Colorectal Cancers Mimic Structural Organization of Normal Colonic Crypts

Colonic crypts are stereotypical structures with distinct stem cell, proliferating, and differentiating compartments. Colorectal cancers derive from colonic crypt epithelia but, in contrast, form morphologically disarrayed glands. In this study, we investigated to which extent colorectal cancers phenocopy colonic crypt architecture and thus preserve structural organization of the normal intestinal epithelium. A subset of colon cancers showed crypt-like compartments with high WNT activity and nuclear β-Catenin at the leading tumor edge, adjacent proliferation, and enhanced Cytokeratin 20 expression in most differentiated tumor epithelia of the tumor center. This architecture strongly depended on growth conditions, and was fully reproducible in mouse xenografts of cultured and primary colon cancer cells. Full crypt-like organization was associated with low tumor grade and was an independent prognostic marker of better survival in a collection of 221 colorectal cancers. Our findings suggest that full activation of preserved intestinal morphogenetic programs in colon cancer requires in vivo growth environments. Furthermore, crypt-like architecture was linked with less aggressive tumor biology, and may be useful to improve current colon cancer grading schemes.     

Differences between cystic fibrosis and normal cells in the degree of satellite association

Summary The degree of satellite association was found to be significantly higher in phytohemagglutinin (PHA)-stimulated lymphocytes from cystic fibrosis (CF) patients than from those of control individuals. PHA-stimulated lymphocytes from obligatory heterozygotes for the CF mutant allele showed an intermediate degree of satellite association. The degree of satellite association was estimated by the frequency of cells exhibiting associations, by the number of associations per cell, and by the number of chromosomes in an association. The differences in the degree of satellite association were dependent on the concentration of colchicine used for cell arrest. These findings may assist in developing a diagnostic method for the early identification of heterozygotes for the CF allele and for prenatal detection of CF homozygous fetuses.This paper is based on a portion of a dissertation to be submitted by Y. Ravia in partial fulfilment of the Ph. D. requirements in the Graduate School of Tel Aviv University     

Propionate and butyrate dependent bacterial sulfate reduction at extremely haloalkaline conditions and description of Desulfobotulus alkaliphilus sp. nov.

Evidence on the utilization of simple fatty acids by sulfate-reducing bacteria (SRB) at extremely haloalkaline conditions are practically absent, except for a single case of syntrophy by Desulfonatronum on acetate. Our experiments with sediments from soda lakes of Kulunda Steppe (Altai, Russia) showed sulfide production with sulfate as electron acceptor and propionate and butyrate (but not acetate) as an electron donor at a pH 10–10.5 and a salinity 70–180 g l^?1. With propionate as substrate, a highly enriched sulfidogenic culture was obtained in which the main component was identified as a novel representative of the family Syntrophobacteraceae. With butyrate as substrate, a pure SRB culture was isolated which oxidized butyrate and some higher fatty acids incompletely to acetate. The strain represents the first haloalkaliphilic representative of the family Desulfobacteraceae and is described as Desulfobotulus alkaliphilus sp. nov.     

Studies of formation of bivalent copper complexes with native and denatured DNA

The formation of Cu2+ complexes with native and denatured DNA is studied by the methods of differential UV spectroscopy, CD spectroscopy, and viscometry. On ion binding to the bases of native DNA the latter transforms into a new conformation. This transition is accompanied with a sharp increase in UV absorption and a decrease in the intrinsic viscosity though the high degree of helicity persists. Possible sites of Cu2+ ion binding on DNA of various conformations are found along with corresponding constants of complex formation.     

Naturally occurring monoterpenoids in needles of Picea abies (L.) Karst

Summary An analysis was made of the effects of different sampling and extraction techniques on the amounts and pattern of monoterpenoids isolated from needles of Norway spruce. The following isolation and analysis procedure was finally adopted: liquid nitrogen-cooled needles were pulverized by a microdismembrator, extracted with pentane overnight at 2°–3°C and concentrated to a volume not less than 3 ml/g fresh weight on a Vigreux column. The crude extract was injected splitless (with solvent split) onto a cold programmed temperature vaporized (PTV) precolumn of a gas chromatograph and the vaporizable compounds heated to a capillary column. This method was tested for production of artefacts and quantitative extraction and applied to needles of eleven 80-year-old spruce trees.     

