Elaboration of immune stimulating lipid-saponin subunit antigen carrier based on glycolipid monogalactosyldiacylglycerol from sea macrophytes and triterpene glycosides from Cucumaria japonica

The ability of some triterpene glycosides of holothurians: holotoxin A₁ from Apostichopus japonicus and a mixture of monosulfated triterpene glycosides from Cucumaria japonica called cucumarioside (CD) to form supramolecular complexes with cholesterol (Chol) and monogalactosyldiacylglycerol (MGDG) or phosphatidylcholine (PC) was studied. A transmission electron microscopy method was used to observe supramolecular lipid-saponin complexes formed by holotoxin A1 and CD with cholesterol in the presence of membrane lipids. The observed supramolecular complexes are tubular nanoparticles with a length of 100–300 nm, an external diameter of 10–16 nm and an internal diameter of 2–6 nm. The formation of tubular nanoparticles was more effective in the presence of MGDG than with PC. Nanoparticles forming in the presence of MGDG are shaped as a tubule, have a constant diameter and a strongly pronounced internal channel. In contrast, PC has no such properties; this lipid is unable to fully integrate in tubular nanoparticles. Based on electron-microscopy data the range of weight ratio of MGDG-Chol-CD was determined as a 1–10: 2: 3 that provided most effective formation of tubular nanoparticles. Different methods of incorporation of model antigens in complex MGDG-Chol-CD were studied. Influenza hemagglutinin and neuraminidase from commercial vaccine “Influvac” and pore forming protein YompF from Yersinia pseudotuberculosis were used as model antigens. From 54 to 72% of protein of “Influvac” vaccine and 88–92% of YompF were incorporated in supramolecular complexes depending on the method of incorporation. The loss of functional activity of hemagglutinin of vaccine “Influvac” was the result of applying ultrasonic disintegration for incorporation of this protein in complex MGDG-Chol-CD. YompF incorporation in MGDG-Chol-CD complex led to the increased diameter of tubular particles, in the same time incorporation of vaccine “Influvac” antigens produced the “cap” formation at the end of tubules. The possibility of a described supramolecular complex MGDG-Chol-CD to be a carrier for subunit bacterial and viral antigens is shown.     

Personalized In Vitro Cancer Modeling — Fantasy or Reality?

With greater technological advancements and understanding of pathophysiology, “personalized medicine” has become a more realistic goal. In the field of cancer, personalized medicine is the ultimate objective, as each cancer is unique and each tumor is heterogeneous. For many decades, researchers have relied upon studying the histopathology of tumors in the hope that it would provide clues to understanding the pathophysiology of cancer. Current preclinical research relies heavily upon two-dimensional culture models. However, these models have had limited success in recreating the complex interactions between cancer cells and the stroma environment in vivo. Thus, there is increasing impetus to shift to three-dimensional models, which more accurately reflect this phenomenon. With a more accurate in vitro tumor model, drug sensitivity can be tested to determine the best treatment option based on the tumor characteristics. Many methods have been developed to create tumor models or “tumoroids,” each with its advantages and limitations. One significant problem faced is the replication of angiogenesis that is characteristic of tumors in vivo. Nonetheless, if three-dimensional models could be standardized and implemented as a preclinical research tool for therapeutic testing, we would be taking a step towards making personalized cancer medicine a reality.     

Lipid-dependent sequential allosteric activation of heat-sensing TRPV1 channels by anchor-stereoselective “hot” vanilloid compounds and analogs

Both a silent resident phosphatidylinositol lipid and a “hot” vanilloid agonist capsaicin or resiniferatoxin have been shown to share the same inter-subunit binding pocket between a voltage sensor like domain and a pore domain in TRPV1. However, how the vanilloid competes off the resident lipid for allosteric TRPV1 activation is unknown. Here, the in silico research suggested that anchor-stereoselective sequential cooperativity between an initial recessive transient silent weak ligand binding site and a subsequent dominant steady-state strong ligand binding site in the vanilloid pocket may facilitate the lipid release for allosteric activation of TRPV1 by vanilloids or analogs upon non-covalent interactions. Thus, the resident lipid may play a critical role in allosteric activation of TRPV1 by vanilloid compounds and analogs.     

Coupling of homotropic and heterotropic interactions in Escherichia coli aspartate transcarbamylase

Several types of conditions allow the disconnection of homotropic and heterotropic interactions in Escherichia coli aspartate transcarbamylase. A model that includes a concerted gross conformational change corresponding to the homotropic cooperative interactions between the catalytic sites and local “site by site” effects promoted by the effectors accounts for this disconnection as well as for the other known properties of the enzyme. However, the substrate concentration influences the extent of stimulation and feedback inhibition of the catalytic activity by the effectors. This result is explained by assuming that these effectors promote a “primary effect”, which is exerted locally “site by site”, and a “secondary effect”, which is mediated by the substrate. As predicted by the model, relaxed (R) forms of the enzyme show only the primary effect. In addition 2-ThioU-aspartate transcarbamylase, a modified form of the enzyme in which the homotropic cooperative interactions between the catalytic sites are selectively abolished, shows the same heterogeneity in CTP binding sites as normal aspartate transcarbamylase.     

