bluuebunny/crossref_metadata_2025 huggingface dataset:
The Emerging Contribution of Director Disqualification in UK Competition Law
Authors: Whelan Peter
Published: June 2021

Peter Whelan assesses a developing and increasingly significant enforcement tool in the UK competition authority’s armoury in Chapter 11. In it, Whelan notes that the enforcement of UK competition law is deterrence-focused and comprises both criminal and non-criminal (i.e. civil/administrative) elements. The chapter concentrates on the non-criminal enforcement apparatus that has been developed over the last twenty years. More specifically, it critically evaluates a particular enforcement mechanism that has been gaining increasing importance throughout the recent development of UK competition enforcement practice: the use of director disqualification. It first establishes the normative role of director disqualification in the UK’s armoury of non-criminal antitrust sanctions (i.e. its complementing of the deterrent function of corporate antitrust fines), following which it highlights their potential for performing this role effectively. It then outlines the legal basis for the use of director disqualification within the UK and evaluates the policy and enforcement practice to date with respect to such orders, before proceeding to outline some of the insights that the UK director disqualification regime can provide to other jurisdictions. Ultimately it concludes that, on the basis of the promising, albeit nascent, UK experience to date, director disqualification should be seriously considered by jurisdictions that wish to operate a robust competition law enforcement regime.
Private Enforcement in the UK
Authors: Rodger Barry J
Published: June 2021

In Chapter 12, Barry Rodger retraces his footsteps in relation to his contributions in both earlier collections on the theme of private enforcement in the UK, with a particular slant on the extent to which consumers have benefited, or may benefit, from statutory and case law developments in the area. Accordingly, this chapter assesses how private enforcement of competition law rights has developed in the UK over the last twenty years. Key legislative developments, inter alia the Competition Act 1998, Enterprise Act 2002 and Consumer Rights Act 2015, have transformed the private enforcement architecture, notably with the introduction, and increasingly significant and enhanced role of the specialist tribunal, Competition Appeal Tribunal, and the availability of an opt-out collective redress mechanism. The chapter assesses the key UK statutory and case law developments, in comparison with the US private antitrust enforcement model, to reflect on the disappointing extent to which effective redress for consumers has been provided to date, despite those legal and institutional developments, although the recent Supreme Court ruling in Merricks should be pivotal in this context.
The Quiet Decline of the UK Cartel OffenceA Principled Victory in the Face of Practical Failure
Authors: MacCulloch Angus
Published: June 2021

In Chapter 13, Angus MacCulloch focuses on criminal enforcement. The UK Cartel Offence was introduced in the Enterprise Act 2002 to challenge hard-core cartels and enhance the deterrent effect of the UK competition regime. In its initial phase of operation there was some success. However, a number of significant cases failed to secure convictions. This damaged confidence in the ability of the UK competition authorities to bring successful prosecutions, and ultimately questioned the usefulness of the Cartel Offence. This chapter examines the problems that beset the original Cartel Offence and the lessons learned from the small number of prosecutions brought before the courts. It goes on to examine the reforms in 2013, that removed the controversial ‘dishonesty’ element from the offence, and replaced it with carve outs for openness and publication. Alongside the practical issues in relation to the development of the UK Cartel Offence consideration is also given to a parallel process which saw a form of consensus developing in the academic literature as to the nature of the wrong at the heart of individual cartel activity. It is suggested that this greater understanding can be used to direct efforts to rebuild confidence in the reformed UK Cartel Offence going forward. Increased importance should be given to the securing of good evidence of individual culpability in relation to cartel activity during the investigation phase. It argues that once good evidence is secured, better prosecution cases can be built on the basis of the new narrative of wrongfulness for hard core cartel activity.
Freedom of Expression as Self-Restraint
Authors: Kramer Matthew H.
Published: June 2021

Freedom of Expression as Self-Restraint rigorously expounds the principle of freedom of expression, and provides a novel justificatory foundation for it. Under that principle, a system of governance in any society can legitimately prohibit various modes of communication but cannot ever legitimately prohibit them qua modes of communication. As the book argues, such a principle is absolute in that it is exceptionless; it imposes general duties that are binding always and everywhere on every system of governance. In addition to injecting a new level of philosophical sophistication into the debates over these matters, the book supplies a novel justification for the principle of freedom of expression. It ties that principle to an ideal of governmental self-restraint, and it shows how that ideal connects to the paramount moral responsibility of every system of governance: the responsibility to bring about the political and social and economic conditions under which every member of a society can be warranted in harboring an ample sense of self-respect. In short, compliance by a system of governance with the principle of freedom of expression is integral to the fulfillment of that paramount responsibility. Kramer lengthily engages with arguments by feminists in favor of legal restrictions on pornography, and with prominent arguments in favor of banning the advocacy of hateful creeds. While accepting that some types or instances of pornography and hatemongering can properly be proscribed, he maintains that most types and instances of those modes of communication are morally protected by the principle of freedom of expression.
Introduction: Philosophical Preliminaries
Authors: Kramer Matthew H.
Published: June 2021

