Geometry strongly influences the response of numerical models of the lumbar spine--a probabilistic finite element analysis

Typical FE models of the human lumbar spine consider a single, fixed geometry. Such models cannot account for potential effects of the natural variability of the spine's geometry. In this study, we performed a probabilistic uncertainty and sensitivity analysis of a fully parameterized, geometrically simplified model of the L3-L4 segment. We examined the impact of the uncertainty in all 40 geometry parameters, estimated lower and upper bounds for the required sample size and determined the most important geometry parameters. The natural variability of the spine's geometry indeed strongly affects intradiscal pressure, range of motion and facet joint contact forces. Deriving generalized statements from fixed-geometry models as well as transferring those results to different cases thus can easily lead to wrong conclusions and should only be performed with extreme caution. We recommend a sample size of ≈ 100 to obtain reasonable accurate point estimates and a sufficient overview of the remaining uncertainties. Yet, only few parameters, especially those determining the disc geometry (disc height, end-plate width and depth) and the facets' position (intra-articular space, pedicle length, facet angles), proved to be truly important. Accurate measurement and modeling of those structures should therefore be prioritized.     

Synthesis of bivalent lactosides and their activity as sensors for differences between lectins in inter- and intrafamily comparisons

The synthesis of nine bivalent lactosides (based on ditriazoles, diamides, a glycocyclophane and an acyclic analogue of the glycocyclophane) and one monovalent lactosyl triazole facilitated the assessment of the sensitivity of plant/animal lectins to this type of ligand display. The inhibitory potency of the compounds was determined in two assays of increasing biorelevance. These were solid-phase and cell binding set-ups. Hereby, the ability of the compounds to inhibit the binding of two plant agglutinins and the entire set of adhesion/growth-regulatory galectins from one organism (chicken) to a glycoprotein or to cell surfaces was systematically evaluated. Differential sensitivities were detected between plant and animal lectins and also between distinct galectin forms within the chicken series. Two of the bivalent probes can be considered as sensors for interlectin differences. Most pronounced were the selectivities of N-glycosyl 1,2,3-triazole derivatives for the chimera-type galectin and its proteolytically truncated version.     

Novel triazines as potent and selective phosphodiesterase 10A inhibitors

The identification of highly potent and orally active triazines for the inhibition of PDE10A is reported. The new analogs exhibit low-nanomolar potency for PDE10A, demonstrate high selectivity against all other members of the PDE family, and show desired drug-like properties. Employing structure-based drug design approaches, we investigated the selectivity of PDE10A inhibitors against other known PDE isoforms, by methodically exploring the various sub-regions of the PDE10A ligand binding pocket. A systematic assessment of the ADME and pharmacokinetic properties of the newly synthesized compounds has led to the design of drug-like candidates with good brain permeability and desirable drug kinetics (t(1/2), bioavailability, clearance). Compound 66 was highly potent for PDE10A (IC(50)=1.4nM), demonstrated high selectivity (>200×) for the other PDEs, and was efficacious in animal models of psychoses; reversal of MK-801 induced hyperactivity (MED=0.1mg/kg) and conditioned avoidance responding (CAR; ID(50)=0.2mg/kg).     

Aggregative response in bats: prey abundance versus habitat

In habitats where prey is either rare or difficult to predict spatiotemporally, such as open habitats, predators must be adapted to react effectively to variations in prey abundance. Open-habitat foraging bats have a wing morphology adapted for covering long distances, possibly use information transfer to locate patches of high prey abundance, and would therefore be expected to show an aggregative response at these patches. Here, we examined the effects of prey abundance on foraging activities of open-habitat foragers in comparison to that of edge-habitat foragers and closed-habitat foragers. Bat activity was estimated by counting foraging calls recorded with bat call recorders (38,371 calls). Prey abundance was estimated concurrently at each site using light and pitfall traps. The habitat was characterized by terrestrial laser scanning. Prey abundance increased with vegetation density. As expected, recordings of open-habitat foragers clearly decreased with increasing vegetation density. The foraging activity of edge- and closed-habitat foragers was not significantly affected by the vegetation density, i.e., these guilds were able to forage from open habitats to habitats with dense vegetation. Only open-habitat foragers displayed a significant and proportional aggregative response to increasing prey abundance. Our results suggest that adaptations for effective and low-cost foraging constrains habitat use and excludes the guild of open-habitat foragers from foraging in habitats with high prey abundance, such as dense forest stands.     

Elevated air carbon dioxide concentrations increase dissolved carbon leaching from a cropland soil

Terrestrial sources of nitrogen (N), particularly N-fixing alder, may be important for sustaining production in headwater streams that typically lack substantial subsidies of marine-derived nutrients from spawning salmon yet support upstream-dispersing juvenile salmonids. However, other physiographic characteristics, such as watershed slope and topographic wetness, also control transport of nutrients to streams and may confound apparent linkages between alder and stream N. Seasonal patterns in precipitation and temperature may interact with watershed characteristics to modulate stream N availability. We empirically modeled the effect of alder cover and other watershed physiographic variables on stream N and contrasted these relationships over the growing season among 25 first-order streams from the lower Kenai Peninsula, Alaska. For each date, percent alder cover, mean topographic wetness, and mean slope were used as watershed predictors of NO _x –N concentration (nitrate?+?nitrite) and daily NO _x –N yield using Generalized Additive Models (GAM) and compared using Akaike’s Information Criterion (AIC_c). Alder cover was the only probable model and explained 75–96% of the variation in NO _x –N concentration and 83–89% of the variation in daily NO _x –N yield. The relationship between alder and both NO _x –N concentration and daily NO _x –N yield changed from constant inputs in May across the range of alder cover (linear fit) to increasing inputs in July and September (non-linear fits) implying that high-alder watersheds were N-saturated. The strong linkage between alder and stream N coupled with the concurrent timing of maximum stream N from alder in the spring to salmon fry emergence indicates the potential importance of this subsidy to headwater stream ecosystems.     