The Excitations and Suppressions of the Times: Locating the Emotions in the Liver in Modern Chinese Medicine

This paper explores how doctors of Chinese medicine have borrowed from a long history of scholarship on the problem of “constraint” to develop treatments for modern emotion-related disorders, such as depression. I argue that this combining of medical practices was made possible by a complex sequence of events. First, doctors in the 1920 and 1930s were engaged in a critical reexamination of the entire corpus of Chinese medical knowledge. Spurred by the encounter with European imperialism, the sudden rise of Japan as a new power in East Asia, and the political struggles to establish a Chinese nation state, these scholars were among the first to speculate on the possible relationship between Chinese medicine and Western medicine. Second, in the 1950 and 1960s, doctors like other intellectuals were focused on national reunification and institution building. They rejected some of the experimental claims of their predecessors to focus on identifying the key characteristics of Chinese medicine, such as the methodology of “pattern recognition and treatment determination bianzheng lunzhi.” The flexibility of the new bianzheng lunzhi paradigm allowed doctors to quietly adopt innovations from their early twentieth century counterparts that they ostensibly rejected, ultimately paving the way for contemporary treatments of depression.     

Supramolecular structure of the OXPHOS system in highly thermogenic tissue of Arum maculatum

The protein complexes of the mitochondrial respiratory chain associate in defined ways forming supramolecular structures called respiratory supercomplexes or respirasomes. In plants, additional oxidoreductases participate in respiratory electron transport, e.g. the so-called “alternative NAD(P)H dehydrogenases” or an extra terminal oxidase called “alternative oxidase” (AOX). These additional enzymes were previously reported not to form part of respiratory supercomplexes. However, formation of respiratory supercomplexes might indirectly affect “alternative respiration” because electrons can be channeled within the supercomplexes which reduces access of the alternative enzymes towards their electron donating substrates. Here we report an investigation on the supramolecular organization of the respiratory chain in thermogenic Arum maculatum appendix mitochondria, which are known to have a highly active AOX for heat production. Investigations based on mild membrane solubilization by digitonin and protein separation by blue native PAGE revealed a very special organization of the respiratory chain in A. maculatum, which strikingly differs to the one described for the model plant Arabidopsis thaliana: (i) complex I is not present in monomeric form but exclusively forms part of a I + III₂ supercomplex, (ii) the III₂ + IV and I + III₂ + IV supercomplexes are detectable but of low abundance, (iii) complex II has fewer subunits than in A. thaliana, and (iv) complex IV is mainly present as a monomer in a larger form termed “complex IVa”. Since thermogenic tissue of A. maculatum at the same time has high AOX and I + III₂ supercomplex abundance and activity, negative regulation of the alternative oxidase by supercomplex formation seems not to occur. Functional implications are discussed.     

Comprehensive phylogenetic analyses of the Ruppia maritima complex focusing on taxa from the Mediterranean

Recent molecular phylogenetic studies reported high diversity of Ruppia species in the Mediterranean. Multiple taxa, including apparent endemics, are known from that region, however, they have thus far not been exposed to phylogenetic analyses aimed at studying their relationships to taxa from other parts of the world. Here we present a comprehensive phylogenetic analyses of the R. maritima complex using data sets composed of DNA sequences of the plastid genome, the multi-copy nuclear ITS region, and the low-copy nuclear phyB gene with a primary focus on the Mediterranean representatives of the complex. As a result, a new lineage, “Drepanensis”, was identified as the seventh entity of the complex. This lineage is endemic to the Mediterranean. The accessions included in the former “Tetraploid” entity were reclassified into two entities: an Asia–Australia–Europe disjunct “Tetraploid_α” with a paternal “Diploid” origin, and a European “Tetraploid_γ” originating from a maternal “Drepanensis” lineage. Another entity, “Tetraploid_β”, is likely to have been originated as a result of chloroplast capture through backcrossing hybridization between paternal “Tetraploid_α” and maternal “Tetraploid_γ”. Additional discovery of multiple tetraploidizations as well as hybridization and chloroplast capture at the tetraploid level indicated that hybridization has been a significant factor in the diversification of Ruppia.     