This chapter expounds some technical philosophical notions that figure prominently in the rest of the book. Among the phenomena elucidated are moral conflicts, strong permissibility versus weak permissibility, overtopping obligations versus non-overtopping obligations, weak absoluteness versus strong absoluteness, physical freedom versus deontic freedom, the Hohfeldian analysis of legal positions (claim-rights and duties, liberties and no-rights, powers and liabilities, and immunities and disabilities), and the distinction between causal relationships and constitutive relationships. Although all of these concepts and distinctions are of great significance in debates over the principle of freedom of expression, they also figure saliently in many other debates. Hence, the purpose of this introductory chapter is to provide the basic philosophical preliminaries that are needed for a rigorous engagement with the principle of freedom of expression. The chapter sets the stage for the subsequent chapters, where that principle is the focus of attention.
Components of a Theory of Freedom of Expression
Authors: Kramer Matthew H.
Published: June 2021

This chapter introduces many of the key concepts that are of distinctive importance in any theory of freedom of expression. It first explores the nature of expression or communication (comprising addressors, addressees, communicative contents, and certain intentions), and it then considers whether the distinction between speech and conduct is tenable. Contending that the distinction between high-value modes and low-value modes of expression does not have any bearing on the applicability of the principle of freedom of expression, the chapter maintains that that principle protects every mode of expression against any governmental restrictions that are directed against it qua mode of expression. Compliance with that principle by a system of governance involves several types of neutrality, which the chapter expounds. Having explored the ways in which a system of governance can fail to achieve those types of neutrality, the chapter then explains how the purposes of various laws or policies are to be ascertained. Thereafter, the chapter concludes by pondering how private parties (such as hecklers and employers and organizations in charge of public fora) are constrained by the principle of freedom of expression.
Legitimate Restrictions
Authors: Kramer Matthew H.
Published: June 2021

Although the principle of freedom of expression is a moral absolute that imposes general duties which bind all systems of governance always and everywhere, it is consistent with various legal or governmental restrictions on modes of expression—provided that those restrictions are directed against the modes of expression not qua modes of expression but instead qua communication-independent misconduct. This chapter explores this aspect of the principle of freedom of expression by cataloguing various types of communicative conduct that can properly be subjected to legal restrictions. Among those types of communicative conduct are perjury, libel, solicitation to commit a crime, true threats, fighting words, incitement (in the sense of the term articulated by the U.S. Supreme court ruling in Brandenburg v Ohio), and fraud. All of these types of communicative conduct, along with several others, can be legally prohibited in accordance with the principle of freedom of expression.
Democratic Speech in Divided Times
Authors: Lepoutre Maxime
Published: June 2021

Democratic Speech in Divided Times offers a comprehensive account of the norms that should govern public discourse in circumstances marked by deep and often unjust social divisions. Part I investigates what forms of democratic speech are desirable in these settings. This part shows, firstly, that some forms of public discourse that are symptomatic of division can nevertheless play a crucial democratic function. In particular, it argues that emotionally charged speech—and most notably, speech voicing deep anger—plays a fundamental role in overcoming entrenched epistemic divisions and in facilitating the exchange of shared reasons. This part also examines how, in contrast, other characteristic features of the public discourse of divided societies endanger democratic life. Here, the argument considers the proliferation of hate speech and misinformation, and examines what forms of democratic speech should be used to combat them. Part II considers how realistic the foregoing account of public discourse is. Specifically, it assesses the complications that arise from intergroup antipathy, pervasive political ignorance, and the fragmentation of the public sphere. The normative picture of public discourse that this book defends can largely withstand these problems. And, while these social conditions do qualify the value of democratic speech in some respects, they are at least as problematic for political ideals that give up on inclusive democratic speech altogether. Accordingly, while realising the ideal of democratic speech that this book outlines is challenging, we should not lose patience with this task.
Financial crises and the limits of reforms
Authors: Keller Eileen
Published: June 2021

The introductory chapter presents the main themes of the book. It illustrates the recurrence of financial crises and discusses the contours and limits of the financial sector reforms implemented in response to the global financial crisis from 2007 to 2009. It spells out the specificities of banking and outlines the implications of the crisis for the future role of banks in so-called bank-based financial systems. The chapter spells out the empirical puzzle and discusses the shortcomings of existing theoretical approaches in accounting for it. The social learning approach and the methodology of the research on which the book builds are presented. The chapter summarizes the book’s main argument and closes with a short chapter outline.
Social learning in response to the crisis
Authors: Keller Eileen
Published: June 2021

This chapter explains the differing conclusions drawn from the financial crisis in France and Germany. The chapter provides original empirical data that shows that the banks’ lending behaviour during the crisis cannot account satisfactorily for the differing reform priorities identified in France and Germany. The second part of the chapter explores the pre-crisis developments that triggered the differing reactions. Simply put, while social learning was about the conditions and the availability of bank lending in France, in Germany it was related to the risks and the benefits associated with innovative market-based banking and financial sector concentration. The chapter confirms the respective narratives by systematically comparing and crosschecking the diverging dynamics in the two cases, building on extensive empirical data.
Comparative lessons
Authors: Keller Eileen
Published: June 2021

This chapter takes stock of the previous chapters and discusses the findings. It summarizes the book’s main argument and examines the nature of social learning with respect to true learning, the institutional dispositions in place, the role of politics, and contingency. The validity of the argument is discussed using Bayesian updating, presenting additional evidence that confirms the findings in a comparative perspective and in the light of competing hypotheses. The relevance of the social learning framework is demonstrated by applying it to the British and the US-American crisis responses and to differing outcomes on bank structural reform. The chapter closes with a brief discussion of the theoretical merits of social learning for ideational scholarship in institutional analysis, public policy-making, and economic sociology.
Efficiency of Dynamic Portfolio Choices: An Experiment
Authors: Magnani Jacopo , Rabanal Jean Paul , Rud Olga A , Wang Yabin
Published: June 2021

