Mass balance characterisation of Al-7Si-Mg alloy microstructures as a function of solution treatment time

Mass balance calculations were performed to model the effect of solution treatment time on A356 and A357 alloy microstructures. Image analysis and electron probe microanalysis were used to characterise microstructures and confirm model predictions. In as-cast microstructures, up to 8 times more Mg is tied up in the pi-phase than in Mg2Si. The dissolution of pi is accompanied by a corresponding increase in the amount of beta-phase. This causes the rate of pi dissolution to be limited by the rate of beta formation. It is predicted that solution treatments of the order of tens of minutes at 540degreesC produce near-maximum T6 yield strengths, and that Mg contents in excess of 0.52 wt% have no advantage.

Duck hepatitis B virus expresses a regulatory HBx-like protein from a hidden open reading frame

Duck hepatitis B viruses (DHBV), unlike mammalian hepadnaviruses, are thought to lack X genes, which encode transcription-regulatory proteins believed to contribute to the development of hepatocellular carcinoma. A lack of association of chronic DHBV infection with hepatocellular carcinoma development supports this belief. Here, we demonstrate that DHBV genomes have a hidden open reading frame from which a transcription-regulatory protein, designated DHBx, is expressed both in vitro and in vivo. We show that DHBx enhances neither viral protein expression, intracellular DNA synthesis, nor virion production when assayed in the full-length genome context in LMH cells. However, similar to mammalian hepadnavirus X proteins, DHBx activates cellular and viral promoters via the Raf-mitogen-activated protein kinase signaling pathway and localizes primarily in the cytoplasm. The functional similarities as,well as the weak sequence homologies of DHBx and the X proteins of mammalian hepadnaviruses strongly suggest a common ancestry of ortho- and avihepadnavirus X genes. In addition, our data disclose similar intracellular localization and transcription regulatory functions of the corresponding proteins, raise new questions as to their presumed role in hepatocarcinogenesis, and imply unique opportunities for deciphering of their still-enigmatic in vivo functions.

Muscle strength and function before and after anterior cruciate ligament reconstruction using semitendonosus and gracilis

This study assessed the quadriceps and hamstring strength before and 6 months after anterior cruciate ligament (ACL) reconstructive surgery using the hamstrings and related the findings to functional performance. Six months after surgery is a critical time for assessment as this is when players are returning to sport. Maximum isokinetic strength of 31 patients with complete unilateral ACL ruptures was measured at speeds of 60 degrees and 120 degrees per second. Functional assessment included the single hop, the triple hop, the shuttle run, side-step and carioca tests. All patients underwent a controlled quadriceps emphasized home-based physiotherapy program both before and after surgery. Results show that before surgery there was a 7.3% quadriceps strength deficit at 60 degrees per second compared to the uninjured leg but no hamstring strength deficit. After surgery there was a statistically significant but relatively small loss of muscle strength. The quadriceps strength deficit had increased to 12% and there was a 10% hamstring deficit. Post-operatively there was an 11% and 6.3% improvement in the hop tests, a 9% (P < 0.01) improvement in the shuttle run, a 15% (P < 0.001) improvement in the side step and a 24% (P < 0.001) improvement in the carioca tests (P < 0.001) despite the loss of muscle strength. (C) 2001 Elsevier Science B.V. All rights reserved.

Development of response-selective agonists of human C5a anaphylatoxin: Conformational, biological, and therapeutic considerations

Numerous studies on the relationship between the structure and function of peptide agonists derived from the biologically active, C-terminal region of human C5a anaphylatoxin have been reported over the past decade. These studies have been performed with the objective of parlaying this structure-function information into the design of peptide/peptidomimetic modulators of C5a receptor (C5aR)-mediated function. In this review, we describe a rational approach for the development of conformationally biased, decapeptide agonists of C5a and described how these stabilized and specific conformational features relate to the expression of specific C5a-like activities in vitro and in vivo. The therapeutic potential of such response-selective C5a agonists is discussed and underscored by the results of one such response-selective C5a agonist that was used in vivo as an effective molecular adjuvant capable of generating antigen-specific humoral and cellular immune responses. Finally, we describe the synthesis of a new generation of highly response-selective, conformationally biased C5a agonist and discuss the in vitro and in vivo biologic results that so indicate this biologic selectivity.

Microtubule integrity is essential for apical polarization and epithelial morphogenesis in the thyroid

In this study, we examined the contribution of microtubules to epithelial morphogenesis in primary thyroid cell cultures. Thyroid follicles consist of a single layer of polarized epithelial cells surrounding a closed compartment, the follicular lumen. Freshly isolated porcine thyroid cells aggregate and reorganize to form follicles when grown in primary cultures. Follicular reorganization is principally a morphogenetic process that entails the assembly of biochemically distinct apical and basolateral membrane domains, delimited by tight junctions. The establishment of cell surface polarity during folliculogenesis coincided with the polarized redistribution of microtubules, predominantly in the developing apical poles of cells. Disruption of microtubule integrity using either colchicine or nocodazole caused loss of defined apical membrane domains, tight junctions and follicular lumina. Apical membrane and tight junction markers became randomly distributed at the outer surfaces of aggregates. In contrast, the basolateral surface markers, E-cadherin and Na+,K+-ATPase, remained correctly localized at sites of cell-cell contact and at the free surfaces of cell aggregates. These findings demonstrate that microtubules play a necessary role in thyroid epithelial morphogenesis. Specifically, microtubules are essential to preserve the correct localization of apical membrane components within enclosed cellular aggregates, a situation that is also likely to pertain where lumina must be formed from solid aggregates of epithelial precursors. (C) 2001 Wiley-Liss, Inc.

