Disease-related Metabolites in Culture Medium of Fibroblasts from Patients with d-2-Hydroxyglutaric Aciduria, l-2-Hydroxyglutaric Aciduria, and Combined d/l-2-Hydroxyglutaric Aciduria

Abstract

Background: d-2-Hydroxyglutaric aciduria (D-2-HGA), l-2-hydroxyglutaric aciduria (L-2-HGA), and the combined d/l-2-hydroxyglutaric aciduria (D/L-2-HGA) are poorly understood organic acidurias. To investigate the usefulness of cultured human skin fibroblasts for both diagnostic and research purposes, we measured disease-related metabolites in the cell culture medium. Methods: We measured d-2-hydroxyglutarate (D-2-HG), l-2-hydroxyglutarate (L-2-HG), succinate, 2-ketoglutarate, and citrate in fibroblast cell medium by stable-isotope-dilution gas chromatography–mass spectrometry and glutamine, glutamic acid, and lysine with an amino acid analyzer. We used six cell lines from patients with D-2-HGA, two from patients with L-2-HGA, three from patients with D/L-2-HGA, and seven control cell lines. Culture medium was analyzed after a 96-h incubation period. Results: Culture media from cell lines from D-2-HGA patients contained D-2-HG at concentrations 5- to 30-fold higher than media from controls, whereas the concentration of L-2-HG in media was not increased. Media from L-2-HGA cell lines showed a fivefold increase in L-2-HG compared with controls. Media containing fibroblasts from D/L-2-HGA patients contained moderately increased amounts of both D-2-HG and L-2-HG. For all cell lines, succinate concentrations in the blank medium were higher than after 96 h of incubation with the exception of two of three D/L-2-HGA cell lines. Media of D-2-HGA cell lines had 2-ketoglutarate concentrations that were 40% of that for controls. Glutamic acid concentrations in media of these cell lines were 60% lower than in controls. Conclusions: Cell culture media from fibroblasts from patients with D-2-HGA, L-2-HGA, or D/L-2-HGA contain increased amounts the corresponding 2-HGs, demonstrating the suitability of fibroblasts for both diagnosis of and research concerning 2-HGAs.

l‐2‐Hydroxyglutaric aciduria, a disorder of metabolite repair

“…The scarcity of L-2-hydroxyglutarate in nature suggests that it is endogenously produced in humans. This conclusion is consistent with the findings that the excretion of L-2-hydroxyglutarate in patients with L-2hydroxyglutaric aciduria is little affected by the diet and that L-2-hydroxyglutarate accumulates in cultured cells deficient in L-2-hydroxyglutarate dehydrogenase (Struys et al 2003).…”

Metabolism of γ-hydroxybutyrate to d-2-hydroxyglutarate in mammals: further evidence for d-2-hydroxyglutarate transhydrogenase

“…Further studies in animals will be required to determine whether the circulating t 1/2 of d-2-HG is significantly longer than that of GHB (approximately 30-45 minutes). In conclusion, our studies provide evidence for the existence of d-2-hydroxyglutarate transhydrogenase beyond the rat [41] and suggest that this enzyme linked to GHB metabolism may be ubiquitous in mammalian species.…”

L-2-hydroxyglutaric aciduria and brain malignant tumors

“…fibroblasts from patients demonstrates the suitability of these cells for diagnostic and research purposes. 8 The association of L-2-OHGA with brain malignancies has been previously reported in three patients. 9 We report on four additional cases belonging to a series of 10 diagnosed in our institute.…”