Evaluation of toxic potential of captan: Induction of hsp70 and tissue damage in transgenic Drosophila melanogaster (hsp70-lacZ) Bg9

The study investigated the working hypothesis that a widely used fungicide captan exerts toxic effects on nontarget organisms. Transgenic Drosophila melanogaster (hsp70-lacZ) was used as a model by assaying stress gene expression as an endpoint for cytotoxicity and also to evaluate whether stress gene expression is sufficient enough to protect and to prevent tissue damage against toxic insult of the chemical. The study was further extended to understand the effect of the pesticide on development, life cycle, and reproduction of the organism and finally to evaluate a concentration of the chemical to be nontoxic to the organism. The study showed that (i) captan causes cytotoxicity at and above 0.015 ppm; (ii) at 0.0015 ppm captan, absence of hsp70 expression in the exposed organism was evaluated as the concentration referred to as no observed adverse effect level (NOAEL) for Drosophila; (iii) emergence pattern of flies was affected only at the highest concentration of captan by 4 days, while hatching and survivorship were unaffected even at this concentration; (iv) reproductive performance was significantly affected only at 125.0 and 1250.0 ppm captan, while in the lower dietary concentrations no such deleterious effects were observed; (v) at 1250.0 ppm, hsp70 failed to protect the cells from toxicant assault after 48 h exposure, thus leading to tissue damage as revealed by Trypan Blue staining. The present study shows the cytotoxic potential of captan and further reveals the application of stress genes in determining NOAEL and its expression as bioindicator of exposure to environmental contaminants.     

Systems Model of T Cell Receptor Proximal Signaling Reveals Emergent Ultrasensitivity

Receptor phosphorylation is thought to be tightly regulated because phosphorylated receptors initiate signaling cascades leading to cellular activation. The T cell antigen receptor (TCR) on the surface of T cells is phosphorylated by the kinase Lck and dephosphorylated by the phosphatase CD45 on multiple immunoreceptor tyrosine-based activation motifs (ITAMs). Intriguingly, Lck sequentially phosphorylates ITAMs and ZAP-70, a cytosolic kinase, binds to phosphorylated ITAMs with differential affinities. The purpose of multiple ITAMs, their sequential phosphorylation, and the differential ZAP-70 affinities are unknown. Here, we use a systems model to show that this signaling architecture produces emergent ultrasensitivity resulting in switch-like responses at the scale of individual TCRs. Importantly, this switch-like response is an emergent property, so that removal of multiple ITAMs, sequential phosphorylation, or differential affinities abolishes the switch. We propose that highly regulated TCR phosphorylation is achieved by an emergent switch-like response and use the systems model to design novel chimeric antigen receptors for therapy.     

Detection of Pancreatic Carcinomas by Imaging Lactose-Binding Protein Expression in Peritumoral Pancreas Using [18F]Fluoroethyl-Deoxylactose PET/CT

Background

Early diagnosis of pancreatic carcinoma with highly sensitive diagnostic imaging methods could save lives of many thousands of patients, because early detection increases resectability and survival rates. Current non-invasive diagnostic imaging techniques have inadequate resolution and sensitivity for detection of small size (∼2–3 mm) early pancreatic carcinoma lesions. Therefore, we have assessed the efficacy of positron emission tomography and computer tomography (PET/CT) imaging with β-O-D-galactopyranosyl-(1,4′)-2′-deoxy-2′-[¹⁸F]fluoroethyl-D-glucopyranose ([¹⁸F]FEDL) for detection of less than 3 mm orthotopic xenografts of L3.6pl pancreatic carcinomas in mice. [¹⁸F]FEDL is a novel radioligand of hepatocarcinoma-intestine-pancreas/pancreatitis-associated protein (HIP/PAP), which is overexpressed in peritumoral pancreatic acinar cells.

Methodology/Principal Findings

Dynamic PET/CT imaging demonstrated rapid accumulation of [¹⁸F]FEDL in peritumoral pancreatic tissue (4.04±2.06%ID/g), bi-exponential blood clearance with half-lives of 1.65±0.50 min and 14.14±3.60 min, and rapid elimination from other organs and tissues, predominantly by renal clearance. Using model-independent graphical analysis of dynamic PET data, the average distribution volume ratio (DVR) for [¹⁸F]FEDL in peritumoral pancreatic tissue was estimated as 3.57±0.60 and 0.94±0.72 in sham-operated control pancreas. Comparative analysis of quantitative autoradiographic images and densitometry of immunohistochemically stained and co-registered adjacent tissue sections demonstrated a strong linear correlation between the magnitude of [¹⁸F]FEDL binding and HIP/PAP expression in corresponding regions (r = 0.88). The in situ analysis demonstrated that at least a 2–4 fold apparent lesion size amplification was achieved for submillimeter tumors and to nearly half a murine pancreas for tumors larger than 3 mm.

Conclusion/Significance

We have demonstrated the feasibility of detection of early pancreatic tumors by non-invasive imaging with [¹⁸F]FEDL PET/CT of tumor biomarker HIP/PAP over-expressed in peritumoral pancreatic tissue. Non-invasive non-invasive detection of early pancreatic carcinomas with [¹⁸F]FEDL PET/CT imaging should aid the guidance of biopsies and additional imaging procedures, facilitate the resectability and improve the overall prognosis.     

