Oncolytic Vesicular Stomatitis Virus in an Immunocompetent Model of MUC1-Positive or MUC1-Null Pancreatic Ductal Adenocarcinoma

Vesicular stomatitis virus (VSV) is a promising oncolytic agent against various malignancies. Here, for the first time, we tested VSV in vitro and in vivo in a clinically relevant, immunocompetent mouse model of pancreatic ductal adenocarcinoma (PDA). Our system allows the study of virotherapy against PDA in the context of overexpression (80% of PDA patients) or no expression of human mucin 1 (MUC1), a major marker for poor prognosis in patients. In vitro, we tested three VSV recombinants, wild-type VSV, VSV-green fluorescent protein (VSV-GFP), and a safe oncolytic VSV-ΔM51-GFP, against five mouse PDA cell lines that either expressed human MUC1 or were MUC1 null. All viruses demonstrated significant oncolytic abilities independent of MUC1 expression, although VSV-ΔM51-GFP was somewhat less effective in two PDA cell lines. In vivo administration of VSV-ΔM51-GFP resulted in significant reduction of tumor growth for tested mouse PDA xenografts (+MUC1 or MUC1 null), and antitumor efficacy was further improved when the virus was combined with the chemotherapeutic drug gemcitabine. The antitumor effect was transient in all tested groups. The developed system can be used to study therapies involving various oncolytic viruses and chemotherapeutics, with the goal of inducing tumor-specific immunity while preventing premature virus clearance.     

Sympathetic support of energy expenditure and sympathetic nervous system activity after gastric bypass surgery

Objective:

This study was designed to determine how gastric bypass affects the sympathetically‐mediated component of resting energy expenditure (REE) and muscle sympathetic nerve activity (MSNA).

Design and Methods:

We measured REE before and after beta‐blockade in seventeen female subjects approximately three years post‐gastric bypass surgery and in nineteen female obese individuals for comparison. We also measured MSNA in a subset of these subjects.

Results:

The gastric bypass subjects had no change in REE after systemic beta‐blockade, reflecting a lack of sympathetic support of REE, in contrast to obese subjects where REE was reduced by beta‐blockade by approximately 5% (P < 0.05). The gastric bypass subjects, while still overweight (BMI = 29.3 vs 38.0 kg·m^?2 for obese subjects, P < 0.05), also had significantly lower MSNA compared to obese subjects (10.9 ± 2.3 vs. 21.9 ± 4.1 bursts·min^?1, P < 0.05). The reasons for low MSNA and a lack of sympathetically mediated support of REE after gastric bypass are likely multifactorial and may be related to changes in insulin sensitivity, body composition, and leptin, among other factors.

Conclusions:

These findings may have important consequences for the maintenance of weight loss after gastric bypass. Longitudinal studies are needed to further explore the changes in sympathetic support of REE and if changes in MSNA or tissue responsiveness are related to the sympathetic support of REE.

     

A Conserved Interaction between a C-Terminal Motif in Norovirus VPg and the HEAT-1 Domain of eIF4G Is Essential for Translation Initiation

Translation initiation is a critical early step in the replication cycle of the positive-sense, single-stranded RNA genome of noroviruses, a major cause of gastroenteritis in humans. Norovirus RNA, which has neither a 5´ m⁷G cap nor an internal ribosome entry site (IRES), adopts an unusual mechanism to initiate protein synthesis that relies on interactions between the VPg protein covalently attached to the 5´-end of the viral RNA and eukaryotic initiation factors (eIFs) in the host cell. For murine norovirus (MNV) we previously showed that VPg binds to the middle fragment of eIF4G (4GM; residues 652–1132). Here we have used pull-down assays, fluorescence anisotropy, and isothermal titration calorimetry (ITC) to demonstrate that a stretch of ~20 amino acids at the C terminus of MNV VPg mediates direct and specific binding to the HEAT-1 domain within the 4GM fragment of eIF4G. Our analysis further reveals that the MNV C terminus binds to eIF4G HEAT-1 via a motif that is conserved in all known noroviruses. Fine mutagenic mapping suggests that the MNV VPg C terminus may interact with eIF4G in a helical conformation. NMR spectroscopy was used to define the VPg binding site on eIF4G HEAT-1, which was confirmed by mutagenesis and binding assays. We have found that this site is non-overlapping with the binding site for eIF4A on eIF4G HEAT-1 by demonstrating that norovirus VPg can form ternary VPg-eIF4G-eIF4A complexes. The functional significance of the VPg-eIF4G interaction was shown by the ability of fusion proteins containing the C-terminal peptide of MNV VPg to inhibit in vitro translation of norovirus RNA but not cap- or IRES-dependent translation. These observations define important structural details of a functional interaction between norovirus VPg and eIF4G and reveal a binding interface that might be exploited as a target for antiviral therapy.     

