Vision Science Program, School of Optometry, Indiana University, Bloomington, Indiana, United States.
Department of Ophthalmology, Duke University, Durham, North Carolina, United States.
Invest Ophthalmol Vis Sci. 2021 Sep 2;62(12):15. doi: 10.1167/iovs.62.12.15.
The Slc4a11 knock out (KO) mouse model recapitulates the human disease phenotype associated with congenital hereditary endothelial dystrophy (CHED). Increased mitochondrial reactive oxygen species (ROS) in the Slc4a11 KO mouse model is a major cause of edema and endothelial cell loss. Here, we asked if autophagy was activated by ROS in the KO mice.
Immortalized cell lines and mouse corneal endothelia were used to measure autophagy and lysosome associated protein expressions using Protein Simple Wes immunoassay. Autophagy and lysosome functions were examined in wild type (WT) and KO cells as well as animals treated with the mitochondrial ROS quencher MitoQ.
Even though autophagy activation was evident, autophagy flux was aberrant in Slc4a11 KO cells and corneal endothelium. Expression of lysosomal proteins and lysosomal mass were decreased along with reduced nuclear translocation of lysosomal master regulator, transcription factor EB (TFEB). MitoQ reversed aberrant lysosomal functions and TFEB nuclear localization in KO cells. MitoQ injections in KO animals reduced corneal edema and decreased the rate of endothelial cell loss.
Mitochondrial ROS disrupts TFEB signaling causing lysosomal dysfunction with impairment of autophagy in Slc4a11 KO corneal endothelium. Our study is the first to identify the presence as well as cause of lysosomal dysfunction in an animal model of CHED, and to identify a potential therapeutic approach.
Slc4a11 敲除（KO）小鼠模型再现了与先天性遗传性血管内皮营养不良（CHED）相关的人类疾病表型。Slc4a11 KO 小鼠模型中增加的线粒体活性氧（ROS）是水肿和内皮细胞丧失的主要原因。在这里，我们想知道 ROS 是否在 KO 小鼠中激活了自噬。
使用蛋白简单 Wes 免疫测定法，在永生化细胞系和小鼠角膜内皮中测量自噬和溶酶体相关蛋白的表达。在 WT 和 KO 细胞以及用线粒体 ROS 清除剂 MitoQ 处理的动物中，检查自噬和溶酶体功能。
尽管自噬激活明显，但 Slc4a11 KO 细胞和角膜内皮中的自噬流异常。溶酶体蛋白的表达和溶酶体质量减少，同时溶酶体主调控因子转录因子 EB（TFEB）的核易位减少。MitoQ 逆转了 KO 细胞中异常的溶酶体功能和 TFEB 核定位。MitoQ 注射到 KO 动物中可减少角膜水肿并降低内皮细胞丢失率。
线粒体 ROS 破坏了 TFEB 信号传导，导致 Slc4a11 KO 角膜内皮中溶酶体功能障碍和自噬受损。我们的研究首次在 CHED 的动物模型中确定了溶酶体功能障碍的存在及其原因，并确定了一种潜在的治疗方法。
