组织金属蛋白酶 3 抑制剂(TIMP3)突变会增加糖酵解活性,并使 RPE 细胞中的谷氨酰胺代谢失调。

IF 7 2区 医学 Q1 ENDOCRINOLOGY & METABOLISM Molecular Metabolism Pub Date : 2024-07-22 DOI:10.1016/j.molmet.2024.101995
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引用次数: 0

摘要

研究目的组织金属蛋白酶抑制剂 3(TIMP3)的突变会导致索斯比眼底营养不良症(SFD),这是一种显性遗传的罕见黄斑变性,会导致视力丧失。TIMP3 主要由视网膜色素上皮细胞(RPE)合成,RPE 细胞构成血液-视网膜外屏障。RPE 的一个主要功能是合成葡萄糖等重要营养物质并将其运输到视网膜。最近,RPE 细胞的代谢功能障碍已成为视网膜变性的一个重要诱因。我们试图确定 RPE 代谢功能障碍是否是导致自发性视网膜病变的发病机制:我们对表达 TIMP3 S179C 变体的小鼠的 RPE 进行了定量蛋白质组学研究。对 S179C ARPE-19 初步进行了[P13C6]葡萄糖同位素示踪,随后在 SFD iRPE 细胞中进行了[U-13C6]葡萄糖和[U-13C5]谷氨酰胺同位素示踪:结果:对表达突变型 S179C TIMP3 的小鼠 RPE 进行定量蛋白质组学和富集分析,发现了富集于代谢相关途径和过程的差异表达蛋白质。值得注意的是,这些结果突显了糖酵解和葡萄糖代谢的失调。用[U-13C6]葡萄糖进行的稳定同位素追踪实验表明,S179C TIMP3 APRE-19 细胞的葡萄糖利用和糖酵解活性增强。同样,在 SFD iRPE 中进行的[U-13C6] 葡萄糖示踪显示,葡萄糖对糖酵解和 TCA 循环的贡献增加。此外,[U-13C5] 谷氨酰胺追踪发现了苹果酸酶活性改变的证据:这项研究提供了有关 SFD 中 RPE 糖代谢失调的重要信息,并揭示了与其他视网膜变性疾病的潜在共性,强调了 RPE 细胞代谢是一个治疗靶点。
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Tissue Inhibitor of Metalloproteinase 3 (TIMP3) mutations increase glycolytic activity and dysregulate glutamine metabolism in RPE cells

Objectives

Mutations in Tissue Inhibitor of Metalloproteinases 3 (TIMP3) cause Sorsby's Fundus Dystrophy (SFD), a dominantly inherited, rare form of macular degeneration that results in vision loss. TIMP3 is synthesized primarily by retinal pigment epithelial (RPE) cells, which constitute the outer blood-retinal barrier. One major function of RPE is the synthesis and transport of vital nutrients, such as glucose, to the retina. Recently, metabolic dysfunction in RPE cells has emerged as an important contributing factor in retinal degenerations. We set out to determine if RPE metabolic dysfunction was contributing to SFD pathogenesis.

Methods

Quantitative proteomics was conducted on RPE of mice expressing the S179C variant of TIMP3, known to be causative of SFD in humans. Proteins found to be differentially expressed (P < 0.05) were analyzed using statistical overrepresentation analysis to determine enriched pathways, processes, and protein classes using g:profiler and PANTHER Gene Ontology. We examined the effects of mutant TIMP3 on RPE metabolism using human ARPE-19 cells expressing mutant S179C TIMP3 and patient-derived induced pluripotent stem cell-derived RPE (iRPE) carrying the S204C TIMP3 mutation. RPE metabolism was directly probed using isotopic tracing coupled with GC/MS analysis. Steady state [U–13C6] glucose isotopic tracing was preliminarily conducted on S179C ARPE-19 followed by [U–13C6] glucose and [U–13C5] glutamine isotopic tracing in SFD iRPE cells.

Results

Quantitative proteomics and enrichment analysis conducted on RPE of mice expressing mutant S179C TIMP3 identified differentially expressed proteins that were enriched for metabolism-related pathways and processes. Notably these results highlighted dysregulated glycolysis and glucose metabolism. Stable isotope tracing experiments with [U–13C6] glucose demonstrated enhanced glucose utilization and glycolytic activity in S179C TIMP3 APRE-19 cells. Similarly, [U–13C6] glucose tracing in SFD iRPE revealed increased glucose contribution to glycolysis and the TCA cycle. Additionally, [U–13C5] glutamine tracing found evidence of altered malic enzyme activity.

Conclusions

This study provides important information on the dysregulation of RPE glucose metabolism in SFD and implicates a potential commonality with other retinal degenerative diseases, emphasizing RPE cellular metabolism as a therapeutic target.

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来源期刊
Molecular Metabolism
Molecular Metabolism ENDOCRINOLOGY & METABOLISM-
CiteScore
14.50
自引率
2.50%
发文量
219
审稿时长
43 days
期刊介绍: Molecular Metabolism is a leading journal dedicated to sharing groundbreaking discoveries in the field of energy homeostasis and the underlying factors of metabolic disorders. These disorders include obesity, diabetes, cardiovascular disease, and cancer. Our journal focuses on publishing research driven by hypotheses and conducted to the highest standards, aiming to provide a mechanistic understanding of energy homeostasis-related behavior, physiology, and dysfunction. We promote interdisciplinary science, covering a broad range of approaches from molecules to humans throughout the lifespan. Our goal is to contribute to transformative research in metabolism, which has the potential to revolutionize the field. By enabling progress in the prognosis, prevention, and ultimately the cure of metabolic disorders and their long-term complications, our journal seeks to better the future of health and well-being.
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