Glucoraphasatin: Chemistry, occurrence, and biological properties

Glucoraphasatin is an atypical glucosinolate mainly found in Raphanus sativus roots and sprouts. This review focuses on the chemistry, the occurrence, and the biological properties of glucoraphasatin.     

Liver Accumulation of Plasmodium chabaudi-Infected Red Blood Cells and Modulation of Regulatory T Cell and Dendritic Cell Responses

It is postulated that accumulation of malaria-infected Red Blood Cells (iRBCs) in the liver could be a parasitic escape mechanism against full destruction by the host immune system. Therefore, we evaluated the in vivo mechanism of this accumulation and its potential immunological consequences. A massive liver accumulation of P. c. chabaudi AS-iRBCs (Pc-iRBCs) was observed by intravital microscopy along with an over expression of ICAM-1 on day 7 of the infection, as measured by qRT-PCR. Phenotypic changes were also observed in regulatory T cells (Tregs) and dendritic cells (DCs) that were isolated from infected livers, which indicate a functional role for Tregs in the regulation of the liver inflammatory immune response. In fact, the suppressive function of liver-Tregs was in vitro tested, which demonstrated the capacity of these cells to suppress naive T cell activation to the same extent as that observed for spleen-Tregs. On the other hand, it is already known that CD4+ T cells isolated from spleens of protozoan parasite-infected mice are refractory to proliferate in vivo. In our experiments, we observed a similar lack of in vitro proliferative capacity in liver CD4+ T cells that were isolated on day 7 of infection. It is also known that nitric oxide and IL-10 are partially involved in acute phase immunosuppression; we found high expression levels of IL-10 and iNOS mRNA in day 7-infected livers, which indicates a possible role for these molecules in the observed immune suppression. Taken together, these results indicate that malaria parasite accumulation within the liver could be an escape mechanism to avoid sterile immunity sponsored by a tolerogenic environment.     

Isolation of high molecular weight DNA from wholeEuglena cells

Summary A method for isolating high quality DNA from wholeEuglena cells is described. The procedure consists in: the weakening of the cell pellicle in glycerol avoiding the mechanical disruption of cells and shearing damage in DNA molecules; the decondensation ofEuglena compact chromatin directly inside the cells; the complete dissociation of cells and nucleoproteins in sarkosyl detergent; the optional digestion of proteins and RNA with DNase-free enzymes and the final purification of DNA by isopycnic banding in CsCl gradients. Degradation of DNA is prevented all along the extraction procedure by glycerol, antioxydants, EDTA and sarkosyl detergent. Using the enzymatic digestion step, DNA containing few single-stranded nicks is obtained with a yield approaching 100%. DNA with no single-stranded nick could be obtained with a 35% yield when the enzymatic digestion step was omitted. In both cases, the double-stranded DNA has an average molecular weight equal or greater than 6×10⁷. It is free of contaminants and could be easily digested with restriction enzymes. After digestion with Eco RI and size-fractionation in agarose gel this DNA has permitted specific hybridization of the rDNA sequences with a radioactive rRNA probe.Abbreviations Kbp kilobasepairs - Kb kilobases     

Comparative mutagenicity testing of a drug candidate, U-48753E: mechanism of induction of gene mutations in mammalian cells and quantitation of potential hazard

U-48753E is a potential human drug which was subjected to a battery of short-term assays for genetic activity. The compound was negative in the Salmonella (Ames) test, the in vitro UDS assay, the mouse bone-marrow micronucleus test and the Drosophila sex-linked recessive lethal assay. However, it was weakly positive in the CHO/HPRT assay in the presence of metabolic activation (S9). The weak positive response might easily have been labeled artifactual since there was no dose response and the dose level producing positive findings varied from experiment to experiment. In addition, the weak positive response was not confirmed in V79 cells. However, a reproducible dose-related increase in mutants was observed in the AS52/XPRT assay in the presence of S9. Metabolism of this drug proceeds through conversion of aliphatic N-methyl groups to formaldehyde. Addition of formaldehyde dehydrogenase to the S9 resulted in elimination of the mutagenicity of the compound in AS52 cells. Thus, the mutants were probably induced by formaldehyde. From the endogenous levels of formaldehyde in human blood, and the limiting potential therapeutic dose levels, the genotoxic hazard associated with U-48753E is marginal. This assessment of risk and its quantitation depend upon an understanding metabolism and exposure limits imposed by known side effects of the drug. This study can serve as a model for quantitative genetic risk assessment when mutagenicity is due to N-demethylation and formation of formaldehyde in situ.     

