Aneurysms after microvascular anastomoses. Incidence and pathogenesis in experimental animals.

This study has demonstrated that aneurysms occur in about 50 percent of the rat femoral arteries subjected to microvascular anastomosis. These aneurysms are consistent histologically--being characterized by medial necrosis, loss of elastic lamellae, and subintimal hyperplasia. Mechanical trauma is implicated as a factor in their pathogenesis, though further study in necessary to define the exact etiology.     

Transbilayer distribution of bromine in fluid bilayers containing a specifically brominated analogue of dioleoylphosphatidylcholine

We describe in this paper the transbilayer distribution of the bromines of the specifically halogenated phospholipid 1-oleoyl-2-(9,10-dibromostearoyl)-sn-glycero-3- phosphocholine (OBPC). The distribution was determined by X-ray diffraction of oriented multilayers of mixtures of OBPC and 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) at 66% relative humidity by the general approach of Franks et al. (1978) [Nature 276, 530-532]. The bromine distribution of OBPC in the fluid L alpha phase is described accurately by a pair of Gaussian functions located 7.97 +/- 0.27 A from the center of the bilayer with l/e half-widths of 4.96 +/- 0.62 A. We find that OBPC bilayers are accurately described as DOPC bilayers with an additional bromine distribution centered at the position of the double bond of DOPC and conclude that OBPC is an excellent structural isomorph for DOPC under the conditions of these experiments. The distribution obtained is the complete and fully resolved transbilayer image of the halogen label because the broad distribution of the bromines is due entirely to thermal disorder and not to experimental limitations [Wiener, M. C., & White, S. H. (1991a) Biophys. J. 59, 162-173]. The observed width of the bromine distribution indicates that virtually all of the hydrocarbon interior is accessible to the bromines. The distance between the bromine/double-bond position and the headgroup phosphate position was determined from one-dimensional Patterson maps and found to be approximately 12 A. The application of accurately determined bromine distributions to the quantitative interpretation of fluorescence quenching experiments is discussed. A method for the self-consistent global analysis of diffraction data from mixtures that permits the use of data sets with different instrumental scale factors is developed in an Appendix.     

Reconstitution of RecBC DNase activity from purified Escherichia coli RecB and RecC proteins

The Escherichia coli RecB protein, normally synthesized in low amounts, has been amplified by linkage of the recB gene to the phage lambda leftward promoter in an expression plasmid. From strains harboring this plasmid, RecB protein has been purified to homogeneity by a simple procedure which includes affinity chromatography on a column of RecC protein bound to agarose. The purified RecB protein has DNA-dependent ATPase activity but no exonuclease activity. RecC protein alone has neither ATPase nor exonuclease activity. However, when combined together, the RecB and RecC proteins show the ATP-dependent double-stranded exonuclease properties characteristic of the RecBC DNase.     

Stereological Determination of Orientations,Second-Order Quantities and Correlations for Random Spatial Fibre Systems

This paper presents methods for the stereological analysis of spatial fibre systems on the base of planar or thin sections. Under the assumption that the cross-section figures of the tubular fibres can be measured, the orientation distribution of the fibre system and its line density L_v can be determined from one section only and without distributional assumptions. A simple way to study the degree of randomness of fibre systems consists in the statistical analysis of the point pattern of centres of intersection figures. More sophisticated methods are of stereological nature and yield the spatial reduced second moment measure. Similarly also correlations between two fibre systems can be quantified. The methods are demonstrated by two examples concerning samples of human brain.     

Xyloside transport by XylP, a member of the galactoside-pentoside-hexuronide family.

This paper describes the functional characterization of the xyloside transporter, XylP, of Lactobacillus pentosus with the aid of a spectroscopy-based assay system. In order to monitor the transport reaction, the natural xyloside isoprimeverose, a building block of hemicellulose, and the analogue methyl-isoprimeverose were chemically synthesized by a new and efficient procedure. The XylP protein was purified by metal affinity chromatography, following high level expression in Lactococcus lactis from the nisin-inducible promoter. The purified XylP protein was incorporated into liposomes, in which the glucose dehydrogenase from Acinetobacter calcoaceticus (sGDH) was entrapped. sGDH can oxidize aldose sugars in the presence of dichlorophenol-indophenol as electron acceptor. The coupled assay thus involves XylP-mediated isoprimeverose uptake followed by internal oxidation of the sugar by sGDH, which can be monitored from the reduction of 2,6-dichlorophenol-indophenol at 600 nm. The uptake of isoprimeverose was stimulated by the presence of the non-oxidizable methyl-isoprimeverose on the trans-side of the membrane, indicating that exchange transport is faster than unidirectional downhill uptake. Unlike other members of the galactoside-pentoside-hexuronide family, XylP does not transport monosaccharides (xylose) but requires a glycosidic linkage at the anomeric carbon position. Consistent with a proton motive force-driven mechanism, the uptake was stimulated by a membrane potential (inside negative relative to outside) and inhibited by a pH gradient (inside acidic relative to outside). The advantages of the here-described transport assay for studies of carbohydrate transport are discussed.     

