为81.71%、78.05%,与透明晶状体组比较染色阳性率及染色强度差异均有统计学意义(P<0.05)。3种类型年龄相关性白内障透射电镜下均可见LECs多表现为肿胀,线粒体嵴缺失、空泡变、髓样化,粗面内质网扩张、颗粒融合和脱颗粒现象,仅1例核性白内障中可见到核膜皱缩向外呈锐角凸起,部分双层核膜融合,模糊不清,细胞核内染色质浓缩,未见凋亡小体。结论Smac、caspase-3蛋白可能参与了年龄相关性白内障的形成。年龄相关性白内障LECs存在凋亡、胀亡等多种细胞死亡方式,细胞凋亡与细胞胀亡的发生可能存在部分分子机制的重叠。LECs超微结构的改变表明线粒体形态改变和功能异常在年龄相关性白内障发生中起重要作用。

前囊下性白内障患者晶状体上皮细胞基质金属蛋白酶-3的表达意义

目的:探讨基质金属蛋白酶-3(mat ri xmet al lopro-teinase-3,MMP-3)在前囊下性白内障患者的晶状体上皮细胞(lensepithelialcells,LECs)中的表达及意义。方法:采用SP免疫组织化学染色法,测定21例前囊下性白内障、18例核性白内障和10例正常晶状体上皮细胞中MMP-3的表达情况,并进行阳性细胞细胞计数。结果:MMP-3在前囊下性白内障晶状体上皮细胞中有表达,而在核性白内障和正常晶状体上皮细胞中均无表达。结论:MMP-3可能是调节细胞外基质(extracellulamatrix,ECM)降解的重要因素而参与前囊膜下性白内障的形成。

氯通道蛋白-3在透明晶状体和老年性白内障患者晶状体上皮细胞中的表达

目的研究透明晶状体和老年性白内障患者晶状体上皮细胞氯通道蛋白-3(chloride channel protein 3,CLC-3)的表达。方法随机收集40例老年性白内障手术患者和5例透明晶状体(来自晶状体脱位患者)的晶状体前囊膜,采用免疫组织化学染色和免疫荧光技术检测晶状体上皮细胞CLC-3的表达。结果CLC-3阳性表达于晶状体上皮细胞的细胞膜。老年白内障组40张切片中,CLC-3表达均为阳性。而透明晶状体组5张切片中,4张阴性,1张可疑阳性。计算机图像分析结果,透明晶状体组平均吸光度(A)值明显低于老年白内障组,差异有统计学意义(P<0.05)。结论老年性白内障患者晶状体上皮细胞CLC-3表达阳性,CLC-3可能参与老年性白内障的形成。

硅油眼晶状体前囊膜及上皮细胞的病理学改变及Caspase-3表达

目的 观察硅油充填眼晶状体前囊膜及上皮细胞的病理学改变及 Caspase- 3表达 ,探讨硅油引起白内障的病因及与凋亡的关系。方法 收集 2 7只晶状体前囊膜 ,通过HE染色和免疫组化方法对硅油眼的晶状体前囊膜和上皮细胞及 Caspase- 3在其上的表达进行观察。结果 前囊膜下许多空间被硅油滴占据 ;Caspase- 3在晶状体上皮细胞有很好的表达 ,2 7例中有 2 3例呈阳性表达。结论 硅油滴与白内障的发生、发展有关 ;

MMP-3和TIMP-1在白内障晶状体上皮细胞中的表达及意义

目的探讨基质金属蛋白酶-3(matrixmetalloproteinase-3,MMP-3)和金属蛋白酶组织抑制因子-1(tissueinhibitorofmetalloproteinase-1,TIMP-1)与白内障发病机制的关系。方法采用免疫组织化学方法测定MMP-3和TIMP-1在前囊下性白内障31眼、核性白内障28眼和正常晶状体12眼上皮细胞(lensepithelialcell,LEC)中的表达。结果MMP-3在前囊下性白内障的表达分别与核性白内障、正常晶状体比较,差异有显著意义(χ2=31.490,34.479;P=0.000),而核性白内障与正常晶状体相比较,差异无显著意义(χ2=2.449,P=0.118);TIMP-1表达在3组LEC中差异均无显著意义(P>0.05);MMP-3与TIMP-1在前囊下性白内障LEC中的表达没有相关性(r=0.121,P=0.516),而在核性白内障中有相关性(r=-0.513,P=0.005)。结论MMP-3表达增强及MMP-3/TIMP-1的表达失衡可能与前囊下性白内障的形成密切有关。

