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Various vaccine quality attributes should be monitored to ensure consistency, potency, purity, and safety of vaccine products prior to lot release. Vaccine particle size and protein antigen aggregation are two important considerations for particle-adsorbed vaccines. In this study, we evaluated the use of imaging flow cytometry as a potential all-in-one platform to measure adjuvant particle size and to detect protein aggregates through a combination of brightfield microscopy, side scatter detection, and fluorescence microscopy. An aluminum phosphate adjuvant was analyzed for size using the brightfield function, and the size measurement was compared against laser diffraction. Heat-induced protein aggregates of either unadsorbed antigens or aluminum phosphate adjuvant-adsorbed antigens were stained with the fluorescent ProteoStat aggregation dye, followed by detection and analysis using a combination of the brightfield and fluorescence microscopy functions. The change in aggregation of unadsorbed antigens was confirmed using dynamic light scattering. These results demonstrate the versatility of the imaging flow cytometry platform for the evaluation of multiple vaccine quality characteristics.

Scaling network packet processing performance to meet the in- creasing speed of network ports requires software programs to carefully leverage the network devices’ hardware features. This is a complex task for network programmers, who need to learn and deal with the heterogeneity of device architectures, and re-think their software to leverage them. In this paper we make first steps to reverse this design process, enabling the automatic generation of tailored hardware designs starting from a network packet processing program. We introduce eHDL, a high-level synthesis tool that automatically generates hardware pipelines from unmodified Linux’s eBPF/XDP programs. eHDL is designed to enable software developers to directly define and implement the hardware functions they need in the NIC. We prototype eHDL targeting a Xilinx Alveo U50 FPGA NIC, and evaluate it with a set of 5 eBPF/XDP programs. Our results show that the generated pipelines are efficient in terms of required hardware resources, using only 6.5%-13.3% of the FPGA, and always achieve the line rate forwarding throughput with about 1 microsecond of per-packet forwarding latency. Compared to other network-specific high-level synthesis tool, eHDL enables software programmers with no hardware expertise to describe stateful functions that operate on the entire packet data. Compared to alternative processor-based solutions that perform eBFP/XDP offloading to a NIC, eHDL provides 10-100x higher throughput.

SummaryAlnus glutinosa response to Frankia alni is driven by several sequential physiological modifications that include calcium spiking, root hair deformation, penetration, induction of primordium, formation and growth of nodule. Here, we have conducted a transcriptomic study to analyse plant responses to Frankia alni at early stages of symbiosis establishment.Forty-two genes were significantly activated by either with a Frankia culture supernatant or with living cells separated from the roots by a dialysis membrane permitted to identify plant genes which expression changes upon early contact with Frankia. Most of these genes encode biological processes, including oxidative stress and response to stimuli. The most upregulated gene is the non-specific lipid transfer protein (nsLTP) encoding gene with a fold change of 141. Physiological experiments showed that nsLTP increases Frankia nitrogen fixation at sub-lethal concentration. Immunohistochemistry experiments conducted at an early infection stage indicated that nsLTP protein is localized at the deformed root hair region after Frankia inoculation and later in nodules, precisely around bacterial vesicles. Taken together, these results suggest that nsLTP acts at early and late stages of symbiosis, probably by increasing nitrogen uptake by Frankia.

Purpose: To investigate the presence of amblyopia risk factors in newborns with congenital nasolacrimal duct obstruction (CNLDO) and age-matched healthy control subjects. Methods: This retrospective case-control study involved newborns aged 30 to 60 days with CNLDO and age-matched healthy control subjects. Amblyopia risk factors were identified in accordance with the American Association for Pediatric Ophthalmology and Strabismus Vision Screening Committee recommendations. The prevalence of amblyopia risk factors was compared in newborns with CNLDO and age-matched healthy control subjects, newborns with unilateral and bilateral CNLDO, and the affected eye and fellow eye of newborns with unilateral CNLDO. Results: Amblyopia risk factors were found in 18 patients (11.9%) with CNLDO and 19 control subjects (8.7%) ( P = .314). Eyes with CNLDO showed a significantly lower spherical equivalent compared to control eyes (2.01 ± 1.21 vs 2.79 ± 1.14 diopters, P < .001). No difference in amblyopia risk factors was found in eyes with unilateral and bilateral CNLDO (11.5% vs 12.1%; P = .908) or in eyes with unilateral CNLDO and fellow eyes (9.8% vs 12.3%; P = .540). Conclusions: CNLDO does not seem to be associated with amblyopia risk factors in newborns. Because anisometropia might develop later on, all patients with CNLDO should be monitored for amblyopia. [ J Pediatr Ophthalmol Strabismus . 2020;57(1):39–43.]

