A comparison of metabolic pathway dynamics in man and other mammals
Baumgarten, Ingrid M
MetadataShow full item record
The object of the present study was to determine whether there was a difference in the flux through two different metabolic pathways, the purine salvage pathway and the urea cycle, in skin fibroblasts from three species. A double label approach was adopted to measure the incorporation of the purine bases, hypoxanthine and adenine, and the amino acids, citrulline and arginine in acid precipitable material. Before examining the between species variation it was necessary to examine different levels of possible variation such as the variation between individuals from each species, the variation between separate experiments and the replication error within experiments. Eight individuals from each of three species, bUffalo, human, and rhinoceros were examined with the labelled purine bases. Skin fibroblasts from four humans and eight rhinoceros individuals were also examined over varying passage numbers until the fibroblasts senesced, to determine the effect of ageing on the uptake of hypoxanthine and adenine. The same four human fibroblast cultures were transformed with a transforming virus and examined to see the effects of transformation on the uptake of the purine bases, these transformed fibroblasts were compared with previously transformed rhinoceros fibroblasts. The uptake of labelled citrulline and arginine was also examined in three individuals from each of the three species. The major part of variation throughout the study was found to be at the between experiment level, despite stringently controlled conditions. This between experiment variation obscured any variation found within individuals from each species. In spite of this major between - experiment variation, the results showed that there was significant variation between the three species in the uptake of hypoxanthine. Adenine uptake was similar in the buffalo and human, but was significantly different between both these species and the rhinoceros. citrulline uptake showed no variation between the three species, whereas arginine showed a significant variation between the rhinoceros and the other two species. Buffalo and human showed no significant variation in arginine uptake. There was a significant increase in the uptake of hypoxanthine and adenine in transformed fibroblasts relative to untransformed fibroblasts. As a consequence of the significant between-experiment variation demonstrated in this study, it is apparent that great care must be taken to standardize the conditions when using a double label approach, especially if the assay is to be used for the diagnosis of inborn errors of metabolism.