We study the efficiency of dynamic portfolio choices using the nonparametric methods of Dybvig (1988) and Post (2003). We compare a dynamic portfolio task against an equivalent static Arrow-Debreu problem under two alternative environments: (1) nonpooled with $2^T$ terminal states and (2) pooled with $T+1$ unique terminal states. The results suggest that, within each environment, efficiency is lower in a static format and when the number of final states is larger. In the nonpooled dynamic task, which allows for path dependent strategies, we find that a form of stop-loss strategy drives efficiency losses.
Investigation of Protein‐Protein Interactions in the Pyruvate Dehydrogenase Complex from Escherichia coli by Hydrogen/Deuterium Exchange Mass Spectrometry
Authors: Wang Junjie , Nemeria Natalia S. , Jordan Frank
Published: June 2021

The pyruvate dehydrogenase multienzyme complex (PDHc) from Escherichia coli is a molecular machine consisting of multiple copies of three enzyme components: pyruvate dehydrogenase (E1p), dihydrolipoyl acetyltransferase (E2p), and dihydrolipoyl dehydrogenase (E3p), which cooperate with each other to perform the oxidative decarboxylation of pyruvate to acetyl‐CoA. To probe the protein‐protein interactions and conformational changes between these components, we used hydrogen/deuterium exchange mass spectrometry (DXMS). Based on our study, the regions of interactions are mapped as follows: 1. The interaction of E2p with the active regions of E1p that were missing from previous X‐ray crystallography (an inner loop formed by residues 401–413, outer loop formed by residues 541–557 and N‐terminal residues 1–55); 2. The binding loci on peripheral‐subunit binding domain (PSBD) of E2p that interact with E1p and E3p; 3. The interaction of the E3p redox‐active region with E1p and E2p. This DXMS method has proved to be fast, accurate and reliable in characterizing the interaction of components in PDHc. The authors thank NIHGM‐050380 for financial support.
The Allosteric Regulation of Phenylalanine Hydroxylase Provides a Foundation for New PKU Therapies
Authors: Jaffe Eileen K , Stith Linda , Lawrence Sarah H , Andrake Mark , Dunbrack Roland L
Published: June 2021

The structural basis for allostery in phenylalanine hydroxylase (PAH), whose dysfunction causes phenylketonuria (PKU), is poorly understood. A dissociative model for PAH allostery is proposed to consist of an equilibrium between architecturally different tetramers whose interconversion requires a ~90Â° rotation between PAH catalytic and regulatory domains, the latter of which contains an ACT domain. This unprecedented model is supported by in vitro data on purified full length mammalian PAH. The conformational change renders the tetramers chromatographically separable using ion exchange methods. A novel aspect of the activated tetramer model is an allosteric Phe binding site at an inter‐subunit interface of ACT domains; this provides a structural rationale for Phe‐induced tetramer stabilization. Ligand‐stabilized ACT domain dimerization follows the multimerization and ligand binding behavior of ACT domains in the PDB. Spectroscopic, chromatographic, and electrophoretic methods demonstrate a PAH equilibrium consisting of architecturally distinct tetramers as well as dimers. We postulate that PKU‐associated mutations may shift the PAH quaternary structure equilibrium in favor of the low activity assemblies. Pharmacological chaperones that bind to and stabilize the ACT:ACT interface can potentially provide PKU patients with a novel small molecule therapeutic. Supported by NIH P30CA006927.
Measurement of Urinary Catecholamines for Mice
Authors: Smith Emily A , Schwartz Abigail L , Garrett Teresa L , Lucot James B
Published: June 2021

A method for extraction of unconjugated catecholamines in 20μL urine samples has been developed using alumina extraction prior to separation by high performance liquid chromatography (HPLC) and electrochemical detection (ECD). Murine experiments tested the application of the procedure. In the first, collection occurred in the morning and evening prior to handling, and in the morning after three days of handling. In the second, passively obtained urine was compared to stressfully obtained urine in the same mice. Urine was extracted alongside 2,3‐dihydroxybenzoic acid internal standard via alumina and brought to pH 8.5 with tris buffer. Two wash steps preceded acid elution for analysis on HPLC with ECD. This method averaged 50 to 70% recovery. Differences were not seen when comparing small variations collection time, handling exposure, or collection technique. This procedure effectively measures sympathetic activity in micro samples, with a limit of detection in the attomole range.
Role of the sphingolipid biostat in pancreatic βcell apoptosis induced by gluco‐lipotoxicity
Authors: COANT Nicolas , Veret Julien , Giussani Paola , Portha Bernard , Berdyshev Evgeny V , Le Stunff Herve
Published: June 2021

Accumulated evidence suggests that Type 2 diabetes is often associated with abnormalities in lipids metabolism. Adverse effects of fatty acids on β cell function and viability are potentiated by the presence of hyperglycemia; this phenomenon is called gluco‐lipotoxicity. Sphingoid base‐1‐phosphates and ceramides are potent bioactive lipids involved in different patho‐physiological processes. This study was undertaken to determine whether these lipids are produced in palmitate‐treated pancreatic β cells and what role they play in palmitate‐induced beta cell apoptosis.

