Internalization and degradation of receptor bound C-reactive protein by U-937 cells: induction of H2O2 production and tumoricidal activity

The presence of a membrane receptor for C-reactive protein (CRP-R) on the human monocytic cell line U-937 was the basis for determining the metabolic fate of the receptor-bound ligand and the functional response of the cells to CRP. Internalized [125I]CRP was measured by removing cell surface-bound [125I]CRP with pronase. Warming cells to 37 degrees C resulted in the internalization of approx. 50% of the receptor-bound [125I]CRP or receptor-bound [125I]CRP-PC-KLH complexes. U-937 cells degraded about 25% of the internalized [125I]CRP into TCA-soluble radiolabeled products. The lysosomotrophic agents (chloroquine, NH4Cl) greatly decreased the extent of CRP degradation without altering binding or internalization. In addition, a pH less than 4.0 resulted in dissociation of receptor-bound [125I]CRP. Treatment of U-937 cell with monensin, a carboxylic ionophore which prevents receptor recycling, resulted in accumulation of internalized [125I]CRP. Therefore, it appears that the CRP-R complex is internalized into an endosomal compartment where the CRP is uncoupled from its receptor and subsequently degraded. CRP initiated the differentiation of the U-937 cells so that they acquired the ability to produce H2O2 and also display in vitro tumoricidal activity. The results support the concept that internalization and degradation of CRP leads to the activation of monocytes during inflammation.     

A classification of compounds in American Sign Language: an evaluation of the Bisetto and Scalise framework

Cross-linguistic comparisons of compounds are difficult because of the varied criteria and terms used by different linguists (Scalise and Bisetto 2009). To address this problem, Scalise and Bisetto proposed a universal three-level classification of compound types. Although several researchers have shown that American Sign Language (ASL) has compound signs, a classification of compound types in ASL has not been completed. All of the potential compounds in an ASL dictionary (Costello 1994) were identified, then verified as compounds with the help of a fluent deaf signer by applying standard tests for composition. These compounds were then classified using the Scalise and Bisetto classification. We found that Scalise and Bisetto??s three-level hierarchical classification successfully captured cross-category relationships among subtypes of compounds but fails to predict the existence of one type of compound attested in ASL. In our revised classification, a?consistent set of criteria is used at each level, resulting in a classification that is both conceptually simpler and empirically more adequate. The second tier category for hierarchical compounds are bifurcated into the categories expressed predicate and unexpressed predicate, according to whether each predicate in a compound??s semantic structure is expressed by one of the overt constituents. The revision has the further advantage of allowing us to avoid any reference to word class/grammatical category in applying our taxonomy, a goal that we show to be desirable on both theoretical and empirical grounds.     

Deletion of ripA alleviates suppression of the inflammasome and MAPK by Francisella tularensis

Francisella tularensis is a facultative intracellular pathogen and potential biothreat agent. Evasion of the immune response contributes to the extraordinary virulence of this organism although the mechanism is unclear. Whereas wild-type strains induced low levels of cytokines, an F. tularensis ripA deletion mutant (LVSΔripA) provoked significant release of IL-1β, IL-18, and TNF-α by resting macrophages. IL-1β and IL-18 secretion was dependent on inflammasome components pyrin-caspase recruitment domain/apoptotic speck-containing protein with a caspase recruitment domain and caspase-1, and the TLR/IL-1R signaling molecule MyD88 was required for inflammatory cytokine synthesis. Complementation of LVSΔripA with a plasmid encoding ripA restored immune evasion. Similar findings were observed in a human monocytic line. The presence of ripA nearly eliminated activation of MAPKs including ERK1/2, JNK, and p38, and pharmacologic inhibitors of these three MAPKs reduced cytokine induction by LVSΔripA. Animals infected with LVSΔripA mounted a stronger IL-1β and TNF-α response than that of mice infected with wild-type live vaccine strain. This analysis revealed novel immune evasive mechanisms of F. tularensis.     

Visual contrast detection by a single channel versus probability summation among channels

It is now generally accepted that the human visual system consists of subsystems (channels) that may be activated in parallel. According to some models of detection, detection is by probability summation among channels, while in other models it is assumed that detection is by a single channel that may even be tuned specifically to the stimulus pattern (detection by a matched filter). So far, arguments in particular for the hypothesis of probbbility summation are based on plausibility considerations and on demonstrations that the data from certain detection experiments are compatible with this hypothesis. In this paper it is shown that linear contrast interrelationship functions together with a property of a large class of distribution functions (strict log-concavity or logconvexity on the relevant set of contrasts/intensities) uniquely point to detection by a single channel. In particular, models of detection by probability summation based on Quick's Model are incompatible with linear contrast interrelationship functions. Sufficient (and observable) conditions for the strict logconcavity/log-convexity of distribution functions are presented.     

