锂治疗可增加 Fuchs 内皮角膜营养不良小鼠模型中内皮细胞的存活和自噬。
Lithium treatment increases endothelial cell survival and autophagy in a mouse model of Fuchs endothelial corneal dystrophy.
机构信息
Cornea and Anterior Segment Division, Wilmer Eye Institute, Johns Hopkins Medical Institutions, Baltimore, Maryland 21231, USA.
出版信息
Br J Ophthalmol. 2013 Aug;97(8):1068-73. doi: 10.1136/bjophthalmol-2012-302881. Epub 2013 Jun 12.
BACKGROUND
Lithium previously has been shown to reduce both endoplasmic reticulum (ER) and oxidative stress in other in vitro and in vivo model systems. We investigated lithium's effects on cultured corneal endothelial cells (CECs) exposed to these types of stress and in a mouse model of Fuchs endothelial corneal dystrophy (FECD).
METHODS
Viability of cultured bovine CECs was determined by CellTiter-Glo. 2-month-old Col8a2(Q455K/Q455K) mutant (Q455K) and C57/Bl6 wild type animals were divided into two groups of 15 mice. Group I received 0.2% lithium carbonate-containing chow and Group II received control chow for 7 months. Confocal microscopy, transmission electron microscopy, real-time PCR (RT-PCR) and western blot were performed.
RESULTS
Pretreatment with lithium increased viability of cultured CECs after H2O2 and thapsigargin exposure compared with untreated controls (p<0.05). In vivo analysis of mouse corneal endothelium showed the following: endothelial cell density of lithium treated Q455K was higher than for untreated Q455K (p<0.01). transmission electron microscopy of lithium treated Q455K showed normal endothelium with enlarged autophagosomes, but untreated Q455K showed dilated ER and guttae. Compared with untreated Q455K endothelium, lithium treated Q455K showed significant upregulation of P62, Tmem74, Tm9sf1 and Tmem166 by RT-PCR and of Atg5-12 conjugate by western blotting indicating that lithium treatment increased autophagy. Although RT-PCR unexpectedly showed increased levels of lithium response genes, caspase 12, Gsk3β, Arrβ2 and Impa1, western blotting showed the expected downregulation of Arrβ2 and Impa1 proteins in response to lithium treatment.
CONCLUSIONS
Lithium increases cultured CEC survival against ER and oxidative stress. Increased autophagy in lithium treated endothelium in a mouse model of FECD suggests autophagy may contribute to increased endothelial cell survival.
背景
锂先前已被证明可减少其他体外和体内模型系统中的内质网（ER）和氧化应激。我们研究了锂对暴露于这些应激类型的培养的角膜内皮细胞（CEC）和 Fuchs 内皮角膜营养不良（FECD）的小鼠模型中的作用。
方法
通过 CellTiter-Glo 测定培养的牛 CEC 的活力。将 2 月龄 Col8a2（Q455K/Q455K）突变（Q455K）和 C57/Bl6 野生型动物分为两组，每组 15 只小鼠。I 组接受含 0.2％碳酸锂的饲料，II 组接受对照饲料 7 个月。进行共聚焦显微镜检查、透射电子显微镜检查、实时 PCR（RT-PCR）和 Western blot。
结果
与未处理的对照组相比，H2O2 和 thapsigargin 暴露后用锂预处理可增加培养的 CEC 的活力（p<0.05）。对小鼠角膜内皮的体内分析表明：锂处理的 Q455K 的内皮细胞密度高于未处理的 Q455K（p<0.01）。锂处理的 Q455K 的透射电子显微镜显示正常的内皮细胞，伴有扩大的自噬体，但未处理的 Q455K 显示扩张的 ER 和 guttae。与未处理的 Q455K 内皮细胞相比，锂处理的 Q455K 通过 RT-PCR 显示 P62、Tmem74、Tm9sf1 和 Tmem166 的显著上调，并且通过 Western blot 显示 Atg5-12 缀合物的上调，表明锂处理增加了自噬。尽管 RT-PCR 出人意料地显示锂反应基因、caspase 12、Gsk3β、Arrβ2 和 Impa1 的水平升高，但 Western blot 显示锂处理后 Arrβ2 和 Impa1 蛋白的预期下调。
结论
锂可增加培养的 CEC 对 ER 和氧化应激的存活能力。在 FECD 的小鼠模型中，锂处理的内皮细胞中自噬的增加表明自噬可能有助于增加内皮细胞的存活。
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