Stable-
isotope tracer experiments performed in vitro are evaluated for their utility in differentiating between
pyruvate dehydrogenase and
cytochrome oxidase deficiencies, two of several
enzyme defects commonly associated with the lactic acidemias. Fibroblasts of
enzyme-deficient individuals and of age-matched controls are grown in medium containing [U-13C]
glucose. Direct analysis of cells and
conditioned culture medium provides only minor differences in the organic
acid/amino acid GC-MS profiles, making differentiation of
enzyme defects difficult by this method. However, differences have been found in the
glucose turnover into various cell metabolites, making differentiation of these two
enzyme defects possible. The cellular pool of
glutamic acid experiences 13C-enrichment in both the control and
cytochrome oxidase deficient lines, but not in the
pyruvate dehydrogenase-deficient line. The cellular pool of an unknown, possibly an aminopentose
sugar, on the other hand, experiences 13C-enrichment in the
pyruvate dehydrogenase and control lines, but not in the
cytochrome oxidase line. These observations, as well as other differences in the extent of enrichment into various metabolite pools, suggest that this stable-
isotope approach, in vitro, is feasible and may allow these two
enzyme defects to be differentiated in a definitive manner. Such stable-
isotope experiments are easy to carry out with cultured cells and are inexpensive. Applications of the technique to other
genetic disorders might be appropriate.