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Nucleotide triphosphatase and RNA chaperone activities of murine norovirus NS3
No full textModulation of RNA structure is essential in the life cycle of RNA viruses. Immediate replication upon infection requires RNA unwinding to ensure that RNA templates are not in intra-or intermolecular duplex forms. The calicivirus NS3, one of the highly conserved nonstructural (NS) proteins, has conserved motifs common to helicase superfamily 3 among six genogroups. However, its biological functions are not fully understood. In this study we report the oligomeric state and the nucleotide triphosphatase (NTPase) and RNA chaperone activities of the recombinant full-length NS3 derived from murine norovirus (MNV). The MNV NS3 has an Mg2+-dependent NTPase activity, and site-directed mutagenesis of the conserved NTPase motifs blocked enzyme activity and viral replication in cells. Further, the NS3 was found via fluorescence resonance energy transfer (FRET)-based assays to destabilize double-stranded RNA in the presence of Mg2+ or Mn2+ in an NTP-independent manner. However, the RNA destabilization activity was not affected by mutagenesis of the conserved motifs of NTPase. These results reveal that the MNV NS3 has an NTPase-independent RNA chaperone-like activity, and that a FRET-based RNA destabilization assay has the potential to identify new antiviral drugs targeting NS3
Fractionation and Characterization of Lycopene-Oxidation Products by LC-MS/MS (ESI)(+): Elucidation of the Chemopreventative Potency of Oxidized Lycopene in Breast-Cancer Cell Lines
No full textLycopene (LYC) has been correlated with the reduction of certain cancers and chronic diseases. However, the existence and biofunctionality of degraded, oxidized, and biotransformed LYC products in vivo have not been revealed. Therefore, this study aimed to screen and elucidate the potential bioactive lycopene-derived products in breast-cancer and noncancerous cells. LYC-oxidation or -cleavage products were generated using KMnO4. These oxidation products were separated as fractions I-III by silica column chromatography using gradient solvent systems. Further, LC-MS/MS (ESI)(+) was used to elucidate their possible fragmentation patterns and structures. Fraction II showed higher cytotoxicity (IC50 value of 64.5 mu M), cellular uptake, and apoptosis-inducing activity in MCF-7 cells. This fraction consists of major peak m/z 323, identified as apo-8,6'-carotendial. The cytotoxicity-inducing activity may be due to partial ROS generation with mitochondrial dysfunction. Further, the role of apo-8,6'-carotendial in the induction of apoptosis is demonstrated for the first time. These results illustrated that LYC-oxidation derivatives or metabolites are involved in growth inhibition of cancer cells. Exploration of specific oxidized-carotenoid products will give further insight into the field of nutritional biochemistry
Circulating HLA-DR(+)CD4(+) effector memory T cells resistant to CCR5 and PD-L1 mediated suppression compromise regulatory T cell function in tuberculosis
No full textChronic T cell activation is a hallmark of pulmonary tuberculosis (PTB). The mechanisms underpinning this important phenomenon are however, poorly elucidated, though known to rely on control of T effector cells (Teff) by regulatory T cells (Treg). Our studies show that circulating natural Treg cells in adults with PTB preserve their suppressive potential but Teff cells from such subjects are resistant to Treg-mediated suppression. We found this to be due to expansion of an activated Teff subset identified by Human Leukocyte Antigen (HLA)DR expression. Sensitivity to suppression was restored to control levels by depletion of this subset. Comparative transcriptome analysis of Teff cells that contain HLA-DR+ cells versus the fraction depleted of this population identified putative resistance mechanisms linked to IFNG, IL17A, IL22, PD-L1 and beta-chemokines CCL3L3, CCL4 expression. Antibody blocking experiments confirmed HLA-DR+ Teff cells, but not the fraction depleted of HLA-DR+ effectors, to be resistant to Treg suppression mediated via CCR5 and PD-L1 associated pathways. In the presence of HLA-DR+ Teff cells, activation of NF kappa B downstream of CCR5 and PD-L1 was perturbed. In addition, HLA-DR+ Teff cells expressed significantly higher levels of Th1/Th17 cytokines that may regulate Treg function through a reciprocal counter-balancing relationship. Taken together, our study provides novel insight on how activated HLA-DR(+)CD4(+) T cells may contribute to disease associated inflammation by compromising Treg-mediated suppression in PTB
On Consistency of Compressive Spectral Clustering
