Citrullination of CXCL8 by peptidylarginine deiminase alters receptor usage, prevents proteolysis, and dampens tissue inflammation

Biological functions of proteins are influenced by posttranslational modifications such as on/off switching by phosphorylation and modulation by glycosylation. Proteolytic processing regulates cytokine and chemokine activities. In this study, we report that natural posttranslational citrullination or deimination alters the biological activities of the neutrophil chemoattractant and angiogenic cytokine CXCL8/interleukin-8 (IL-8). Citrullination of arginine in position 5 was discovered on 14% of natural leukocyte-derived CXCL8(1–77), generating CXCL8(1–77)Cit5. Peptidylarginine deiminase (PAD) is known to citrullinate structural proteins, and it may initiate autoimmune diseases. PAD efficiently and site-specifically citrullinated CXCL5, CXCL8, CCL17, CCL26, but not IL-1β. In comparison with CXCL8(1–77), CXCL8(1–77)Cit5 had reduced affinity for glycosaminoglycans and induced less CXCR2-dependent calcium signaling and extracellular signal-regulated kinase 1/2 phosphorylation. In contrast to CXCL8(1–77), CXCL8(1–77)Cit5 was resistant to thrombin- or plasmin-dependent potentiation into CXCL8(6–77). Upon intraperitoneal injection, CXCL8(6–77) was a more potent inducer of neutrophil extravasation compared with CXCL8(1–77). Despite its retained chemotactic activity in vitro, CXCL8(1–77)Cit5 was unable to attract neutrophils to the peritoneum. Finally, in the rabbit cornea angiogenesis assay, the equally potent CXCL8(1–77) and CXCL8(1–77)Cit5 were less efficient angiogenic molecules than CXCL8(6–77). This study shows that PAD citrullinates the chemokine CXCL8, and thus may dampen neutrophil extravasation during acute or chronic inflammation

The development of therapeutic proteins can be hindered by poor decision-making strategies in the early stage

In this study we address two major issues related to the current development process of therapeutic proteins and their characterization. First, due to limited samples amounts, the selection of lead molecules in the early stages is often based on the results from a limited physicochemical characterization. The latter can be based on measurements of only 2-3 parameters, e.g. protein melting temperature, protein aggregation temperature, and is usually performed in only one buffer, e.g. PBS. The hypothesis we present is that such approach can lead to the rejection of lead candidates that can still be manufacturable and can move on to clinical trials. The second matter we address are the often-reported correlations between protein physicochemical parameters in the literature. We propose that such correlations can be found only in a small sample population, e.g. one protein in different solution conditions or different proteins from the same class. However, we expect that such correlations would not be valid in a large population, including various protein structures and solution conditions. In order to address the above-mentioned issues, we created the PIPPI consortium (http://www.pippi.kemi.dtu.dk) and applied systematic approach to map the physicochemical properties of a wide range of proteins and extensively study their stability as a function of the solution conditions. We show that promising therapeutic protein lead candidate can appear as non-manufacturable when only limited physicochemical characterization is performed, e.g. a few methods are used and only a few solution conditions are tested. Therefore, the rejection rate during early-stage development can be improved by more thorough physicochemical characterization. Moreover, only weak linear correlations between biophysical properties of proteins are observed in a large populations. This suggests that the often-reported correlations between parameters describing the protein stability are not representative of a global population. Understanding the connections between various physiochemical parameters would require a systematic database which is currently in development by the PIPPI consortium

Holmium Nanoparticles: Preparation and In Vitro Characterization of a New Device for Radioablation of Solid Malignancies

# The Author(s) 2010. This article is published with open access at Springerlink.com Purpose The present study introduces the preparation and in vitro characterization of a nanoparticle device comprising holmium acetylacetonate for radioablation of unresectable solid malignancies. Methods HoAcAc nanoparticles were prepared by dissolving holmium acetylacetonate in chloroform, followed by emulsification in an aqueous solution of a surfactant and evaporation of W. Bult: R. Varkevisser: P. R. Luijten: A. D. van het Schip

