Mas-related G protein-coupled receptor type D antagonism improves portal hypertension in cirrhotic rats

Splanchnic vasodilatation contributes to the development and aggravation of portal hypertension (PHT). We previously demonstrated that in cirrhosis, angiotensin‐ mediates splanchnic vasodilatation through the Mas receptor (MasR). In this study, we investigated whether the recently characterized second receptor for angiotensin‐(1–7), Mas‐related G protein‐coupled receptor type D (MrgD), contributes to splanchnic vasodilatation in cirrhotic and noncirrhotic PHT. Splanchnic vascular hemodynamic and portal pressure were determined in two rat models of cirrhotic PHT and a rat model with noncirrhotic PHT, treated with either MrgD blocker D‐Pro(7)‐Ang‐(1‐7) (D‐Pro) or MasR blocker A779. Gene and protein expression of MrgD and MasR were measured in splanchnic vessels and livers of cirrhotic and healthy rats and in patients with cirrhosis and healthy subjects. Mesenteric resistance vessels isolated from cirrhotic rats were used in myographs to study their vasodilatory properties. MrgD was up‐regulated in cirrhotic splanchnic vessels but not in the liver. In cirrhotic rats, treatment with D‐Pro but not A779 completely restored splanchnic vascular resistance to a healthy level, resulting in a 33% reduction in portal pressure. Mesenteric vessels pretreated with D‐Pro but not with A779 failed to relax in response to acetylcholine. There was no splanchnic vascular MrgD or MasR up‐regulation in noncirrhotic PHT; thus, receptor blockers had no effect on splanchnic hemodynamics. Conclusion: MrgD plays a major role in the development of cirrhotic PHT and is a promising target for the development of novel therapies to treat PHT in cirrhosis. Moreover, neither MrgD nor MasR contributes to noncirrhotic PHT

Alzheimer’s disease AdvaxCpG- adjuvanted MultiTEP-based dual and single vaccines induce high-titer antibodies against various forms of tau and Aβ pathological molecules

This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/Although β-amyloid (Aβ) may be the primary driver of Alzheimer's disease (AD) pathology, accumulation of pathological tau correlates with dementia in AD patients. Thus, the prevention/inhibition of AD may require vaccine/s targeting Aβ and tau simultaneously or sequentially. Since high antibody titers are required for AD vaccine efficacy, we have decided to generate vaccines, targeting Aβ (AV-1959R), Tau (AV-1980R) or Aβ/tau (AV-1953R) B cell epitopes, based on immunogenic MultiTEP platform and evaluate the immunogenicity of these vaccines formulated with Advax(CpG), delta inulin, Alhydrogel(®), Montanide-ISA51, Montanide-ISA720, MPLA-SM pharmaceutical grade adjuvants. Formulation of AV-1959R in Advax(CpG) induced the highest cellular and humoral immune responses in mice. The dual-epitope vaccine, AV-1953R, or the combination of AV-1959R and AV-1980R vaccines formulated with Advax(CpG) induced robust antibody responses against various forms of both, Aβ and tau pathological molecules. While anti-Aβ antibody titers after AV-1953R immunization were similar to that in mice vaccinated with AV-1959R or AV-1959R/AV-1980R combination, anti-tau titers were significantly lower after AV-1953R injection when compared to the AV-1980R or AV-1959R/AV-1980R. In silico 3D-modeling provided insight into the differences in immunogenicity of these vaccine constructs. In sum, AV-1959R and AV-1980R formulated with Advax(CpG) adjuvant were identified as promising immunogenic vaccines for ongoing pre-clinical assessment and future human clinical trials

Proteomic identification of galectin-11 and 14 ligands from Haemonchus contortus

Haemonchus contortus is the most pathogenic nematode of small ruminants. Infection in sheep and goats results in anaemia that decreases animal productivity and can ultimately cause death. The involvement of ruminant-specific galectin-11 (LGALS-11) and galectin-14 (LGALS-14) has been postulated to play important roles in protective immune responses against parasitic infection; however, their ligands are unknown. In the current study, LGALS-11 and LGALS-14 ligands in H. contortus were identified from larval (L4) and adult parasitic stages extracts using immobilised LGALS-11 and LGALS-14 affinity column chromatography and mass spectrometry. Both LGALS-11 and LGALS-14 bound more putative protein targets in the adult stage of H. contortus (43 proteins) when compared to the larval stage (two proteins). Of the 43 proteins identified in the adult stage, 34 and 35 proteins were bound by LGALS-11 and LGALS-14, respectively, with 26 proteins binding to both galectins. Interestingly, hematophagous stage-specific sperm-coating protein and zinc metalloprotease (M13), which are known vaccine candidates, were identified as putative ligands of both LGALS-11 and LGALS- 14. The identification of glycoproteins of H. contortus by LGALS-11 and LGALS-14 provide new insights into host-parasite interactions and the potential for developing new interventions

