锌对培养的人视网膜色素上皮细胞中半胱天冬酶-3及DNA降解的影响

The influence of zinc on caspase-3 and DNA breakdown in cultured human retinal pigment epithelial cells.

作者信息

Wood J P, Osborne N N

机构信息

The Nuffield Laboratory of Ophthalmology, University of Oxford, Walton Street Oxford, OX2 6AW, England.

出版信息

Arch Ophthalmol. 2001 Jan;119(1):81-8.

PMID:11146730

Abstract

OBJECTIVES

To investigate the role of extracellular zinc on the death process of cultured human retinal pigment epithelial (RPE) cells.

METHODS

Confluent cells on borosilicate glass coverslips were treated with substances in serum-free growth medium for various times and were analyzed for death by means of changes in morphologic features, numbers of attached cells, and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) procedure. Some cultures were also exposed to experimental ischemia (defined as a lack of oxygen, glucose, and serum). Electrophoresis and Western blotting and enzyme assays were used to investigate changes in expression of the protease enzyme, caspase-3.

RESULTS

Experimental ischemia caused death of RPE cells. Zinc sulfate had no effect on these cells at low concentrations (100 pmol/L to 10 nmol/L), but protected them at higher concentrations (< or = 10 micromol/L) and appeared to exacerbate cell death at still greater concentrations. Moreover, zinc compounds (>10 micromol/L) also induced death of cells in control cultures that could be blocked by zinc chelators and partially by the caspase-3 inhibitor, DEVD-FMK. Zinc also increased the amount of the active form of caspase-3 in RPE cells.

CONCLUSIONS

Zinc salts protect RPE cells from experimental ischemia-induced death at low concentrations (100 pmol/L-10 nmol/L). However, at higher concentrations, zinc causes cell death and alters the cellular level of caspase-3. These observations are consistent with the death process being apoptosis.

CLINICAL RELEVANCE

Zinc supplements are taken by many individuals. Low doses of zinc can protect RPE cells against ischemic-type insults as may occur in certain ocular complaints. Furthermore, high concentrations of zinc can damage RPE cells. Because zinc ions are known to be taken up by RPE cells from the choroidal circulation, the actual therapeutic dose taken by patients is critical.

摘要

目的

研究细胞外锌在培养的人视网膜色素上皮（RPE）细胞死亡过程中的作用。

方法

将硼硅酸盐玻璃盖玻片上的汇合细胞在无血清生长培养基中用各种物质处理不同时间，并通过形态学特征变化、贴壁细胞数量以及末端脱氧核苷酸转移酶介导的脱氧尿苷三磷酸缺口末端标记（TUNEL）法分析细胞死亡情况。一些培养物还暴露于实验性缺血（定义为缺氧、缺葡萄糖和无血清）环境中。采用电泳、蛋白质免疫印迹和酶分析来研究蛋白酶caspase-3表达的变化。

结果

实验性缺血导致RPE细胞死亡。低浓度（100 pmol/L至10 nmol/L）的硫酸锌对这些细胞无影响，但高浓度（≤10 μmol/L）时可保护细胞，而更高浓度时似乎会加剧细胞死亡。此外，锌化合物（>10 μmol/L）也可诱导对照培养物中的细胞死亡，这种死亡可被锌螯合剂阻断，部分可被caspase-3抑制剂DEVD-FMK阻断。锌还增加了RPE细胞中活性形式的caspase-3的量。