Currently, processed meats appear increasingly as a nonhealthy food because of their content of fat, salt, nitrite, and particularly in red meat, for the heme-iron concerning oxidant effect and radicals formation in human gut. Polygonum cuspidatum and rosemary extract has been tested to counteract these effects and experiments were carried out to add polyphenols into whole meat cuts with the aims to improve their healthiness. The addition of these extracts can reduce the oxidation-reduction potential of products, increasing the antioxidant power. Nevertheless, a low percentage of polyphenols were found in the products because of the process effects and an interaction with the ascorbate residue was observed. In addition, some drawbacks were a decrease in of yield and a worsening of some sensorial properties when large amount of extract was added. The balance between the improvement of the wholesomeness content and the acceptability of products is the challenge the current research have to overcome.

Background: Ayurveda, the traditional Indian therapeutic system, involves herbs and spices as drug ingredients and a kind of food intake regulation. The health and curative aspects of plant products are often ascribed to their antimicrobial and antioxidant activities (AA). However, it seems somewhat possible to correlate the AA of herbal extracts or plant isolates with their phenolic contents (PC). Objective: Indian researchers have carried out some tests for the determination of AA and PC of plant products that are worth mentioning. Methods: Among the herbal analysis techniques, the important contributions of Indians and people of Indian origin in this field include the remarkable separation of phenolic compounds, including the use of reversed-phase columns, on-line HPLC, the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay to effectively screen and identify antioxidant compounds from herbal extracts, and other chromatographic, spectrographic, and spectroscopic techniques, in the analysis of complex biological matrices. Results: The aim of this paper is to present an overview of such arguments with reference to herbal drugs and food intake regulation for specific ailments. Conclusions: The lack of uniformity in the use of test protocols for antioxidant assays (particularly using the DPPH free radical) and differences in in vitro mechanisms of antiradical activity and in vivo metabolism of polyphenols have been the issues associated with these experiments that have been raised by researchers in the period from 2000 onward.

P4 is a domain-specific programming language for expressing how packets are processed by network devices. P4 is used to program P4-enabled devices with a custom forwarding pipeline. SRv6 is a network architecture that encodes a list of instructions in the IPv6 packet header to define a network-wide packet processing program. Each instruction defines a node to process the packet and the behavior to be applied to that packet by that node. Recently, the SRv6 architecture has been extended to support a set of new instructions, known as uSID instructions, that provide a better scalability and MTU efficiency. In this demo paper, we provide a P4 forwarding pipeline that supports the SRv6 uSID instructions. Our implementation leverages the P4 BMV2 behavioral model and extends the ONOS controller to support the new SRv6 behaviors. Finally, we show two scenarios where we use our implementation to provide a fully programmable network fabric.

The Segment Routing (SR) architecture is based on loose source routing. A list of instructions, called segments can be added to the packet headers, to influence the forwarding and the processing of the packets in an SR enabled network. In SRv6 (Segment Routing over IPv6 data plane) the segments are represented with IPv6 addresses, which are 16 bytes long. There are some SRv6 service scenarios that may require to carry a large number of segments in the IPv6 packet headers. This can be inefficient due to the overheads of the segment list, or can even cause processing issues in the crossed nodes due to the very large header to be processed. In this paper we present the Micro SID solution for the efficient representation of segment identifiers. With this solution, the length of the segment list can be drastically reduced., Comment: Extended version (v1) of submitted paper - October 2020