Electron paramagnetic resonance study of ferrous cytochrome P-450scc-nitric oxide complexes: effects of cholesterol and its analogues

Electron paramagnetic resonance (EPR) spectra of nitric oxide (NO) complexes of ferrous cytochrome P-450scc were measured at 77 K for the first time without using the rapid-mixing and freeze-quenching technique. Without substrate the EPR spectra were very similar to those of cytochrome P-450cam (from Pseudomonas putida) and cytochrome P-450LM (from rat liver microsomes) with rhombic symmetry; gx = 2.071, gz = 2.001, gy = 1.962, and Az = 2.2 mT for 14NO complexes. Upon addition of substrates [such as cholesterol, 22(S)-hydroxycholesterol, 22(R)-hydroxycholesterol, 25-hydroxycholesterol, and 22-ketocholesterol], the EPR spectra exhibited many variations having rhombic symmetry in the major component and an additional minor component with less rhombic symmetry. Furthermore, addition of 20(S)-hydroxycholesterol caused a striking change in the EPR spectrum. The component with rhombic symmetry disappeared completely, and the component with less rhombic symmetry dominated (gx = 2.027, gz = 2.007, gy = 1.984, and Az = 1.76 mT for 14NO complexes). These observations suggest the existence of the following physiologically important natures: (1) the conformational flexibility of the active site of the enzyme due to the steric interaction between the substrate and the heme-bound ligand molecule and (2) the importance of the hydroxylation of the cholesterol side chain at the 20S position to proceed the side-chain cleavage reaction in cytochrome P-450scc.     

Development of Four Populations of Meloidogyne hapla on Two Cultivars,of Cucumber at Different Temperatures

The infectivity and development of four populations of Meloidogyne hapla were compared, at three temperatures, on tomato and two varieties of cucumber. A population from Canada produced few root-galls on cucumber and, except at 24 C, no larvae developed into adult females and produced egg masses. In contrast, a population with 45 chromosomes from America produced many galls on cucumber and small proportions of larvae became females and produced egg masses at 20 and 24 C. At 18 C this population produced no egg masses on cucumber, but a population from Britain and one from America with 17 chromosomes produced more egg masses at this temperature than at 20 or 24 C. Dissection of the galls showed that on cucumber many larvae died or their growth and development was slowed.     

A single-cell assay of beta-galactosidase activity in Saccharomyces cerevisiae

A novel assay of single-cell exogenous beta-galactosidase activity in Saccharomyces cerevisiae has been developed. Intracellular fluorescence due to the hydrolysis of resorufin-beta-D-galactopyranoside attains a steady state between production of resorufin and its subsequent leakage from the cell. The cells are permeabilized with Triton X-100, and the assay is performed at 0 degrees C. These conditions were chosen to minimize intercellular fluorescence communication. Free resorufin in the extracellular space is bound by bovine serum albumin to prevent its uptake by cells. Two regimes of fluorescence accumulation are observed, one limited by the rate of diffusion of substrate into the cell, and one limited by the rate of enzymatic cleavage of the substrate. A quantitative correlation between fluorescence and beta-galactosidase activity is obtained under optimized assay conditions.     

Serum retinol and the inverse relationship between serum cholesterol and cancer.

Several human studies have shown an inverse relation between vitamin A intake (and serum concentrations of retinol and carotene) and cancer. Serum cholesterol concentrations have also been reported in inverse relation to cancer. In a study of 3102 people in Evans County, Georgia, who were followed for over 12-14 years to assess the incidence of cancer there was an inverse association between the risk of cancer and both serum retinol and serum cholesterol concentrations. The data also showed an unexpectedly strong correlation between serum retinol and total cholesterol concentrations. The inverse relationship with cancer was stronger with serum retinol than with cholesterol, which suggested that the association with cholesterol might be secondary. This suggestion may also explain the cholesterol-cancer association reported in several other cohort studies. Further studies of the relation between serum concentrations of cholesterol, retinol, and carotene and the incidence of cancer are needed.