Modulation of protein synthesis and secretion by substratum in primary cultures of rat hepatocytes

Hepatocytes isolated by perfusion of adult rat liver and cultured on substrata consisting of one or more of the major components of the liver biomatrix (fibronectin, laminin, type IV collagen) have been examined for the synthesis of defined proteins. Under these conditions, tyrosine amino transferase, a marker of hepatocyte function, is maintained at similar levels in response to dexamethasone over 5 days in culture on each substratum, and total cellular protein synthesis remains constant. By contrast, there is a rapid decrease in synthesis and secretion of albumin and a 3-7-fold increase in synthesis and secretion of alpha-fetoprotein which are most marked on a laminin substratum, but least evident on type IV collagen, and an increased synthesis of fibronectin and type IV collagen. The newly synthesized matrix proteins are present in the cell layer as well as in cell secretions. The enhanced synthesis of fibronectin is less in cells seeded onto a fibronectin substratum than on laminin or type IV collagen substrata, and its synthesis by hepatocytes seeded onto a mixed substratum of laminin and fibronectin is down-regulated by fibronectin in a dose-related manner. Similarly, type IV collagen synthesis is less when the cells are seeded on the homologous matrix protein substratum than on heterologous substrata. These results indicate that hepatocytes cultured in serum-free medium on substrata composed of components of the liver biomatrix maintain certain functions of the differentiated state (tyrosine amino transferase), lose others (albumin secretion) and switch to increased synthesis of matrix components as well as fetal markers such as alpha-fetoprotein. The magnitude of these effects depends on the substratum on which the hepatocytes are cultured.     

P2 pyridine N-oxide thrombin inhibitors: a novel peptidomimetic scaffold

In this study, we have demonstrated that the critical hydrogen bonding motif of the established 3-aminopyrazinone thrombin inhibitors can be effectively mimicked by a 2-aminopyridine N-oxide. As this peptidomimetic core is more resistant toward oxidative metabolism, it also overcomes the metabolic liability associated with the pyrazinones. An optimization study of the P(1) benzylamide delivered the potent thrombin inhibitor 21 (K(i) = 3.2 nM, 2xaPTT = 360 nM), which exhibited good plasma levels and half-life after oral dosing in the dog (C(max) = 2.6 microM, t(1/2) = 4.5 h).     

Microbial-invertebrate interactions and potential for biotechnology

Conclusion As the interactions between marine invertebrates and their bacterial commensals and symbionts are better understood, the application of biotechnology will enhance both environmental and economic benefit. In the immediate future, marine bacteria, either selected or genetically engineered, will play a significant role in enhancing the development of selected invertebrates in aquaculture and in the field. Luck may also favor discovery of mechanisms to suppress the development of biofouling species, perhaps by making it possible to coat submerged surfaces with bacterial films designed to repell larvae and/or interfere with larval morphogenesis. In any case, the future is appealing.     

Validation of an algorithm for real-time measurement of sheep activity in confinement by recording movement within a commercial weighing crate

This study reports a method of measuring sheep activity in confinement based on recording movement of sheep during weighing. The aim was to show that an algorithm to analyse continuous weight records could be used to measure activity quickly and efficiently using commercial weighing equipment and that this measurement is related to animal productivity, specifically liveweight. A prototype method required magnetic switches on the weighing crate to record gate positioning and enclosure of the animal, with data analysis subsequent to the completion of weighing. The method reported here does not require additional equipment, instead using a computer program to analyse weights in real-time to record the degree of sheep movement. The result is summarised as the coefficient of variation of the weight over a period of 8–20 s and reported as an activity score.The real-time method had a higher repeatability (P < 0.05) than the prototype method and gave a stronger association with liveweight with heavier lambs and heavier young sheep being less active under confinement at 6 months and 18 months, respectively (P < 0.05).These findings indicate that a simple system could be developed enabling real-time reporting of activity scores during normal weighing.     

