RESUMEN
The control of phosphatidylcholine biosynthesis in the hamster liver was examined. Livers of hamsters fasted for 24 and 48 h were perfused with labelled choline. Under both fasting conditions, the incorporation of labelled choline into phosphatidylcholine was reduced. After 48 h of fasting, the 52% reduction in phosphatidylcholine biosynthesis was caused by changes in several factors including a diminishing rate of choline uptake and severe reductions in the pool sizes of ATP and CTP (to 33-37% control values) which resulted in a decrease in the pools of choline-containing metabolites. The activation of cytidylyltransferase after 48 h of fasting might be regarded as a compensatory mechanism for the maintenance of phosphatidylcholine biosynthesis. After 24 h of fasting, a 25% reduction in phosphatidylcholine biosynthesis was observed. The ATP and CTP levels were decreased but the reduction was not severe enough to affect the choline uptake or the labelling of the phosphocholine fraction. The activities of the cytidylyltransferase remained unchanged but an accumulation of labelled CDP-choline was detected. Although choline-phosphotransferase activity was not changed in the microsomes, the enzyme activity was attenuated in the postmitochondrial fraction. Further analysis revealed that cholinephosphotransferase in the liver was inhibited by an endogenous inhibitor in the cytosol which was later identified as argininosuccinate. The level of argininosuccinate was elevated during fasting and the change quantitatively accounted for the attenuation of cholinephosphotransferase activity. The inhibition of choline-phosphotransferase by argininosuccinate, coupled with a substantial decrease in the diacylglycerol level, would provide the hamster liver with an immediate mechanism for the transient modulation of phosphatidylcholine biosynthesis during short-term fasting.
Asunto(s)
Ácido Argininosuccínico/farmacología , Diacilglicerol Colinafosfotransferasa/metabolismo , Privación de Alimentos/fisiología , Hígado/enzimología , Fosfatidilcolinas/metabolismo , Animales , Ácido Argininosuccínico/aislamiento & purificación , Compartimento Celular , Colina/metabolismo , Colina Quinasa/análisis , Citidililtransferasa de Colina-Fosfato , Cricetinae , Diacilglicerol Colinafosfotransferasa/antagonistas & inhibidores , Diacilglicerol Colinafosfotransferasa/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Masculino , Mesocricetus , Nucleotidiltransferasas/análisis , Fracciones Subcelulares/enzimologíaRESUMEN
Argininosuccinate (ASA) is determined in a few minutes with little manipulation by reversed phase high-performance liquid chromatography (HPLC) using O-phthaldialdehyde. The two cyclic anhydrides of ASA are not formed during analysis but, if present, can be determined simultaneously. As little as 1 nl urine from a patient with argininosuccinic aciduria was sufficient for analysis; the ASA/creatinine ratio was 50.8 mmol g-1 and daily excretion was 5-7g ASA. We found small amounts of the two anhydrides in the patient's urine and we give factors to estimate, from their peaks, the corresponding amount of ASA. Urine from normal children showed a small acid-labile (at 100 degrees C) peak at the ASA position, which we tentatively assign to genuine ASA. From this peak less than 2mg ASA day-1 were excreted in our controls. Procedures for collection and storage of samples and the potential of this method for heterozygote detection are discussed.