紫外线照射后人晶状体上皮细胞的iTRAQ定量蛋白质组学研究

目的:探索氧化损伤状态下人晶状体上皮细胞(SRA01/04)的蛋白表达变化,以期为年龄相关性白内障(ARC)发病机制的研究提供新的蛋白靶点。方法:将SRA01/04细胞分为实验组和对照组,实验组即细胞通过紫外线(UVB)照射以构建细胞氧化损伤模型,照射时长为10min;对照组即细胞未经任何处理。应用同位素标记相对和绝对定量(iTRAQ)技术对这两组细胞进行蛋白组测序,以差异倍数>1.2且p<0.05的标准筛选差异表达蛋白(DEPs),并通过基因本体(GO)和京都基因和基因组百科全书(KEGG)数据库分别对上调和下调显著性前50的DEPs进行功能富集分析,利用Pathway commons构建蛋白-蛋白相互作用(PPI)网络。结果:本研究共筛选出552个DEPs。相比于对照组,实验组中上调的DEPs有176个,包括HMGB1、USP1等,下调的DEPs有376个,包括POLR2A、POLR2B等。GO和KEGG富集分析显示上调和下调显著性前50的DEPs参与了多种重要的生物学过程和信号通路。PPI网络提示氧化损伤修复(ODR)相关蛋白在UVB诱导的氧化损伤中可能起关键作用。结论:UVB照射SRA01/04细胞可致多种蛋白,尤其是ODR相关蛋白的表达发生改变,这些研究结果为深入探索ARC发病机制以及研究与治疗靶点相关的蛋白质或通路提供了细胞层面的参考。

年龄对水合氯醛诱导的小鼠可逆性晶状体混浊及Na^(+)-K^(+)-ATP酶表达的影响

目的:探究年龄对水合氯醛诱导的小鼠急性可逆性晶状体混浊及Na^(+)-K^(+)-ATP酶表达的影响。方法:青年(3月龄)和老年(24月龄)C57BL/6小鼠各12只,4%水合氯醛(400mg/kg)腹腔注射诱导急性可逆性晶状体混浊。在注射后10、20、30、45、60、90、120、150min分别应用裂隙灯观察晶状体混浊程度,根据晶状体混浊评价系统记录混浊程度分级情况并拍照。苏木素-伊红(HE)染色观察晶状体病理改变,免疫组织化学染色检测晶状体Na^(+)-K^(+)-ATP酶表达。结果:水合氯醛注射后两组小鼠晶状体混浊和消退过程相似,但青年组小鼠晶状体混浊出现早、持续时间略长,混浊厚重,呈乳白色;老年组小鼠晶状体混浊出现晚,持续时间略短,混浊轻薄,呈薄雾状。HE染色显示水合氯醛注射后晶状体上皮细胞(LECs)下皮质聚集大量水泡,浅层晶状体纤维细胞结构紊乱。免疫组织化学染色显示LECs及纤维Na^(+)-K^(+)-ATP酶的表达呈现阳性,水合氯醛注射前青年组和老年组小鼠LECs中Na^(+)-K^(+)-ATP酶表达较弱,注射后45min LECs中Na^(+)-K^(+)-ATP酶的表达上调,且老年组小鼠LECs的Na^(+)-K^(+)-ATP酶上调更为显著。结论:年龄影响水合氯醛诱导小鼠急性可逆性晶状体混浊,Na^(+)-K^(+)-ATP酶参与水合氯醛诱导的晶状体混浊。

江苏省阜宁县农村50岁及以上人群年龄相关性黄斑变性的流行病学调查

目的:分析江苏省阜宁县50岁及以上农村人群中年龄相关性黄斑变性(age-related macular degeneration,ARMD)的患病率情况。方法:本项调查研究采用随机整群抽样的方法,在江苏省盐城阜宁县设置30个调查点,对50岁及以上农村人群进行视力和眼部相关检查。计量资料采用均数±标准差表示,组间比较采用独立样本t检验;计数资料采用率或构成比表示,组间比较采用χ~2检验。应用Logistic回归分析法分析年龄、性别等因素对ARMD患病率的影响。结果:随机抽样6145例,实际纳入5947例,受检率为96.78%。检出ARMD患者448例633眼,患病率为7.53%。按日常生活视力(PVA)和视力损伤标准计算,在中重度视力损伤和盲眼中,由ARMD引起的中重度视力损失和盲分别占11.96%和4.13%;按最佳矫正视力(BCVA)和视力损伤标准计算,在中重度视力损伤和盲眼中,由ARMD引起的中重度视力损伤和盲分别占7.79%和4.45%。高龄是影响ARMD的相关因素(OR=1.01,P=0.040)。结论:江苏省阜宁县农村50岁及以上人群中,ARMD患病率较高,是影响老年人视力的主要原因之一。

