Vasoactive Intestinal Polypeptide Promotes Intestinal Barrier Homeostasis and Protection Against Colitis in Mice

Inflammatory bowel disease is a chronic gastrointestinal inflammatory disorder associated with changes in neuropeptide expression and function, including vasoactive intestinal peptide (VIP). VIP regulates intestinal vasomotor and secretomotor function and motility; however, VIP’s role in development and maintenance of colonic epithelial barrier homeostasis is unclear. Using VIP deficient (VIPKO) mice, we investigated VIP’s role in epithelial barrier homeostasis, and susceptibility to colitis. Colonic crypt morphology and epithelial barrier homeostasis were assessed in wildtype (WT) and VIPKO mice, at baseline. Colitic responses were evaluated following dinitrobenzene sulfonic acid (DNBS) or dextran-sodium sulfate (DSS) exposure. Mice were also treated with exogenous VIP. At baseline, VIPKO mice exhibited distorted colonic crypts, defects in epithelial cell proliferation and migration, increased apoptosis, and altered permeability. VIPKO mice also displayed reduced goblet cell numbers, and reduced expression of secreted goblet cell factors mucin 2 and trefoil factor 3. These changes were associated with reduced expression of caudal type homeobox 2 (Cdx2), a master regulator of intestinal function and homeostasis. DNBS and DSS-induced colitis were more severe in VIPKO than WT mice. VIP treatment rescued the phenotype, protecting VIPKO mice against DSS colitis, with results comparable to WT mice. In conclusion, VIP plays a crucial role in the development and maintenance of colonic epithelial barrier integrity under physiological conditions and promotes epithelial repair and homeostasis during colitis.     

Suppression of a Natural Killer Cell Response by Simian Immunodeficiency Virus Peptides

Natural killer (NK) cell responses in primates are regulated in part through interactions between two highly polymorphic molecules, the killer-cell immunoglobulin-like receptors (KIRs) on NK cells and their major histocompatibility complex (MHC) class I ligands on target cells. We previously reported that the binding of a common MHC class I molecule in the rhesus macaque, Mamu-A1*002, to the inhibitory receptor Mamu-KIR3DL05 is stabilized by certain simian immunodeficiency virus (SIV) peptides, but not by others. Here we investigated the functional implications of these interactions by testing SIV peptides bound by Mamu-A1*002 for the ability to modulate Mamu-KIR3DL05⁺ NK cell responses. Twenty-eight of 75 SIV peptides bound by Mamu-A1*002 suppressed the cytolytic activity of primary Mamu-KIR3DL05⁺ NK cells, including three immunodominant CD8⁺ T cell epitopes previously shown to stabilize Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. Substitutions at C-terminal positions changed inhibitory peptides into disinhibitory peptides, and vice versa, without altering binding to Mamu-A1*002. The functional effects of these peptide variants on NK cell responses also corresponded to their effects on Mamu-A1*002 tetramer binding to Mamu-KIR3DL05. In assays with mixtures of inhibitory and disinhibitory peptides, low concentrations of inhibitory peptides dominated to suppress NK cell responses. Consistent with the inhibition of Mamu-KIR3DL05⁺ NK cells by viral epitopes presented by Mamu-A1*002, SIV replication was significantly higher in Mamu-A1*002⁺ CD4⁺ lymphocytes co-cultured with Mamu-KIR3DL05⁺ NK cells than with Mamu-KIR3DL05^- NK cells. These results demonstrate that viral peptides can differentially affect NK cell responses by modulating MHC class I interactions with inhibitory KIRs, and provide a mechanism by which immunodeficiency viruses may evade NK cell responses.     

Optimization of Heavy Chain and Light Chain Signal Peptides for High Level Expression of Therapeutic Antibodies in CHO Cells

Translocation of a nascent protein from the cytosol into the ER mediated by its signal peptide is a critical step in protein secretion. The aim of this work was to develop a platform technology to optimize the signal peptides for high level production of therapeutic antibodies in CHO cells. A database of signal peptides from a large number of human immunoglobulin (Ig) heavy chain (HC) and kappa light chain (LC) was generated. Most of the HC signal peptides contain 19 amino acids which can be divided into three domains and the LC signal peptides contain 22 amino acids. The signal peptides were then clustered according to sequence similarity. Based on the clustering, 8 HC and 2 LC signal peptides were analyzed for their impacts on the production of 5-top selling antibody therapeutics, namely, Herceptin, Avastin, Remicade, Rituxan, and Humira. The best HC and LC signal peptides for producing these 5 antibodies were identified. The optimized signal peptides for Rituxan is 2-fold better compared to its native signal peptides which are available in the public database. Substitution of a single amino acid in the optimized HC signal peptide for Avastin reduced its production significantly. Mass spectrometry analyses revealed that all optimized signal peptides are accurately removed in the mature antibodies. The results presented in this report are particularly important for the production of these 5 antibodies as biosimilar drugs. They also have the potential to be the best signal peptides for the production of new antibodies in CHO cells.     