Promoters active in mammalian cells (pSV40) and in plants (pNOS) permit expression in yeast of the Tn5 APH(3') II gene

Abstract Four plasmids were constructed by associating Escherichia coli and yeast selection markers and replication origins to a structural gene coding for aminoglycoside phosphotransferase (APH(3')) controlled by different flanking sequences. We used the two bacterial genes of Tn5 (APH(3')II) and Tn903 (APH(3')I) as such and the chimeric pSVneo (APH(3')II) and pNOSneo (APH(3')II) constructs, functional in mammalian and plant cells, respectively. Yeast clones resistant to G418 were obtained with all plasmids except with that bearing the bacterial APH(3')II gene. The three plasmids harbouring the functional APH genes, however, conferred different levels of G418 resistance to yeast.     

Transferrin endocytosis and iron uptake in developing myogenic cells in culture: effects of microtubular and metabolic inhibitors, sulphydryl reagents and lysosomotrophic agents

The experiments described in this study were designed to investigate receptor-mediated endocytosis of transferrin and its role in iron uptake by cultured chick presumptive myoblasts (dividing and non-dividing) and myotubes. The effects of a variety of inhibitors on the internalization of transferrin and iron were investigated and three main effects were found: (i) sulphydryl reagents and microtubular inhibitors reduced the rate of transferrin and iron internalization to similar degrees, (ii) metabolic inhibitors reduced the rate of iron uptake more than that of transferrin endocytosis, and (iii) lysosomotrophic agents almost completely abolished iron accumulation by the cells without any effect on the rate of transferrin internalization. The results suggest that metabolic energy is required not only for the endocytosis of transferrin but also for subsequent steps in the iron uptake process, and that iron release from transferrin occurs in acidified endosomes. Overall, these experiments show that all or virtually all of the iron taken up by developing muscle cells from transferrin occurs as a consequence of receptor-mediated endocytosis of the protein.     

Demonstration of an intact complex of epidermal growth factor and its receptor in the endosomes of A-431 cells

The compartmentalization of the epidermal growth factor (EGF) receptors in A-431 cells was studied using centrifugation of the microsomal fraction of these cells in continuous Percoll gradient. The existence of an intact (non-degraded) EGF receptor in plasma membrane and endosome fraction was demonstrated by electrophoretic analysis of in vitro phosphorylated Percoll fractions. No phosphorylated receptor was revealed in lysosomal fraction by this method. The existence of non dissociated EGF-receptor complexes in intracellular compartments 30 minutes after the start of internalization was proven using a synthesized photoreactive labeled EGF derivative (125I-EGF-SANAH). The removing of pH gradient in organellar membranes by 10 mkM of monensin did not affect dissociation from its receptor. The data obtained proved the existence of non-dissociated and non-degraded EGF-receptor complexes in the endosomal compartment of A-431 cells.     

A universal instinct for designing thermoregulated promotors in gram-positive bacteria]

The construction of plasmid pVKH300, which is useful for modifying any promoter into the thermoregulated form in B. subtilis cells, is presented. The main features of the plasmid are the presence of effectively expressed in B. subtilis lambda C1857 gene and recognition site of BglII restriction enzyme between OR2 and OR3 lambda phage operator sites. Promoterless alpha-amylase gene of B. amyloliquefaciens is used as a reporter gene for promoter cloning into BglII site of pVKH300. Examples of promoter-containing DNA fragments cloning with pVKH300 as vector are presented. It was found that the best regulated promoter, in a plasmid named pVKH332, was cloned in such a way that the distance between central nucleotides of OR2 and OR3 is equal to integer number of DNA helix turns (84 b.p. in the case).