Our lipidomic analysis revealed that palmitate at low and high glucose supplementation increased (dihydro)‐sphingosine‐1‐phosphate (S1P) and (dihydro)‐ceramides levels in INS‐1 β cells. This increase is associated with an increase of SphK1 and Cers4 proteins. Interestingly, we show that the over‐expression of Cers4 or the inhibition of SphK1 induces an increase of apoptosis induced by palmitate with 30 mM glucose, whereas down‐regulation of CerS4 by siRNA or over‐expression of SphK1 reduced apoptosis.

Taken together, our results suggest an important role of the balance between S1P and ceramide (i.e. sphingolipid biostat) in β cell death induced by gluco‐lipotoxicity. These data suggest that targeting specific sphingolipid metabolites could be a novel therapeutic strategy to reduce the development of TD2.
Cystathionine‐γ‐lyase (CSE) activation and H2S formation mediate the effect of vitamin D on glucose transporter 4 (GLUT4) translocation and glucose utilization in high glucose treated 3T3L1 adipocytes
Authors: Manna Prasenjit , Jain Sushil K.
Published: June 2021

This study examined the biochemical mechanism by which vitamin D supplementation may regulate glucose metabolism in diabetes. 3T3L1 adipocytes were treated with high glucose (HG, 25 mM) in the presence or absence of 1,25‐dihydroxyvitamin D3 [1,25(OH)2D3] (25 or 50 nM), the active form of vitamin D. 1,25(OH)2D3‐treatment up‐regulated GLUT4 translocation, glucose uptake as well as glucose utilization, CSE activation and H2S formation in HG‐treated adipocytes. CSE catalyzes the H2S formation. The effect of 1,25(OH)2D3 on GLUT4 translocation, glucose utilization, and H2S formation was prevented by propargylglycine (an inhibitor of CSE) or in CSE‐knockdown (using antisense CSE) adipocytes compared to controls. 1,25(OH)2D3 treatment along with insulin enhanced GLUT4 translocation and glucose utilization compared to either insulin or 1,25(OH)2D3 alone in HG‐treated cells. 1,25(OH)2D3 supplementation also inhibited MCP‐1 and stimulated adiponectin secretion in HG‐treated adipocytes and this positive effect was prevented in CSE‐inhibitor‐treated or CSE‐knockdown adipocytes. These results demonstrate the essential role of CSE activation and H2S formation in mediating the effect of vitamin D on GLUT4 translocation, glucose utilization, and decrease in inflammatory markers in HG‐treated adipocytes. (Supported by NIDDK RO1 DK072433)
Mouse Models of Insulin Resistance: What Have We Learned?
Authors: Bunner Anne E , Barnard Neal D
Published: June 2021

Type 2 diabetes (T2D) affects about 26 million adults in the US, and is a leading cause of cardiovascular mortality. Insulin resistance is a core pathology of T2D, and often arises years before diagnosis. In an effort to understand and treat insulin resistance and T2D, basic researchers have used genetically modified mice to identify insulin signaling pathways and attempt to determine the effects of their perturbation. However, despite decades of these inquiries, the pathophysiology of insulin resistance remains controversial, and no pharmacological cure exists. In light of these facts, and the costs of animal research, we reviewed the uses and limitations of genetically engineered mouse models of insulin resistance. We examined the features of several widely employed mouse models and the relevance of results to human patients. These genetically modified mice have important differences from the human patients they model at the molecular, cellular, organ, and whole‐organism level. Much has been learned about signaling pathways in mice, but the mechanistic insights gained are often limited to the species and strain of the experimental animal, rather than generalizable to the human condition. We conclude that the significant barriers to translatability point out the need to transition from mouse models to human‐based studies of T2D.
Nucleotide Binding Stabilizes Na,K‐ATPase Quaternary Structure and Facilitates Trafficking to Plasma Membrane
Authors: Galva Charitha , Louis Kathleen R , Gatto Craig
Published: June 2021

Na,K‐ATPase (NKA) is an integral membrane protein that plays a central role in ionic homeostasis in animals by mediating transport of Na+ and K+ ions against their electrochemical gradients. NKA is a heterodimer of α and β subunits. It is not yet clear if the native structure of NKA consists of αβ‐protomers or higher order oligomeric states. α‐ α associations were earlier demonstrated using in vitro pull down assays with isolated ATP‐binding domain (ABD) and full‐length α (Costa et al., 2002). Our objective is to elucidate the oligomeric state of NKA throughout maturation and trafficking and the role it plays in protein‐protein interactions. Consistent with previous observations, results from our co‐immunoprecipitation experiments using lysates from HEK 293 cells stably expressing α‐subunit with v5‐ or myc‐tagged NKA α‐subunits demonstrate α‐α interaction. Moreover, this self‐association is stabilized by including 2mM Mg‐ATP in the assay. Our preliminary results from confocal imaging of HEK 293 cells co‐expressing GFP‐tagged ABD and mcherry‐tagged full length α‐subunit revealed GFP‐ABD in the plasma membrane (PM), consistent with the isolated ABD being “carried” to the PM with full‐length α‐subunits. We also found that the α‐subunit lacking the M4M5 cytoplasmic loop (αΔ45LL), and thus unable to bind ATP, was retained in ER. Studies are underway to see if expressing the isolated ABD will rescue αΔ45LL and facilitate its delivery to the PM. This work was supported by NIH grant GM061583.
Mechanisms of non‐random sphingolipid organization in the plasma membranes of fibroblast cells
Authors: Kraft Mary L , Klitzing Haley A , Frisz Jessica F , Kou Kaiyan , Weber Peter K , Zimmerberg Joshua
Published: June 2021

