Monte Carlo Study of Structural and Thermodynamic Properties of Liquid Chloroform Using a Five Site Model

A five site potential model combining Lennard–Jones plus Coulomb potential functions has been developed for chloroform molecule. The partial charges needed for Coulombic interactions were derived using the chelpg procedure implemented in the gaussian 92 program. These calculations were performed at the MP2 level with MC-311G* basis set for Cl and 6-311G** for C and H atoms. The parameters for the Lennard–Jones potentials were optimized to reproduce experimental values for the density and enthalpy of vaporization of the pure liquid at 298 K and 1 atm. The statistical mechanics calculations were performed with the Monte Carlo method in the isothermic and isobaric (NpT) ensemble. Besides the values obtained for density, ρ, and molar enthalpy of vaporization at constant pressure, Δ H_V, for liquid chloroform, results for molar volume, V_m, molar heat capacity, C_p, isobaric thermal expansivity, α_p, and isothermal compressibility, κ_T, for this pure liquid are also in very good agreement with experimental observations. Size effects on the values of thermodynamic properties were investigated. The potential model was also tested by computing the free energy for solvating one chloroform molecule into its own liquid at 298 K using a statistical perturbation approach. The result obtained compares well with the experimental value. Site–site pair correlation functions were calculated and are in good accordance with theoretical results available in the literature. Dipole–dipole correlation functions for the present five site model were also calculated at different carbon–carbon distances. These correlations were compared to those obtained using the four site model reported in the literature. An investigation of the solvent dependence of the relative free energy for cis/trans conversion of a hypothetical solute in TIP4P water and chloroform was accomplished. The results show strong interaction of water and chloroform molecules with the gauche conformer. The value obtained for the free energy barrier for cis/trans rotation in TIP4P water is higher than that for chloroform. This result is in agreement with the continuous theory for solvation as the conformer with higher dipole moment is more favoured by the solvent with higher dieletric constant. The results also show an increase in entropy as the solute goes from the cis to the trans geometry and this result is more appreciable in the aqueous solution. This revised version was published online in June 2006 with corrections to the Cover Date.     

In Vivo and In Vitro Effects of Some Plant Hormones on Rat Erythrocyte Carbonic Anhydrase and Glucose-6-Phosphate Dehydrogenase Activities

The present study was undertaken to determine in vivo and in vitro effects of some plant growth regulators on rat erythrocyte carbonic anhydrase (CA) and glucose-6-phosphate dehydrogenase (G6PD) activities. Both in vivo and in vitro, spermidine and kinetin did not affect enzymatic activities of CA and G6PD, whereas putrescine decreased these activities, and abscisic acid increased them. Since plants use such growth regulators, their effects should be considered on mammals consuming them since they may possess important biological effects.     

Basic fibroblast growth factor, neurofilament, and glial fibrillary acidic protein immunoreactivities in the myenteric plexus of the rat esophagus and colon

The enteric nervous system consists of a number of interconnected networks of neuronal cell bodies and fibers as well as satellite cells, the enteric glia. Basic fibroblast growth factor (bFGF) is a mitogen for a variety of mesodermal and neuroectodermal-derived cells and its presence has been described in many tissues. The present work employs immunohistochemistry to analyze neurons and glial cells in the esophageal and colic enteric plexus of the Wistar rat for neurofilament (NF) and glial fibrillary acidic proteins (GFAP) immunoreactivity as well as bFGF immunoreactivity in these cells. Rats were processed for immunohistochemistry; the distal esophagus and colon were opened and their myenteric plexuses were processed as whole-mount preparations. The membranes were immunostained for visualization of NF, GFAP, and bFGF. NF immunoreactivity was seen in neuronal cell bodies of esophageal and colic enteric ganglia. GFAP-immunoreactive enteric glial cells and processes were present in the esophageal and colic enteric plexuses surrounding neuronal cell bodies and axons. A dense net of GFAP-immunoreactive processes was seen in the ganglia and connecting strands of the myenteric plexus. bFGF immunoreactivity was observed in the cytoplasm of the majority of the neurons in the enteric ganglia of esophagus and colon. The two-color immunoperoxidase and immunofluorescence methods revealed bFGF immunoreactivity also in the nucleus of GFAP-positive enteric glial cells. The results suggest that immunohistochemical localization of NF and GFAP may be an important tool in the study of the plasticity in the enteric nervous system. The presence of bFGF in neurons and glia of the myenteric plexus of the esophagus and the colon indicates that this neurotrophic factor may exert autocrine and paracrine actions in the enteric nervous system.     