In-network computation based on programmable data plane hardware provides a tremendous opportunity to improve throughput, latency and reduce congestion in data center scenarios. However, a judicious use of these network devices must be done based on their limited resources and the specific features of the application to be offloaded. This paper promotes FlowBlaze, a stateful hardware programmable data plane, as a candidate for offloading online MapReduce tasks. Above all, tasks with strict time requirements can benefit from in-network computing since it can significantly lower their latency. Given that MapReduce is a generic programming paradigm, in this paper we first try to identify which subset of MapReduce operations can be transparently offloaded to a specific hardware architecture and which are the limitations of this offloading in terms of memory and computational resources. After, we show how the FlowBlaze architecture can match the partition/aggregation paradigm and we discuss a set of primitives exposed by the FlowBlaze abstraction to perform mapping and aggregation. Finally, we prove the feasibility of this approach applying it to a click-stream analysis use case.

Protein concentration is an important attribute in the production of subunit or component-based vaccine antigens. Rigorous monitoring of protein concentration is required to identify potential areas for yield improvement. The current GMP method for quantitation is the plate-based ELISA which requires numerous hands-on steps and has low sensitivity in comparison to new microfluidic systems. To address this issue, a sensitive automated microCapillary Electrophoresis ImmunoAssay (mCE IA) method was developed to accurately separate and quantitate pertactin (PRN), an important antigen of the modern acellular Pertussis (aP) vaccine. PRN is reported to be a low-yielding antigen; thus, it is critical to observe its concentration throughout its manufacturing process. First, a primary antibody for PRN was identified to establish suitable immunoprobing conditions for detection of PRN over a wide linear dynamic range that spans 3 orders of magnitude. Next, the pre-adsorbed PRN Drug Substance (DS) was used as a reference standard to quantitate PRN samples against a calibration curve with adequate accuracy and precision. Four representative samples including three in-process steps and final adjuvanted drug product: Quadracel®, were examined to demonstrate the capability of mCE IA to quantitate PRN with high sensitivity and specificity. The matrices of the selected samples contain additional components (e.g. other proteins, growth factors, cell culture media, residual ammonium sulfate, and aluminum adjuvant) often making the quantitation of PRN challenging. The specificity and method linearity were demonstrated by spiking pre-adsorbed PRN DS into the four representative samples. In addition, it was shown that reportable concentrations of PRN for nine downstream process steps as analyzed by our method is comparable to concentrations obtained with ELISA. Most importantly, this study demonstrated that our method's quantitative accuracy is independent of matrix components, as each sample undergoes extensive dilution. This allows for seamless end-to-end analysis of PRN from fermenter harvest, through to complex downstream process samples to adjuvanted drug products. Finally, for the first time the developed and qualified mCE IA method was shown to quantify PRN throughout the entire manufacturing process to provide rapid feedback for process optimizations allowing for accurate yield and step-loss calculations.

One of the most important challenges in the modern world of food and beverage is the persistence of world hunger. Also, the surplus production of food in richer countries has determined important environmental problems. The problem should not be only the identification of the best food for human beings, but also the possibility of supplying this food in adequate amounts, with good nutritional intakes and in a sustainable way. In the ambit of crops, cereals are always the first choice. However, other vegetables can be considered with favour, including grain legumes. The use of domesticated legumes and pulses in human history is well documented worldwide. One of the most interesting grain legumes (because of its intrinsic resistance to drastic environmental conditions) is Lathyrus sativus (grass pea). Similarly to other grain legumes, it is historically reported to be used for the production of soups, unleavened breads, and other meals. On the other hand, grass pea may be questioned with concern to human and animal nutrition because of a peculiar neurotoxin associated with neurolathyrism disease. This chapter concerns chemical and biochemical properties of grass pea, with nutritional evaluations.

The production of certain recipes based on both cereals and grain legumes may be supposed to be a good deal when speaking of authenticity and traceability issues. Apparently, grain legumes—and specifically Lathyrus sativus—may be included in these discussions when speaking of economically motivated adulteration. Actually, safety and health concerns related to grass pea should be good arguments against food frauds linked with the commerce of L. sativus. However, the Italian perspective suggests some counterargument. Several cultivars are well known and considered ‘Traditional Agricultural Food Products’ in Italy, meaning that the historical tradition should be correlated with the geographical importance. Anyway, two different viewpoints have to be considered when speaking of authenticity and traceability issues in connection with grass pea: first of all, the birth or growth of ‘market niches’ with a strong interest in grass pea, in countries with high income; and secondly, the possible use of this grain legume, with the intention of adulterating other flours, in some Asian country above all. For these reasons, grass pea can have some important role when speaking of food frauds.

