Synthesis of lysine-containing fragments of the Proteus mirabilis O27 O-specific polysaccharide and neoglycoconjugates therefrom.

Amide-linked lysine mono- and di-uronic acid fragments of the O-specific polysaccharide from P. mirabilis O27 have been synthesised. N epsilon-Boc-L-lysine tert-butyl ester was condensed with 2-azidoethyl glycosides of glucuronic acid and beta-D-GlcpNAc-(1----3)-beta-D-GlcpA. Transformation of the products into 2-acrylamidoethyl glycosides, followed by deprotection using trifluoroacetic acid, gave the target monomers that were converted into high-molecular-weight copolymer-type neoglycoconjugates.     

Redial generations of Fasciola gigantica in the pulmonate snail Lymnaea truncatula.

Lymnaea truncatula, 4 mm in height, were subjected to infection by a single miracidium of Fasciola gigantica, then raised at 23 degrees C until day 60 of the experiment. Histological study of these snails demonstrated a mean redial burden of 34 parasites at day 60, of which one third were degenerating forms. The mean number of living independent rediae did not exceed 5 for the first and second generations. Conversely, in subsequent generations there were as many as 18 rediae per snail at day 60. The first living redia of the first generation in particular gave rise to daughter rediae. Mature rediae appeared at day 35 and especially concerned the first and second generations at day 60. The authors conclude that development of the first and second redial generations occurs during the same period, and that the forms of the first cohort of the second generation are produced from the first redia of the first generation which originated from the sporocyst.     

Reconstruction of a complex defect of the hand with two distinct segments of the scapula and a scapular fascial flap transferred as a single transplant.

To repair multiple defects of the metacarpal bones in combination with skin defects, a combined scapular free flap on a single vascular pedicle--the subscapular artery and the accompanying veins--can be used successfully. This flap includes two distinct bone segments of the lateral border of the scapula and a scapular fascial flap. Because of its long and mobile vascular pedicle, such a transplant can be used for one-stage reconstruction of the first and other metacarpal bones of the long fingers of the hand. The scapular fascial flap included in the transplant provides good functional and aesthetic results in combined injury of the skin of the hand.     

Stellate cells of aortic intima: II. Arborization of intimal cells in culture.

The present study analyzed effects of different cAMP-elevators on cell morphology in primary culture of human intimal and medial cells from grossly normal and atherosclerotic areas. In primary culture of human aortic cells adenylate cyclase activator forskolin and other cAMP elevators induced arborization of cells, i.e. they reversibly changed the shape of cells. This resulted in the formation of thin branching processes and in the concentration of cytoplasm around the nucleus. In the culture, the shape of the arborized cells resembled that of stellate ones detected in the aortic intima in situ. The arborization of cells was accompanied by destruction of myofilaments. Due to cAMP elevators' effect, most of the arborized cells were exhibited in the cultures isolated from the elastic-hyperplastic layer of the intima. The number of arborized cells was significantly less in the cultures isolated from the musculo-elastic layer and still lesser in those isolated from media. We failed to reveal any significant difference in the number of arborized cells cultured from fatty streaks, atherosclerotic plaques and grossly normal aortic areas. Obtained results suggest that the previously revealed polymorphism of human aortic intimal cells may be accounted for by the cell shape transformations underlined by the mechanism similar to that of arborization in vitro.     

NADPH-dependent reduction of amaranch in liver microsomes characterized the quantity of low spin forms of cytochrome P-450.

We confirmed that NADPH-dependent anaerobic amaranch reduction in rat liver microsomes is compatible with the interaction of the dye with Fe(III) heme of cytochrome P-450 as the type II substrate. This process is rate-limiting in the whole reaction. High positive correlation (r = 0.949) between the values of Vmax for reaction of NADPH-dependent anaerobic amaranch reduction and the relative content low spin forms of cytochrome P-450 determined by ESR in microsomes from liver of control and induced by PB, BP, IS and 4-MP rats was observed. Relative content of low spin forms of cytochrome P-450 determined by ESR was increased according to BP less than PB less than control less than IS approximately 4-MP; Vmax values increased according to BP less than PB less than control less than IS less than 4-MP. Thus, reaction of NADPH-dependent anaerobic amaranch reduction may be used for determination of low spin forms of cytochrome P-450 at physiological conditions.     

Interaction of apolipoprotein B-containing lipoprotein secreted by Hep G2 cells with receptors for low-density lipoprotein.

Radioligand and immunoenzymatic techniques were used to characterize the receptor binding properties of apolipoprotein B-containing lipoprotein produced by HepG2 cell line (H-LpB). It was found that compared to plasma low-density lipoprotein (LDL), the interaction of H-LpB nonseparated from conditioned medium with normal fibroblasts was 6-8-times lower and only slightly exceeded the nonspecific binding of LDL modified by malondialdehyde, while the uptake of the indicated lipoproteins by LDL receptor-negative strain of fibroblasts were identical. The uptake of H-LpB by normal fibroblasts increased 1.5-2-times after isolation from the culture medium by immunoaffinity chromatography. The effect of isolation could be explained by the finding that apolipoprotein E-containing lipoprotein secreted by HepG2 cells effectively competed for the binding with LDL-receptors. The obtained results suggest that H-LpB produced by HepG2 cells is poorly recognized by the LDL-receptors.     

