Crystal structure of the lectin from Dioclea grandiflora complexed with core trimannoside of asparagine-linked carbohydrates

The seed lectin from Dioclea grandiflora (DGL) has recently been shown to possess high affinity for 3, 6-di-O-(alpha-D-mannopyranosyl)-alpha-D-mannopyranose, the core trimannoside of asparagine-linked carbohydrates, but lower affinity for biantennary complex carbohydrates. In the previous paper, the thermodynamics of DGL binding to deoxy analogs of the core trimannoside and to a biantennary complex carbohydrate were determined by isothermal titration microcalorimetry. The data suggest that DGL recognizes specific hydroxyl groups of the trimannoside similar to that of the jack bean lectin concanavalin A (ConA) (Gupta, D. Dam, T. K., Oscarson, S., and Brewer, C. F. (1997) J. Biol. Chem. 272, 6388-6392). However, the thermodynamics of DGL binding to certain deoxy analogs and to the complex carbohydrate are different from that of ConA. In the present paper, the x-ray crystal structure of DGL complexed to the core trimannoside was determined to a resolution of 2.6 A. The overall structure of the DGL complex is similar to the structure of the ConA-trimannoside complex (Naismith, J. H., and Field, R. A. (1996) J. Biol. Chem. 271, 972-976). The location and conformation of the bound trimannoside as well as its hydrogen-bonding interactions in both complexes are nearly identical. However, differences exist in the location of two loops outside of the respective binding sites containing residues 114-125 and 222-227. The latter residues affect the location of a network of hydrogen-bonded water molecules that interact with the trisaccharide. Differences in the arrangement of ordered water molecules in the binding site and/or protein conformational differences outside of the binding site may account for the differences in the thermodynamics of binding of the two lectins to deoxy analogs of the trimannoside. Molecular modeling studies suggest how DGL discriminates against binding the biantennary complex carbohydrate relative to ConA.     

Interrelationships of clinical outcome, length of resection, and energy cost of walking after prosthetic knee replacement following resection of a malignant tumor of the distal aspect of the femur

An automated method for analysis of in vivo proton magnetic resonance (MR) spectra and reconstruction of metabolite distributions from MR spectroscopic imaging (MRSI) data is described. A parametric spectral model using acquisition specific, a priori information is combined with a wavelet-based, nonparametric characterization of baseline signals. For image reconstruction, the initial fit estimates were additionally modified according to a priori spatial constraints. The automated fitting procedure was applied to four different examples of MRS data obtained at 1.5 T and 4.1 T. For analysis of major metabolites at medium TE values, the method was shown to perform reliably even in the presence of large baseline signals and relatively poor signal-to-noise ratios typical of in vivo proton MRSI. Identification of additional metabolites was also demonstrated for short TE data. Automated formation of metabolite images will greatly facilitate and expand the clinical applications of MR spectroscopic imaging.     

Carotid rupture and intraplaque hemorrhage: immunophenotype and role of cells involved

BACKGROUND: A complete immunohistochemical characterization in complicated carotid plaques is still lacking. The cellular components of 165 carotid endarterectomy specimens were analyzed to assess their role in the pathogenesis of plaque rupture and intraplaque hemorrhage without rupture. METHODS AND RESULTS: The fibrous caps at the sites of plaque rupture showed CD68+ macrophages, T-lymphocytes, and scarce B-lymphocytes. Ruptured plaques showed mononuclear infiltrates in the caps, shoulders, and bases of the plaques in 85% of the cases. Only 46% of nonruptured plaques showed such infiltrates (P <.0001). Two types of lipid cores were recognized: avascular or mildly vascularized and highly vascularized. The vessels of the latter type reacted with CD31 and CD34. In 57.5% of the cases, the base and the shoulders of the plaques showed neoformed, CD34+ vessels, often surrounded by mononuclear infiltrates. Intraplaque hemorrhage without rupture had highly vascularized lipid cores in all cases. T-lymphocytes and macrophages were in close contact with neoformed vessels. CONCLUSIONS: Plaque rupture is characterized by mononuclear cell infiltration of the caps, whereas intraplaque hemorrhage without rupture is characterized by extensive vascularization of the plaque.     