Caspase-3及Bax在人晶状体上皮细胞中的表达

目的 :观察Caspase 3及Bax在人晶状体上皮细胞中的表达 .方法 :收集 30只白内障晶状体前囊膜以及胚胎晶状体 (6只 )和成人透明晶状体 (4只 )的前囊膜 ,采用免疫组织化学的方法观察Caspase 3及Bax的表达情况及染色强度 .结果 :Caspase 3及Bax在各型白内障中均有很好的表达 ,在年龄相关性白内障中Caspase 3及Bax的表达为 11/ 12 ,10 / 12 ,在高度近视性白内障中Caspase 3及Bax的表达为 7/ 8,6 / 8,在外伤性白内障中Caspase 3及Bax的表达为 5 / 6 ,5 / 6 ,在并发性白内障中Caspase 3及Bax的表达为 3/ 4 ,4 / 4 ;在各型白内障中Caspase 3及Bax的表达没有显著性差异 (P >0 .0 5 ) .结论

紫外线B照射对人晶状体上皮细胞质膜钙ATP酶3表达的影响

背景 细胞质膜钙ATP酶3（PMCA3）参与维持晶状体上皮细胞（LECs） Ca2＋的平衡,可能与白内障的病理过程有关,紫外线B（UVB）是引起白内障的重要因素之一,但UVB对LECs中PMCA3表达的影响少有报道. 目的 研究UVB对人LECs系B-3（HLE B-3） PMCA3表达的影响. 方法 对HLE B-3细胞进行培养和传代,当细胞达80％以上融合时分别暴露于用不同剂量（0、5、10、20 mJ·s/cm2）的UVB分别照射0、20、40和80 s,然后继续培养24、48和72 h,用MTT法检测不同剂量UVB照射后细胞的生存率;用JC-1染色法检测UVB照射后细胞线粒体膜电位（△ψm）的变化;以DCFH-DA染色法检测UVB照射后细胞内活性氧簇（ROS）的变化;采用annexin V-FITC/PI染色法检测UVB照射后细胞的凋亡情况;用Fluo-3/AM染色法检测细胞内Ca2＋浓度的变化;分别采用实时荧光定量PCR（real-time PCR）法和Western blot法检测UVB照射后细胞中PMCA3 mRNA及PMCA蛋白的表达变化. 结果 随着UVB照射剂量的增加和照射时间的延长,细胞生存率均明显下降,差异均有统计学意义（F分组=72.411,P=0.000; F时间=36.588,P=0.000）,其中10 mJ·s/cm2、20 mJ·s/cm2 UVB照射后24 h HLE B-3细胞的生存率分别为（75.3±2.2）％和（48.7±4.5）％,明显低于对照组的（100.0±0.0）％,差异均有统计学意义（P=0.001、0.000）;5、10、20 mJ·s/cm2 UVB照射后48 h细胞的生存率分别为（84.9±1.2）％、（69.3±17.4）％和（32.8±4.5）％,均明显低于对照组的（100.0±0.0）％,差异均有统计学意义（P=0.047、0.000、0.000）;5、10、20 mJ·s/cm2 UVB照射后72 h细胞的生存率分别为（55.1±3.0）％、（42.1±1.9）％和（26.1±4.7）％,与对照组的（100.0±0.0）％相比生存率明显降低,差异均有统计学意义（均P=0.000）.JC-1染色法检测表明,对照组细胞内可见红色荧光,5mJ·s/cm2 UVB照射后细胞内出现绿色荧光,10 mJ·s/cm2及20 mJ·s/cm2 UVB照射组出现绿色荧光增强,红色荧光减弱.5、10、20 mJ· s/cm2 UVB照射后细胞内的ROS由0.4％分别增加至35.8％、51.9％和76.7％.0 mJ·s/cm2 UVB照射组凋亡和坏死细胞比例为2.0％,5、10、20 mJ·s/cm2 UVB照射组凋亡和坏死细胞率分别为4.2％、7.6％和15.1％.10 mJ·s/cm2和20 mJ·s/cm2 UVB照射组细胞内Ca2＋水平分别为0 mJ·s/cm2照射组的（1.2±0.1）倍和（1.3±0.1）倍,差异均有统计学意义（P=0.039、0.004）.5、10和20 mJ·s/cm2 UVB照射组HLE B-3细胞PMCA3 mRNA表达量均明显低于对照组,差异均有统计学意义（P=0.001、0.004、0.000）,各组细胞中的PMCA蛋白相对表达量也均明显低于对照组,差异均有统计学意义（P=0.000、0.000、0.001）. 结论 UVB照射可导致白内障发生的机制可能与人LECs中PMCA3表达量下降和诱导LECs凋亡有关,UVB的作用呈剂量和时间依赖性.