Cost-Effectiveness of HBV and HCV Screening Strategies – A Systematic Review of Existing Modelling Techniques

Introduction

Studies evaluating the cost-effectiveness of screening for Hepatitis B Virus (HBV) and Hepatitis C Virus (HCV) are generally heterogeneous in terms of risk groups, settings, screening intervention, outcomes and the economic modelling framework. It is therefore difficult to compare cost-effectiveness results between studies. This systematic review aims to summarise and critically assess existing economic models for HBV and HCV in order to identify the main methodological differences in modelling approaches.

Methods

A structured search strategy was developed and a systematic review carried out. A critical assessment of the decision-analytic models was carried out according to the guidelines and framework developed for assessment of decision-analytic models in Health Technology Assessment of health care interventions.

Results

The overall approach to analysing the cost-effectiveness of screening strategies was found to be broadly consistent for HBV and HCV. However, modelling parameters and related structure differed between models, producing different results. More recent publications performed better against a performance matrix, evaluating model components and methodology.

Conclusion

When assessing screening strategies for HBV and HCV infection, the focus should be on more recent studies, which applied the latest treatment regimes, test methods and had better and more complete data on which to base their models. In addition to parameter selection and associated assumptions, careful consideration of dynamic versus static modelling is recommended. Future research may want to focus on these methodological issues. In addition, the ability to evaluate screening strategies for multiple infectious diseases, (HCV and HIV at the same time) might prove important for decision makers.     

Harnessing Case Isolation and Ring Vaccination to Control Ebola

As a devastating Ebola outbreak in West Africa continues, non-pharmaceutical control measures including contact tracing, quarantine, and case isolation are being implemented. In addition, public health agencies are scaling up efforts to test and deploy candidate vaccines. Given the experimental nature and limited initial supplies of vaccines, a mass vaccination campaign might not be feasible. However, ring vaccination of likely case contacts could provide an effective alternative in distributing the vaccine. To evaluate ring vaccination as a strategy for eliminating Ebola, we developed a pair approximation model of Ebola transmission, parameterized by confirmed incidence data from June 2014 to January 2015 in Liberia and Sierra Leone. Our results suggest that if a combined intervention of case isolation and ring vaccination had been initiated in the early fall of 2014, up to an additional 126 cases in Liberia and 560 cases in Sierra Leone could have been averted beyond case isolation alone. The marginal benefit of ring vaccination is predicted to be greatest in settings where there are more contacts per individual, greater clustering among individuals, when contact tracing has low efficacy or vaccination confers post-exposure protection. In such settings, ring vaccination can avert up to an additional 8% of Ebola cases. Accordingly, ring vaccination is predicted to offer a moderately beneficial supplement to ongoing non-pharmaceutical Ebola control efforts.     

Molecular Dynamic Simulations Reveal the Structural Determinants of Fatty Acid Binding to Oxy-Myoglobin

The mechanism(s) by which fatty acids are sequestered and transported in muscle have not been fully elucidated. A potential key player in this process is the protein myoglobin (Mb). Indeed, there is a catalogue of empirical evidence supporting direct interaction of globins with fatty acid metabolites; however, the binding pocket and regulation of the interaction remains to be established. In this study, we employed a computational strategy to elucidate the structural determinants of fatty acids (palmitic & oleic acid) binding to Mb. Sequence analysis and docking simulations with a horse (Equus caballus) structural Mb reference reveals a fatty acid-binding site in the hydrophobic cleft near the heme region in Mb. Both palmitic acid and oleic acid attain a “U” shaped structure similar to their conformation in pockets of other fatty acid-binding proteins. Specifically, we found that the carboxyl head group of palmitic acid coordinates with the amino group of Lys45, whereas the carboxyl group of oleic acid coordinates with both the amino groups of Lys45 and Lys63. The alkyl tails of both fatty acids are supported by surrounding hydrophobic residues Leu29, Leu32, Phe33, Phe43, Phe46, Val67, Val68 and Ile107. In the saturated palmitic acid, the hydrophobic tail moves freely and occasionally penetrates deeper inside the hydrophobic cleft, making additional contacts with Val28, Leu69, Leu72 and Ile111. Our simulations reveal a dynamic and stable binding pocket in which the oxygen molecule and heme group in Mb are required for additional hydrophobic interactions. Taken together, these findings support a mechanism in which Mb acts as a muscle transporter for fatty acid when it is in the oxygenated state and releases fatty acid when Mb converts to deoxygenated state.     