Fuzzy modeling in symptomatic HIV virus infected population

This paper introduces a model for the evolution of positive HIV population and manifestation of AIDS (acquired immunideficiency syndrome). The focus is on the nature of the transference rate of HIV to AIDS. Expert knowledge indicates that the transference rate is uncertain and depends strongly on the viral load and the CD4+ level of the infected individuals. Here, we suggest to view the transference rate as a fuzzy set of the viral load and CD4+ level values. In this case the dynamic model results in a fuzzy model that preserves the biological meaning and nature of the transference rate λ. Its behavior fits the natural history of HIV infection reported in the medical science domain. The paper also includes a comparison between the fuzzy model and a classic Anderson’s model using data reported in the literature.     

FGF-2 and S100beta immunoreactivities increase in reactive astrocytes, but not in microglia, in ascending dopamine pathways following a striatal 6-OHDA-induced partial lesion of the nigrostriatal system

Partial lesions were induced in rat midbrain dopamine ascending pathways by intrastriatal injection of 6-hydroxydopamine (6-OHDA), and after two weeks changes were observed in the immunoreactivities of S100beta, a calcium-binding protein, and basic fibroblast growth factor (FGF-2), which is neurotrophic. Semiquantitative microdensitometric image analysis revealed increased intensities of FGF-2 and S100beta immunostaining in putative glial profiles of the ipsilateral neostriatum, pars compacta (SNc) and reticulata (SNr) of the substantia nigra and ventral tegmental area (VTA). Double immunofluorescence and immunoperoxidase procedures, using antibodies against glial fibrillary acidic protein and OX-42, showed that these increased immunoreactivities were restricted to reactive astrocytes; they were not observed in reactive microglia. These results indicate that reactive astrocytes may exert paracrine trophic actions through S100beta and FGF-2 in the midbrain dopamine ascending pathways after striatal 6-OHDA treatment. Interactions between S100beta and FGF-2 may be relevant to neuronal maintenance and repair following dopamine injury.     

A biologically inspired neural network for dynamic programming

An artificial neural network with a two-layer feedback topology and generalized recurrent neurons, for solving nonlinear discrete dynamic optimization problems, is developed. A direct method to assign the weights of neural networks is presented. The method is based on Bellmann's Optimality Principle and on the interchange of information which occurs during the synaptic chemical processing among neurons. The neural network based algorithm is an advantageous approach for dynamic programming due to the inherent parallelism of the neural networks; further it reduces the severity of computational problems that can occur in methods like conventional methods. Some illustrative application examples are presented to show how this approach works out including the shortest path and fuzzy decision making problems.     

Optimizing Human Synovial Fluid Preparation for Two-Dimensional Gel Electrophoresis

Background

Prenatal screening for Down Syndrome (DS) would benefit from an increased number of biomarkers to improve sensitivity and specificity. Improving sensitivity and specificity would decrease the need for potentially risky invasive diagnostic procedures.

Results

We have performed an in depth two-dimensional difference gel electrophoresis (2D DIGE) study to identify potential biomarkers. We have used maternal plasma samples obtained from first and second trimesters from mothers carrying DS affected fetuses compared with mothers carrying normal fetuses. Plasma samples were albumin/IgG depleted and expanded pH ranges of pH 4.5 - 5.5, pH 5.3 - 6.5 and pH 6 - 9 were used for two-dimensional gel electrophoresis (2DE). We found no differentially expressed proteins in the first trimester between the two groups. Significant up-regulation of ceruloplasmin, inter-alpha-trypsin inhibitor heavy chain H4, complement proteins C1s subcomponent, C4-A, C5, and C9 and kininogen 1 were detected in the second trimester in maternal plasma samples where a DS affected fetus was being carried. However, ceruloplasmin could not be confirmed as being consistently up-regulated in DS affected pregnancies by Western blotting.

Conclusions

Despite the in depth 2DE approach used in this study the results underline the deficiencies of gel-based proteomics for detection of plasma biomarkers. Gel-free approaches may be more productive to increase the number of plasma biomarkers for DS for non-invasive prenatal screening and diagnosis.     