Organization of the plasma membranes’ of mammalian cells into domains of differing protein in lipid composition is likely important for coordinating cell signaling and the numerous other processes that occur at the cell surface. By using a high‐resolution secondary ion mass spectrometry (SIMS) approach to chemically image the distributions of metabolically incorporated 15N‐sphingolipids in the plasma membranes of intact fibroblast cells, we previously showed that sphingolipids are non‐randomly organized in the plasma membrane. We now use this approach to test the mechanisms that are most often hypothesized to be responsible for sphingolipid distribution in the plasma membrane. We show that the non‐random sphingolipid organization within the plasma membrane cannot be attributed to favorable interactions with cholesterol. Instead, our results indicate that intracellular vesicle traffic and the cytoskeleton are responsible for the sphingolipid domains we observed.
Muscle Enriched A‐Type Lamin Interacting Protein (MLIP) a novel regulator cardiac growth and function
Authors: Cattin Marie‐Elodie , Mak‐Washburn Esther , Burgon Patrick G
Published: June 2021

The newly discovered Muscle‐enriched A‐type lamin interacting protein (MLIP) is most abundantly expressed in the heart. Its biological function remains elusive but it may be of high relevance for cardiac function as it has no paralogous homologue suggesting no functional redundancy.

To elucidate MLIP's function, we generated both hemizygous (MLIP+/‐) and homozygous (MLIP−/−) null mice. At 8 weeks of age, they both develop dilated cardiomyopathy characterized by enlarged hearts (heart to body weights MLIP+/+ 5.62mg/g vs MLIP+/− 10.73mg/g & MLIP −/− 11.03mg/g), functionally dilated (LVDD : +/+ 2.89mm; +/− 3.93mm; −/− 4.11mm) with reduced function (LVFS : +/+ 47%; +/− 31%; −/− 29%). Histological analysis shows no overt abnormalities of both MLIP+/− and MLIP−/− hearts. Preliminary data indicate a preserved cardiomyocyte size in MLIP+/− and MLIP−/− mice, suggesting that the increase in heart size might be due to hyperplasia rather than cellular hypertrophy. Moreover, cardiac gene expression of 12 week‐old MLIP+/− and MLIP−/− mice reveals an upregulation of genes involved in cardiac development and control of cell cycle. Cardiomyocytes proliferation of post‐natal MLIP+/− and MLIP−/− hearts and cardiac response to pro‐hypertrophic stimulus are currently under investigation.

In conclusion, MLIP is a newly discovered protein that impacts genes involved in cardiac development, growth and function.
A BRAF‐MEK complex reveals the molecular basis of oncogenic mutations
Authors: Hymowitz Sarah Gillmor , Sudhamsu Jawahar , Haling Jacob , Peck Ariana , Yen Ivana , Morales Tony , Brandhuber Barbara , Malek Shiva
Published: June 2021

The Ras/RAF/MEK/ERK pathway (MAPK pathway) controls intracellular signal transduction downstream of extracellular growth factors and is frequently activated in cancer. Using a combination of crystallographic, biochemical and cellular approaches we show that BRAF forms a stable complex with MEK prior to pathway activation. The BRAF‐MEK kinase domains interact in a tetrameric complex dependent on BRAF dimerization. Formation of this complex is mutually exclusive with either MEK‐KSR or MEK‐MEK interactions. Structural and biochemical evidence illustrate that the molecular determinants for B‐RAF enzymatic activity is distinct from those required for interaction with MEK. Our work also reveals the molecular basis for the paradoxical ability of both BRAF‐activating and –inactivating oncogenic mutations to stimulate the MAPK pathway. Similarly, this data rationalizes the diverse effects of BRAF inhibition by pathway priming of ATP‐competitive BRAF inhibitors.
A‐kinase anchoring protein (AKAP)‐Lbc coordinates protein kinase A (PKA) phosphorylation and inhibition of Src homology 2 domain‐containing phosphatase 2 (Shp2)
Authors: Burmeister Brian T. , Taglieri Domenico M. , Wang Li , Carnegie Graeme K.
Published: June 2021

Pathological cardiac hypertrophy (an increase in cardiac mass resulting from stress‐induced cardiac myocyte growth) is a major factor underlying heart failure. Shp2 is a tyrosine phosphatase that is critical for cardiac function. Mutations resulting in loss of Shp2 catalytic activity are associated with cardiac hypertrophy and congenital heart defects.

We have identified a novel mechanism of Shp2 inhibition that may promote cardiac hypertrophy. We demonstrate that Shp2 is a component of the A‐kinase anchoring protein (AKAP)‐Lbc complex. AKAP‐Lbc facilitates PKA phosphorylation of Shp2, which inhibits its protein tyrosine phosphatase activity.