This Chapter concerns the use of grass pea (Lathyrus sativus) in human and animal nutrition by the viewpoint of safety and health. Domesticated grass pea is documented and historically demonstrated. This grain legume can be found worldwide. Substantially, Lathyrus genus—and grass pea in particular—seems to be a constant presence in the human history in association with cultivated cereals and other legumes. From the nutritional viewpoint, L. sativus is reported to have a demonstrated deficiency of vitamins A, B1, C, D, and E, some amino acids and elements. However, these deficiencies are not important worries. Health and safety problems are linked to the β-ODAP neurotoxin, responsible for neurolathyrism, and other antinutritional factors (condensed tannins and total phenolics; some enzymatic inhibitory activities). Actually, ‘neurolathyrism is associated with osteolathyrism (under the common ‘lathyrism’ name). Anyway, the main problem is the presence of β-ODAP neurotoxin, and a safe use of grass pea cannot be assured in these conditions. This Chapter also explains common culinary methods for the safe consumption of grass pea-based foods by the chemical angle.

This Chapter concerns the traditional use of a peculiar crop legume, grass pea (Lathyrus sativus) as a food product or ingredient for typical food products. At present, this legume is known worldwide and widely used in relation to human nutrition. In fact, grass pea has very interesting nutritional features, including the abundance in proteins, and peculiar organoleptic properties. On the other hand, grass pea may be questioned with concern to human and animal nutrition because of a peculiar neurotoxin associated with neurolathyrism and occurs if grass pea-based foods are consumed in notable quantities. However, these food products are an interesting legacy for many geographical and ethnic cultures. At present, many food varieties are reported to be correlated with grass pea worldwide. An overview of different food versions ascribed to L. sativus in the world is offered here in relation to continental areas and taking into mind the existence of a peculiar culinary pattern in Ethiopia (soups/sauces/processed seeds/unleavened breads) which can be partially found in other non-African nations.

The use of grain legumes in various European regions has been observed with peculiar interest in the Middle Age, although the presence of seeds such as Lathyrus sativus (grass pea) is demonstrated since the Neolithic Era. The cereal/legume association has been reported in Andorra, Italy, and France at least from the late Roman Empire to the late Middle Age. Cereals and other plants such as Pisum sativum, Vicia faba, and L. sativus were often present and probably cultivated in good amounts. Interestingly, the same localisation of known recipes based on L. sativus in certain areas found in Morocco, Spain, France, and Italy (as an ideal line from the Maghreb area to Central Europe) can be seen as the heritage of past memories. Anyway, grass pea is a known and cultivated grain legume in Europe and specifically in Italy. The presence and persistence of grass pea is demonstrated also because of the current existence of certain recipes in some areas with strong medieval traditions. This chapter concerns regional culinary traditions associated with grass pea in Italy, with some consideration concerning historical reasons for the localisation of polenta-like dishes in some circumscribed areas only.

Offloading to the edge a subset of the cloud services requested by users is a very appealing solution to reduce the bandwidth pressure, minimize latency and improve the overall quality of experience of mobile users. Achieving this goal is technically challenging, since the network functionalities needed to manage this offloading are not trivial, and present significant requirements in terms of scalability and speed. We propose to realize these network functionalities directly in the dataplane, exploiting the characteristic of FlowBlaze, a novel stateful programmable dataplane. Furthermore, we show how the network functions for the traffic offload can be easily expressed using an ad-hoc domain specific language developed for the description of per-flow stateful network functions.