白内障晶状体上皮细胞中miR-126和caspase-3的表达关系及意义

目的检测白内障晶状体上皮细胞(LECs)中miR-126、caspase-3的表达情况,探讨二者在白内障晶状体上皮细胞凋亡中作用及表达关系。方法选取2017年5月至2018年6月收治的78例白内障患者为观察组,同期选取52例正常人(角膜移植术供体)为对照组,采用实时荧光PCR(qRT-PCR)法检测晶状体上皮细胞中miR-126、Caspase-3 mRNA表达水平;Western blot检测晶状体上皮细胞中Caspase-3蛋白表达水平;流式细胞仪测定细胞凋亡率;分析miR-126、caspase-3与晶状体上皮细胞凋亡率的相关性;分析miR-126与Caspase-3在白内障晶状体上皮细胞中表达的相关性。结果观察组白内障患者晶状体上皮细胞中miR-126、Caspase-3 mRNA及蛋白水平、晶状体上皮细胞凋亡率均高于对照组(P<0.05);白内障患者晶状体上皮细胞中miR-126、caspase-3表达与晶状体上皮细胞凋亡率呈显著正相关(r=0.669,P<0.05;r=0.633,P<0.05);白内障患者晶状体上皮细胞中miR-126水平与caspase-3表达水平呈显著正相关(r=0.599,P<0.05)。结论miR-126、caspase-3在白内障晶状体上皮细胞中高表达,二者与晶状体上皮细胞凋亡有关,提示二者可能通过影响晶状体上皮细胞凋亡参与白内障疾病发生及发展。

Cyt C和Caspase 3在STZ诱导糖尿病大鼠晶状体上皮细胞中的表达

目的研究Cyt C和Caspase 3在STZ诱导糖尿病大鼠晶状体上皮细胞(lens epithelial cells,LEC)中的表达,为糖尿病性白内障的发生机制提供新的线索。方法 60只健康雄性SD大鼠分为实验组和对照组,每组各30只,实验组大鼠腹腔注射STZ溶液,对照组腹腔注射柠檬酸缓冲液。每4周监测大鼠血糖并用裂隙灯观察大鼠晶状体混浊情况,12周末取大鼠晶状体前囊膜,应用免疫组织化学和Western blot检测LEC中Cyt C和Caspase 3的表达。结果 12周末实验组大鼠空腹血糖为(24.04±2.40)mmol·L-1,对照组为(5.13±0.37)mmol·L-1。裂隙灯观察显示实验过程中实验组大鼠晶状体出现混浊,而对照组大鼠晶状体均透明。免疫组织化学结果显示:实验组大鼠LEC中Cyt C和Caspase 3表达呈阳性,而对照组则为阴性。Western blot结果显示实验组胞浆内Cyt C含量(0.928 0±0.029 7)大幅度上升,远高于对照组(0.525 0±0.025 8),差异有统计学意义(P<0.05),而实验组大鼠线粒体内Cyt C含量(0.510 5±0.026 4)则大幅度下降,远低于对照组(0.846 0±0.033 5),差异有统计学意义(P<0.05);实验组大鼠LEC中Caspase 3的表达量(0.906 0±0.027 6)明显高于对照组(0.430 5±0.025 2),差异有统计学意义(P<0.05)。结论 STZ诱导糖尿病大鼠LEC中Cyt C由线粒体释放入胞浆且Caspase 3表达量增加,高血糖可能通过刺激LEC释放Cyt C和激活Caspase 3,诱导LEC凋亡从而形成白内障。