Molecular characterization of a novel short-chain dehydrogenase/reductase that reduces all-trans-retinal

The reduction of all-trans-retinal in photoreceptor outer segments is the first step in the regeneration of bleached visual pigments. We report here the cloning of a dehydrogenase, retSDR1, that belongs to the short-chain dehydrogenase/reductase superfamily and localizes predominantly in cone photoreceptors. retSDR1 expressed in insect cells displayed substrate specificities of the photoreceptor all-trans-retinol dehydrogenase. Homology modeling of retSDR1 using the carbonyl reductase structure as a scaffold predicted a classical Rossmann fold for the nucleotide binding, and an N-terminal extension that could facilitate binding of the enzyme to the cell membranes. The presence of retSDR1 in a subset of inner retinal neurons and in other tissues suggests that the enzyme may also be involved in retinol metabolism outside of photoreceptors.     

Unexpected expression of glucose transporter 4 in villous stromal cells of human placenta

Glucose transporter 4 (GLUT4) protein expression was characterized in human and rodent term placentas. A 50-kDa protein was detected, by immunoblotting, in term human placenta at levels averaging 25% of those found in white adipose tissue. It was also present, albeit at lower levels, in mouse and rat placentas. The specificity of the 50-kDa signal was established by using skeletal muscle and placental tissues obtained from GLUT4-null mice as controls. Indirect immunohistochemistry, performed in human placentas, showed that intravillous stromal cells were conspicuously labeled by GLUT4 and revealed colocalization of GLUT4 transporters with insulin receptors. This study provides the first evidence that the insulin-responsive GLUT4 glucose transporter is present in human and rodent hemochorial placentas. Placental GLUT4 gene and protein levels were not modified in human pregnancy complicated by insulin-dependent diabetes mellitus. The significance of the high level of GLUT4 protein in human placenta remains to be elucidated, because, so far, this organ was not considered to be insulin-sensitive, with regard to glucose transport.     

Influence of magnesium substitution on a collagen-apatite biomaterial on the production of a calcifying matrix by human osteoblasts

The induction of a calcifying matrix is of great interest in the restoration of bone defects. In a previous in vitro study we demonstrated that a collagen sponge constituted of type I collagen fibrils, chondroitin sulfates, and hydroxyapatite crystals induces an earlier and a more abundant synthesis of a new extracellular calcifying matrix than do other biomaterials such as collagen or hydroxyapatite alone. Bone mineral contains various amounts of magnesium ions, either adsorbed at the surface of apatite crystals or incorporated inside the crystal structure. Magnesium is known to reduce the degradation rate of tricalcium phosphate ceramics and to influence the crystallization of mineral substance. Thus we evaluated two sponges modified with different substituted apatites. The substituted low magnesium-containing apatite sample decreased the osteoinductive properties of the sponge whereas the substituted high magnesium-containing apatite sample had a toxic effect on bone cells and prevented the formation of any extracellular matrix. Such a toxic effect can be explained by the presence of large numbers of magnesium ions released into the culture medium even though at physiological level magnesium is able to promote bone mineralization and to control the growth of hydroxyapatite crystals. Thus collagen sponges containing hydroxyapatite remain one of the most appropriately evaluated biomaterials used for the restoration of periodontal pockets and bone defects.     

