Receptors for carbohydrate ligands including heparin on the cell surface of Leishmania and other trypanosomatids

By employing neoglycoproteins (NGP) and glycosaminoglycans, the detection of endogenous glycoligand-binding sites has become possible. Monitoring specific binding of 11 of these sugar receptor-specific tools, 13 trypanosomatids of monogenetic genera Blastocrithidia, Crithidia, Herpetomonas , and Leptomonas and digenetic genera Endotrypanum , Leishmania , and Sauroleishmania were analysed by agglutination and fluorescence assays. NGP showed agglutination reactions only with the digenetic but not with the monogenetic species. Sensitive flow cytofluorimetric investigations revealed that the apparently different reactivity to NGP is due to a pronounced quantitative difference in expression of binding sites between mono- and digenetic flagellates. Moreover, flow cytofluorimetry was used to demonstrate the occurrence of receptor sites for heparin on the cell surfaces of all trypanosomatids. An indication for a correlation of the binding capacity for the NGP N-acetyl-β-d-glucosamine and heparin to differences in the pathogenicity of parasites was observed for Leishmania donovani as well as Leishmania enriettii . Infective populations of these species contained a significantly higher number of cells which had bound N-acetyl-β-d-glucosamine and heparin than noninfective (long-term in vitro -cultured) populations. The results of the present report additionally support the hypothesis that lectin–carbohydrate interactions with neutral sugar moieties and heparin or heparin-like molecules participate in the interactions between trypanosomatids and host (cells), and that the detected binding sites for carbohydrates and heparin may thus be referred to as potential virulence factors.     

Sequential Role of Hippocampus and Amygdala, Entorhinal Cortex and Parietal Cortex in Formation and Retrieval of Memory for Inhibitory Avoidance in Rats

The hippocampus and amygdala, the entorhinal cortex and the parietal cortex participate, in that sequence, both in the formation and in the expression of memory for a step-down inhibitory avoidance task in rats. Bilateral infusion of AP5 or muscimol caused retrograde amnesia when given O min after training into both hippocampus and amygdala, when given or 180 min after training into the entorhinal cortex, or when given 180 min after training into the parietal cortex. Therefore, memory formation requires the sequential and integrated activity of all these areas mediated by glutamate NMDA receptors in each case. Pre-test administration of CNQX 1 day after training into hippocampus and amygdala, 1 or 31 days after training in entorhinal cortex, or 1, 31 or 60 days after training in the parietal cortex temporarily blocked retention test performance. Therefore, 1 day after training, all these brain structures are necessary for retrieval; 1 month later, the hippocampus and amygdala are no longer necessary for retrieval but the entorhinal and parietal cortex still are; and 60 days after training only the parietal cortex is needed. In all cases the mechanisms of retrieval require intact glutamate AMPA receptors.     

Disposition of leucovorin and its metabolites in dietary folic acid-deplete mice – comparison between tumor, liver and plasma

Purpose: A comprehensive pharmacokinetic study of leucovorin (5-formyltetrahydrofolate, 5-HCO-FH₄) and its metabolites was conducted in plasma, liver and implanted tumor tissue from mice maintained on a low folic acid diet. While it has been previously demonstrated that the antitumor activity of fluorouracil (FU) can be potentiated by 5-HCO-FH₄, the optimum time for administration of FU after 5-HCO-FH₄, to maximally elevate the active folate metabolite methylenetetrahydrofolate in tumor has not been established. Human plasma studies have defined the pharmacokinetics of circulating 5-HCO-FH₄ and its metabolites, but comparison with human tumor accumulation has not been practicable because of sampling difficulties. As an alternative, a mouse model system, based on low dietary folic acid, was used to evaluate plasma, liver and implanted tumor reduced folates after administration of 5-HCO-FH₄.Methods: Plasma and tissue samples were collected from folate-deplete mice over a 12-h period after intraperitoneal administration of 90 mg/kg [R, S ] 5-HCO-FH₄. Reduced folates were evaluated using a ternary complex assay. Results: The time at which max‐imal accumulation of parent compound and all metabolites, except 5-methyltetrahydrofolate (5-CH₃FH₄), occurred in tumor was the same as in plasma. Alternatively, peak liver accumulation was delayed relative to plasma for all folates except 5-CH₃FH₄. Conclusions: The results suggest that mouse plasma accumulation of reduced folates, with the exception of 5-CH₃FH₄, can predict tumor accumulation. Hence, evaluation of human plasma folate accumulation may potentially provide a means to improve the timing of the administration of FU relative to 5-HCO-FH₄ to achieve a superior therapeutic outcome. Received: 24 July 1996 / Accepted: 29 October 1996     

Low dose intradermal vaccination against hepatitis B in mentally retarded patients