Conformational control in structure-based drug design

In structure-based drug design, the basic goal is to design molecules that fit complementarily to a given binding pocket. Since such computationally modeled molecules may not adopt the intended bound conformation outside the binding pocket, one challenge is to ensure that the designed ligands adopt similar low energy conformations both inside and outside of the binding pocket. Computational chemistry methods and conformational preferences of small molecules from PDB and Cambridge Structural Database (CSD) can be used to predict the bound structures of the designed molecules. Herein, we review applications of conformational control in structure-based drug design using selected examples from the recent medicinal chemistry literature. The main purpose is to highlight some intriguing conformational features that can be applied to other drug discovery programs.     

Investigation and documentation of hybridization betweenParkinsonia aculeata andCercidium praecox (Leguminosae:Caesalpinioideae)

Morphometric, cytogenetic, geographical and ecological evidence for hybridization betweenParkinsonia aculeata andCercidium praecox is presented. Morphometric investigation using the character count procedure and cytogenetic observations confirm hybrid status. All diagnostic morphometric characters were intermediate in the hybrid. Both parents (2n = 28) show regular tetrad formation and pollen fertility greater than 94%. Hybrids have a chromosome number of 2n = 28 or 2n = 30, and display meiotic abnormalities including lagging chromosomes and micronucleus formation; less than 21% of hybrid pollen was fertile. Ecological and geographical information suggests that hybridization is occurring at increasing frequency due to the expanding range ofP. aculeata associated with cultivation as an ornamental, coupled with ecological disturbance and weediness, and the cultivation ofC. praecox and hybrids as fodder, ornamental and shade trees. Hybrid fertility and phenological observations, in conjunction with F-weighted principal component analysis, suggest that the progeny of F₁ hybrids are established. The hybrid is formally described asP. ×carterae.     

Stress survival islet 1 (SSI-1) survey in Listeria monocytogenes reveals an insert common to listeria innocua in sequence type 121 L. monocytogenes strains

Listeria monocytogenes strains (n = 117) were screened for the presence of stress survival islet 1 (SSI-1). SSI-1(+) strains (32.5%) belonged mainly to serotypes 1/2c, 3b, and 3c. All sequence type 121 (ST-121) strains included (n = 7) possessed homologues to Listeria innocua genes lin0464 and lin0465 instead of SSI-1.     

Comparisons of Allergenic and Metazoan Parasite Proteins: Allergy the Price of Immunity

Allergic reactions can be considered as maladaptive IgE immune responses towards environmental antigens. Intriguingly, these mechanisms are observed to be very similar to those implicated in the acquisition of an important degree of immunity against metazoan parasites (helminths and arthropods) in mammalian hosts. Based on the hypothesis that IgE-mediated immune responses evolved in mammals to provide extra protection against metazoan parasites rather than to cause allergy, we predict that the environmental allergens will share key properties with the metazoan parasite antigens that are specifically targeted by IgE in infected human populations. We seek to test this prediction by examining if significant similarity exists between molecular features of allergens and helminth proteins that induce an IgE response in the human host. By employing various computational approaches, 2712 unique protein molecules that are known IgE antigens were searched against a dataset of proteins from helminths and parasitic arthropods, resulting in a comprehensive list of 2445 parasite proteins that show significant similarity through sequence and structure with allergenic proteins. Nearly half of these parasite proteins from 31 species fall within the 10 most abundant allergenic protein domain families (EF-hand, Tropomyosin, CAP, Profilin, Lipocalin, Trypsin-like serine protease, Cupin, BetV1, Expansin and Prolamin). We identified epitopic-like regions in 206 parasite proteins and present the first example of a plant protein (BetV1) that is the commonest allergen in pollen in a worm, and confirming it as the target of IgE in schistosomiasis infected humans. The identification of significant similarity, inclusive of the epitopic regions, between allergens and helminth proteins against which IgE is an observed marker of protective immunity explains the ‘off-target’ effects of the IgE-mediated immune system in allergy. All these findings can impact the discovery and design of molecules used in immunotherapy of allergic conditions.