Antibiotic susceptibility profile of group B streptococcus acquired vertically

OBJECTIVE: To determine the contemporary antibiotic susceptibility profile of vertically acquired group B streptococcal isolates. METHODS: Susceptibility to ampicillin, penicillin G, erythromycin, clindamycin, cefazolin, and gentamicin was assessed by two methods, minimal inhibitory concentration and disc diffusion. RESULTS: The susceptibility profiles of 119 colonizing and eight invasive strains of group B streptococcus isolated from January 1996 to September 1997 at two hospitals in Birmingham, Alabama-University of Alabama at Birmingham and Cooper Green-were studied. Minimal inhibitory concentration determinations indicated that all colonizing strains were susceptible or moderately susceptible to ampicillin and penicillin G. Resistance was noted by at least one strain to each of the other antibiotics; all were resistant to gentamicin, whereas 27 (21%) were resistant to erythromycin, five (4%) to clindamycin, and one (1%) to cefazolin. All of the eight invasive strains were susceptible or moderately susceptible to ampicillin, penicillin G, clindamycin, and cefazolin; one (13%) was resistant to erythromycin, and all were resistant to gentamicin. Disc diffusion results generally were concordant with minimal inhibitory concentration results, although by disc diffusion fewer isolates were classified as susceptible, and more as moderately susceptible, to ampicillin and penicillin G than by minimal inhibitory concentration. CONCLUSION: Universal susceptibility of group B streptococcus to members of the penicillin family supports the continued use of penicillin G or ampicillin for early onset neonatal group B streptococcal disease prevention. For patients allergic to beta-lactam agents, clindamycin (4% resistance) may be a better alternative than erythromycin (21% resistance).     

Sulfasalazine in the treatment of juvenile chronic arthritis: a randomized, double-blind, placebo-controlled, multicenter study. Dutch Juvenile Chronic Arthritis Study Group

AIMS: To determine whether left ventricular volumes and ejection fractions calculated from single plane two-dimensional echocardiograms using the algorithm (0.85A2L) correlate with those calculated using the biplane Simpson's method, and whether small changes in volumes and ejection fraction occurring post-infarction could be detected from single-plane as well as from biplane two-dimensional echocardiograms. METHODS AND RESULTS: Serial two-dimensional echocardiograms were obtained in 371 patients from the DEFIANT II trial a mean of 2 days, 1 week and 6 months post-infarction. Single plane volumes from the apical four chamber and apical long axis correlated closely with biplane Simpson's left ventricular volumes. Both single-plane left ventricular volumes significantly over-estimated biplane Simpson's volumes. Biplane Simpson's ejection fractions were consistently slightly under-estimated from the single-plane images. Differences between biplane Simpson's and single-plane volumes increased independently with increasing left ventricular size and distortion. The small changes in left ventricular volumes and ejection fraction over time were as reliably detected from single plane as from biplane images. CONCLUSION: Single-plane left ventricular volumes over-estimate biplane Simpson's volumes and under-estimate ejection fraction, and these discrepancies are amplified in dilated hearts with abnormal shape.     

A27L protein mediates vaccinia virus interaction with cell surface heparan sulfate

Vaccinia virus has a wide host range and infects mammalian cells of many different species. This suggests that the cell surface receptors for vaccinia virus are ubiquitously expressed and highly conserved. Alternatively, different receptors are used for vaccinia virus infection of different cell types. Here we report that vaccinia virus binds to heparan sulfate, a glycosaminoglycan (GAG) side chain of cell surface proteoglycans, during virus infection. Soluble heparin specifically inhibits vaccinia virus binding to cells, whereas other GAGs such as condroitin sulfate or dermantan sulfate have no effect. Heparin also blocks infections by cowpox virus, rabbitpox virus, myxoma virus, and Shope fibroma virus, suggesting that cell surface heparan sulfate could be a general mediator of the entry of poxviruses. The biochemical nature of the heparin-blocking effect was investigated. Heparin analogs that have acetyl groups instead of sulfate groups also abolish the inhibitory effect, suggesting that the negative charges on GAGs are important for virus infection. Furthermore, BSC40 cells treated with sodium chlorate to produce undersulfated GAGs are more refractory to vaccinia virus infection. Taken together, the data support the notion that cell surface heparan sulfate is important for vaccinia virus infection. Using heparin-Sepharose beads, we showed that vaccinia virus virions bind to heparin in vitro. In addition, we demonstrated that the recombinant A27L gene product binds to the heparin beads in vitro. This recombinant protein was further shown to bind to cells, and such interaction could be specifically inhibited by soluble heparin. All the data together indicated that A27L protein could be an attachment protein that mediates vaccinia virus binding to cell surface heparan sulfate during viral infection.