Growth patterns in brooding dinosaurs reveals the timing of sexual maturity in non-avian dinosaurs and genesis of the avian condition

The timing of sexual maturation in non-avian dinosaurs is not known. In extant squamates and crocodilians it occurs in conjunction with the initial slowing of growth rates as adult size is approached. In birds (living dinosaurs) on the other hand, reproductive activity begins well after somatic maturity. Here we used growth line counts and spacing in all of the known brooding non-avian dinosaurs to determine the stages of development when they perished. It was revealed that sexual maturation occurred well before full adult size was reached-the primitive reptilian condition. In this sense, the life history and physiology of non-avian dinosaurs was not like that of modern birds. Palaeobiological ramifications of these findings include the potential to deduce reproductive lifespan, fecundity and reproductive population sizes in non-avian dinosaurs, as well as aid in the identification of secondary sexual characteristics.     

Evaluation of organic insecticides for management of spotted‐wing drosophila (Drosophila suzukii) in berry crops

Spotted‐wing drosophila, Drosophila suzukii (Matsumura), is an invasive pest affecting fruit production in many regions of the world. Insecticides are the primary tactic for controlling D. suzukii in organic as well as conventional production systems. Organic growers have a greater challenge because fewer insecticides are approved for use in organic agriculture. The most effective organically approved product is spinosad, but alternatives are needed because of label restrictions limiting the number of applications per year, toxicity to beneficial arthropods and the risk of developing resistance. We evaluated several organically approved insecticides against D. suzukii in laboratory assays and field trials conducted on organic blueberry and raspberry farms. Spinosad was consistently the most effective insecticide, but a few other insecticides such as azadirachtin + pyrethrins, Chromobacterium subtsugae and sabadilla alkaloids showed moderate activity. None of the treatments had long residual activity. Mortality started to decline by 3 days after treatment, and by 5 days after application, the treatments were not different from the controls. These products may be useful in rotation programmes, necessary for reducing reliance on spinosad and mitigating resistance. Cultural and biological control approaches are needed in fruit production for D. suzukii management, but insecticides will likely continue to be the dominant management tactic while these other approaches are being optimized and adopted.     

Impact of phagostimulants on effectiveness of OMRI‐listed insecticides used for control of spotted‐wing drosophila (Drosophila suzukii Matsumura)

Spotted‐wing drosophila, Drosophila suzukii Matsumura, is an invasive pest in the United States that causes considerable damage to fruit crops. It is responsible for many millions of dollars of revenue loss. The female D. suzukii has a heavily sclerotized ovipositor and can lay eggs in ripening or ripe fruit. The arrival of this invasive species has disrupted existing integrated pest management programmes, and growers rely on repeated insecticide applications to protect fruit. Organic growers have few chemical control options, and their reliance on spinosad increases the risk of developing insecticide resistance. We hypothesized that combining phagostimulants with insecticides would increase insecticide efficacy by prompting flies to spend more time in contact with residues. Therefore, the objective of this study was to evaluate the effectiveness of sucrose and the yeast Saccharomyces cerevisiae as phagostimulants in combination with organic biopesticides against D. suzukii in blueberries. Adding sucrose with or without yeast did not improve insecticide efficacy in terms of adult fly mortality or fruit infestation. Spinosad was very effective in all experiments, and for this product, there is little room for improvement. The phagostimulants had no effect on residual activity of any insecticide. The addition of sucrose with or without yeast did not improve the effectiveness of organic insecticides for D. suzukii. Concentrations of these phagostimulants in our experiments (0.36%) may have been too low to elicit a response. Further research is recommended to test different types and concentrations of phagostimulants.     