Relationships of APOE genotypes with small RNA and protein cargo of brain tissue extracellular vesicles from patients with late-stage AD

Background and Objectives Variants of the apolipoprotein E (APOE) gene are the greatest known risk factors for sporadic Alzheimer disease (AD). Three major APOE isoform alleles, ϵ2, ϵ3, and ϵ4, encode and produce proteins that differ by only 1-2 amino acids but have different binding partner interactions. Whereas APOE ϵ2 is protective against AD relative to ϵ3, ϵ4 is associated with an increased risk for AD development. However, the role of APOE in gene regulation in AD pathogenesis has remained largely undetermined. Extracellular vesicles (EVs) are lipid bilayer-delimited particles released by cells to dispose of unwanted materials and mediate intercellular communication, and they are implicated in AD pathophysiology. Brain-derived EVs (bdEVs) could act locally in the tissue and reflect cellular changes. To reveal whether APOE genotype affects EV components in AD brains, bdEVs were separated from patients with AD with different APOE genotypes for parallel small RNA and protein profile. Methods bdEVs from late-stage AD brains (BRAAK stages 5-6) from patients with APOE genotypes ϵ2/3 (n = 5), ϵ3/3 (n = 5), ϵ3/4 (n = 6), and ϵ4/4 (n = 6) were separated using our published protocol into a 10,000g pelleted extracellular fraction (10K) and a further purified EV fraction. Counting, sizing, and multiomic characterization by small RNA sequencing and proteomic analysis were performed for 10K, EVs, and source tissue. Results Comparing APOE genotypes, no significant differences in bdEV total particle concentration or morphology were observed. Overall small RNA and protein profiles of 10K, EVs, and source tissue also did not differ substantially between different APOE genotypes. However, several differences in individual RNAs (including miRNAs and tRNAs) and proteins in 10K and EVs were observed when comparing the highest and lowest risk groups (ϵ4/4 and ϵ2/3). Bioinformatic analysis and previous publications indicate a potential regulatory role of these molecules in AD. Discussion For patients with late-stage AD in this study, only a few moderate differences were observed for small RNA and protein profiles between APOE genotypes. Among these, several newly identified 10K and EV-associated molecules may play roles in AD progression. Possibly, larger genotype-related differences exist and are more apparent in or before earlier disease stages

In silico structural homology modelling and docking for assessment of pandemic potential of a novel H7N9 influenza virus and its ability to be neutralized by existing anti-hemagglutinin antibodies.

The unpredictable nature of pandemic influenza and difficulties in early prediction of pandemic potential of new isolates present a major challenge for health planners. Vaccine manufacturers, in particular, are reluctant to commit resources to development of a new vaccine until after a pandemic is declared. We hypothesized that a structural bioinformatics approach utilising homology-based molecular modelling and docking approaches would assist prediction of pandemic potential of new influenza strains alongside more traditional laboratory and sequence-based methods. The newly emerged Chinese A/Hangzhou/1/2013 (H7N9) influenza virus provided a real-life opportunity to test this hypothesis. We used sequence data and a homology-based approach to construct a 3D-structural model of H7-Hangzhou hemagglutinin (HA) protein. This model was then used to perform docking to human and avian sialic acid receptors to assess respective binding affinities. The model was also used to perform docking simulations with known neutralizing antibodies to assess their ability to neutralize the newly emerged virus. The model predicted H7N9 could bind to human sialic acid receptors thereby indicating pandemic potential. The model also confirmed that existing antibodies against the HA head region are unable to neutralise H7N9 whereas antibodies, e.g. Cr9114, targeting the HA stalk region should bind with high affinity to H7N9. This indicates that existing stalk antibodies initially raised against H5N1 or other influenza A viruses could be therapeutically beneficial in prevention and/or treatment of H7N9 infections. The subsequent publication of the H7N9 HA crystal structure confirmed the accuracy of our in-silico structural model. Antibody docking studies performed using the H7N9 HA crystal structure supported the model's prediction that existing stalk antibodies could cross-neutralise the H7N9 virus. This study demonstrates the value of using in-silico structural modelling approaches to complement physical studies in characterization of new influenza viruses

Hangzhou-H7 neutralizing antibody binding.

<p>(<b>A</b>) The structures of neutralizing antibodies are shown either binding to the Hangzhou-H7 (shown in blue) globular head region (left figure) or stalk region (right figure). Different colours represent different antibodies. HA residues interacting with stalk-binding neutralizing antibodies are shown in the right hand figure highlighted in yellow. (B) HA-neutralizing antibodies binding the head or stalk regions were docked to the predicted Hangzhou-H7 structure and binding energies were calculated and presented as an absolute value. PDB IDs of antibodies against H1 are 3LZF, 3SM5, 4M5Z, 3GBN, 3ZTN, PDB IDs for H3 are 1EO8, 1KEN, 1QFU, 2VIR, 4FQR, 3SDY, 4NM8, H5 PDB IDs are 3FKU and 3GBM and H7 ID is 4FQV.</p

Binding affinity of Hangzhou-H7 and H7N7 for human and avian receptors.