Bio-mediated soil improvement

New, exciting opportunities for utilizing biological processes to modify the engineering properties of the subsurface (e.g. strength, stiffness, permeability) have recently emerged. Enabled by interdisciplinary research at the confluence of microbiology, geochemistry, and civil engineering, this new field has the potential to meet society's ever-expanding needs for innovative treatment processes that improve soil supporting new and existing infrastructure. This paper first presents an overview of bio-mediated improvement systems, identifying the primary components and interplay between different disciplines. Geometric compatibility between soil and microbes that restricts the utility of different systems is identified. Focus is then narrowed to a specific system, namely bio-mediated calcite precipitation of sands. Following an overview of the process, alternative biological processes for inducing calcite precipitation are identified and various microscopy techniques are used to assess how the pore space volume is altered by calcite precipitation, the calcite precipitation is distributed spatially within the pore space, and the precipitated calcite degrades during loading. Non-destructive geophysical process monitoring techniques are described and their utility explored. Next, the extent to which various soil engineering properties is identified through experimental examples. Potential advantages and envisioned applications of bio-mediated soil improvement are identified. Finally, the primary challenges that lie ahead, namely optimization and upscaling of the processes and the education/training of researchers/practitioners are briefly discussed.     

Production estimates of the benthic invertebrate community in a small Danish stream

Quantitative samples were used to investigate density, biomass and annual production of the benthic invertebrate fauna in a small Danish stream. Forty-eight taxa were found and the total invertebrate densities varied from 3 810 m^?2 in July to 20 040 m^?2 in December. The total mean annual biomass of the invertebrate fauna was 6.1 g ash-free dry wt m^?2. The annual production of the invertebrates was estimated from their mean annual biomass and their annual P/B ratio. Production of the primary consumers (herbivores and detritivores) was 21.4 g ash-free dry wt m^?2 y^?1 and of secondary consumers (carnivores) 1.1 g m^?2 y^?1. The amount of invertebrate production available to the trout population and the importance of the species as food for trout are discussed.     

Characterization and isolation of a C-reactive protein receptor from the human monocytic cell line U-937

Human C-reactive protein (CRP) is an acute phase reactant that is opsonic and an activator of macrophage tumoricidal function. CRP also activates the classical C cascade. These activities suggest that CRP might interact with monocytes/macrophages via specific receptors in a manner analogous to the interaction of IgG with FcR. With the use of radio-labeled human CRP, we have observed specific binding of CRP to human blood monocytes and the human monocytic cell line U-937. Binding was saturable at a pathophysiologic concentration of CRP, with an estimated KD of 9.5 x 10(-8) M and 3.6 x 10(5) binding sites/cell. Specific binding was inhibited by polyclonal human IgG as well as an IgG1 myeloma. In the converse experiment, CRP failed to inhibit specific [125I]IgG binding. The mAb IV.3, which inhibits binding of IgG immune complexes to FcRII, did not inhibit CRP binding. A 100-fold excess of phosphorylcholine or the phosphorylcholine binding peptide of CRP (residues 47-63) failed to inhibit binding. Although human rIFN-gamma and PMA increased FcRI expression, these reagents had no affect on CRP receptor expression. A single membrane protein of 38 to 41 kDa from U-937 cells was chemically cross-linked to [125I]CRP; the cross-linking was inhibited by human IgG1 but not the IV.3 mAb. Furthermore, two membrane proteins with a Mr of 38 to 40 kDa and 58 to 60 kDa were isolated by CRP ligand-affinity chromatography. These proteins were of a distinct size from those isolated for FcRI from an IgG ligand matrix. These studies demonstrate specific binding of human CRP to a human monocytic cell line via receptors that are distinct from the IgG FcR and implicate CRP in nonspecific, preimmune host defense reaction mediated by cells of the monocytic lineage.     

Synchronization of the human promyelocytic cell line HL 60 by thymidine

Cultures of the promyelocytic cell line HL 60 were synchronized with thymidine. A concentration of 0.05 mM thymidine and an exposure time of 24 hr was found optimal for blocking about 90% of the cells in S phase. Following release from the thymidine block the cell cultures were followed intermittently over 40 hr for fluctuation in cell numbers, labelling with radioactive thymidine and nuclear DNA distributions. Mathematical evaluation of the results revealed a cycling time of 18.6 hr and a duration of specific cell phases of 8.6 hr, 7.1 hr and 2.9 hr for G1, S and G2 + M, respectively. The doubling time was 26 hr and the growth fraction was estimated as 1.     

Eukaryotic topoisomerase I-DNA interaction is stabilized by helix curvature.

The influence of DNA structure on topoisomerase I-DNA interaction has been investigated using a high affinity binding site and mutant derivatives thereof. Parallel determinations of complex formation and helix structure in the absence of superhelical stress suggest that the interaction is intensified by stable helix curvature. Previous work showed that a topoisomerase I binding site consists of two functionally distinct subdomains. A region located 5' to the topoisomerase I cleavage site is essential for binding. The region 3' to the cleavage site is covered by the enzyme, but not essential. We report here that the helix conformation of the latter region is an important modulator of complex formation. Thus, complex formation is markedly stimulated, when an intrinsically bent DNA segment is installed in this region. A unique pattern of phosphate ethylation interferences in the 3'-part of the binding site indicates that sensing of curvature involves backbone contacts. Since dynamic curvature in supercoiled DNA may substitute for stable curvature, our findings suggest that topoisomerase I is able to probe DNA topology by assessment of writhe, rather than twist.