Genomic and Metabolic Responses to Methionine-Restricted and Methionine-Restricted, Cysteine-Supplemented Diets in Fischer 344 Rat Inguinal Adipose Tissue, Liver and Quadriceps Muscle

Abstract

Background/Aims: Methionine restriction (MR) is a dietary intervention that increases lifespan, reduces adiposity and improves insulin sensitivity. These effects are reversed by supplementation of the MR diet with cysteine (MRC). Genomic and metabolomic studies were conducted to identify potential mechanisms by which MR induces favorable metabolic effects, and that are reversed by cysteine supplementation. Methods: Gene expression was examined by microarray analysis and TaqMan quantitative PCR. Levels of selected proteins were measured by Western blot and metabolic intermediates were analyzed by mass spectrometry. Results: MR increased lipid metabolism in inguinal adipose tissue and quadriceps muscle while it decreased lipid synthesis in liver. In inguinal adipose tissue, MR not only caused the transcriptional upregulation of genes associated with fatty acid synthesis but also of Lpin1, Pc, Pck1 and Pdk1, genes that are associated with glyceroneogenesis. MR also upregulated lipolysis-associated genes in inguinal fat and led to increased oxidation in this tissue, as suggested by higher levels of methionine sulfoxide and 13-HODE + 9-HODE compared to control-fed (CF) rats. Moreover, MR caused a trend toward the downregulation of inflammation-associated genes in inguinal adipose tissue. MRC reversed most gene and metabolite changes induced by MR in inguinal adipose tissue, but drove the expression of Elovl6, Lpin1, Pc, and Pdk1 below CF levels. In liver, MR decreased levels of a number of long-chain fatty acids, glycerol and glycerol-3-phosphate corresponding with the gene expression data. Although MR increased the expression of genes associated with carbohydrate metabolism, levels of glycolytic intermediates were below CF levels. MR, however, stimulated gluconeogenesis and ketogenesis in liver tissue. As previously reported, sulfur amino acids derived from methionine were decreased in liver by MR, but homocysteine levels were elevated. Increased liver homocysteine levels by MR were associated with decreased cystathionine β-synthase (CBS) protein levels and lowered vitamin B6 and 5-methyltetrahydrofolate (5MeTHF) content. Finally, MR upregulated fibroblast growth factor 21 (FGF21) gene and protein levels in both liver and adipose tissues. MRC reversed some of MR’s effects in liver and upregulated the transcription of genes associated with inflammation and carcinogenesis such as Cxcl16, Cdh17, Mmp12, Mybl1, and Cav1 among others. In quadriceps muscle, MR upregulated lipid metabolism-associated genes and increased 3-hydroxybutyrate levels suggesting increased fatty acid oxidation as well as stimulation of gluconeogenesis and glycogenolysis in this tissue. Conclusion: Increased lipid metabolism in inguinal adipose tissue and quadriceps muscle, decreased triglyceride synthesis in liver and the downregulation of inflammation-associated genes are among the factors that could favor the lean phenotype and increased insulin sensitivity observed in MR rats.

The Mitochondrial Free Radical Theory of Aging

“…58 In this line, a recent metabolomic and genomic MetR study found changes in the expression of a large number of genes and proteins that led the authors to conclude that MetR increases lipid metabolism in adipose tissue and muscle, whereas it decreases lipid synthesis in the liver. 59 Therefore, these changes in lipid metabolism seem to be involved in the strong decrease in adiposity and increased insulin sensitivity observed in isocaloric restriction of dietary methionine.…”

“…Interestingly, adding cysteine to the MetR diet reverses most of the studied beneficial changes on adiposity and insulin resistance 60 and increases the transcription of various genes associated with inflammation and carcinogenesis. 59 Therefore, the beneficial changes of MetR diet have been attributed to the decrease of cysteine in serum 60 or liver 59 observed in animals subjected to MetR. On the other hand, excessive intake of dietary methionine is toxic.…”

“…Since it is known that GSSG thiolization of isolated complex I increases its rate of ROS production, 88 a similar reaction of methanethiol or cysteine (which also has a free thiol group available for direct reaction) with complex I thiol groups could be involved in the decrease in mtROSp in MetR. This dietary manipulation decreases hepatic methionine and cysteine 59 and likely methanetiol levels, which can decrease thiolization of complex I subunits and then their rates of mtROSp. Alternatively, cysteine could also interact with the protein cysteines of some of the FeS clusters of the hydrophilic arm of complex I, leading to iron release or availability for reaction and then ROS generation.…”

“…59 In addition, modifications of DNA methylation could be also involved. 89,90 Methionine is an essential amino acid with many key roles in mammalian metabolism including protein synthesis and function, as well as protein and DNA methylation.…”

Dietary l-methionine restriction decreases oxidative stress in porcine liver mitochondria

“…Accumulating evidence suggests that improved mitochondrial function and decrease in mitochondrial reactive oxygen species (ROS) generation might be critical factors that contribute to the effects observed (Perrone et al, 2012(Perrone et al, , 2013. Mitochondria are the major intracellular organelles responsible for biological oxidation in mammalian cells.…”