Overall, our data indicate that AKAP‐Lbc integrates PKA and Shp2 signaling in the heart and that AKAP‐Lbc‐associated Shp2 activity is reduced in hypertrophic hearts in response to chronic β‐adrenergic stimulation and PKA activation. Thus, while induction of cardiac hypertrophy is a multifaceted process, inhibition of Shp2 activity through AKAP‐Lbc‐anchored PKA is a previously unrecognized mechanism that may promote compensatory cardiac hypertrophy. We are currently mapping the PKA phosphorylation site(s) on Shp2 and investigating downstream hypertrophic signaling mechanisms.
Multiple Solvent Crystal Structures of Rap1a GTPase
Authors: Donohue Patrick , Mattos Carla
Published: June 2021

Rap1a is a member of the Rap family of Ras‐like small G‐proteins. Rap1a is the closest homologue of the canonical onco‐protein Ras, sharing >;50% sequence identity. Like Ras, it cycles between an active GTP‐bound form and an inactive GDP‐bound form, allowing for associated signaling pathways to be switched on or off. Despite their structural and functional similarity, Ras and Rap1 are regulated by different sets of GTPase‐activating proteins and guanine nucleotide exchange factors and interact with a different group of effector molecules. Rap1 is also important in it's own right as a controller of cell adhesion and junction formation. Additionally, it has recently been indicated as playing a role in the metastasis of some cancers. Multiple Solvent Crystal Structures (MSCS) is a crystallographic method that utilizes organic solvents as functional group probes to protein binding surfaces. Our group has used MSCS to identify binding site hot spots in H‐Ras and we now aim to obtain the binding site hot spots on the surface of Rap1A. The goal is to do a detailed binding surface comparison between the two GTPases and identify their specificity determinants. MSCS also gives a picture of hydration effects on the structure and function of Rap1a, and this will be compared with the water analysis recently done for Ras in our group.
An Oxygen‐chelate Precious Metal‐based Complex, Palladium bis‐Acetylacetonate, Induces Apoptosis in Lung Cancer H460 cells via Endoplasmic Reticulum Stress Pathway rather than interacting with DNA
Authors: Wang Yi , Li Xiaokun , Zhou Huiping , Liang Guang
Published: June 2021

Although some of palladium (Pd) complexes possess promising anti‐tumor activities, they are all chelated with N‐containing ligands and function by interacting with DNA. Few studies on the bio‐activities and mechanism of Pd‐O chelates have been conducted. In this study, a Pd complex containing four O‐donor ligands, Pd(acac)2, has been synthesized and evaluated as an anticancer agent. Solution stability test through 1H‐NMR spectra reveals that Pd(acac)2 maintained chemical stability being in the S‐donor containing DMSO solution. MTT assay shows that Pd(acac)2 exhibited significant cytotoxicity against BGC 823, HL‐60, KB, LS‐174T, and Hela cells. Flow cytometric assay reveals that treatment with Pd(acac)2 significantly induced apoptosis in H460 cells. Interestingly, gel electrophoresis and ultraviolet absorption assay show that Pd(acac)2 did not act directly on DNA. Immunoblotting assay indicates that the anti‐tumor effects of Pd(acac)2 are associated with ER stress‐mediated apoptosis pathway, followed by caspase activation. CHOP knockdown by specific siRNA significantly attenuated Pd(acac)2‐induced cell apoptosis, indicating that the apoptotic pathway is partly ER stress‐dependent. In summary, these data may provide us a novel mechanistic strategy for the development of metal‐based anticancer drugs.

This work was supported by National Natural Science Funding of China (81102452).
Wagonin inhibits LPS‐induced expression of inflammatory cytokines by promoting mRNA degradation in macrophages
Authors: Wan Xiaoshan , Huang Yi , Liang Guang , Xiao Jian , Li Xiaokun , Gurley Emily C , Zhou Huiping
Published: June 2021

Wagonin is a major active component of Scutellaria baicalensis, a widely used Chinese herbal medicine for treatment of inflammatory diseases. It has been well‐documented that wagonin inhibits inflammatory cytokine gene expression. However, the underlying cellular/molecular mechanisms remain to be fully identified. Our previous studies demonstrated that LPS upregulates inflammatory cytokines by modulating RNA binding proteins and increasing mRNA stability. The aim of current study was to examine the effect of wagonin on LPS‐induced expression of inflammatory cytokines and further identify its underlying cellular/molecular mechanisms in macrophages.

Methods

Cultured mouse RAW264.7 macrophages and human THP‐1‐derived macrophages were used in this study. The mRNA and protein levels of IL‐1β, IL‐6, TNF‐α, MCP‐1 were detected by real time RT‐PCR and ELISA, respectively. The expression and intracellular translocation of RNA binding proteins, HuR and CUGBP‐1, were determined by Western blot and immunofluorescence staining.

Results

Wagonin significantly inhibited LPS‐induced expression of IL‐1β, IL‐6, TNF‐α, and MCP‐1 by promoting mRNA degradation through regulating intracellular translocation of HuR and CUGBP‐1 in macrophages.