外泌体在眼科疾病中的研究进展

外泌体(exosomes)是一类直径50~150nm的细胞外膜泡,可以递送生物活性分子(如蛋白质、脂质、DNA、miRNA等)至靶细胞中,以发挥细胞间通讯作用。外泌体介导的细胞间通讯影响靶细胞的凋亡、侵袭、迁移、免疫应答及氧化损伤修复等功能。近年来外泌体研究在眼科学领域迅速开展,本文总结了在眼科疾病中外泌体相关研究的最新进展。

LSS和HMGCR基因的单核苷酸多态性与年龄相关性白内障的关系

目的:探讨汉族人群羊毛甾醇合成酶(LSS)和3-羟基-3-甲基戊二酰辅酶A还原酶(HMGCR)基因的单核苷酸多态性(SNPs)与年龄相关性白内障(ARC)的风险是否相关。方法:采用病例对照研究。研究对象来自流调人群以及临床病例。用TaqMan RT-PCR法检测基因的SNPs;用qRT-PCR检测晶状体上皮细胞(LECs)的LSS mRNA水平;用卡方检验比较ARC组与对照组SNPs的差异,并计算比值比。结果:发现LSS-rs2968在ARC组与对照组人群存在差异(P=0.018),但是Bonferroni校正后该差异消失(P=0.072)。进行分层分析后发现LSS-rs2968 A等位基因与江苏人群的核型ARC危险相关(P=0.003),Bonferroni校正后差异依然存在(P=0.012)。我们进一步发现了各型ARC组LECs中LSS的mRNA表达均低于对照组(P<0.05)。结论:LSS-rs2968 A等位基因可能参与了江苏人群在核型ARC的发生发展,其中A等位基因为易感基因。

Activation of autophagy in the retina after optic nerve crush injury in rats

AIM: To investigate the activation of autophagy in rat retina after optic nerve crush(ONC) and evaluate its relationship with apoptosis of retinal ganglion cells(RGCs).METHODS: The ONC model was established. Western blots were performed to investigate expression of p62, LC3 and Beclin-1. Transmission electron microscopy was performed to discover the autophagosomes in the retina after ONC. Immunohistochemistry was used to confirm the distribution of LC3. TUNEL was performed to confirm the relationship between autophagy and RGC apoptosis. RESULTS: p62/Beclin-1 ratio was declined shortly after ONC until to day 7 after ONC and then restored to a normal level at day 21. There was an opposite change in the LC3-II/LC3 I ratio in the retina compared to the p62/Beclin-1 ratio. Increased autophagosomes were found after ONC using transmission electron microscopy, and most of the LC3-stained cells were colocalized with RGCs and Müller cells. More LC3-immunoreactive cells and apoptotic RGCs were found on day 7 following ONC. CONCLUSION: Possible activation of autophagy in RGCs after ONC;autophagy mainly occurred in RGCs and Müller cells, and the apoptosis of RGCs after ONC may be partly associated with autophagic activation.

Differentially expressed long noncoding RNAs and regulatory mechanism of LINC02407 in human gastric adenocarcinoma

BACKGROUND Long noncoding RNAs(lncRNAs)have been identified to play important roles in the development and progression of various tumors,including gastric cancer(GC).However,the molecular role of lncRNAs in GC progression remains unclear.AIM To investigate the differential expression of lncRNAs in human GC and elucidate the function and regulatory mechanism of LINC02407.METHODS The Cancer Genome Atlas database was used to investigate the involvement of lncRNAs in GC.Quantitative real-time polymerase chain reaction was used to estimate the relative expression level of LINC02407 in GC tissues and cells.Functional experiments including CCK8 assay,apoptosis assay,wound healing assay,and transwell assay were used to investigate the effect of LINC02407 on GC cells.Some microRNAs were predicted and verified via bioinformatics analysis and the luciferase reporter system.Predictive analysis and Western blot assay were used to analyze the expression of related proteins.RESULTS Many differentially expressed lncRNAs were identified in GC,and some of them including LINC02407 can affect the survival.LINC02407 was upregulated in tumor tissues compared with adjacent tissues.HGC-27 cells showed the highest LINC02407 expression and HaCaT cells exhibited the lowest expression.Different experiment groups were constructed using LINC02407 overexpressing plasmids and related siRNAs.The results of functional experiments showed that LINC02407 can promote the proliferation,migration,and invasion of GC cells but inhibit apoptosis.Luciferase reporter assay showed that hsa-miR-6845-5p and hsa-miR-4455 was downstream regulated by LINC02407.Western blot analysis showed that adhesion G protein-coupled receptor D1(ADGRD1)was regulated by the LINC02407-miR-6845-5p/miR-4455-ADGRD1 pathways.CONCLUSION LINC02407 plays a role in GC through the LINC02407-miR-6845-5p/miR-4455-ADGRD1 pathways,and thus,it may be an important oncogene and has potential value in GC diagnosis and treatment.