No full textSpectral clustering is one of the most popular methods for community detection in graphs. A key step in spectral clustering algorithms is the eigen decomposition of the nxn graph Laplacian matrix to extract its k leading eigenvectors, where k is the desired number of clusters among n objects. This is prohibitively complex to implement for very large datasets. However, it has recently been shown that it is possible to bypass the eigen decomposition by computing an approximate spectral embedding through graph filtering of random signals. In this paper, we analyze the working of spectral clustering performed via graph filtering on the stochastic block model. Specifically, we characterize the effects of sparsity, dimensionality and filter approximation error on the consistency of the algorithm in recovering planted clusters
Opto-Electronic System for Intravenous Infusion Monitoring
No full textIntravenous (IV) infusion is the administration of fluids directly into the blood stream through the vein. Gravity fed IV-infusion can be affected by errors and requires constant monitoring. The opto-electronic system presented here accurately tracks the fluid flow and assists the user in monitoring the infusion sessions by generating alerts on detection of errors. The system generates alerts upon detecting significant deviation from set drip rate. The system keeps track of total volume infused and alerts when a desired volume is about to be administered. Prototype system was tested in laboratory conditions and was found to be accurate within 4% of independently measured infusion volume. Such a device offers a potential solution to reduce the risks associated with the IV infusion therapy especially in low-resource setting or during delivery of IV therapy at home
Experimental investigation of effect of orientation and surface roughness on drying of porous media consisting of rods
No full textStudies of drying from a conventional porous medium (CPM), consisting of spheres, have shown the existence of three periods. In the first period drying rate is high and essentially depends on the atmospheric demand while in the last stage mass transfer happens from within the porous medium and is solely governed by the internal properties. In a previous study (Kumar and Arakeri) it was shown, using a cluster of closely packed smooth rods stacked vertically, that stage 1 was sustained till nearly the whole liquid gets evaporated. Near-zero radii contacts, between the rods, offering infinite height rise was shown to be the reason behind such high sustained evaporation rates and elongated duration of stage 1. Here we show that the orientation of the rods dramatically changes the evaporation process from this vertically oriented rod-based porous medium (VRBPM). Unlike the vertical case, in the horizontal case (HRBPM) smooth rods do not support stage 1. In this case transition to stage 2 was due to the liquid-vapour meniscus receding continuously in the network from the start of the experiment Rough rods pinned the liquid along its rough surfaces and stage 1 is sustained although its duration is much smaller compared to the vertical case. Maintained hydraulic connections against gravity along the roughness of the rods thus provides a better insight towards understanding stage 1 in a conventional porous medium consisting of complicated pore geometry and their connectivity. In all the experiments, infra-red heating at about 1000 W/m(2) causes evaporation from an initially saturated RBPM kept in an acrylic box. (C) 2018 Elsevier Ltd. All rights reserved
Drug treatment induces phenotypic switch and stemness during acquisition of drug resistance
No full textThe breast tumor heterogeneity and development of drug resistance has become a challenge for anti-cancer therapy. Tumor cells acquire resistance due to intra-tumor heterogeneity imposed by epithelial to mesenchymal transition. The drugs which target epithelial cells may fail to target mesenchymal and the intermediate phenotypes due to which the cancer cells acquire drug resistance. This concept provoked us to understand the phenotypic diversity among the tumor heterogeneity which might lead to drug resistance. Hence the aim of our study is to identify the mechanisms of anti-cancer drug resistance which would ultimately help in identification of new clinical targets.
Treatment of breast cancer cell lines MCF7 and MDA MB 231 with Doxorubicin showed a loss of epithelial markers and gain of mesenchymal markers in MCF7 cells, and vice versa in MDA MB 231 cells, thereby shifting them into an intermediate phenotype. Moreover, Doxorubicin treatment of MDA MB 231-Nanog stable cells led to an increase in Nanog positive cells confirming that the treatment-induced increase in stemness property of cancer cells. We predict that phosphoproteomic dataset of the anti-cancer drug-resistant cells let us analyze the possible molecular players involved in the phenotypic switch and hence drug resistance (work in progress). Thus, our current data suggest that
the anti-cancer therapy induces cancer cells to undergo a phenotypic switch and increases the stemness property which might be responsible to gain resistance. Identification of the critical regulators of the phenotypic switch by phosphoproteomics analysis is likely to help us investigate the possible mechanisms underlying anti-cancer drug resistance. Previous reports also suggest that conventional therapies fail to eliminate cancer stem cells thereby allowing tumor relapse.