Ninety-two patients and 28 staff members of an institute for mentally retarded patients were immunized intradermally with 2 microgram doses hepatitis B vaccine (HB-Vax), initially at month 0 and 1, and after evaluation followed by two booster injections at month 5 and 10, respectively. Blood samples were taken at months 2 and 11. Seroconversion rates of the two groups were similar on both occasions. Titre distributions for mentally retarded patients and staff members corrected for age, side effects and sex using multiple regression were significantly different at month 2 (p = 0.05) but not at month 11. Geometric mean titres for mentally retarded patients and staff members were 39 and 67 at month 2, respectively, and 1082 and 672 IU l-1 at month 11, respectively. Percentage anti-HBs/a(w) differed significantly between patients and staff members. Side effects, largely local (pigmentation), were observed with a frequency of about 35% in both mentally retarded patients and staff members. Since 92% of the mentally retarded patients had anti-HBs titres greater than 10 IU l-1 and approximately equal to 80% greater than 100 IU l-1, low dose vaccination, four injections by intradermal route, seems to be a suitable cost-reducing immunization system in institutes for mentally retarded patients.     

Sequence analysis of L RNA of Lassa virus

The L RNA of three Lassa virus strains originating from Nigeria, Ghana/Ivory Coast, and Sierra Leone was sequenced and the data subjected to structure predictions and phylogenetic analyses. The L gene products had 2218-2221 residues, diverged by 18% at the amino acid level, and contained several conserved regions. Only one region of 504 residues (positions 1043-1546) could be assigned a function, namely that of an RNA polymerase. Secondary structure predictions suggest that this domain is very similar to RNA-dependent RNA polymerases of known structure encoded by plus-strand RNA viruses, permitting a model to be built. Outside the polymerase region, there is little structural data, except for regions of strong alpha-helical content and probably a coiled-coil domain at the N terminus. No evidence for reassortment or recombination during Lassa virus evolution was found. The secondary structure-assisted alignment of the RNA polymerase region permitted a reliable reconstruction of the phylogeny of all negative-strand RNA viruses, indicating that Arenaviridae are most closely related to Nairoviruses. In conclusion, the data provide a basis for structural and functional characterization of the Lassa virus L protein and reveal new insights into the phylogeny of negative-strand RNA viruses.     

Treatment of motoneuron degeneration by intracerebroventricular delivery of VEGF in a rat model of ALS

Neurotrophin treatment has so far failed to prolong the survival of individuals affected with amyotrophic lateral sclerosis (ALS), an incurable motoneuron degenerative disorder. Here we show that intracerebroventricular (i.c.v.) delivery of recombinant vascular endothelial growth factor (Vegf) in a SOD1(G93A) rat model of ALS delays onset of paralysis by 17 d, improves motor performance and prolongs survival by 22 d, representing the largest effects in animal models of ALS achieved by protein delivery. By protecting cervical motoneurons, i.c.v. delivery of Vegf is particularly effective in rats with the most severe form of ALS with forelimb onset. Vegf has direct neuroprotective effects on motoneurons in vivo, because neuronal expression of a transgene expressing the Vegf receptor prolongs the survival of SOD1(G93A) mice. On i.c.v. delivery, Vegf is anterogradely transported and preserves neuromuscular junctions in SOD1(G93A) rats. Our findings in preclinical rodent models of ALS may have implications for treatment of neurodegenerative disease in general.     

Serum concentrations of oestradiol-17beta, progesterone, relaxin and chorionic gonadotrophin during blastocyst implantation in natural pregnancy cycle and in embryo transfer cycle in the rhesus monkey

The present study was undertaken to assess the temporal association between the profiles of serum concentrations of oestradiol-17beta, progesterone, chorionic gonadotrophin (CG) and relaxin in pregnancies established naturally, and after embryo transfer, as well as in failed pregnancies in rhesus monkeys. In naturally mated cycles (group 1) a conception rate of 75% was obtained. In group 1, the mean day of CG detection in serum was 11.5 +/- 1.9 day post-ovulation, and for relaxin, 9.0 +/- 2.5 day post-ovulation. In group 2, embryo transfer to synchronous, non-mated surrogate recipients was performed; seven embryo transfer cycles yielded three pregnancies which were allowed to continue to term and normal infants were delivered. In embryo transfer cycles the mean day of CG detection was 14.8 +/- 1.8 day post- ovulation, and for relaxin, 11.4 +/- 2.6 day post-ovulation. A delay of about 3 days was observed in the appearance in circulation of CG (P < 0.05) and also of relaxin (P < 0.05) between natural mated and embryo transfer conception cycles. Significant differences (P < 0.05 for progesterone and P < 0.03 for oestradiol) were obtained for the areas under the curves for progesterone and oestradiol between days 12 and 16 in conception cycles compared with failed pregnancies. These data provide the first observation of the normal hormonal signals associated with maternal recognition of transferred embryos during the peri- implantation period, and suggest that the use of such an experimental primate embryo transfer model may help to elucidate components of maternal and embryonic signal-response mechanisms during embryo implantation.