Cyclic changes in the matrix metalloproteinase system in the ovary and uterus

With each estrous or menstrual cycle, extensive alterations occur in the extracellular matrix and connective tissue of the ovary and uterus. In the ovary, these changes occur during follicular development, breakdown of the follicular wall and extrusion of the oocyte, as well as during the formation and regression of the corpus luteum. In the uterus, the endometrium undergoes dramatic connective tissue turnover associated with tissue breakdown and subsequent regrowth during each menstrual cycle. These changes in the ovarian and uterine extracellular architecture are regulated, in part, by the matrix metalloproteinase (MMP) system. This system is comprised of both a proteolytic component, the MMPs, and associated inhibitors, and it is involved in connective tissue remodeling processes throughout the body. The current review highlights the key features of the MMP system and focuses on the changes in the MMPs and the tissue inhibitors of metalloproteinases during the dynamic remodeling that takes place in the ovary and uterus during the estrous and menstrual cycles.     

The N-terminal 17% of apoB binds tightly and irreversibly to emulsions modeling nascent very low density lipoproteins

The N-terminal 17% of apolipoprotein B (apoB-17) readily associates with dimyristoylphosphatidylcholine (DMPC) multilamellar vesicles (MLV) to form large (240-A diameter) discoidal particles. Because apoB is normally secreted with triacylglycerol (TAG)-rich lipoproteins, we studied the binding of apoB-17 to triolein-rich emulsions modeling nascent TAG-rich very low density-like lipoproteins. Emulsions with the following composition (by weight) were prepared: 85--89% triolein, 1.1--1.4% cholesterol, and 10--14% phosphatidylcholines (PC) including either egg yolk (EY)-, dimyristoyl (DM)-, or dipalmitoyl (DP)-PC representing (at 25 degrees C), respectively, a fluid surface, a surface at transition, and a mainly solid surface. The respective sizes were 1,260 +/- 500, 1,070 +/- 450, and 830 +/- 300 A mean diameter. The emulsions were incubated with conditioned medium containing apoB-17, and then reisolated by ultracentrifugation. Analysis of the emulsion-bound proteins by gel electrophoresis showed that all three emulsions bound primarily apoB-17. The DPPC emulsions bound more apoB-17 than EYPC or DMPC emulsions. Immunoaffinity-purified apoB-17 exhibited saturable, high affinity binding to EYPC and DPPC emulsions. The respective K(d) values were 32 +/- 23 and 85 +/- 27 nM and capacities (N) were 10 and 58 molecules of apoB-17 per particle. When apoB-17 bound to emulsions was incubated with DMPC MLV at 26 degrees C for 18 h, it remained bound to the emulsions, indicating that once bound to these emulsions it is unable to exchange off and solubilize DMPC into discs. In contrast, apoE-3 bound to emulsions dissociated from the emulsions when incubated with DMPC MLV and formed discs.Thus, apoB-17 binds strongly and irreversibly to emulsions modeling nascent lipoproteins. It therefore may play an important role in the stabilization of nascent VLDL and chylomicrons.- Herscovitz, H., A. Derksen, M. T. Walsh, C. J. McKnight, D. L. Gantz, M. Hadzopoulou-Cladaras, V. Zannis, C. Curry, and D. M. Small. The N-terminal 17% of apoB binds tightly and irreversibly to emulsions modeling nascent very low density lipoproteins. J. Lipid Res. 2001. 42: 51;-59.