Flash fluorometer made from off-the-shelf photographic equipment to measure tissue levels of fluorescein

A device to measure fluorescein in tissue has been constructed from standard photographic equipment--an electronic strobe and a flashmeter both covered with interference filters. The instrument works well in the light and need not touch the area being measured, an advantage over existing fluorometers. The instrument has been used to measure the amount of dye in flaps in rats, pigs, and three humans. The results revealed that the amount of dye in a freshly made flap was rarely as much as in normal skin, and skin with less than 20 percent of the dye of control areas usually sloughed, although there were exceptions. In the future the instrument will be improved, and its readings will be compared to those obtained from radioactive microspheres, the present "gold standard" of techniques to measure vascularity. The instrument can be used to estimate the blood supply to any tissue and seems to be as reliable as the dermofluorometers already on the market.     

Sertoli and Leydig cell numbers and gonadotropin receptors in rat testis from birth to puberty

Summary In testes of rats from 2 to 60 days of age, we examined the number of Sertoli cells (SC) and Leydig cells (LC) as well as the binding of radioiodinated gonadotropins to frozen sections and homogenates. The number of SC per testis increased only during the first 2 postnatal weeks, whereas that of LC was stable up to days 7–10 and increased thereafter. The uptake of ¹²⁵I-labelled human follicle-stimulating hormone (¹²⁵I-FSH) to frozen sections was confined to sex cords or seminiferous tubules, while that of ¹²⁵I-labelled human choriogonadotropin (¹²⁵I-hCG) matched the distribution of LC in the interstitium. High affinity receptors for FSH and hCG were found in homogenates at all stages studied. The number of FSH receptors per testis increased steadily, whereas that of hCG receptors was low until days 7–10 and rose afterwards. Thus, SC in rat testis appear to proliferate in the presence of fetal LC during the first 2 postnatal weeks and to differentiate concomitantly with the emergence of the adult LC generation after day 10. The complement of FSH receptors in SC remains constant as they proliferate and increases after day 21 as they differentiate. The hCG receptor number is relatively fixed in each LC generation, being higher in adult compared to fetal LC.     

Suramin, a novel antitumor compound

Suramin, a polyanionic compound originally synthesized for use as an antiparasitic agent, has recently entered clinical trials for the treatment of a variety of human cancers refractory to conventional modalities of therapy. This is based on suramin's ability to bind and to inactivate growth factor and enzyme systems critical to cellular homeostasis and proliferation. In addition, this compound possesses adrenocorticolytic properties in vivo and exerts significant cytostatic and cytocidal effects against a variety of human tumor cell lines in vitro. Pilot studies using suramin have thus far been conducted in adrenocortical carcinoma, prostate cancer refractory to conventional hormonal manipulation and nodular lymphomas.     

Incidence of sensitisation to the pollens of urticaceae (Parietaria), Poaceae and Oleaceae (Olea europea) and pollen rain in Liguria (Italy)

Summary The authors examined 734 sensitised patients living in four localities in Liguria (Genoa, Savona, Pietra Ligure and Sanremo). The commonest source of sensitisation (62.7%) was Urticaceae (Parietaria), followed by Poaceae (52.5%) andOlea europaea L. (24.0%). A survey of airborne pollens revealed a greater presence of Urticaceae and Poaceae in Genoa and of Oleaceae in Pietra Ligure and Sanremo.     

1H- and13C-NMR spectroscopic studies of lipid extracts of the red algaGracilaria longa

Lipid extracts of the red algaGracilaria longa were studied by¹H- and¹³C-NMR spectroscopy. Peaks in the¹³C-NMR spectra attributable to sterols, chlorophylls and carotenoids allowed free and acylated cholesterol, chlorophylla and lutein to be identified as the most abundant components of these classes. A content of 0.5 ± 0.1 μmoles of total cholesterol/g wet alga was estimated from the¹H-NMR spectrum, which also allowed the determination of the phosphatidylcholine/total lipid molar ratio (9.5 ± 0.5%). The¹³C-NMR spectroscopic experiments provided information on the position of the double bonds on the fatty acid residues. A comparison between NMR spectra of lipid extracts obtained for wet and dried alga showed that the alga undergoes both a dramatic peroxidation and some glycolipid degradation during the drying process.     

Localization of the high affinity calcium-binding site on tubulin molecule

Tubulin is a calcium-binding protein. Two different modes of interaction of calcium with tubulin have been described: a high affinity interaction to one or two binding sites and lower affinity interactions to several other binding sites. In the present study, we have used limited proteolysis of tubulin with trypsin, chymotrypsin, and subtilisin to localize the high affinity calcium-binding sites. Our results indicate that two sites are located in the carboxyl-terminal region of both tubulin subunits, and that tubulin deprived of its carboxyl-terminal region is able to polymerize in the presence of 0.5 mM calcium.