EFFECTS OF MAPLE SYRUP URINE DISEASE METABOLITES ON MOUSE l-FIBROBLASTS IN VITRO: A FINE STRUCTURAL AND BIOCHEMICAL STUDY

—The branched-chain amino and ketoacids i.e. l -leucine, l -isoleucine, l -valine, alpha-ketoisocaproic acid (AKICA), alpha-keto-beta-methylvaleric acid (AKBMVA) and alpha-ketoisovaleric acid (AKIVA)] were administered to mouse strain l fibroblasts in tissue culture in an attempt to study the effects of increased levels of the compounds in an in vitro system. All of these compounds are found to be elevated in the blood of patients with Maple Syrup Urine Disease (MSUD). With AKICA, l -leucine, AKIVA and AKBMVA, there was a decreased growth rate at concentrations of 10 to 30 times the levels found in Maple Syrup Urine Disease. Combined administration of the above six compounds at the maximum blood levels noted in MSUD produced a significantly decreased growth rate. Electron microscopic studies revealed numerous annulate lamellae in cells treated with AKICA and in those treated with a combination of all six MSUD compounds. AKICA-treated cells contained elevated concentrations per cell of free fatty acids, triglycerides, sterols and some classes of phospholipids. Isotope labelling experiments were performed using U-¹⁴C] AKICA and ³H]isoleucine, which were added to l -cell suspension cultures containing various levels of unlabelled AKICA. Labelled AKICA and isoleucine were both taken up by the cells. The net uptake of isoleucine was inhibited by AKICA in concentrations found in MSUD. Folch-Lees extraction of cells treated with labelled AKICA revealed increased ¹⁴C counts only in the lower lipid phase. The growth inhibition and annulate lamellae observed with AKICA treatment may be due to an arrest of the cells in phase G₁ of the cell growth cycle, possibly due to decreased isoleucine uptake. It is proposed that a similarly-mediated arrest in the proliferation of oligodendroglial cells during the critical period of myelination gliosis might account for the myelination abnormalities reported in MSUD.