Curcumin alleviates matrix metalloproteinase-3 and -9 activities during eradication of Helicobacter pylori infection in cultured cells and mice

Current therapy-regimens against Helicobacter pylori (Hp) infections have considerable failure rates and adverse side effects that urge the quest for an effective alternative therapy. We have shown that curcumin is capable of eradicating Hp-infection in mice. Here we examine the mechanism by which curcumin protects Hp infection in cultured cells and mice. Since, MMP-3 and -9 are inflammatory molecules associated to the pathogenesis of Hp-infection, we investigated the role of curcumin on inflammatory MMPs as well as proinflammatory molecules. Curcumin dose dependently suppressed MMP-3 and -9 expression in Hp infected human gastric epithelial (AGS) cells. Consistently, Hp-eradication by curcumin-therapy involved significant downregulation of MMP-3 and -9 activities and expression in both cytotoxic associated gene (cag)(+ve) and cag(-ve) Hp-infected mouse gastric tissues. Moreover, we demonstrate that the conventional triple therapy (TT) alleviated MMP-3 and -9 activities less efficiently than curcumin and curcumin's action on MMPs was linked to decreased pro-inflammatory molecules and activator protein-1 activation in Hp-infected gastric tissues. Although both curcumin and TT were associated with MMP-3 and -9 downregulation during Hp-eradication, but unlike TT, curcumin enhanced peroxisome proliferator-activated receptor-γ and inhibitor of kappa B-α. These data indicate that curcumin-mediated healing of Hp-infection involves regulation of MMP-3 and -9 activities.     

Comparative Biochemical and Structural Properties of an Industrially Important Biocatalyst Cellobiohydrolase Cel7A from Thermophilic Aspergillus fumigatus

Applied Biochemistry and Microbiology - Cellobiohydrolase GH7 (Cel7A) is a crucial component of an efficient cellulase system. High cellulase activity was identified in the secretome of a locally...     

Effects of GM1 on brain spectrin-aminophospholipid interactions

Spectrin, a major component of the membrane skeletal meshwork of metazoan cells, is implicated to associate with membrane domains and is known to act as a scaffold for stabilization and activation of different signalling modules. We have studied the effect of GM1 (monosialotetrahexosyl ganglioside), a well-known model ganglioside and a signalling moiety, on the interaction of non-erythroid brain spectrin with both saturated and unsaturated aminophospholipids by spectroscopic methods. We observe that GM1 modulates brain spectrin-aminophospholipid interaction to the greatest degree whereas its effect on erythroid spectrin is not as pronounced. Fluorescence quenching studies show that brain spectrin interacts with DMPC/DMPE-based vesicles with a 10-fold increased affinity in presence of very low amounts of 2% and 5% GM1, and the extent of quenching decreases progressively in presence of increasing amounts of GM1. Interaction of brain spectrin with unsaturated membrane systems of DOPC/DOPE weakens in presence GM1. Increase in the mean lifetime of the Trp residues of brain spectrin in presence of GM1 indicates change in the microenvironment of spectrin, without affecting the secondary structure of the protein significantly. Studies on pressure – area isotherm of Langmuir-Blodgett monolayer and Brewster's angle microscopy show that GM1 has an expanding effect on the aminophospholipid monolayers, and ordered regions in DMPC/DMPE mixed monolayers are formed and are stabilized at higher pressure. GM1-induced fluidization of the phospholipid membranes and probable physical contact between bulky sugar head group of GM1 and spectrin, may explain the modulatory role of GM1 on aminophospholipid interactions with nonerythroid brain spectrin.     

Probing Conformational Stability and Dynamics of Erythroid and Nonerythroid Spectrin: Effects of Urea and Guanidine Hydrochloride

We have studied the conformational stability of the two homologous membrane skeletal proteins, the erythroid and non-erythroid spectrins, in their dimeric and tetrameric forms respectively during unfolding in the presence of urea and guanidine hydrochloride (GuHCl). Fluorescence and circular dichroism (CD) spectroscopy have been used to study the changes of intrinsic tryptophan fluorescence, anisotropy, far UV-CD and extrinsic fluorescence of bound 1-anilinonapthalene-8-sulfonic acid (ANS). Chemical unfolding of both proteins were reversible and could be described as a two state transition. The folded erythroid spectrin and non-erythroid spectrin were directly converted to unfolded monomer without formation of any intermediate. Fluorescence quenching, anisotropy, ANS binding and dynamic light scattering data suggest that in presence of low concentrations of the denaturants (up-to 1M) hydrogen bonding network and van der Waals interaction play a role inducing changes in quaternary as well as tertiary structures without complete dissociation of the subunits. This is the first report of two large worm like, multi-domain proteins obeying twofold rule which is commonly found in small globular proteins. The free energy of stabilization (ΔG_u ^H ₂ ⁰) for the dimeric spectrin has been 20 kcal/mol lesser than the tetrameric from.     