Alimentary prion infections: Touchdown in the intestine

Neurodegenerative diseases are caused by proteinaceous aggregates, usually consisting of misfolded proteins which are often typified by a high proportion of β-sheets that accumulate in the central nervous system. These diseases, including Morbus Alzheimer, Parkinson disease and Transmissible Spongiform Encephalopathies (TSEs)—also termed prion disorders—afflict a substantial proportion of the human population and, as such, the etiology and pathogenesis of these diseases has been the focus of mounting research. Although many of these diseases arise from genetic mutations or are sporadic in nature, the possible horizontal transmissibility of neurodegenerative diseases poses a great threat to population health. In this article we discuss recent studies that suggest that the “non-transmissible” status bestowed upon Alzheimer and Parkinson diseases may need to be revised as these diseases have been successfully induced through tissue transplants. Furthermore, we highlight the importance of investigating the “natural” mechanism of prion transmission including peroral and perenteral transmission, proposed routes of gastrointestinal uptake and neuroinvasion of ingested infectious prion proteins. We examine the multitude of factors which may influence oral transmissibility and discuss the zoonotic threats that Chronic Wasting disease (CWD), Bovine Spongiform Encephalopathy (BSE) and Scrapie may pose resulting in vCJD or related disorders. In addition, we suggest that the 37 kDa/67 kDa laminin receptor on the cell surface of enterocytes, a major cell population in the intestine, may play an important role in the intestinal pathophysiology of alimentary prion infections.Key words: prion, 37 kDa/67 kDa laminin receptor, CJD, BSE, CWD, scrapie, Alzheimer disease, Parkinson disease, intestine, enterocytesMany different mechanisms exist which underlie the etiology of the numerous neurodegenerative diseases affecting the human population. Amongst the most prominent are Morbus Alzheimer, prion disorders, Parkinson disease, Chorea Huntington, frontotemporal dementia and amylotrophic lateral sclerosis. The molecular mechanisms underlying these diseases vary; however, all neurodegenerative diseases share a common feature: they are caused by protein aggregation. The only neurodegenerative diseases proven to be transmissible are prion disorders. In contrast to frontotemporal dementia, recent evidence suggests that Alzheimer and Parkinson diseases may also be transmissible. Pre-symptomatic Alzheimer disease (APP23) mice exhibited an increase in the Alzheimer phenotype when brain homogenate of autopsied human Alzheimer disease patients and older, amyloid beta- (Aβ-) laden APP23 mice was injected into their hippocampi.¹ These findings suggest that the Aβ-abundant brain homogenate of Alzheimer disease patients may possess the ability to induce or supplement the overproduction of Aβ, possibly leading to the onset of Alzheimer disease.The pathological feature associated with Parkinson disease is the formation of Lewy bodies in cell bodies and neuronal processes in the brain.² The main component of these protein aggregates is α-synuclein (reviewed in ref. ²). Autopsies of Parkinson disease patients revealed that Lewy bodies had formed on healthy embryonic neurons that had been grafted onto the brain tissue of the patients several years before (prior to said examination).^3–5 It may thus be proposed that α-synuclein transmission is possible from diseased to healthy neurons, suggesting that Parkinson disease may be transmissible from a Parkinson disease patient to a healthy individual. These findings imply that Alzheimer and Parkinson diseases may be transmissible through tissue transplants and the use of contaminated surgical tools.⁶Prion disorders, also termed Transmissible Spongiform Encephalopathies (TSEs), are fatal neurodegenerative diseases that affect the central nervous system (CNS) of multiple animal species. In lieu of the social, economic and political ramifications of such infections, as well as the possible intra- and interspecies transmissibility of such disorders, various routes of experimental transmission have been investigated including intracerebral, intraperitoneal, intraventricular, intraocular, intraspinal and subcutaneous injections (reviewed in ref. ^7–9). However, such routes of transmission are not representative of the “natural” mechanism as the majority of prion disorders are contracted through ingestion of infectious prion (PrP^Sc) containing material. Thus, the peroral and perenteral prion transmission is of greatest consequence with respect to TSE disease establishment. Moreover, the presence of PrP^Sc in the buccal cavity of scrapie-infected sheep¹⁰ (reviewed in ref. ¹¹) and the possible horizontal transfer as a result hereof, as may be similarly proposed for animals suffering from other TSEs, may further contribute to the oral transmissibility of TSEs.A number of model systems have been employed to study TSE transmissibility. Owing to ethical constraints, TSE transmissibility to humans via the oral route may not be directly investigated and as a result hereof, alternative model systems are needed. These may include the use of transgenic mice, cell lines which are permissive to infection¹² and experimental animals such as sheep, calves, goats, minks, ferrets and non-human primates (reviewed in ref. ⁹).Intestinal entry of PrP^Sc has been proposed to occur via two pathways, the membranous (M) cell-dependent and M cell-independent pathways (Fig. 1).^13,14 The former involves endocytic M (microfold)-cells, which cover the intestinal lymphoid follicles (Peyer''s patches)¹⁴ and may take up prions and thereby facilitate the translocation of these proteins across the intestinal epithelium into the lymphoid tissues (reviewed in ref. ⁹) as has been demonstrated in a cellular model.¹³ Following such uptake by the M cells, the prions may subsequently pass to the dendritic cells and follicular dendritic cells (FDCs) (Fig. 1), which allow for prion transport to the mesenteric lymph nodes and replication, respectively.¹⁵ The prion proteins may subsequently gain access to the enteric nervous system (ENS) and ultimately the central nervous system (CNS).¹⁵Open in a separate windowFigure 1Proposed routes of gastrointestinal entry of ingested infectious prions (PrP^Sc) as well as possible pathways of amplification and transport to the central nervous system.However, prion intestinal translocation has been observed in the absence of M cells and has been demonstrated to be as a result of the action of polar, 37 kDa/67 kDa LRP/LR (non-integrin laminin receptor; reviewed in ref. ^16–18) expressing enterocytes. Enterocytes are the major cell population of the intestinal epithelium and due to their ability to endocytose pathogens, nutrients and macromolecules,¹⁹ it has been proposed that these cells may represent a major entry site for alimentary prions (Fig. 1).Since enterocyte prion uptake has been demonstrated to be dependent on the presence of LRP/LR on the apical brush border of the cells,^14,20 the interaction between varying prion protein strains and the receptor^21–23 may be employed as a model system to study possible oral transmissibility of prion disorders across species as well as the intestinal pathophysiology of alimentary prion infections.