<p>Hangzhou-H7 and the closely related H7N7 virus (A/Netherlands/219/2003) were docked to human and avian sialic acid receptors and affinities calculated. (<b>A</b>) The predicted Hangzhou-H7 model receptor affinities are shown by the black bars and the A/Netherlands/219/2003 affinities by the white bars. (<b>B</b>) The highest affinity docking conformation of Hangzhou-H7 on the human (brown), and avian (purple), receptors. HA mutations that influence the receptor binding specificity are highlighted in red and other residues that interact with the receptor in yellow. Residues that form the receptor-binding pocket are highlighted in green.</p

Panblok-H1+advax H1N1/2009pdm vaccine: Insights into rapid development of a delta inulin adjuvanted recombinant pandemic influenza vaccine

Timely vaccine supply is critical during influenza pandemics but is impeded by current virus-based manufacturing methods. The 2009 H1N1/2009pdm ‘swine flu’ pandemic reinforced the need for innovation in pandemic vaccine design. We report on insights gained during rapid development of a pandemic vaccine based on recombinant haemagglutinin (rHA) formulated with Advax™ delta inulin adjuvant (Panblok-H1/Advax). Panblok-H1/Advax was designed and manufactured within 1 month of the pandemic declaration by WHO and successfully entered human clinical testing in under 3 months from first isolation and sequencing of the novel pandemic virus, requiring several major challenges to be overcome. Panblok-H1/Advax successfully induced neutralising antibodies against the pandemic strain, but also induced cross-neutralising antibodies in a subset of subjects against an H1N1 strain (A/Puerto Rico/8/34) derived from the 1918 Spanish flu, highlighting the possibility to use Advax to induce more broadly cross-protective antibody responses. Interestingly, the rHA from H1N1/2009pdm exhibited variants in the receptor binding domain that had a major impact on receptor binding and hemagglutination ability. We used an in silico structural modeling approach to better understand the unusual behavior of the novel hemagglutinin, thereby demonstrating the power of computational modeling approaches for rapid characterization of new pandemic viruses. While challenges remain in ensuring ultrafast vaccine access for the entire population in response to future pandemics, the adjuvanted recombinant Panblok-H1/Advax vaccine proved its utility during a real-life pandemic situation

Teladorsagia Circumcincta Galectin-Mucosal Interactome in Sheep

Teladorsagia circumcincta is the most important gastrointestinal parasite in the livestock industry in temperate regions around the world, causing great economic losses. The infective third-stage larvae (L3) of Teladorsagia circumcincta secrete a large number of excretory-secretory (E/S) molecules, some of which are likely to play critical roles in modulating the host immune response. One of the most abundant E/S molecules is a protein termed Tci-gal-1, which has similarity to mammalian galectins. Galectins are a family of carbohydrate-binding molecules, with characteristic domain organisation and affinity for β-galactosids that mediate a variety of important cellular functions including inflammation and immune responses. To understand the role of Tci-gal-1 at the host–parasite interface, we used a proteomics pull-down approach to identify Tc-gal-1 interacting proteins from sheep abomasal scrapes and whole tissue. A total of 135 unique proteins were identified from whole abomasal tissue samples, while 89 proteins were isolated from abomasal scrape samples. Of these proteins, 63 were present in both samples. Many of the host proteins identified, such as trefoil factors and mucin-like proteins, play critical roles in the host response. The identification of Tci-gal-1 binding partners provides new insights on host–parasite interactions and could lead to the development of new control strategies

Revealing the proteome of motor cortex derived extracellular vesicles isolated from amyotrophic lateral sclerosis human postmortem tissues

Amyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disease characterized by the deposition of misfolded proteins in the motor cortex and motor neurons. Although a multitude of ALS-associated mutated proteins have been identified, several have been linked to small extracellular vesicles such as exosomes involved in cell-cell communication. This study aims to determine the proteome of extracellular vesicles isolated from the motor cortex of ALS subjects and to identify novel ALS-associated deregulated proteins. Motor cortex extracellular vesicles (MCEVs) were isolated from human postmortem ALS (n = 10) and neurological control (NC, n = 5) motor cortex brain tissues and the MCEVs protein content subsequently underwent mass spectrometry analysis, allowing for a panel of ALS-associated proteins to be identified. This panel consists of 16 statistically significant differentially packaged proteins identified in the ALS MCEVs. This includes several upregulated RNA-binding proteins which were determined through pathway analysis to be associated with stress granule dynamics. The identification of these RNA-binding proteins in the ALS MCEVs suggests there may be a relationship between ALS-associated stress granules and ALS MCEV packaging, highlighting a potential role for small extracellular vesicles such as exosomes in the pathogenesis of ALS and as potential peripheral biomarkers for ALS