Conclusion

Wagonin exerts its anti‐inflammatory effect through regulating intracellular translocation of HuR and CUGBP‐1 in macrophages.
CTP synthase, a smooth muscle‐sensitive therapeutic target for effective vascular repair after injury
Authors: Tang rui
Published: June 2021

Neointima formation due to smooth muscle cell (SMC) proliferation following injury is a major medical challenge in cardiovascular intervention. However, anti–neointimal drugs often block re‐endothelialization as a side effect. Identification of SMC sensitive therapeutic targets will potentially conquer this medical barrier. In the present study, we identified CTP synthase (CTPS) as one of these potential targets. Blockade of CTPS function by shRNA or its specific inhibitor suppressed SMC proliferation and neointima formation. In contrast, targeting CTPS treatments had no side effects on endothelial cell (EC) in vitro, and promoted reendothelialization in vivo. Mechanistically, blockade of CTPS pathway activated CTP synthesis salvage pathway in ECs but not in SMCs, which sustained EC proliferation. Meanwhile, blockade of CTPS function induced SMC phenotypic modulation into the contractile phenotype, which characterized by the expression of pro‐angiogenic factors while inhibiting anti‐angiogenic factors in SMCs, leading to enhanced EC proliferation. In summary, for the first time, our study discovered a fundamental difference between SMC and EC in their CTP biosynthesis during pathological proliferation which provided a novel strategy to selectively block SMC proliferation while not disturbing or even promoting reendothelialization for effective vascular repair after injury.
Phosphorylation of Tau‐4R and Microtubule Binding Region of MAP‐2C by PKA and GSK‐3β Inhibits their O‐Glycosylation by OGT
Authors: Khatra Balwant S , Juarez Melina , Johnson Daisy , DeTure Michael
Published: June 2021

Microtubule associated proteins (MAPs) play an important role in microtubule assembly. In an earlier study we showed that phosphorylation of microtubule binding region of MAP‐2 (MTBR; a peptide containing conserved repeat region of MAPs) by Glycogen Synthase Kinase‐3β (GSK‐3β) inhibits its ability to assemble microtubules (Teel et. el. The FASEB Journal 11, A1352, 1997). In the present study we studied the effects of phosphorylation of MAPs on their O‐glycosylation. Two isomers of β‐N‐acetylglucosaminyl transferase (OGT); short OGT (sOGT), and nucleoplasmic (ncOGT) were used. The ncOGT showed greater activity toward MAP‐2c, Tau‐4R, and MTBR. The MTBR and Tau‐4R were pre‐incubated with protein kinase A (PKA) or PKA followed by GSK‐3β, and then incubated with ncOGT and UDP‐[3H]N‐Acetylglucosamine. The incorporation of 3H into these pre‐phosphorylated proteins show that phosphorylation by PKA inhibits their O‐glycosylation by ~50%. Subsequent phosphorylation by GSK‐3β decreases their O‐glycosylation further by ~90%. We conclude that phosphorylation and O‐glycosylation may be competing for the same sites on MTBR and Tau‐4R and thus may play an important role in the regulation of microtubule assembly. (Supported by NIH‐RISE Grant #07298409)
Role of DLC1 in Regulating Cellular and Focal Adhesion Dynamics
Authors: Kaushik Shelly , Low Boon Chuan
Published: June 2021

Deleted in Liver Cancer‐1 (DLC1) is a RhoGTPase‐activating protein that is down‐regulated in a number of cancers. It specifically acts on RhoA and to some extent Cdc42, but has no discernible effect on Rac1 activity. DLC1 has been shown to localize to the focal adhesions and its absence leads to enhanced cell migration. However, its role at the focal adhesions and its subsequent effect on cell migration is still unclear. Using immunofluorescence techniques, Total Internal Reflection Fluorescence Microscopy (TIRFM) and quantitative image analysis we examined the effect of DLC1 on cell migration and cell spreading and determined its localization and dynamics at focal complexes, focal adhesions and fibrillar adhesions to understand how these cellular dynamics are affected. We found that DLC1 inhibits cell spreading in a RhoA‐dependent, substrate‐independent manner whereas it inhibits cell migration in a RhoA‐independent manner. DLC1 also localizes to all three adhesion types, with a specific spatial distribution in the focal adhesions during early and late stages of cell spreading when compared to focal adhesion proteins such as paxillin and Focal Adhesion Kinase (FAK). In summary, DLC1 regulates cellular dynamics by its down‐regulation of RhoA as well as its effects on focal adhesion dynamics.

Supported by Mechanobiology Institute and Ministry of Education, Singapore (Academic Research Fund T208A3121)
Cortical femur response to synbiotics in aging mice
Authors: Blanton Cynthia , Tyler Morgan , Gabaldon Annette
Published: June 2021

Age‐related dysbiosis and bone loss might be causally related through mechanisms involving micronutrient absorption and immune function. Previous work in our laboratory showed that skeletal structure is protected by dietary synbiotics in rats exposed to simulated weightlessness. This project investigated whether these prior findings extend to a mouse model of aging. Three groups of male mice were studied: 10‐mo‐old, fed a synbiotic diet for 4 months (SYN); 10‐mo‐old, fed a control diet for 4 months (CON); 10‐mo‐old, sacrificed at time zero (BSL). The synbiotic diet contained fructooligosaccharides (31.5 g/kg diet) and Lactobacillis acidophilus + Lactococcus lactis lactis (1012 CFUs/kg diet). Femurs were sliced in 5 μm cross‐sections at the proximal diaphysis and analyzed using digital imaging. Results showed a significant decrease in cortical wall thickness for the SYN vs. BSL mice (P<0.01). Total cross‐sectional area (CSA) increased in both SYN and CON vs. BSL mice (P<0.001), primarily due to enlargement of medullary CSA (SYN and CON vs. BSL, P<0.05), with SYN showing a greater area vs. CON mice (P<0.01). The ratio of medullary:total CSA was higher and cortical:total CSA lower in SYN vs. CON and BSL mice (P<0.01). These data suggest that synbiotics exert no benefit on cortical femur in aging mice.
Iron deficiency and anemia are not associated with food insecurity in pregnant women in the United States: NHANES 1999–2008
Authors: Park Clara , Eicher‐Miller Heather A
Published: June 2021