²⁴ Moreover, the blockage of such interactions through the use of anti-LRP/LR specific antibodies has been reported to reduce PrP^Sc endocytosis¹⁹ and thus these antibodies may serve as potential therapeutics to prevent infectious prion internalization and thereby prevent prion infections. It must be emphasized that the adhesion of prion proteins to cells is not solely dependent on the LRP/LR-PrP^Sc interactions;²⁴ however, this interaction is of importance with regards to internalization and subsequent pathogenesis.We applied the aforementioned cell model to study the possible oral transmission of PrP^BSE, PrP^CWD and ovine PrP^Sc to cervids, cattle, swine and humans.²⁴ The direct transmission of the aforementioned animal prion disorders to humans as a result of dietary exposure and the possible establishment of zoonotic diseases is of great public concern. It must however be emphasized that the study investigated the co-localization of LRP/LR and various prion strains and not the actual internalization process.PrP^BSE was shown to co-localize with LRP/LR on human enterocytes²⁴, thereby suggesting that PrP^BSE is transmissible to humans via the oral route which is widely accepted as the manner by which variant CJD originated. This suspicion was previously investigated using a macaque model, which was successfully perorally infected by BSE-contaminated material and subsequently lead to the development of a prion disorder that resembles vCJD.²⁵ These results, due to the evolutionary relatedness between macaques and humans, allowed researchers to confirm the oral transmissibility of PrP^BSE to humans. PrP^BSE may also potentially lead to prion disorder establishment in swine,²⁴ livestock of great economic and social importance.The prion disorder affecting elk, mule deer and white-tailed deer is termed CWD. Cases of the disease are most prevalent in the US but are also evident in Canada and South Korea.^26,27 As the infectious prion isoform is reported to be present in the blood²⁸ and skeletal muscle,²⁹ hunting, consumption of wild venison and contact with other animal products derived from CWD-infected elk and deer may thereby pose a public health risk. Our studies demonstrate that PrP^CWD co-localizes with LRP/LR on human enterocytes²⁴ thereby suggesting a possible oral transmissibilty of this TSE to humans. This is, however, inconsistent with results obtained during intra-cerebral inoculation of the brains and spinal cords of transgenic mice overexpressing the human cellular prion protein (PrP^c),^26,27 which is essential for TSE disease establishment and progression. Further, discrepancies have also been reported with respect to non-human primates, as squirrel monkeys have been successfully intracerebrally inoculated with mule-deer prion homogenates,³⁰ while cynolmolgus macaques were resistant to infection.³¹ CWD has been transmitted to ferrets, minks and goats³² and as these animals may serve as domestic animals or livestock, secondary transmission from such animals to humans, through direct contact or ingestion of infected material, may be an additional risk factor that merits further scientific investigation.Ovine PrP^Sc co-localization with LRP/LR on human and bovine enterocytes may be indicative of the infectious agents'' ability to effect cross-species infections. The oral transmissibility of Scrapie has been confirmed in hamsters fed with sheep-scrapie-infected material.³³The discrepancies with regards to the transmissibility of certain infectious prion proteins when assessed by different model systems may be due to the experimental transmission route employed. Oral exposure often results in significantly prolonged incubation times when compared to intracerebral inoculation techniques and thus failure of transgenic mice and normal experimental animals to develop disease phenotypes after being fed TSE-contaminated material may not necessarily indicate that the infection process failed.¹⁴ Apart from the route of infection, numerous other factors may influence transmission between species, including dose, PrP polymorphisms and genetic factors, the prion strain employed as well as the efficacy of prion transport to the CNS.³⁴ The degree of homology between the PrP^c protein in the animals serving as the infectious prion source and recipient has also been described as a feature limiting cross-species transmission.³⁴ The negative results, as referred to above, obtained upon prion-protein inoculation of animal models may have resulted due to the slow rate at which the infectious prion induces conformational conversion of the endogenous PrP^c in the animal cells and this in turn results in low levels of infectious prion replication and symptom development.²⁷Furthermore, even in the event that certain prion disorders are not directly transmissible to humans, most are transmissible to at least a single species of domestic animal or livestock. The infectious agents properties may be altered in the secondary host such that it becomes transmissible to humans (reviewed in ref. ³⁵). Thus, interspecies transmission between animals may indirectly influence human health.It is noteworthy to add that although the oral route of PrP^Sc transmission may result in prolonged incubation times, it may broaden the range of susceptible hosts. A common constituent of food is ferritin, a protein that is resistant to digestive enzyme hydrolysis and, due to its homology across species, it may serve as co-transporter of PrP^Sc and facilitate enterocyte internalization of the infectious prion.³⁶ It may thus be proposed that prion internalization may occur via a ferritin-PrP^Sc complex even in the absence of co-localization between the infectious agent and LRP/LR such that many more cross-species infections (provided that the other infection factors are favorable) may be probable.³⁷ In addition, digestive enzymes in the gastrointestinal tract facilitate PrP^Sc binding to the intestinal epithelium and subsequent intestinal uptake³⁶ and thus depending on the individuals'' digestive processes, the susceptibility to infection and the rate of disease development may vary accordingly. As a result hereof, though laboratory experiments in cell-culture and animal models may render a particular prion disorder non-infectious to humans, this may not be true for all individuals.In lieu of the above statements, with particular reference to inconsistencies in reported results and the multiple factors influencing oral transmissibility of TSEs, further transmission studies are required to evaluate the zoonotic threat which CWD, BSE and Scrapie may pose through ingestion.     