Urban security (or public safety), rather than a “social problem” tackled neutrally, is a issue of political contestation, owing to its threefold gist as right to not be victims of crime, policy goal and social demand. This article, highlighting how planning research has neglected to engage with contemporary paradoxes of security, makes the case for a critical approach to crime prevention and explores the embeddedness of urban security in planning practice in the Lisbon Metropolitan Area. We debate the relations of urban security with changing planning paradigms and political approaches around the vertical (multilevel/multi-scale) and horizontal distribution of planning practices., The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: Funded by Fundação para a Ciência e Tecnologia, grants SFRH/ BPD/86394/2012 and UID/SOC/50013/2013.

Programmable data planes recently emerged as a prominent innovation in Software Defined Networking (SDN), by permitting support of stateful flow processing functions over hardware network switches specifically designed for network processing. Unlike early SDN solutions such as OpenFlow, modern stateful data planes permit to keep (and dynamically update) local per-flow states inside network switches, thus dramatically improving reactiveness of network applications to state changes. Still, also in stateful data planes, the control and update of non-local states is assumed to be completely delegated to a centralized controller and thus accessed only at the price of extra delay. Our LOADER proposal aims at contrasting the apparent dichotomy between local states and global states. We do so by introducing a new possibility: permit to take localized (in-switch) decisions not only on local states but also on replicated global states, thus providing support for network-wide applications without incurring the drawbacks of classical approaches. To this purpose, i) we provide high-level programming abstractions devised to define the states and the update logic of a generic network-wide application, and ii) we detail the underlying low level state management and replication mechanisms. We then show LOADER's independence of the stateful data plane technology employed, by implementing it over two distinct stateful data planes (P4 switches and OPP - Open Packet Processor - switches), and by experimentally validating both implementations in an emulated testbed using a simple distributed Deny-of-Service (DoS) detection application.

XTRA (XFSM for Transport) aims at providing a first attempt towards a “ code-once-port-everywhere ” platform-agnostic programming abstraction tailored to the deployment of transport layer functions. XTRA’s programming abstraction not only fits SW platforms, but is specifically designed to harness, with no re-coding effort, the offloading opportunities offered by CPU-less HW boards or smart NICs. We demonstrate the viability of XTRA with three completely different implementations of the underlying execution engine (HW proof-of-concept on a NetFPGA board, User-space SW over Linux’ Open Data Plane, and NS3 emulator). Flexibility is shown via a number of example applications, ranging from a variety of congestion control algorithms, to a middlebox-type TCP proxy functionality, up to a customized “Timer-Based” (TB) TCP which leverages the native reliance of XTRA on timers, so as to produce a loss recovery operation which, despite being formalized only via a handful of code lines, performs almost comparable with the highly optimized Linux and FreeBSD implementations.

Tyrosyl-DNA phosphodiesterase 2 (TDP2) repairs abortive topoisomerase II cleavage complexes. Here, we identify a novel short isoform of TDP2 (TDP2S) expressed from an alternative transcription start site. TDP2S contains a mitochondrial targeting sequence, contributing to its enrichment in the mitochondria and cytosol, while full-length TDP2 contains a nuclear localization signal and the ubiquitin-associated domain in the N-terminus. Our study reveals that both TDP2 isoforms are present and active in the mitochondria. Comparison of isogenic wild-type (WT) and TDP2 knockout (TDP2-/-/-) DT40 cells shows that TDP2-/-/- cells are hypersensitive to mitochondrial-targeted doxorubicin (mtDox), and that complementing TDP2-/-/- cells with human TDP2 restores resistance to mtDox. Furthermore, mtDox selectively depletes mitochondrial DNA in TDP2-/-/- cells. Using CRISPR-engineered human cells expressing only the TDP2S isoform, we show that TDP2S also protects human cells against mtDox. Finally, lack of TDP2 in the mitochondria reduces the mitochondria transcription levels in two different human cell lines. In addition to identifying a novel TDP2S isoform, our report demonstrates the presence and importance of both TDP2 isoforms in the mitochondria.