Broadsheet - Dendritic cells and their emerging clinical applications

Dendritic cells (DC) are now recognised as a unique leukocyte type, consisting of two or more subsets. The origins and functional inter-relationships of these cells are the subject of intense basic scientific investigation. They play important roles in initiating and directing immune responses, defending the host from pathogens and maintaining self tolerance. Fundamental studies are defining new molecules and mechanisms associated with DC function. The first methods for counting these rare blood cell populations are already providing interesting new clinical data. Indeed, abnormal DC function may contribute to deficiencies in the immune response against malignancies. Phase I trial data suggests that DC-based cancer vaccination protocols may contribute an important new biological approach to cancer therapy. Manipulation of DC to facilitate allogeneic transplantation and even to manage autoimmune disease are likely developments.

A soluble form of CD83 is released from activated dendritic cells and B lymphocytes, and is detectable in normal human sera

CD83 is an inducible glycoprotein expressed predominantly by dendritic cells (DC) and B lymphocytes. Expression of membrane CD83 (mCD83) is widely used as a marker of differentiated/ activated DC but its function and ligand(s) are presently unknown. We report the existence of a soluble form of CD83 (sCD83). Using both a sCD83-specific ELISA and Western blotting, we could demonstrate the release of sCD83 by mCD83(+) B cell and Hodgkin's disease-derived cell lines, but not mCD83(-) cells. Inhibition of de novo protein synthesis did not affect the release of sCD83 during short-term (2 h) culture of cell lines although mCD83 expression was significantly reduced, suggesting sCD83 is generated by the release of mCD83. Isolated tonsillar B lymphocytes and monocyte-derived DC, which are mCD83(low), released only low levels of sCD83 during culture. However, the differentiation/activation of these populations both up-regulated mCD83 and increased sCD83 release significantly. Analysis of sera from normal donors demonstrated the presence of low levels (121 +/- 3.6 pg/ml) of circulating sCD83. Further studies utilizing purified sCD83 and the analysis of sCD83 levels in disease may provide clues to the function and ligand(s) of CD83.

The murine chaperonin 10 gene family contains an intronless, putative gene for early pregnancy factor, Cpn10-rs1

Early pregnancy factor (EPF) is a secreted protein with growth regulatory and immunomodulatory properties. Human platelet-derived EPF shares amino acid sequence identity with chaperonin 10 (Cpn10), a mitochondrial matrix protein which functions as a molecular chaperone. The striking differences in cellular localization and function of the two proteins suggest differential regulation of production reflecting either alternative transcription of the same gene or transcription from different genes. In mammals and more distantly related genera, there is a large gene family with homology to CPN 10 cDNA, which includes intronless copies of the coding sequence. To determine whether this could represent the gene for EPF, we have screened a mouse genomic library and sequenced representative Cpn10 family members, looking for a functional gene distinct from that of Cpn 10, which could encode EPF. Eight distinct genes were identified. Cpn10 contains introns, while other members are intronless. Six of these appear to be pseudogenes, and the remaining member, Cpn10-rs1, would encode a full-length protein. The 309-bp open reading frame (ORF) is identical to that of mouse Cpn10 cDNA with the exception of three single-base changes, two resulting in amino acid changes. Only one further single nucleotide difference between the Cpn10-rs1 and Cpn10 cDNAs is observed, located in the 3' UTR. Single nucleotide primer extension was applied to discriminate between Cpn10-rs1 and Cpn10 expression. Cpn10, which is ubiquitous, was detected in all tissue samples tested, whereas Cpn10-rs1 was expressed selectively. The pattern was completely coincident with known patterns of EPF activity, strongly suggesting that Cpn10-rs1 does encode EPF. The complete ORF of Cpn10-rs1 was expressed in E. coli. The purified recombinant protein was found to be equipotent with native human platelet-derived EPF in the bioassay for EPF, the rosette inhibition test.