Studying the genetic basis of drought tolerance in sorghum by managed stress trials and adjustments for phenological and plant height differences

Managed environments in the form of well watered and water stressed trials were performed to study the genetic basis of grain yield and stay green in sorghum with the objective of validating previously detected QTL. As variations in phenology and plant height may influence QTL detection for the target traits, QTL for flowering time and plant height were introduced as cofactors in QTL analyses for yield and stay green. All but one of the flowering time QTL were detected near yield and stay green QTL. Similar co-localization was observed for two plant height QTL. QTL analysis for yield, using flowering time/plant height cofactors, led to yield QTL on chromosomes 2, 3, 6, 8 and 10. For stay green, QTL on chromosomes 3, 4, 8 and 10 were not related to differences in flowering time/plant height. The physical positions for markers in QTL regions projected on the sorghum genome suggest that the previously detected plant height QTL, Sb-HT9-1, and Dw2, in addition to the maturity gene, Ma5, had a major confounding impact on the expression of yield and stay green QTL. Co-localization between an apparently novel stay green QTL and a yield QTL on chromosome 3 suggests there is potential for indirect selection based on stay green to improve drought tolerance in sorghum. Our QTL study was carried out with a moderately sized population and spanned a limited geographic range, but still the results strongly emphasize the necessity of corrections for phenology in QTL mapping for drought tolerance traits in sorghum.     

Using linear algebra for protein structural comparison and classification

In this article, we describe a novel methodology to extract semantic characteristics from protein structures using linear algebra in order to compose structural signature vectors which may be used efficiently to compare and classify protein structures into fold families. These signatures are built from the pattern of hydrophobic intrachain interactions using Singular Value Decomposition (SVD) and Latent Semantic Indexing (LSI) techniques. Considering proteins as documents and contacts as terms, we have built a retrieval system which is able to find conserved contacts in samples of myoglobin fold family and to retrieve these proteins among proteins of varied folds with precision of up to 80%. The classifier is a web tool available at our laboratory website. Users can search for similar chains from a specific PDB, view and compare their contact maps and browse their structures using a JMol plug-in.