{"title":"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。","authors":"","doi":"10.1016/j.molmet.2024.101995","DOIUrl":null,"url":null,"abstract":"<div><h3>Objectives</h3><p>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.</p></div><div><h3>Methods</h3><p>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–<sup>13</sup>C<sub>6</sub>] glucose isotopic tracing was preliminarily conducted on S179C ARPE-19 followed by [U–<sup>13</sup>C<sub>6</sub>] glucose and [U–<sup>13</sup>C<sub>5</sub>] glutamine isotopic tracing in SFD iRPE cells.</p></div><div><h3>Results</h3><p>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–<sup>13</sup>C<sub>6</sub>] glucose demonstrated enhanced glucose utilization and glycolytic activity in S179C TIMP3 APRE-19 cells. Similarly, [U–<sup>13</sup>C<sub>6</sub>] glucose tracing in SFD iRPE revealed increased glucose contribution to glycolysis and the TCA cycle. Additionally, [U–<sup>13</sup>C<sub>5</sub>] glutamine tracing found evidence of altered malic enzyme activity.</p></div><div><h3>Conclusions</h3><p>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.</p></div>","PeriodicalId":18765,"journal":{"name":"Molecular Metabolism","volume":null,"pages":null},"PeriodicalIF":7.0000,"publicationDate":"2024-07-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2212877824001261/pdfft?md5=f27c912d6cfdad145dbd8ae7a0e9f38f&pid=1-s2.0-S2212877824001261-main.pdf","citationCount":"0","resultStr":"{\"title\":\"Tissue Inhibitor of Metalloproteinase 3 (TIMP3) mutations increase glycolytic activity and dysregulate glutamine metabolism in RPE cells\",\"authors\":\"\",\"doi\":\"10.1016/j.molmet.2024.101995\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><h3>Objectives</h3><p>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.</p></div><div><h3>Methods</h3><p>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–<sup>13</sup>C<sub>6</sub>] glucose isotopic tracing was preliminarily conducted on S179C ARPE-19 followed by [U–<sup>13</sup>C<sub>6</sub>] glucose and [U–<sup>13</sup>C<sub>5</sub>] glutamine isotopic tracing in SFD iRPE cells.</p></div><div><h3>Results</h3><p>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–<sup>13</sup>C<sub>6</sub>] glucose demonstrated enhanced glucose utilization and glycolytic activity in S179C TIMP3 APRE-19 cells. Similarly, [U–<sup>13</sup>C<sub>6</sub>] glucose tracing in SFD iRPE revealed increased glucose contribution to glycolysis and the TCA cycle. Additionally, [U–<sup>13</sup>C<sub>5</sub>] glutamine tracing found evidence of altered malic enzyme activity.</p></div><div><h3>Conclusions</h3><p>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.</p></div>\",\"PeriodicalId\":18765,\"journal\":{\"name\":\"Molecular Metabolism\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":7.0000,\"publicationDate\":\"2024-07-22\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.sciencedirect.com/science/article/pii/S2212877824001261/pdfft?md5=f27c912d6cfdad145dbd8ae7a0e9f38f&pid=1-s2.0-S2212877824001261-main.pdf\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Molecular Metabolism\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S2212877824001261\",\"RegionNum\":2,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"ENDOCRINOLOGY & METABOLISM\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Molecular Metabolism","FirstCategoryId":"3","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S2212877824001261","RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"ENDOCRINOLOGY & METABOLISM","Score":null,"Total":0}
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.
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.