The McKenzie method for the management of acute non-specific low back pain: design of a randomised controlled trial [ACTRN012605000032651]

BACKGROUND: Low back pain (LBP) is a major health problem. Effective treatment of acute LBP is important because it prevents patients from developing chronic LBP, the stage of LBP that requires costly and more complex treatment. Physiotherapists commonly use a system of diagnosis and exercise prescription called the McKenzie Method to manage patients with LBP. However, there is insufficient evidence to support the use of the McKenzie Method for these patients. We have designed a randomised controlled trial to evaluate whether the addition of the McKenzie Method to general practitioner care results in better outcomes than general practitioner care alone for patients with acute LBP. METHODS/DESIGN: This paper describes the protocol for a trial examining the effects of the McKenzie Method in the treatment of acute non-specific LBP. One hundred and forty eight participants who present to general medical practitioners with a new episode of acute non-specific LBP will be randomised to receive general practitioner care or general practitioner care plus a program of care based on the McKenzie Method. The primary outcomes are average pain during week 1, pain at week 1 and 3 and global perceived effect at week 3. DISCUSSION: This trial will provide the first rigorous test of the effectiveness of the McKenzie Method for acute non-specific LBP

The circulating SARS-CoV-2 spike variant N439K maintains fitness while evading antibody-mediated immunity

SARS-CoV-2 can mutate to evade immunity, with consequences for the efficacy of emerging vaccines and antibody therapeutics. Herein we demonstrate that the immunodominant SARS-CoV-2 spike (S) receptor binding motif (RBM) is the most divergent region of S, and provide epidemiological, clinical, and molecular characterization of a prevalent RBM variant, N439K. We demonstrate that N439K S protein has enhanced binding affinity to the hACE2 receptor, and that N439K virus has similar clinical outcomes and in vitro replication fitness as compared to wild- type. We observed that the N439K mutation resulted in immune escape from a panel of neutralizing monoclonal antibodies, including one in clinical trials, as well as from polyclonal sera from a sizeable fraction of persons recovered from infection. Immune evasion mutations that maintain virulence and fitness such as N439K can emerge within SARS-CoV-2 S, highlighting the need for ongoing molecular surveillance to guide development and usage of vaccines and therapeutics

25th annual computational neuroscience meeting: CNS-2016

The same neuron may play different functional roles in the neural circuits to which it belongs. For example, neurons in the Tritonia pedal ganglia may participate in variable phases of the swim motor rhythms [1]. While such neuronal functional variability is likely to play a major role the delivery of the functionality of neural systems, it is difficult to study it in most nervous systems. We work on the pyloric rhythm network of the crustacean stomatogastric ganglion (STG) [2]. Typically network models of the STG treat neurons of the same functional type as a single model neuron (e.g. PD neurons), assuming the same conductance parameters for these neurons and implying their synchronous firing [3, 4]. However, simultaneous recording of PD neurons shows differences between the timings of spikes of these neurons. This may indicate functional variability of these neurons. Here we modelled separately the two PD neurons of the STG in a multi-neuron model of the pyloric network. Our neuron models comply with known correlations between conductance parameters of ionic currents. Our results reproduce the experimental finding of increasing spike time distance between spikes originating from the two model PD neurons during their synchronised burst phase. The PD neuron with the larger calcium conductance generates its spikes before the other PD neuron. Larger potassium conductance values in the follower neuron imply longer delays between spikes, see Fig. 17.Neuromodulators change the conductance parameters of neurons and maintain the ratios of these parameters [5]. Our results show that such changes may shift the individual contribution of two PD neurons to the PD-phase of the pyloric rhythm altering their functionality within this rhythm. Our work paves the way towards an accessible experimental and computational framework for the analysis of the mechanisms and impact of functional variability of neurons within the neural circuits to which they belong