PEDF Improves Mitochondrial Function in RPE Cells During Oxidative Stress
PEDF Improves Mitochondrial Function in RPE Cells During Oxidative Stress
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DOI：
10.1167/iovs.14-14696
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发表时间：
2014-10-01
影响因子：
4.4
通讯作者：
Tombran-Tink, Joyce
中科院分区：
文献类型：
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作者：
He, Yuan;Leung, Kar Wah;Tombran-Tink, Joyce
目的.氧化应激在健康和衰老中起着重要作用。我们已经表明，氧化应激损害线粒体功能，并促进RPE细胞死亡的年龄依赖性的方式。本研究探讨色素上皮衍生因子（PEDF）通过线粒体途径在限制氧化应激诱导的RPE细胞损伤中的作用。将来自50至55岁、60至65岁和70至75岁（yo）供体的三组早期传代RPE细胞用PEDF预处理，然后暴露于亚致死剂量的过氧化氢（H2O2），或在H2O2处理后用PEDF后处理。PEDF对线粒体功能和细胞活力的影响进行了检查。在人RPE细胞中，氧化应激诱导LDH释放、活性氧（ROS）水平和细胞死亡的年龄依赖性增加以及三磷酸腺苷（ATP）产生和线粒体膜电位（Δ Psim）的减少。用PEDF预处理或后应激处理导致细胞活力增加，细胞色素c释放和半胱天冬酶3裂解的抑制，以及由ROS产生减少和ATP产生和Δ Psi m增加表示的线粒体功能改善。氧化应激也破坏了线粒体的网状网络、运输和分布，并阻断了细胞中磷脂酰肌醇3激酶（PI3K）、Akt和Erk信号的激活。与50至55岁年龄组相比，这些作用在来自> 60岁个体的RPE细胞中更明显。色素上皮衍生因子减轻了氧化应激对线粒体重塑和细胞分布的负面影响，并解除了其对PI3K/Akt和丝裂原活化蛋白激酶（MAPK）信号的控制。尽管PEDF增强了细胞中PI3K/Akt和MAPK信号传导，但线粒体网络和功能的稳定依赖于其PI3K/Akt的激活。PEDF活性的特异性使用药理学抑制剂LY294002、SH6和U0126证实。我们还表明，在没有氧化应激的情况下，单独的PI3K/Akt通路的药理学抑制足以破坏线粒体的结构和功能。此外，PEDF阻断氧化应激对亲环素D和UCP2表达的影响，这些基因控制线粒体功能，以及凋亡基因caspase 3、Bax和Bcl 2。PEDF对活性氧水平的控制与UCP 2调节特异性相关，因为PEDF诱导的UCP 2缺陷细胞中该基因的表达与活性氧产生的减少有关。结论。我们提供的证据表明，PEDF通过稳定线粒体网络和功能促进衰老RPE细胞对氧化应激的恢复力，并且人类RPE细胞中的线粒体动力学部分通过PI3K/Akt途径进行控制。
PURPOSE. Oxidative stress plays an important role in health and aging. We have shown that oxidative stress impairs mitochondrial function and promotes RPE cell death in an age-dependent manner. This study investigates the role of pigment epithelium-derived factor (PEDF) in limiting oxidative stress-induced damage to RPE cells through mitochondrial pathways.METHODS. Three groups of early-passaged RPE cells from donors 50 to 55, 60 to 65, and 70 to 75 years old (yo) were either preconditioned with PEDF followed by exposure to sublethal doses of hydrogen peroxide (H2O2) or post-treated with PEDF after H2O2 treatment. Effects of PEDF on mitochondrial function and cell viability were examined.RESULTS. Oxidative stress induced an age-dependent increase in LDH release, reactive oxygen species (ROS) levels, and cell death and a decrease in adenosine triphosphate (ATP) production and mitochondrial membrane potential (Delta Psi m) in human RPE cells. Preconditioning or poststressed treatment with PEDF resulted in increased cell viability, inhibition of cytochrome c release and caspase 3 cleavage, and improved mitochondria function denoted by a decrease in ROS generation and increases in ATP production and Delta Psi m. Oxidative stress also disrupted the reticular network, trafficking, and distribution of the mitochondria and blocked activation of phosphatidylinositol 3 kinase (PI3K), Akt, and Erk signaling in the cells. These effects were more pronounced in RPE cells from individuals >60 yo compared to the 50 to 55 yo age group. Pigment epithelium-derived factor mitigated negative effects of oxidative stress on mitochondrial remodeling and cellular distribution and unblocked its control of PI3K/Akt and mitogen-activated protein kinase (MAPK) signaling. Although PEDF potentiated both PI3K/Akt and MAPK signaling in the cells, stabilization of mitochondrial networks and function was dependent on its activation of PI3K/Akt. Specificity of PEDF's activity was confirmed using the pharmacological inhibitors LY294002, SH6, and U0126. We also show that in the absence of oxidative stress, pharmacological inhibition of the PI3K/Akt pathway alone was sufficient to disrupt mitochondrial structure and function. In addition, PEDF blocked effects of oxidative stress on expression of cyclophilin D and UCP2, genes controlling mitochondrial function, and the apoptotic genes caspase 3, Bax, and Bcl2. Control of ROS levels by PEDF was specifically linked to UCP2 regulation since PEDF-induced expression of this gene in UCP2-deficient cells was associated with a decrease in ROS production.CONCLUSIONS. We provide evidence that PEDF promotes resilience of aging RPE cells to oxidative stress by stabilizing mitochondrial networks and function and that mitochondrial dynamics in human RPE cells are controlled, in part, through the PI3K/Akt pathway.