MMP-3,OPN和TIMP-1在白内障形成中的作用

目的:研究基质金属蛋白酶-3(matrixmetalloproteinase-3,MMP-3)、骨桥蛋白(osteopontin,OPN)和金属蛋白酶组织抑制因子-1(tissueinhibitorofmetalloproteinase-1,TIMP-1)在白内障晶状体上皮细胞中的表达及意义。方法:采用免疫组织化学方法测定MMP-3,OPN和TIMP-1在31例前囊下性白内障,28例核性白内障和12例正常晶状体上皮细胞中的表达。结果:MMP-3在前囊下性白内障的表达高于核性白内障、正常晶状体(χ2=31.49,34.479;均P=0.000),而在核性白内障与正常晶状体中无明显差别(χ2=2.449,P=0.118);OPN在前囊下性白内障的表达高于核性白内障和正常晶状体(χ2=29.450,15.889;均P=0.000);TIMP-1表达在3组晶状体上皮细胞中差异均无统计学意义(P>0.05);前囊下性白内障MMP-3和OPN的表达水平相关(r=0.381,P=0.035),而MMP-3和OPN与TIMP-1的表达水平不相关(r=0.121,-0.289;P=0.516,0.114)。结论:MMP-3和OPN的表达增强及MMP-3/TIMP-1的表达失衡可能与前囊下性白内障的形成密切相关。

EGCG抗兔晶状体上皮细胞增殖机制中p38激酶作用的研究

目的:研究p38激酶在绿茶提取物中的表没食子儿茶素没食子酸酯[(-)-Epigallocatechin-3-gallate,EGCG]抗晶状体上皮细胞增殖机制中的作用。 方法:用噻唑蓝比色法(MTT比色法)研究p38的特异性抑制剂SB203580对EGCG抑制晶状体上皮细胞增殖作用的影响;用Western Blot法研究EGCG对p38激酶的磷酸化和非磷酸化水平的影响。 结果:(1)预先加入25μmol/L,50μmol/L的SB203580孵育1h后,100μmol/L EGCG对晶状体上皮细胞增殖的抑制率低于对照组,但这种差别没有统计学上的意义(P>0.05);200μmol/L EGCG组对晶状体上皮细胞的抑制率低于对照组,有统计学的意义(P<0.05);(2)在晶状体上皮细胞,磷酸化的p38基础水平很低。p38的磷酸化水平随EGCG浓度的增加逐渐增加,而非磷酸化的p38的水平与基础水平一样保持不变。以200μmol/L EGCG作用15min来研究其对p38的磷酸化和非磷酸化影响的时-效规律发现,加药后初期,p38的磷酸化水平很高,随后逐渐下降。同时,非磷酸化p38的水平保持不变。 结论:(1)EGCG对晶状体上皮细胞的增殖抑制作用可能通过p38通路;(2)EGCG对p38影响主要在于调节蛋白激酶的磷酸化与去磷酸化水平,不影响总蛋白含量。眼科学报 2003;19:236-238。