Peripheral administration of oxytocin increases social affiliation in the naked mole-rat (Heterocephalus glaber)

The neuropeptide oxytocin regulates a wide variety of social behaviors across diverse species. However, the types of behaviors that are influenced by this hormone are constrained by the species in question and the social organization that a particular species exhibits. Therefore, the present experiments investigated behaviors regulated by oxytocin in a eusocial mammalian species by using the naked mole-rat (Heterocephalus glaber). In Experiment 1, adult non-breeding mole-rats were given intraperitoneal injections of either oxytocin (1 mg/kg or 10 mg/kg) or saline on alternate days. Animals were then returned to their colony and behavior was recorded for minutes 15–30 post-injection. Both doses of oxytocin increased huddling behavior during this time period. In Experiment 2, animals received intraperitoneal injections of either oxytocin (1 mg/kg), an oxytocin-receptor antagonist (0.1 mg/kg), a cocktail of oxytocin and the antagonist, or saline across 4 testing days in a counterbalanced design. Animals were placed in either a 2-chamber arena with a familiar conspecific or in a small chamber with 1 week old pups from their home colony and behaviors were recorded for minutes 15–30 post-injection. Oxytocin increased investigation of, and time spent in close proximity to, a familiar conspecific; these effects were blocked by the oxytocin antagonist. No effects were seen on pup-directed behavior. These data suggest that oxytocin is capable of modulating affiliative-like behavior in this eusocial species.     

Antagonism of Beclin 1‐dependent autophagy by BCL‐2 at the endoplasmic reticulum requires NAF‐1

In addition to mitochondria, BCL‐2 is located at the endoplasmic reticulum (ER) where it is a constituent of several distinct complexes. Here, we identify the BCL‐2‐interacting protein at the ER, nutrient‐deprivation autophagy factor‐1 (NAF‐1)—a bitopic integral membrane protein whose defective expression underlies the aetiology of the neurodegenerative disorder Wolfram syndrome 2 (WFS2). NAF‐1 contains a two iron–two sulphur coordinating domain within its cytosolic region, which is necessary, but not sufficient for interaction with BCL‐2. NAF‐1 is displaced from BCL‐2 by the ER‐restricted BH3‐only protein BIK and contributes to regulation of BIK‐initiated autophagy, but not BIK‐dependent activation of caspases. Similar to BCL‐2, NAF‐1 is found in association with the inositol 1,4,5‐triphosphate receptor and is required for BCL‐2‐mediated depression of ER Ca²⁺ stores. During nutrient deprivation as a physiological stimulus of autophagy, BCL‐2 is known to function through inhibition of the autophagy effector and tumour suppressor Beclin 1. NAF‐1 is required in this pathway for BCL‐2 at the ER to functionally antagonize Beclin 1‐dependent autophagy. Thus, NAF‐1 is a BCL‐2‐associated co‐factor that targets BCL‐2 for antagonism of the autophagy pathway at the ER.     

Organic acid production and plant growth promotion as a function of phosphate solubilization by Acinetobacter rhizosphaerae strain BIHB 723 isolated from the cold deserts of the trans-Himalayas

An efficient phosphate-solubilizing plant growth–promoting Acinetobacter rhizosphaerae strain BIHB 723 exhibited significantly higher solubilization of tricalcium phosphate (TCP) than Udaipur rock phosphate (URP), Mussoorie rock phosphate (MRP) and North Carolina rock phosphate (NCRP). Qualitative and quantitative differences were discerned in the gluconic, oxalic, 2-keto gluconic, lactic, malic and formic acids during the solubilization of various inorganic phosphates by the strain. Gluconic acid was the main organic acid produced during phosphate solubilization. Formic acid production was restricted to TCP solubilization and oxalic acid production to the solubilization of MRP, URP and NCRP. A significant increase in plant height, shoot fresh weight, shoot dry weight, root length, root dry weight, and root, shoot and soil phosphorus (P) contents was recorded with the inoculated treatments over the uninoculated NP₀K or NP_TCPK treatments. Plant growth promotion as a function of phosphate solubilization suggested that the use of bacterial strain would be a beneficial addition to the agriculture practices in TCP-rich soils in reducing the application of phosphatic fertilizers.