The paper complements Abu-Orf's theory about violent settings by setting out a theory of fear in urban planning in ordinary urban contexts around three arguments: spatialization of fear; (modernist) spatialities and the encounter and political economies of urban fear. The three theoretical arguments are used to re-frame the planning history of Chelas, an affordable housing district in Lisbon, Portugal, and debate the way fear shapes, and is shaped in turn by, planning practice. Confirming that (growing) fear in ordinary urban contexts is not just an effect of the contemporary organization of cities, the paper argues for a theorization of fear that combines global (hegemonic) and a local (discursive/contingent) perspectives in the theorization of urban fear, and advocates for the need to put fear, and its capacity to create a crisis in urban policy, at the heart of planners' agendas.

Stateful programmable dataplanes are emerging as a disruptive technology and an enabler factor for network function virtualization in SDN environments. In this demo paper we show three real world use cases with stateful forwarding requirements to validate both the hardware and software implementations of the Open Packet Processor data plane.

The mitochondria within human cells play a major role in a variety of critical processes involved in cell survival and death. An understanding of mitochondrial involvement in various human diseases has generated an appreciable amount of interest in exploring this organelle as a potential drug target. As a result, a number of strategies to probe and combat mitochondria-associated diseases have emerged. Access to mitochondria-specific delivery vectors has allowed the study of biological processes within this intracellular compartment with a heightened level of specificity. In this review, we summarize the features of existing delivery vectors developed for targeting probes and therapeutics to this highly impermeable organelle. We also discuss the major applications of mitochondrial targeting of bioactive molecules, which include the detection and treatment of oxidative damage, combating bacterial infections, and the development of new therapeutic approaches for cancer. Future directions include the assessment of the therapeutic benefit achieved by mitochondrial targeting for treatment of disease in vivo. In addition, the availability of mitochondria-specific chemical probes will allow the elucidation of the details of biological processes that occur within this cellular compartment.

Soprattutto nell'ultimo decennio, i sistemi di videosorveglianza sono divenuti una presenza costante nello spazio pubblico urbano. Molto è stato detto a riguardo di questi processi e delle loro relazioni con le pressioni globali per la sicurezza: manca, invece, un dibattito sul ruolo dell'urbanistica e delle micro-politiche urbane. In questo articolo, l'autore muove dalla lettura foucaultiana del panopticon di Jeremy Bentham per discutere la consistenza dei sistemi di sorveglianza in due città del sud Europa, Lisbona e Palermo, e lo spazio della pianificazione urbana nei processi in corso.

Spatial planning and environmental restoration are essential corollaries to the management of protected natural areas; however, without a sound awareness of the evolutionary consistency of biocoenoses, the harmonious integration between human activities and ecosystem preservation remains an unattainable utopia. The theorisation of a balanced welfare, inspired by the universal tendency of ecosystems to reach a steady state, has to go along with the defection from any economic greed. © Springer International Publishing Switzerland 2015.

Something has been happening since the early 1990s: it is not necessary to be a scholar to have noticed that, in cities, in the media or in public and political discourses—in contemporary Western societies, in short—the use of the terms safety and security is becoming more prevalent everyday. Western citizens are more concerned with security than members of most other societies, both present and past. This chapter questions why citizens feel unsafe and constantly under threat despite living in the safest societies ever. Some recent trends in the global and Western arena are debated with the aim of unpacking the connections between security as a social demand and a political goal, and the ways these may have been boosting public fear of crime and violence. Specifically the role of political and media discourses (and misinformation) for growing perceptions of insecurity is unravelled—and two examples of discourses that have been making use of fear to influence urban and social policies are presented (Charles Murray’s ‘underclass’ theory and the discourse on the decline of the American city). Finally, the chapter discusses the cases of Italy and Portugal, describing the processes that have recently brought about a restructuring of the security policymaking in these countries, and suggests a reconsideration of Western trends from a Southern European perspective.