3-氨基苯甲酰胺影响糖尿病大鼠晶状体上皮细胞NF-κB表达的免疫组化研究

目的观察聚腺苷二磷酸核糖聚合酶(PARP)抑制剂3-氨基苯甲酰胺(3-AB)对早期糖尿病(DM)大鼠模型晶状体上皮细胞中核因子κB(NF-κB)表达的影响。方法采用一次性腹腔注射1%链脲佐菌素(STZ)50 mg/kg建立DM大鼠模型。成模后将大鼠随机分为正常对照组(N组)、糖尿病组(DM组)和干预组(DM+P组)。自建立模型后第3天起,DM+P组每日给予3-AB 30 mg/kg灌胃,N组和DM组每天给予等体积0.9%生理盐水灌胃。分别于给药后2、4、8周处死各组大鼠,取出晶状体,免疫组化SP法检测晶状体上皮细胞中NF-κB P65的表达情况。结果 DM 2、4、8周组大鼠晶状体上皮细胞NF-κB P65表达较相应时间点的N组高;DM+P 2、4、8周组大鼠晶状体上皮细胞NF-κB P65表达较相应时间点的N组高;DM+P 2、4、8周组与相应时间点DM组比较,晶状体上皮细胞NF-κB P65表达降低。结论 PARP抑制剂3-AB可以抑制早期糖尿病大鼠晶状体上皮细胞中NF-κB的表达。

Caspase-3抑制剂Ac-DEVD-CHO对氧化损伤诱导晶状体上皮细胞凋亡的防护作用

目的 探讨半胱氨酸天冬氨酸酶(caspase-3)及其抑制剂Ac-DEVD-CHO对过氧化氢(H_2O_2)诱导大鼠晶状体上皮细胞凋亡的影响,为进一步研究晶状体氧化损伤机制及其防护提供实验依据。方法 采用大鼠离体晶状体器官培养的方法,用Western blot法分析氧化损伤后晶状体上皮细胞内caspase-3蛋白酶活性,流式细胞AnnexinV-PI双染色法定量检测caspase-3抑制剂Ac-DEVD-CHO对晶状体上皮细胞凋亡率的影响。结果 氧化损伤诱导晶状体上皮细胞凋亡过程中,细胞内caspase-3酶活性明显增强,caspase-3特异性抑制剂Ac-DEVD-CHO可有效降低晶状体上皮细胞凋亡率。结论 caspase-3蛋白酶在氧化损伤诱导晶状体上皮细胞凋亡过程中占有重要地位,caspase-3抑制剂Ac-DEVD-CHO可有效防护体外晶状体上皮细胞氧化损伤。

老年性白内障患者晶状体上皮细胞CNN3、YAP表达与晶状体上皮细胞凋亡的相关性研究

目的 探讨老年性白内障患者晶状体上皮细胞中钙调节蛋白3(CNN3)、Yes相关蛋白(YAP)信使核糖核酸(mRNA)和蛋白表达水平与晶状体上皮细胞凋亡的关系。方法 选择2019年2月至2021年9月该院收治的67例白内障患者作为老年性白内障组,另选取同期于该院进行角膜移植术并排除白内障的42例患者作为对照组。手术获得患者晶状体前囊膜(晶状体前囊膜主要由上皮细胞构成),检测晶状体上皮细胞中CNN3、YAP mRNA和蛋白表达水平及晶状体上皮细胞凋亡率,分析老年性白内障患者晶状体上皮细胞中CNN3 mRNA与YAP mRNA表达水平的相关性及CNN3 mRNA、YAP mRNA与细胞凋亡率的相关性。结果 与对照组比较,老年性白内障组晶状体上皮细胞中CNN3 mRNA和蛋白表达水平升高,YAP mRNA和蛋白表达水平降低(P<0.05);老年性白内障患者晶状体上皮细胞中CNN3 mRNA与YAP mRNA表达水平呈负相关(P<0.05);老年性白内障组晶状体上皮细胞凋亡率高于对照组(P<0.05);老年性白内障患者晶状体上皮细胞中CNN3 mRNA表达水平与晶状体上皮细胞凋亡率呈正相关(P<0.05),YAP mRNA表达水平与晶状体上皮细胞凋亡率呈负相关(P<0.05)。结论 老年性白内障患者晶状体上皮细胞中CNN3呈高表达、YAP呈低表达,二者与晶状体上皮细胞凋亡率有关,可能参与老年性白内障疾病的发生。