Norbert Blanckaert

Differential regulation of UDP-GlcUA transport in endoplasmic reticulum and in Golgi membranes.

BACKGROUND In the endoplasmic reticulum (ER), the stimulation of UDP-glucuronosyltransferase (UGT) by UDP-GlcNAc is based on the interaction of transport across the ER membrane of UDP-GlcUA with UDP-GlcNAc. Intramicrosomal UDP-GlcNAc stimulates influx of UDP-GlcUA and thereby enhances delivery of UDP-GlcUA to the catalytic center of UGT in the ER lumen. AIM The aim of this study is to investigate whether the interactions between nucleotide sugars for transport across the ER membrane also occur in the Golgi apparatus, and thereby affect UGT activity in Golgi membranes. RESULTS We found that Golgi membrane preparations display UGT activity which, unlike in ER membranes, is not stimulated by UDP-GlcNAc. Efflux of intravesicular UDP-GlcNAc and UDP-Xyl marginally enhanced uptake of UDP-GlcUA in Golgi vesicles; such trans-stimulation was much more pronounced in the ER. Efflux of intravesicular UDP-GlcNAc was strongly trans-stimulated by cytosolic UDP-GlcUA in ER-derived vesicles but less so in Golgi-derived vesicles. CONCLUSION The interaction between transport of UDP-GlcUA and transport of UDP-GlcNAc or UDP-Xyl is different in Golgi vesicles compared with ER vesicles. This finding is consistent with the different effects of UDP-GlcNAc on glucuronidation in Golgi and ER.

Serum Bilirubin Mono- and Diconjugates in Human Newborns

Several alterations of bilirubin metabolism are present during the neonatal period, all essentially leading to an increased load of unconjugated bilirubin and a decreased hepatic handling of the pigment. The resulting neonatal jaundice is a classical example of unconjugated hyperbilirubinemia(1). The presence of conjugated bilirubin in neonatal plasma has often been investigated, and considered as an index of function of hepatic conjugation or secretion(2–5). Unfortunately, several methodological problems are met when trying to measure serum conjugated bilirubin in the presence of high concentrations of the unconjugated pigment. Most procedures for measurement of bilirubins in biological samples rely on diazo-cleavage, resulting in splitting of the tetrapyrrole moiety and producing dipyrrolic red azoderivatives. After the discovery that bilirubin in bile reacts rapidly, without requiring the presence of an “accelerator”, the distinction between direct-reacting and total bilirubin was introduced. However, “direct-reacting” bilirubin is just operationally defined, and it is therefore dependent on reaction conditions and sample composition, while it does not accurately reflect conjugated bilirubin(6). The ethyl-anthranilate method is more specific, and allows to separate and identify the dipyrrole azoderivatives(6). However, it is impossible to relate the sugar-conjugated azodipyrroles to the parent rubins. A similar problem is present with the isotope-dilution method(2). Direct analysis of the tetrapyrroles has been tried by various solvent-partition and chromatographic procedures, but none of these methods proved to be truly specific and accurate(6).

Clinical Significance of Recent Developments in Serum Bilirubins

Normal serum contains unconjugated bilirubin and small concentrations of bilirubin glucuronides (approximately 4% of total bilirubins). A second conjugated form of bilirubin, tentatively identified as bilirubin-protein conjugates, with pharmacokinetic properties that are clearly distinct from those of bilirubin glucuronides, occurs in serum of patients in whom hepatic elimination of bilirubin glucuronides is, or recently has been, impaired.