This chapter focuses on concrete urban space with the aim of exploring the spatialities interlinked with feelings and discourses of fear. An impressive amount of scholarly work has recently depicted processes of fortification, privatisation, polarisation, exclusion, segregation and control. Several attempts have been made to produce comprehensive theoretical understandings of such processes: ‘geographies of fear’, ‘military urbanism’, ‘end of public space’, ‘integral urbanism’, ‘divided cities’, to name some. This chapter sets out a fourfold taxonomy of the spatial processes connected with feelings and discourses of fear, with the aim of organising the knowledge available in literature from the perspective of macro-scale effects over urban territories. Each category is characterised by a specific spatial effect of urban restructuring: Enclosure, spaces of exclusion/seclusion; Barrier, infrastructural nets, with their longitudinal ‘splintering’ effect; Post-Public Space, privatisation and fortification of public space(s) and buildings; Control, the politics of surveillance over urban space. The theoretical discussion of each category is followed by the exemplification of a case from Palermo and/or Lisbon. The conclusions of the chapter, building on findings from Southern Europe, suggest a reframing of mainstream theories and advocate for a conceptual approach to the spatialisation of urban fear more attuned to local characterisations.

Urban fear is imbued with rational and irrational dimensions, some interlinked with, others independent from, urban life or the discourses of fear analysed in the previous chapter. This chapter, acknowledging that fear is not a simple ‘effect’ of specific ‘causes’, deepens the understanding of fear: it looks at the way fear is generated at the intersection of urban space and otherness, to build a theoretical framework supported by the work done by David Sibley on the geographies of exclusion and Iris Marion Young on the politics of difference. Diversity in urban (and especially public) space is debated, exploring encounters within contemporary, multicultural cities, with the aim of unpacking the role of feelings in the creation of relations between identity and otherness. Then, the chapter debates that the way the creation of such relations is necessary for the self-representation of societies and how the politics of exclusion are embedded in the construction of dichotomies such as ‘we’ versus ‘the others’ and the misrepresentation of (minority) groups. In conclusion, the chapter links such themes with urban studies and policy: it critiques Jane Jacobs’ theories on urbanism and exemplifies the processes of stigmatisation/removal through the case of ‘nomad camps’ in Italy.

This chapter critically questions how urban planning as a practice and a discipline, and as a political and technical praxis, is informed by, and has in turn been shaping, urban fear. This is done through two arguments stemming from the multiplex way in which the dimensions of (geo)politics, otherness and (urban) space interrelate: (modernist) spatialities and the encounter, at the intersection of space and otherness; and the political economies of urban fear, at the intersection of otherness and (geo)politics. These arguments are used to explore the role of mainstream paradigms of planning practice: the modernist/rationalist paradigm and the variety of paradigms emerging in recent times from post-Fordist/neoliberal restructuring. Rather than outlining a conclusive and universal definition of the way fear operates, the chapter builds an exploratory theoretical framework, based on the critical evaluation of the discussions presented in the previous chapters, which can be used to unravel the relationship between fear and planning in specific cases at the intersection of global trends and local contextual characterisations. To do this, the chapter reviews existing critiques and then uses them to reconsider the histories of two council housing districts in Palermo (the Zen) and Lisbon (Chelas)—i.e. it sets out, and then tests, a theoretical/exploratory framework.

This book has two objectives: to set out a comprehensive, critical and exploratory theory of fear, space and urban planning, while unravelling the contradictions and paradoxes of their mutual relations; and to enrich recent studies about urban geopolitics and the geopolitics of fear, taking the research done from the point of view of global cities and looking at it from the perspective of ordinary cities. We shall thus use the term ‘fearscapes’, or landscapes of fear, as a linguistic trick with the aim of taking a critical approach to the spatial transformations directly/indirectly connected with, or produced by, discourses and feelings of fear. In short, the book debates whether, and to what extent, the production of landscapes of fear constitutes an (emergent) urban political economy. This chapter sets out the objectives, conceptual background and empirical context of the book. The introduction outlines the object of study, research questions and structure of the book. This will be followed by the summary of some theories about recent socio-spatial urban transformation, before focusing on the transformation in the institutional practice of urban planning, and more especially on the changing patterns of consensus building. The concept of misinformation is introduced as the main instrument for the inquiry of relations between discourses of fear and planning policymaking. In conclusion, the reasons for the election of Southern Europe as a field of study are presented, together with some notes about methodology and the empirical objects of study (the cities of Lisbon and Palermo).

