Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field

Iron Behaving Badly: Inappropriate Iron Chelation as a Major Contributor to the Aetiology of Vascular and Other Progressive Inflammatory and Degenerative Diseases

The production of peroxide and superoxide is an inevitable consequence of aerobic metabolism, and while these particular "reactive oxygen species" (ROSs) can exhibit a number of biological effects, they are not of themselves excessively reactive and thus they are not especially damaging at physiological concentrations. However, their reactions with poorly liganded iron species can lead to the catalytic production of the very reactive and dangerous hydroxyl radical, which is exceptionally damaging, and a major cause of chronic inflammation. We review the considerable and wide-ranging evidence for the involvement of this combination of (su)peroxide and poorly liganded iron in a large number of physiological and indeed pathological processes and inflammatory disorders, especially those involving the progressive degradation of cellular and organismal performance. These diseases share a great many similarities and thus might be considered to have a common cause (i.e. iron-catalysed free radical and especially hydroxyl radical generation). The studies reviewed include those focused on a series of cardiovascular, metabolic and neurological diseases, where iron can be found at the sites of plaques and lesions, as well as studies showing the significance of iron to aging and longevity. The effective chelation of iron by natural or synthetic ligands is thus of major physiological (and potentially therapeutic) importance. As systems properties, we need to recognise that physiological observables have multiple molecular causes, and studying them in isolation leads to inconsistent patterns of apparent causality when it is the simultaneous combination of multiple factors that is responsible. This explains, for instance, the decidedly mixed effects of antioxidants that have been observed, etc...Comment: 159 pages, including 9 Figs and 2184 reference

Impaired T cell-mediated hepatitis in peroxisome proliferator activated receptor alpha (PPARα)-deficient mice

Copyright © The Author(s) 2018. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License, (http://creativecommons.org/licenses/by/4.0/). The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/ publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.Background Peroxisome proliferator activated receptor alpha (PPARα), a regulator of enzymes involved in β oxidation, has been reported to influence lymphocyte activation. The purpose of this study was to determine whether PPARα plays a role in T cell-mediated hepatitis induced by Concanavalin A (ConA). Methods Wild type (wt) or PPARα-deficient (PPARα−/−) mice were treated with ConA (15 mg/kg) by intravenous injection 0, 10 or 24 h prior to sacrifice and serum and tissue collection for analysis of tissue injury, cytokine response, T cell activation and characterization. Results Ten and 24 h following ConA administration, wt mice had significant liver injury as demonstrated by serum transaminase levels, inflammatory cell infiltrate, hepatocyte apoptosis, and expression of several cytokines including interleukin 4 (IL4) and interferon gamma (IFNγ). In contrast, PPARα−/− mice were protected from ConA-induced liver injury with significant reductions in serum enzyme release, greatly reduced inflammatory cell infiltrate, hepatocellular apoptosis, and IFNγ expression, despite having similar levels of hepatic T cell activation and IL4 expression. This resistance to liver injury was correlated with reduced numbers of hepatic natural killer T (NKT) cells and their in vivo responsiveness to alpha-galactosylceramide. Interestingly, adoptive transfer of either wt or PPARα−/− splenocytes reconstituted ConA liver injury and cytokine production in lymphocyte-deficient, severe combined immunodeficient mice implicating PPARα within the liver, possibly through support of IL15 expression and/or suppression of IL12 production and not the lymphocyte as the key regulator of T cell activity and ConA-induced liver injury. Conclusion Taken together, these data suggest that PPARα within the liver plays an important role in ConA-mediated liver injury through regulation of NKT cell recruitment and/or survival.ECU Open Access Publishing Support Fun

Centrality dependence of the pseudorapidity density distribution for charged particles in Pb\u2013Pb collisions at 1asNN = 2.76 TeV

We present the first wide-range measurement of the charged-particle pseudorapidity density distribution, for different centralities (the 0\u20135%, 5\u201310%, 10\u201320%, and 20\u201330% most central events) in Pb\u2013Pb collisions at 1asNN = 2.76 TeV at the LHC. The measurement is performed using the full coverage of the ALICE detectors, 125.0 < \u3b7 < 5.5, and employing a special analysis technique based on collisions arising from LHC \u2018satellite\u2019 bunches. We present the pseudorapidity density as a function of the number of participating nucleons as well as an extrapolation to the total number of produced charged particles (Nch = 17 165 \ub1 772 for the 0\u20135% most central collisions). From the measured dNch/d\u3b7 distribution we derive the rapidity density distribution, dNch/dy, under simple assumptions. The rapidity density distribution is found to be significantly wider than the predictions of the Landau model. We assess the validity of longitudinal scaling by comparing to lower energy results from RHIC. Finally the mechanisms of the underlying particle production are discussed based on a comparison with various theoretical models

Multiplicity dependence of pion, kaon, proton and lambda production in p–Pb collisions at √sNN = 5.02 TeV

Inthis Letter, comprehensive results on π±,K±,K0S, p(pbar) and Λ(Λbar) production at mid-rapidity (0< yCMS < 0.5) in p–Pb collisions at √sNN = 5.02 TeV, measured by the ALICE detector at the LHC, are reported. The transverse momentum distributions exhibit a hardening as a function of event multiplicity, which is stronger for heavier particles. This behavior is similar to what has been observed in pp and Pb–Pb collisions at the LHC. The measured pT distributions are compared to d–Au, Au–Au and Pb–Pb results at lower energy and with predictions based on QCD-inspired and hydrodynamic models