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Placental protein 13 (PP13) was cloned from human term placenta. As sequence analyses, alignments and computational modelling showed its conserved structural and functional homology to members of the galectin family, the protein was designated galectin‐13. Similar to human eosinophil Charcot–Leyden crystal protein/galectin‐10 but not other galectins, its weak lysophospholipase activity was confirmed by ³¹P‐NMR. In this study, recombinant PP13/galectin‐13 was expressed and specific monoclonal antibody to PP13 was developed. Endogenous lysophospholipase activity of both the purified and also the recombinant protein was verified. Sugar binding assays revealed that N‐acetyl‐lactosamine, mannose and N‐acetyl‐glucosamine residues widely expressed in human placenta had the strongest binding affinity to both the purified and recombinant PP13/galectin‐13, which also effectively agglutinated erythrocytes. The protein was found to be a homodimer of 16 kDa subunits linked together by disulphide bonds, a phenomenon differing from the noncovalent dimerization of previously known prototype galectins. Furthermore, reducing agents were shown to decrease its sugar binding activity and abolish its haemagglutination. Phosphorylation sites were computed on PP13/galectin‐13, and phosphorylation of the purified protein was confirmed. Using affinity chromatography, PAGE, MALDI‐TOF MS and post source decay, annexin II and beta/gamma actin were identified as proteins specifically bound to PP13/galectin‐13 in placenta and fetal hepatic cells. Perinuclear staining of the syncytiotrophoblasts showed its expression in these cells, while strong labelling of the syncytiotrophoblasts' brush border membrane confirmed its galectin‐like externalization to the cell surface. Knowing its colocalization and specific binding to annexin II, PP13/galectin‐13 was assumed to be secreted to the outer cell surface by ectocytosis, in microvesicles containing actin and annexin II. With regard to our functional and immunomorphological results, PP13/galectin‐13 may have special haemostatic and immunobiological functions at the lining of the common feto‐maternal blood‐spaces or developmental role in the placenta.

ABCG2, a transporter of the ATP-binding cassette family, is known to play a prominent role in the absorption, distribution, metabolism, and excretion of xenobiotics. Drug-transporter interactions are commonly screened by high-throughput systems using transfected insect and/or human cell lines. The determination of ABCG2-ATPase activity is one method to identify ABCG2 substrate and inhibitors. We demonstrate that the ATPase activities of the human ABCG2 transfected Sf9 cell membranes (MXR-Sf9) and ABCG2-overexpressing human cell membranes (MXR-M) differ. Variation due to disparity in the glycosylation level of the protein had no effect on the transporter. The influence of cholesterol on ABCG2-ATPase activity was investigated because the lipid compositions of insect and human cells are largely different from each other. Differences in cholesterol content, shown by cholesterol loading and depletion experiments, conferred the difference in stimulation of basal ABCG2-ATPase of the two cell membranes. Basal ABCG2-ATPase activity could be stimulated by sulfasalazine, prazosin, and topotecan, known substrates of ABCG2 in cholesterol-loaded MXR-Sf9 and MXR-M cell membranes. In contrast, ABCG2-ATPase could not be stimulated in MXR-Sf9 or in cholesterol-depleted MXR-M membranes. Moreover, cholesterol loading significantly improved the drug transport into inside-out membrane vesicles prepared from MXR-Sf9 cells. MXR-M and cholesterol-loaded MXR-Sf9 cell membranes displayed similar ABCG2-ATPase activity and vesicular transport. Our study indicates an essential role of membrane cholesterol for the function of ABCG2.

Objective – Excitatory amino acid receptors are involved in the normal physiology of the brain, and may play a role in the pathogenesis of neurological disorders such as Huntington's disease, Parkinson's disease, amyotrophic lateral sclerosis, etc. It has been demonstrated that the blockade of one of these receptors ameliorates the symptoms of experimental allergic encephalomyelitis, an animal model of multiple sclerosis (MS). In a recent study, a decreased level of kynurenic acid was found in the cerebrospinal fluid of patients with MS. The only known endogenous excitotoxin receptor antagonist is the tryptophan metabolite kynurenic acid. Another metabolite is quinolinic acid, which exerts different action: it is an excitotoxin receptor agonist. The ratio of these two metabolites is determined by the activities of kynurenine aminotransferase I and II (KAT I and KAT II). In this study, we measured the activities of these enzymes and the concentration of kynurenic acid in the red blood cells (RBC) and in the plasma of patients with MS. KAT activities were detected both in the RBC and in the plasma. As compared with the control subjects, the KAT I and KAT II activities were significantly higher in the RBC of the patients. The concentration of kynurenic acid is elevated in the plasma of MS patients, and there is a tendency to an elevation in the RBC. These changes may indicate a compensatory protective mechanism against excitatory neurotoxic effects. Our data demonstrate the involvement of the kynurenine system in the pathogenesis of MS, which may predict a novel therapeutic intervention.