Food insecurity (FI) has been associated with low iron (Fe) intake in US children, adolescents, and non‐pregnant adults. Pregnant women may be at higher risk of FI and Fe deficiency (ID) as nutritional needs increase throughout pregnancy and more resources are needed to secure food. Maternal ID and anemia are associated with low birth weight and life‐long cognitive impairment in the child. The purpose of this study was to investigate the relationship of ID and FI among pregnant women ages 13 to 54 y (n=1,023) using data from NHANES 1999–2008. Food security was classified using the US Food Security Survey Module. ID was indicated using total body Fe, soluble transferrin receptor, or ferritin, and anemia by hemoglobin or hematocrit. The odds of ID, indicated by ferritin status, were 2.9 times higher for FI compared with food secure (FS) pregnant women. However, after excluding for inflammation, indicated by C‐reactive protein, the odds of ID were not significantly greater for FI compared with FS pregnant women. Models created for other indicators of ID and anemia were not significant. In conclusion, evidence for a significant association of food insecurity with iron deficiency or anemia in pregnant US women is insufficient, yet efforts to reduce the prevalence of ID and anemia should continue among both FS and FI pregnant women.

Grant Funding Source: Purdue University Department of Nutrition Science
Iron and diabetes risk among Filipino women
Authors: Feranil Alan , Lee Nanette , Bas Isabelita , Largado Fe , Adair Linda
Published: June 2021

Some studies suggest that increased iron intake may lead to diabetes risk possibly thru oxidative stress, inflammation, free fatty acid oxidation, insulin extraction and secretion. But, this has not yet been established in less developed countries where iron deficiency is likely to occur. We examined the association of iron intake with impaired fasting glucose/diabetes mellitus (IFG/DM: FG>;=110 mg/dL) using data from 1,871 women (ages 35–68 yr) from the Cebu Longitudinal Health and Nutrition Survey in 2005. About 9% of women had IFG/DM. On average, women consumed 11.25±7.04 mg of iron daily. Logistic regression analyses adjusted for current BMI, age, total energy intake and other confounders showed that the odds of having IFG/DM increased with iron intake. Compared to the lowest quartile of iron intake, those in the highest quartile were 78% more likely to have IFG/DM [OR(95% CI): 1.78 (1.01–3.12)]. Results of this study suggest the need to further examine the link between iron intake and diabetes risk in less developed countries like the Philippines where iron deficiency anemia is still prevalent but diabetes has also become one of the major causes of mortality and morbidity. R01TW008288
Using principal component analysis to characterize dietary patterns in children from Uruguay
Authors: Lee Yujin , Hsiao Pao Ying , Queirolo Elena , Peregalli Fabiana , Kordas Katarzyna
Published: June 2021

Few studies have investigated dietary patterns of people in Uruguay, especially children. The purpose of this study was to identify dietary patterns of 217 children aged 6–8 y in Montevideo, Uruguay (Mean±SD BMI=21.2± 4.5 kg/m2). Diet data were collected via 2 non‐consecutive 24‐hour recalls completed by mothers. Principal component analysis with Varimax rotation was used to identify dietary patterns. Three patterns were identified (explaining 29.1% of variance): 1) ‘Less healthy’ characterized by high consumption of sweetened beverages, sweets, fats and oils, breads, and processed meats, 2) ‘Refined grains’ characterized by high consumption of pizza and dinner pies, pastries, pasta, milk and fruits, 3) ‘More healthy’ characterized by dark green, red and orange vegetables, beans and peas, eggs and white meats. For each component, each child was allocated a score derived from the component's coefficients for food groups and the reported amount of consumption. Spearman correlation coefficients were used to test correlations between total calorie consumption and component scores. Total calorie consumption was correlated with ‘Less healthy’ and ‘Refined grains’ patterns (r= 0.56, r=0.39, respectively). Further studies need to investigate possible associations between dietary patterns and biomarkers of nutritional status in this population.

Grant Funding Source: NIEHS 4231251X1
Inclusion of peers in a school‐based obesity intervention
Authors: Johnston Craig Allen , Moreno Jennette , El‐Mubasher Abeer , Papaioannou Maria A , Woehler Deborah
Published: June 2021

The increasing prevalence of childhood obesity and the comorbid health problems highlight a pressing need to identify effective treatments that address this public health problem during the childhood years. The current study evaluated a school‐based pediatric obesity program for middle‐school children. A total of 163 adolescents between the ages of 11 and 12 were recruited, with 74 adolescents being at or above the 85th percentile for body mass index (BMI). Students were randomized to a peer‐mentor intervention (PM, n=38) or treatment as usual (TAU, n=36) condition. The PM condition received nutrition education that included lifestyle change techniques once a week and participated in physical activity the other 4 days of the week. The PM condition was led by peer mentors and program staff. The TAU condition received a typical health and physical education class. Twelve month measurements have been collected on 55 students (PM, n=26, TAU, n=29). Repeated Measures ANOVA was used to evaluate weight changes between the groups using standardized BMIs at baseline, 3, and 6 months. Adolescents in the PM condition significantly reduced their zBMI when compared to the adolescents in the TAU condition (F=4.3, p<.05). This study suggests that an integrated health program supported by peers is a promising approach for school‐based pediatric obesity prevention. This project was funded by a grant from the Oliver Foundation.