Studying backbone torsional dynamics of intrinsically disordered proteins using fluorescence depolarization kinetics

Intrinsically disordered proteins (IDPs) do not autonomously adopt a stable unique 3D structure and exist as an ensemble of rapidly interconverting structures. They are characterized by significant conformational plasticity and are associated with several biological functions and dysfunctions. The rapid conformational fluctuation is governed by the backbone segmental dynamics arising due to the dihedral angle fluctuation on the Ramachandran ?–ψ conformational space. We discovered that the intrinsic backbone torsional mobility can be monitored by a sensitive fluorescence readout, namely fluorescence depolarization kinetics, of tryptophan in an archetypal IDP such as α-synuclein. This methodology allows us to map the site-specific torsional mobility in the dihedral space within picosecond-nanosecond time range at a low protein concentration under the native condition. The characteristic timescale of ~?1.4 ns, independent of residue position, represents collective torsional dynamics of dihedral angles (? and ψ) of several residues from tryptophan and is independent of overall global tumbling of the protein. We believe that fluorescence depolarization kinetics methodology will find broad application to study both short-range and long-range correlated motions, internal friction, binding-induced folding, disorder-to-order transition, misfolding and aggregation of IDPs.     

Genetic deletion of the tumor necrosis factor receptor p60 or p80 abrogates ligand-mediated activation of nuclear factor-kappa B and of mitogen-activated protein kinases in macrophages.

Tumor necrosis factor (TNF) is a pleiotropic cytokine known to regulate cell growth, viral replication, inflammation, immune system functioning, angiogenesis, and tumorigenesis. These effects are mediated through two different receptors, TNFR1 and TNFR2 (also called p60 and p80, respectively), with p60 receptor being expressed on all cell types and p80 receptor only on cells of the immune system and on endothelial cells. Although the role of p60 receptor in TNF signaling is well established, the role of p80 is less clear. In this report, by using macrophages derived from wild-type mice (having both receptors) and mice in which the gene for either p60 (p60(-/-)), or p80 (p80(-/-)), or both (p60(-/-) p80(-/-)) receptor have been deleted, we have redefined the role of these receptors in TNF-induced activation of nuclear factor (NF)-kappa B and of mitogen-activated protein kinases. TNF activated NF-kappa B in a dose- and time-dependent manner in wild-type macrophages but not in p60(-/-), p80(-/-), or p60(-/-) p80(-/-) macrophages. These results correlated with the I kappa B alpha degradation needed for NF-kappa B activation. We also found that TNF activated c-Jun N-terminal protein kinase in a dose- and time-dependent manner in wild-type macrophages but not in p60(-/-), p80(-/-), or p60(-/-) p80(-/-) macrophages. TNF activated p38 MAPK and p44/p42 MAPK in wild-type but not in p60(-/-), p80(-/-), or p60(-/-) p80(-/-) macrophages. TNF induced the proliferation of wild-type macrophages, but for p60(-/-) and p80(-/-) macrophages proliferation was lower, and in p60(-/-) p80(-/-) it was absent. Overall, our studies suggest that both types of TNF receptors are needed in macrophages for optimum TNF cell signaling.     

Leishmania major LmACR2 is a pentavalent antimony reductase that confers sensitivity to the drug pentostam

Arsenicals and antimonials are first line drugs for the treatment of trypanosomal and leishmanial diseases. To create the active form of the drug, Sb(V) must be reduced to Sb(III). Because arsenic and antimony are related metalloids, and arsenical resistant Leishmania strains are frequently cross-resistant to antimonials, we considered the possibility that Sb(V) is reduced by a leishmanial As(V) reductase. The sequence for the arsenate reductase of Saccharomyces cerevisiae, ScAcr2p, was used to clone the gene for a homologue, LmACR2, from Leishmania major. LmACR2 was able to complement the arsenate-sensitive phenotype of an arsC deletion strain of Escherichia coli or an ScACR2 deletion strain of Saccharomyces cerevisiae. Transfection of Leishmania infantum with LmACR2 augmented Pentostam sensitivity in intracellular amastigotes. LmACR2 was purified and shown to reduce both As(V) and Sb(V). This is the first report of an enzyme that confers Pentostam sensitivity in intracellular amastigotes of Leishmania. We propose that LmACR2 is responsible for reduction of the pentavalent antimony in Pentostam to the active trivalent form of the drug in Leishmania.     

Ion Transport Nanotube Assembled with Vertically Aligned Metallic MoS2 for High Rate Lithium‐Ion Batteries

Metallic phase molybdenum disulfide (MoS₂) is well known for orders of magnitude higher conductivity than 2H semiconducting phase MoS₂. Herein, for the first time, the authors design and fabricate a novel porous nanotube assembled with vertically aligned metallic MoS₂ nanosheets by using the scalable solvothermal method. This metallic nanotube has the following advantages: (i) intrinsic high electrical conductivity that promotes the rate performance of battery and eliminates the using of conductive additive; (ii) hierarchical, hollow, porous, and aligned structure that assists the electrolyte transportation and diffusion; (iii) tubular structure that avoids restacking of 2D nanosheets, and therefore maintains the electrochemistry cycling stability; and (iv) a shortened ion diffusion path, that improves the rate performance. This 1D metallic MoS₂ nanotube is demonstrated to be a promising anode material for lithium‐ion batteries. The unique structure delivers an excellent reversible capacity of 1100 mA h g^?1 under a current density of 5 A g^?1 after 350 cycles, and an outstanding rate performance of 589 mA h g^?1 at a current density of 20 A g^?1. Furthermore, attributed to the material's metallic properties, the electrode comprising 100% pure material without any additive provides an ideal system for the fundamental electrochemical study of metallic MoS₂. This study first reveals the characteristic anodic peak at 1.5 V in cyclic voltammetry of metallic MoS_2. This research sheds light on the fabrication of metallic 1D, 2D, or even 3D structures with 2D nanosheets as building blocks for various applications.