The ribosome modulates the structural dynamics of the conserved GTPase HflX and triggers tight nucleotide binding

The universally conserved GTPase HflX is a putative translation factor whose GTPase activity is stimulated by the 70S ribosome as well as the 50S but not the 30S ribosomal subunit. However, the details and mechanisms governing this interaction are only poorly understood. In an effort to further elucidate the functional mechanism of HflX, we examined its interaction with the 70S ribosome, the two ribosomal subunits (50S and 30S), as well as its ability to interact with guanine nucleotides in the respective ribosomal complexes using a highly purified in vitro system. Binding studies reported here demonstrate that HflX not only interacts with 50S and 70S particles, but also with the 30S subunit, independent of the nucleotide-bound state. A detailed pre-steady-state kinetic analysis of HflX interacting with a non-hydrolyzable analog of mant-GTP, coupled with an enzymatic probing assay utilizing limited trypsinolysis, reveal that HflX·GTP exists in a structurally distinct 50S- and 70S-bound form that stabilizes GTP binding up to 70 000-fold and that may represent the “GTPase-activated” state. This activation is likely required for efficient GTP-hydrolysis, and may be similar to that observed in elongation factor G. Results reported here address the surprising low affinity of free HflX for GTP and suggest that cellular HflX will mainly exist in the HflX·GTP·ribosome-bound form. A minimal model for the functional cycle of HflX is proposed.     

alpha2,3/alpha2,6-Sialylation of N-glycans: non-synonymous signals with marked developmental regulation in bovine reproductive tracts

The glycan part endows cellular glycoconjugates with significant potential for biological recognition. N-Glycan branches often end with alpha2,3/alpha2,6-sialylation, posing the question whether and how placement of the sialic acid at 3 - or 6 -acceptor positions of galactose has cell biological relevance. As attractive model to study developmental regulation we monitored the expression of alpha2,3/alpha2,6-sialylated determinants in fetal and adult bovine testes and ovaries by lectin histochemistry. Distinct expression patterns were detected in both organ types. Oocyte staining, as a prominent example, was restricted to the presence of alpha2,6-sialylated glycans. Treatment with sialidase abolished binding and thus excluded sulfate esters as lectin targets. We added computer simulations to rationalize the observed evidence for non-random expression of the two closely related sialylgalactose isomers. Extensive molecular mechanics and molecular dynamics calculations reveal that the seemingly minor shift of the glycosidic bond from the alpha2,3 position to the alpha2,6 configuration causes significant shape and flexibility changes. They give each disaccharide its own characteristic meaning as signal in the sugar code.     

Tumor-associated antigen 90K/Mac-2-binding protein: possible role in colon cancer

The tumor-associated antigen 90K (TAA90K)/Mac-2-binding protein implicated in cancer progression and metastasis is modified by beta1-6 branched N-linked oligosaccharides in colon cancer cells, glycans shown to contribute to cancer metastasis. To elucidate the role of TAA90K in colon cancer, we examined its expression and function in human colon tumors and colon carcinoma cell lines. Immunohistochemical analyses of colon tumors revealed elevated expression of TAA90K in all samples analyzed compared to normal colon. To examine the function of TAA90K in colon cancer, we carried out protein and cell binding assays using TAA90K-His purified from HT-29 cells colon carcinoma cells infected with recombinant vaccinia virus expressing TAA90K containing a C-terminal poly-histidine tag. TAA90K-His bound to fibronectin, collagen IV, laminins-1, -5, and -10 and galectin-3 (Mac-2) but poorly to collagen I and galectin-1. As expected, binding of TAA90K to galectin-3 was dependent on carbohydrate since it was inhibitable by lactose and asiolofetuin, and a TAA90K-His glycoform purified from HT-29 cells treated with the glycosylation inhibitor 1-deoxymannojirimycin bound poorly to galectin-3. Unlike TAA90K isolated from other cell types, TAA90K-His isolated from colon cancer cells failed to mediate adhesion of colon cancer and normal cell lines, possibly due to cell-type specific glycosylation of TAA90K-His and/or its putative cellular receptor. However, at low concentrations, TAA90K-His enhanced galectin-3-mediated HT-29 cell adhesion while at high concentrations, it inhibited cell adhesion. Thus, a possible mechanism by which TAA90K may contribute to colon cancer progression is by modulating tumor cell adhesion to extracellular proteins, including galectin-3.     

DNA uracil repair initiated by the archaeal ExoIII homologue Mth212 via direct strand incision

No genes for any of the known uracil DNA glycosylases of the UDG superfamily are present in the genome of Methanothermobacter thermautotrophicus ΔH, making it difficult to imagine how DNA-U repair might be initiated in this organism. Recently, Mth212, the ExoIII homologue of M. thermautotrophicus ΔH has been characterized as a DNA uridine endonuclease, which suggested the possibility of a novel endonucleolytic entry mechanism for DNA uracil repair. With no system of genetic experimentation available, the problem was approached biochemically. Assays of DNA uracil repair in vitro, promoted by crude cellular extracts, provide unequivocal confirmation that this mechanism does indeed operate in M. thermautotrophicus ΔH.