The “Pre-Molten Globule,” a New Intermediate in Protein Folding

In vitro folding studies of several proteins revealed the formation, within 2–4 msec, of transient intermediates with a large far-UV ellipticity but no amide proton protection. To solve the contradiction between the secondary structure contents estimated by these two methods, we characterized the isolated C-terminal fragment F2 of the tryptophan synthase β₂ subunit. In β₂, F2 forms its tertiary interactions with the F1 N-terminal region. Hence, in the absence of F1, isolated F2 should remain at an early folding stage with no long-range interactions. We shall show that isolated F2 folds into, and remains in, a “state” called the pre-molten globule, that indeed corresponds to a 2- to 4-msec intermediate. This condensed, but not compact, “state” corresponds to an array of conformations in rapid equilibrium comprising native as well as nonnative secondary structures. It fits the “new view” on the folding process.     

A new conception of the shoot of higher plants

According to the classical model, the “shoot” consists only of the categories “caulome” (“stem” sensu lato) and “phyllome” (“leaf” sensu lato), (and “root” in cases of “adventitious” root formation). If lateral shoots are present, their position is axillary. Consequently, caulome as well as phyllome are inserted on the caulome and only on the caulome. This classical model of the shoot has two disadvantages of great consequence: (1) Intermediate organs cannot be accepted as such, but have to be interpreted (i.e. categorized) as either caulome or phyllome (or root) by distortion of the actual similarity. (2) Certain positional changes of organs cannot be accepted as such, but have to be “explained” by congenital fusion. The new conception of the shoot will have the advantages of the classical model but not its disadvantages. Hence, the shoot may consist of the following parts: (main and lateral) shoot, caulome, phyllome, root, emergence, and structures intermediate between (i.e. partially homologous to) any of the preceding. Thus, the five categories of the classical model, namely “shoot”, “caulome”, “phyllome”, “root” and “emergence” are no longer mutually exclusive; they may merge into each other due to an actual or potential continuum. Intermediate organs are therefore accepted as such; for example, an organ may be characterized as an intermediate form between a caulome and a phyllome. Besides intermediate forms, all changes in position are accepted as such. Hence, the following positional relations are possible: caulome and phyllome may be inserted on the caulome, caulome and phyllome may be inserted on the phyllome; roots may be inserted on caulome or phyllome; intermediate forms may be inserted on the caulome, phyllome, or other intermediate forms. Consequences of the new conception for morphological research are pointed out, especially for homologization, evolutionary considerations, and the direction in which research progresses.     

Actividad de la micafungina contra las biopelículas de Candida

BackgroundMost recalcitrant infections are associated to colonization and microbial biofilm development. These biofilms are difficult to eliminate by the immune response mechanisms and the current antimicrobial therapy.AimTo describe the antifungal of micafungin against fungal biofilms based in the scientific and medical literature of recent years.MethodsWe have done a bibliographic retrieval using the scientific terms “micafungin”, “activity”, “biofilm”, “Candida”, “Aspergillus”, “fungi”, “mycos”*, susceptibility, in PubMed/Medline from the National Library of Medicine from 2006 to 2009.ResultsMost current antifungal agents (amphotericin B and fluconazole) and the new azole antifungals have no activity against fungal biofilms. However, micafungin and the rest of echinocandins are very active against Candida albicans, Candida dubliniensis, Candida glabrata, and Candida krusei biofilms but their activities are variable and less strong against Candida tropicalis and Candida parapsilosis biofilms. Moreover, they have not activities against the biofilms of Cryptococcus y Trichosporon.ConclusionsThe activity of micafungin against Candida biofilms gives more strength to its therapeutic indication for candidaemia and invasive candidiasis associated to catheter, prosthesis and other biomedical devices.     

Qualitative dynamics of a network model of regulation of the immune system: A rationale for the IgM to IgG switch

A mathematical model has been developed that simulates some of the main features of a network theory of regulation of the immune system. According to the network viewpoint, the V regions (idiotypes) on antibodies and lymphocytes are self-antigens, to which other lymphocytes of the system can respond specifically, just as they respond to foreign antigens. The resultant couplings between the lymphocytes are considered to be basic for the regulation of the system.The present mathematical model simulates the interactions between cells that recognize the antigen (“positive cells”), and “negative cells” that have receptors that specifically recognize the V regions of the positive cells. The mathematical model incorporates only the interactions that are postulated to be important in the four steady states of the theory, and includes neither the antigen nor any accessory (“A”) cells. The effects of both antigen-specific and anti-idiotypic T and B cells are included, as well as antigen-specific and anti-idiotypic T cell factors, and the two main classes of antibodies. The model is a first order autonomous ordinary differential equation in two variables. We describe a geometric technique that gives strong information on the model, without explicitly solving the ordinary differential equation. This technique proves to be powerful in permitting us to systematically scan the parameter space of the model. The detailed analysis leads to support for the idea that the model provides a rationale for the switch observed in the immune system from the production of one major class of antibody (IgM) to the other major class (IgG). The analysis also leads to a new, previously unsuspected possibility for the nature of the suppressed state within the context of the postulates of the symmetrical network theory.     