Induced Pluripotent Stem Cell Derived Macrophages as a Cellular System to Study Salmonella and Other Pathogens

A number of pathogens, including several human-restricted organisms, persist and replicate within macrophages (Mφs) as a key step in pathogenesis. The mechanisms underpinning such host-restricted intracellular adaptations are poorly understood, in part, due to a lack of appropriate model systems. Here we explore the potential of human induced pluripotent stem cell derived macrophages (iPSDMs) to study such pathogen interactions. We show iPSDMs express a panel of established Mφ-specific markers, produce cytokines, and polarise into classical and alternative activation states in response to IFN-γ and IL-4 stimulation, respectively. iPSDMs also efficiently phagocytosed inactivated bacterial particles as well as live Salmonella Typhi and S. Typhimurium and were able to kill these pathogens. We conclude that iPSDMs can support productive Salmonella infection and propose this as a flexible system to study host/pathogen interactions. Furthermore, iPSDMs can provide a flexible and practical cellular platform for assessing host responses in multiple genetic backgrounds.     

Analyzing Large Gene Expression and Methylation Data Profiles Using StatBicRM: Statistical Biclustering-Based Rule Mining

Microarray and beadchip are two most efficient techniques for measuring gene expression and methylation data in bioinformatics. Biclustering deals with the simultaneous clustering of genes and samples. In this article, we propose a computational rule mining framework, StatBicRM (i.e., statistical biclustering-based rule mining) to identify special type of rules and potential biomarkers using integrated approaches of statistical and binary inclusion-maximal biclustering techniques from the biological datasets. At first, a novel statistical strategy has been utilized to eliminate the insignificant/low-significant/redundant genes in such way that significance level must satisfy the data distribution property (viz., either normal distribution or non-normal distribution). The data is then discretized and post-discretized, consecutively. Thereafter, the biclustering technique is applied to identify maximal frequent closed homogeneous itemsets. Corresponding special type of rules are then extracted from the selected itemsets. Our proposed rule mining method performs better than the other rule mining algorithms as it generates maximal frequent closed homogeneous itemsets instead of frequent itemsets. Thus, it saves elapsed time, and can work on big dataset. Pathway and Gene Ontology analyses are conducted on the genes of the evolved rules using David database. Frequency analysis of the genes appearing in the evolved rules is performed to determine potential biomarkers. Furthermore, we also classify the data to know how much the evolved rules are able to describe accurately the remaining test (unknown) data. Subsequently, we also compare the average classification accuracy, and other related factors with other rule-based classifiers. Statistical significance tests are also performed for verifying the statistical relevance of the comparative results. Here, each of the other rule mining methods or rule-based classifiers is also starting with the same post-discretized data-matrix. Finally, we have also included the integrated analysis of gene expression and methylation for determining epigenetic effect (viz., effect of methylation) on gene expression level.     

Altered expression and editing of miRNA-100 regulates iTreg differentiation

RNA editing of miRNAs, especially in the seed region, adds another layer to miRNA mediated gene regulation which can modify its targets, altering cellular signaling involved in important processes such as differentiation. In this study, we have explored the role of miRNA editing in CD4⁺ T cell differentiation. CD4⁺ T cells are an integral component of the adaptive immune system. Naïve CD4⁺ T cells, on encountering an antigen, get differentiated either into inflammatory subtypes like Th1, Th2 or Th17, or into immunosuppressive subtype Treg, depending on the cytokine milieu. We found C-to-U editing at fifth position of mature miR-100, specifically in Treg. The C-to-U editing of miR-100 is functionally associated with at least one biologically relevant target change, from MTOR to SMAD2. Treg cell polarization by TGFβ1 was reduced by both edited and unedited miR-100 mimics, but percentage of Treg in PBMCs was only reduced by edited miR-100 mimics, suggesting a model in which de-repression of MTOR due to loss of unedited mir-100, promotes tolerogenic signaling, while gain of edited miR-100 represses SMAD2, thereby limiting the Treg. Such delicately counterbalanced systems are a hallmark of immune plasticity and we propose that miR-100 editing is a novel mechanism toward this end.