Traumatic Brain Injury Alters Methionine Metabolism: Implications for Pathophysiology

Abstract

Methionine is an essential proteinogenic amino acid that is obtained from the diet. In addition to its requirement for protein biosynthesis, methionine is metabolized to generate metabolites that play key roles in a number of cellular functions. Metabolism of methionine via the transmethylation pathway generates S-adenosylmethionine (SAM) that serves as the principal methyl (-CH3) donor for DNA and histone methyltransferases (MTs) to regulate epigenetic changes in gene expression. SAM is also required for methylation of other cellular proteins that serve various functions and phosphatidylcholine synthesis that participate in cellular signaling. Under conditions of oxidative stress, homocysteine (which is derived from SAM) enters the transsulfuration pathway to generate glutathione, an important cytoprotective molecule against oxidative damage. As both experimental and clinical studies have shown that traumatic brain injury (TBI) alters DNA and histone methylation and causes oxidative stress, we examined if TBI alters the plasma levels of methionine and its metabolites in human patients. Blood samples were collected from healthy volunteers (HV; n = 20) and patients with mild TBI (mTBI; GCS > 12; n = 20) or severe TBI (sTBI; GCS < 8; n = 20) within the first 24 h of injury. The levels of methionine and its metabolites in the plasma samples were analyzed by either liquid chromatography-mass spectrometry or gas chromatography-mass spectrometry (LC-MS or GC-MS). sTBI decreased the levels of methionine, SAM, betaine and 2-methylglycine as compared to HV, indicating a decrease in metabolism through the transmethylation cycle. In addition, precursors for the generation of glutathione, cysteine and glycine were also found to be decreased as were intermediate metabolites of the gamma-glutamyl cycle (gamma-glutamyl amino acids and 5-oxoproline). mTBI also decreased the levels of methionine, α-ketobutyrate, 2 hydroxybutyrate and glycine, albeit to lesser degrees than detected in the sTBI group. Taken together, these results suggest that decreased levels of methionine and its metabolic products are likely to alter cellular function in multiple organs at a systems level.

Keywords: S-adenosylmethionine; concussion; epigenetic changes; metabolomics; protein methylation; transsulfuration.

Figure 1

Severe traumatic brain injury (sTBI) reduces plasma methionine and metabolism through the transmethylation pathway. (A) Schematic showing the metabolism of methionine via the transmethylation pathway. Molecules detected and measured are presented in bold text. Key enzymes are indicated by italic text. The plasma levels of (B) methionine were significantly reduced as a result of TBI. (C) S-adenosylmethionine (SAM) was significantly reduced in sTBI compared to healthy volunteers (HV) and patients with a mild TBI (mTBI). When the data were segregated by gender, similar changes in both (D) methionine and (E) SAM were observed in males and females. MAT: methionine adenosyltransferase; MTs: methyltransferases; SAHase: S-adenosyl-L-homocysteine hydrolase. Horizontal bar indicates mean. *p < 0.05.

Figure 2

sTBI reduces the levels of methyl donors for methionine remethylation. (A) Schematic showing the methylation of homocysteine to generate methionine. Molecules detected and measured are presented in bold text. Key enzymes are indicated by italic text. The plasma levels of the methyl donors (B) choline and (C) betaine used in methionine remethylation were significantly reduced in sTBI. (D) The side-product of methionine regeneration, dimethylglycine, is significantly reduced as a result of sTBI. BHMT: betaine-homocysteine S-methyltransferase. Horizontal bar indicates mean. *p < 0.05.

Figure 3

TBI alters methionine metabolism through the transsulfuration pathway. (A) Drawing of the transsulfuration pathway in which homocysteine is converted to cysteine and alpha-ketobutyrate. Alpha-ketobutyrate is reduced to generate 2-hydroxybutyrate. Molecules detected and measured in the current study are presented in bold text. Key enzymes are indicated by italic text. Although the plasma levels of both (B) alpha-ketobutyrate and (C) 2-hydroxybutyrate were significantly elevated in the plasma of sTBI patients compared to other groups, the plasma levels of (D) cysteine were significantly decreased in sTBI patients. In contrast, the levels of cysteine were significantly increased in the mTBI group. HV: healthy volunteers; mTBI: mild TBI patients; CBS: cystathionine-β-synthase; CSE: cystathionine γ-lyase; α-HBDH: alpha-hydroxybutyrate dehydrogenase. Horizontal bar indicates mean. *p < 0.05.

Figure 4

Plasma glycine, but not glutamate levels are reduced as a result of sTBI. (A) Simplified drawing showing the production of glutathione from cysteine, glycine and glutamate. Glutamate cysteine ligase (GCL) catalyzes the reaction between glutamate and cysteine to produce gamma-glutamylcysteine. Glutathione synthase (GS) converts gamma-glutamylcysteine and glycine to generate glutathione. Molecules detected and measured in the current study are presented in bold text. Key enzymes are indicated by italic text. (B) Glutamate levels did not change across any of the subject groups. Horizontal bar indicates mean. (C) Plasma glycine levels in individual subjects from HV mTBI, and sTBI are shown. The levels of glycine are significantly reduced in the plasma of sTBI patients compared to all other groups. *p < 0.05.

Figure 5

TBI reduces glutathione recycling through the gamma-glutamyl cycle. (A) Simplified schematic showing the enzymes and intermediates for the gamma-glutamyl cycle. Molecules detected and measured in the current study are presented in bold text. Key enzymes are indicated by italic text. The levels of the gamma-glutamyl amino acids (B) gamma-glutamylvaline and (C) gamma-glutamylleucine, and the intermediate (D) 5-oxoproline are significantly reduced in the plasma of sTBI patients compared to HV, and mTBI groups. GGT: gamma-glutamyl transpeptidase; GGCT: gamma-glutamyl cyclotransferase. Horizontal bar indicates mean. *p < 0.05.

Figure 6

Summary illustration of methionine metabolic pathways. Metabolites of the transmethylation pathway are indicated in red, the transsulfuration pathway is indicated in blue, and the gamma-glutamyl cycle is shown in green. Black arrowhead (▴) indicates changes in metabolite levels in the plasma of sTBI patients. Open arrows (▵) indicate significant changes measured in the mTBI patients as compared to HV. Up arrowheads indicate increased levels, whereas down arrowheads indicate decreased levels.