A knowledge-based halogen bonding scoring function for predicting protein-ligand interactions

Halogen bonding, a non-covalent interaction between the halogen σ-hole and Lewis bases, could not be properly characterized by majority of current scoring functions. In this study, a knowledge-based halogen bonding scoring function, termed XBPMF, was developed by an iterative method for predicting protein-ligand interactions. Three sets of pairwise potentials were derived from two training sets of protein-ligand complexes from the Protein Data Bank. It was found that two-dimensional pairwise potentials could characterize appropriately the distance and angle profiles of halogen bonding, which is superior to one-dimensional pairwise potentials. With comparison to six widely used scoring functions, XBPMF was evaluated to have moderate power for predicting protein-ligand interactions in terms of “docking power”, “ranking power” and “scoring power”. Especially, it has a rather satisfactory performance for the systems with typical halogen bonds. To the best of our knowledge, XBPMF is the first halogen bonding scoring function that is not dependent on any dummy atom, and is practical for high-throughput virtual screening. Therefore, this scoring function should be useful for the study and application of halogen bonding interactions like molecular docking and lead optimization.

Native mass spectrometry for understanding dynamic protein complex

Biomolecules have evolved to perform specific and sophisticated activities in a highly coordinated manner organizing into multi-component complexes consisting of proteins, nucleic acids, cofactors or ligands. Understanding such complexes represents a task in earnest for modern bioscience. Traditional structural techniques when extrapolating to macromolecules of ever increasing sizes are confronted with limitations posed by the difficulty in enrichment, solubility, stability as well as lack of homogeneity of these complexes. Alternative approaches are therefore prompted to bridge the gap, one of which is native mass spectrometry. Here we demonstrate the strength of native mass spectrometry, used alone or in combination with other biophysical methods such as analytical ultracentrifugation, small-angle neutron scattering, and small-angle X-ray scattering etc., in addressing dynamic aspects of protein complexes including structural reorganization, subunit exchange, as well as the assembly/disassembly processes in solution that are dictated by transient non-covalent interactions. We review recent studies from our laboratories and others applying native mass spectrometry to both soluble and membrane-embedded assemblies. This article is part of a Special Issue entitled “Biophysical Exploration of Dynamical Ordering of Biomolecular Systems” edited by Dr. Koichi Kato.     

Invasive species contribute to biotic resistance: negative effect of caprellid amphipods on an invasive tunicate

As the number of introductions of non-indigenous species (NIS) continues to rise, ecologists are faced with new and unique opportunities to observe interactions between species that do not naturally co-exist. These interactions can have important implications on the invasion process, potentially determining whether NIS become widespread and abundant, survive in small numbers, or fail to establish and disappear. Although many studies have naturally focused on the interactions between NIS and native species to examine their effects and the biological resistance of the recipient community to invasion, few have examined the effects that NIS have on each other. In some cases, interactions can facilitate the invasion process of one or both species (i.e., “invasional meltdowns”), but competition or predation can lead to negative interactions as well. The introduction of the vase tunicate, Ciona intestinalis, in Prince Edward Island (Canada) has harmed mussel aquaculture via heavy biofouling of equipment and mussels. Through both a broad-scale survey and small-scale field experiments, we show that Ciona recruitment is drastically reduced by caprellid amphipods, including the NIS Caprella mutica. This study provides an exciting example of how established invasive species can negatively impact the recruitment of a secondary invader, highlighting the potential for non-additive effects of multiple invasions.     

A model for the genetic control of differential adhesion in insect epidermis

Adhesive relations among cells are believed to play a major role in determining patterns of serial homology, of intercalary regeneration, and of neuronal connectivity. Models for the genetic control of adhesion during development can provide a framework for further analysis of these phenomena. Investigators studying development of Drosophila have proposed that differentiation of segments and of imaginal discs is controlled by a set of bistable “selector genes”. In each region the settings of the selector genes form a binary “word” which determines the properties of cells in the region, including their adhesiveness. I have made an explicit proposal for the relation between binary words and adhesiveness, by assuming that active selector genes repress synthesis of “adhesor” macromolecules, which promote adhesion. This hypothesis correctly predicts the relative cohesiveness of cells in four pupal tissues of the moth Manduca. Works of cohesion and adhesion among the four cell types are deduced from published results of grafting experiments by modelling insect epidermis as a viscoelastic fluid.Further comparisons between deductions from the genetic and fluid models suggest that selector genes, or the adhesor molecules they regulate, interact within single cells in determining adhesiveness between cells. From a specific version of the genetic model I deduce that pairwise interactions between selector genes or adhesor molecules can determine many, though not all, of the relative works of adhesion between unlike cells in Manduca. The genetic and fluid models thus provide a set of working hypotheses for predicting patterns of intercellular adhesion in insect epidermis and for analyzing results of experiments designed to test such predictions.