100ps molecular dynamic simulation of d(TATCACC)2.

A heptanucleotide sequence d(TATCACC)2 from OR3 region of bacteriophage lambda is considered sufficient for the recognition of Cro protein. We present here results on molecular dynamic simulations on this sequence for 100 ps in 0.02 ps interval. The simulations are done using computer program GROMOS. The conformational results are averaged over each ps. The IUPAC torsional parameters for 100 conformations are illustrated using a wheal and a dial systems. Several other stereochemical parameters such as H-bonding lengths and angles, sugar puckers, helix twist and roll angles as also distances between opposite strand phosphorus are depicted graphically. We find that there is rupture of terminal H-bonds. The bases are tilted and shifted away from the helix axis giving rise to bifurcated H-bonds. H-bonds are seen even in between different base pairs. The role of these dynamic structural changes in the recognition of OR3 operator by Cro protein is discussed in the paper.     

Possible repetitive DNA markers for Eusorghum and Parasorghum and their potential use in examining phylogenetic hypotheses on the origin of Sorghum species

Genomic structures of two major species in section Eusorghum (Sorghum), Sorghum bicolor and Sorghum halepense, and their phylogenetic relationships with a species in section Parasorghum, Sorghum versicolor, were studied by using cloned repetitive DNA sequences from the three species. Of the five repetitive DNA clones isolated from S. bicolor and S. halepense, four produced qualitatively similar hybridization patterns with detectable variations in copy numbers of some of the restriction fragments on the Southern blots of the two genomic DNAs. One clone was shown to be diagnostic for S. halepense. Molecular analysis at the DNA level indicates that S. bicolor and S. halepense have similar but not identical genomes, consonant with differences in karyotypes, meiotic chromosome behaviors, morphology, and physiology of the species. In addition to five repetitive clones isolated from S. bicolor and S. halepense, eight more sequences were cloned from S. versicolor. Nine clones were found to be specific for either S. bicolor and S. halepense or S. versicolor. The remaining four had a moderate to strong homology with sequences present in all Sorghum species studied. We speculate that the genome in the common ancestor of Sorghum has differentiated to give rise to genomes of at least three major chromosome sizes; large, medium, and small, as seen at present. Amplifications, eliminations, rearrangements, and new syntheses of repetitive sequences may have been involved in genome differentiation of these species. The results also suggest that the S. versicolor genome has strongly diverged from the genomes of the two species in section Eusorghum.     

Reversed-phase chromatographic method for specific determination of glutathione in cultured malignant cells

A chromatographic method for the specific determination of glutathione in malignant cell lines is described. The method is based on the ability of glutathione-S-transferase to specifically and quantitatively conjugate glutathione to 1-chloro-2,4-dinitrobenzene and chromatographic quantitation of the resultant conjugate, dinitrophenyl-S-glutathione, by reversed-phase liquid chromatography. The assay can be performed on 20 000 g supernatants of cell homogenates without acid extraction. 2-Mercaptoethanol, a sulfhydryl compound often used as a thiol-protective agent to preserve enzymatic activities of a number of enzymes, did not interfere with glutathione determination by this method. The dinitrophenyl-S-glutathione isolated from either standard glutathione samples or from cell homogenates was shown to be identical to authentic dinitrophenyl-S-glutathione using mass spectrometry. Recovery of glutathione in standard samples by the current method was identical to that determined using 5,5′-dithiobis(2-nitrobenzoic acid). Exogenous glutathione added to supernatants of cell homogenate in the presence or absence of 2-mercaptoethanol was also completely recovered.     

Preparation of a fluorescent derivative of benzoylecgonine, and preliminary studies of its application to the analysis of urine

A sensitive high-performance liquid chromatographic method using 3-bromomethyl-6,7-dimethoxy-1-methyl-2(1H)-quinoxalinone (Br-DMEQ) as a fluorescent labeling reagent is described for the determination of benzoylecgonine (BE) and ecgonine (EC). The Br-DMEQ derivatives of BE and EC were separated on a C₁₈ column and detected at 455 nm with excitation at 370 nm. The detection limits of the proposed method were 18.7 fmol for BE and 12.5 pmol for EC at a signal-to-noise ratio of 3. Relative standard deviations of five replicate measurements were 1.94% (10 pmol) and 2.98% (50 pmol) for BE and 6.3% (250 pmol) and 5.62% (1.25 pmol) for EC. This method was applied to the determination of BE in human urine. BE was extracted from urine by solvent extraction with chloroform—isopropyl alcohol (9:1, v/v) solution. Levels of 2.5 · 10⁻⁸ M BE in urine (25 pmol/ml) could be determined.