Effect of the toxic milk mutation (tx) on the function and intracellular localization of Wnd, the murine homologue of the Wilson copperATPase

Wilson disease is an autosomal recessive copper transport disorder resulting from defective biliary excretion of copper and subsequent hepatic copper accumulation and liver failure if not treated. The disease is caused by mutations in the ATP7B (WND) gene, which is expressed predominantly in the liver and encodes a copper-transporting P-type ATPase that is structurally and functionally similar to the Menkes protein (MNK), which is defective in the X-linked copper transport disorder Menkes disease. The toxic milk (tx) mouse has a clinical phenotype similar to Wilson disease patients and, recently, the tx mutation within the murine WND homologue (Wnd) of this mouse was identified, establishing it as an animal model for Wilson disease. In this study, cDNA constructs encoding the wild-type (Wnd-wt) and mutant (Wnd-tx) Wilson proteins (Wnd) were generated and expressed in Chinese hamster ovary (CHO) cells. The fx mutation disrupted the copper-induced relocalization of Wnd in CHO cells and abrogated Wnd-mediated copper resistance of transfected CHO cells. In addition, co-localization experiments demonstrated that while Wnd and MNK are located in the trans-Golgi network in basal copper conditions, with elevated copper, these proteins are sorted to different destinations within the same cell, Ultrastructural studies showed that with elevated copper levels, Wnd accumulated in large multivesicular structures resembling late endosomes that may represent a novel compartment for copper transport. The data presented provide further support for a relationship between copper transport activity and the copper-induced relocalization response of mammalian copper ATPases, and an explanation at a molecular level for the observed phenotype of fx mice.

Analysis of the microbial community structure and function of a laboratory scale enhanced biological phosphorus removal reactor

A laboratory scale sequencing batch reactor (SBR) operating for enhanced biological phosphorus removal (EBPR) and fed with a mixture of volatile fatty acids (VFAs) showed stable and efficient EBPR capacity over a four-year-period. Phosphorus (P), poly-beta-hydroxyalkanoate (PHA) and glycogen cycling consistent with classical anaerobic/aerobic EBPR were demonstrated with the order of anaerobic VFA uptake being propionate, acetate then butyrate. The SBR was operated without pH control and 63.67+/-13.86 mg P l(-1) was released anaerobically. The P% of the sludge fluctuated between 6% and 10% over the operating period (average of 8.04+/-1.31%). Four main morphological types of floc-forming bacteria were observed in the sludge during one year of in-tensive microscopic observation. Two of them were mainly responsible for anaerobic/aerobic P and PHA transformations. Fluorescence in situ hybridization (FISH) and post-FISH chemical staining for intracellular polyphosphate and PHA were used to determine that 'Candidatus Accumulibacter phosphatis' was the most abundant polyphosphate accumulating organism (PAO), forming large clusters of coccobacilli (1.0-1.5 mum) and comprising 53% of the sludge bacteria. Also by these methods, large coccobacillus-shaped gammaproteobacteria (2.5-3.5 mum) from a recently described novel cluster were glycogen-accumulating organisms (GAOs) comprising 13% of the bacteria. Tetrad-forming organisms (TFOs) consistent with the 'G bacterium' morphotype were alphaproteobacteria , but not Amaricoccus spp., and comprised 25% of all bacteria. According to chemical staining, TFOs were occasionally able to store PHA anaerobically and utilize it aerobically.

A clonal culture system demonstrates that IL-4 induces a subpopulation of noncytolytic T cells with low CD8, perforin, and granzyme expression

Immune deviation of cytolytic T cell function, induced by type 2 cytokines like IL-4, is an attractive concept to explain failure of the immune system in some diseases. However, this concept is challenged by previous conflicting results on whether type 2 cytokine-producing CD8(+) T cells are cytolytic. Therefore, we have analyzed the relationship between cytolytic activity and cytokine production among large numbers of primary CD8(+) T cell clones. Single murine CD8(+) T cells of naive phenotype were activated at high efficiency with immobilized Abs to CD3, CD8, and CD11a in the presence of IL-2 (neutral conditions) or IL-2, IL-4, and anti-IFN-gamma Ab (type 2-polarizing conditions) for 8-9 days. Under neutral conditions, most clones produced IFN-gamma without IL-4 and were cytolytic. Under type 2-polarizing conditions, most clones produced IFN-gamma and IL-4 but displayed variable cytolytic activity and CD8 expression. Separation on the basis of surface CD8 levels revealed that, compared with CD8(high) cells from the same cultures, CD8(low) cells were poorly cytolytic and expressed low levels of perforin mRNA and protein and granzyme A, B, and C mRNA. A similar, smaller population of noncytolytic CD8(low) cells was identified among CD8(low) T cells activated in mixed lymphocyte reaction with IL-4. Variable efficiency of generation of the noncytolytic cells may account for the differing results of earlier studies. We conclude that IL-4 promotes the development of a noncytolytic CD8(low) T cell phenotype that might be important in tumor- or pathogen-induced immune deviation.

Existence of a semicontinuous or continuous utility function: a unified approach and an elementary proof

In this paper, we present a new unified approach and an elementary proof of a very general theorem on the existence of a semicontinuous or continuous utility function representing a preference relation. A simple and interesting new proof of the famous Debreu Gap Lemma is given. In addition, we prove a new Gap Lemma for the rational numbers and derive some consequences. We also prove a theorem which characterizes the existence of upper semicontinuous utility functions on a preordered topological space which need not be second countable. This is a generalization of the classical theorem of Rader which only gives sufficient conditions for the existence of an upper semicontinuous utility function for second countable topological spaces. (C) 2002 Elsevier Science B.V. All rights reserved.