A specific method for determination of peroxisomal β-oxidation activity in cultured human skin fibroblasts using a specific substrate, C 9: a possible application for screening of peroxisomal disorders

Abstract

We developed a specific method for direct determination of peroxisomal β-oxidation activity in cultured human skin fibroblasts. When control fibroblasts were incubated with N-(α-methylbenzyl)azelaamic acid (C 9), a specific peroxisomal substrate, C 5 and C 7, the chain-shortened products, were detected with cell concentration and incubation time dependencies and no other products including C 3 were detected. In glutaric aciduria type I and type II fibroblasts, the formation rates of C 2 units liberated from C 9 were almost similar to that in control cells. In contrast to these cell types, the fibroblasts from patient of Zellweger syndrome, in which peroxisomal β-oxidation was impaired, showed no conversion of C 9 to C 5 and C 7. The lack of the C 2 units liberation in Zellweger fibroblasts was not due to an impairment of mitochondrial β-oxidation and/or activation of C 9 to C 9-CoA derivative for subsequent β-oxidation reaction, but rather, appeared to be due to the specific defect of peroxisomal β-oxidation system. These results indicate that C 9 is a useful substrate for the estimation of peroxisomal β-oxidation activity in cultured human skin fibroblasts.