These results suggest that understanding the protein abundance and phosphorylation state of key signaling molecules would elucidate specific signaling interactions or/and pathways differentially active in drug-resistant tumors would help us predict novel kinases, and hence identify drug targets
Whispered speech to neutral speech conversion using bidirectional LSTMs
No full textWe propose a bidirectional long short-term memory (BLSTM) based whispered speech to neutral speech conversion system that employs the STRAIGHT speech synthesizer. We use a BLSTM to map the spectral features of whispered speech to those of neutral speech. Three other BLSTMs are employed to predict the pitch, periodicity levels and the voiced/unvoiced phoneme decisions from the spectral features of whispered speech. We use objective measures to quantify the quality of the predicted spectral features and excitation parameters, using data recorded from six subjects, in a four fold setup. We find that the temporal smoothness of the spectral features predicted using the proposed BLSTM based system is statistically more compared to that predicted using deep neural network based baseline schemes. We also observe that while the performance of the proposed system is comparable to the baseline scheme for pitch prediction, it is superior in terms of classifying voicing decisions and predicting periodicity levels. From subjective evaluation via listening test, we find that the proposed method is chosen as the best performing scheme 26.61% (absolute) more often than the best baseline scheme. This reveals that the proposed method yields a more natural sounding neutral speech from whispered speech
Growth of Interfacial Intermetallic Compound Layer in Diffusion-Bonded SAC-Cu Solder Joints During Different Types of Thermomechanical Excursion
No full textThe effects of mechanical strain on the growth kinetics of interfacial intermetallic compounds (IMCs) sandwiched between Cu substrate and Sn-1.0 wt.%Ag-0.5 wt.%Cu (SAC105) solder have been investigated. Isothermal aging (IA) at 70A degrees C and 125A degrees C, and thermal cycling (TC) as well as thermomechanical cycling (TMC) with shear strain of 12.8% per cycle between -25A degrees C and 125A degrees C were applied to diffusion-bonded solder joints to study the growth behavior of the interfacial IMC layer under various types of thermomechanical excursion (TME). The microstructure of the solder joint tested under each TME was observed at regular intervals. It was observed that the growth rate of the IMC layer was higher in the case of TMC compared with TC or IA. This increased growth rate of the IMC layer in the presence of mechanical strain suggests an additional driving force that enhances the growth kinetics of the IMC. Finite element analysis was performed to gain insight into the effect of TC and TMC on the stress field in the solder joint, especially near the interface between the solder and the substrate. Finally, an analytical model was developed to quantify the effect of strain on the effective diffusivity and express the growth kinetics for all three types of TME using a single expression
Optically Immersed Bolometer IR Detectors Based on V2O5 Thin Films with Polyimide Thermal Impedance Control Layer for Space Applications
No full textOptically immersed bolometer IR detectors were fabricated using electron beam evaporated vanadium oxide as the sensing material. Spin-coated polyimide was used as medium to optically immerse the sensing element to the flat surface of a hemispherical germanium lens. This optical immersion layer also serves as the thermal impedance control layer and decides the performance of the devices in terms of responsivity and noise parameters. The devices were packaged in suitable electro-optical packages and the detector parameters were studied in detail. Thermal time constant varies from 0.57 to 6.0 ms and responsivity from 75 to 757 V W-1 corresponding to polyimide thickness in the range 2 to 70 mu m for a detector bias of 9 V in the wavelength region of 14-16 mu m. Highest D* obtained was 1.2x10(8) cmHz(1/2) W-1. Noise equivalent temperature difference (NETD) of 20 mK was achieved for devices with polyimide thickness more than 32 mu m. The figure of merit, NETD x tau product which describes trade-off between thermal time constant and sensitivity is also extensively studied for devices having different thickness of thermal impedance layers