While our understanding of the folding and structure of water-soluble proteins has progressed to the point where they can be artificially designed and produced from first principles, there has been only limited work toward the de novo design of membrane proteins. Such studies have been hindered in large part due to the practical challenges in the production and characterization of multispanning transmembrane (TM) proteins that arise from their highly hydrophobic character. In this work, we used molecular biology cloning techniques to produce a library of partially randomized Ala- and Ile-rich de novo helix-loop-helix (hairpin) TM constructs as models for tertiary TM-TM folding. From this plasmid DNA library, we selected sequences corresponding to hairpins with 0, 1, or 2 putative TM segments. While purification protocols could be adapted for application with a broad range of designed protein hairpins, bacterial expression of constructs with multiple predicted TM segments was limited as it is with native membrane proteins. Examples of the peptide hairpins obtained were characterized by circular dichroism spectroscopy, tryptophan fluorescence, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). We found that hairpins composed of two TM segments display characteristic behavior on detergent solubilization, such as an increase in helical structure (vs. that in aqueous buffer), and sequence-dependent migration rates in SDS-PAGE analysis-features that may serve as structural hallmarks to verify dual TM topology in hairpin sequences.

α-Helical transmembrane (TM) segments in membrane proteins are comprised primarily of hydrophobic amino acids that accommodate insertion from water into the nonpolar membrane bilayer. In many such segments, however, polar residues are also present for structural or functional reasons. These latter residues impair the local favorable acyl interactions required for solvation by hydrophobic media such as phospholipids in native bilayers or detergents used for in vitro characterization. Using a series of Lys-tagged designed TM-like peptides (typified by KK-YAAAIAAIAWAIAAIAAAIAA-KKK) in which single-Asn residue substitutions (from Ile or Ala) were made successively from the center of the hydrophobic region toward the C-terminus, we demonstrate that polar residues strongly alter the nature of the interaction between TM segments and the solvating detergent. Through the application of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, circular dichroism spectroscopy, and tryptophan fluorescence, we observed drastic differences in the structures of the detergent-peptide complexes that contain relatively minor sequence differences. For example, the blue shift of the Trp fluorescence (indicating local detergent solvation at this location) differs by as much as ~10 nm depending upon the position of a single Asn substitution in an otherwise identical segment. The overall results suggest that polar point mutations occurring in a biological membrane will elicit comparable effects, placing a significant refolding burden on the local protein structure and potentially leading to disease states through altered protein--lipid interactions in membrane proteins.

Given the central roles of membrane proteins in cellular processes ranging from nutrient uptake to cell-cell communication, as well as the importance of these proteins as drug targets, efforts to understand and control their structures are vital in human health and disease. The rational design of membrane proteins with modified properties is thus a highly desirable goal in molecular medicine and biotechnology. However, experimental data showing how individual transmembrane (TM) residues and/or segments direct the packing and folding of membrane proteins into biologically functional entities remain sparse. To address these questions in a systematic manner, helix-helix interactions between two (or more) TM segments must be identified and analyzed. Here we present an overview of the utilization of peptides as models of the TM segments of alpha-helical membrane proteins in uncovering the amino acid sequence motifs and interactions that build these molecules. TM peptide design and production strategies are discussed, and specific examples of the application of TM peptides to the study of membrane proteins are presented. We demonstrate that TM peptides can be routinely produced in sufficient quantities for biophysical analysis, are amenable to a variety of experimental techniques, and can effectively replicate the native helix-helix contacts and key aspects of the natural biological structures of membrane proteins.

Tyrosyl‐DNA phosphodiesterase 2 (TDP2) repairs abortive topoisomerase II cleavage complexes. Here, we identify a novel short isoform of TDP2 (TDP2(S)) expressed from an alternative transcription start site. TDP2(S) contains a mitochondrial targeting sequence, contributing to its enrichment in the mitochondria and cytosol, while full‐length TDP2 contains a nuclear localization signal and the ubiquitin‐associated domain in the N‐terminus. Our study reveals that both TDP2 isoforms are present and active in the mitochondria. Comparison of isogenic wild‐type (WT) and TDP2 knockout (TDP2 (−/−/−)) DT40 cells shows that TDP2 (−/−/−) cells are hypersensitive to mitochondrial‐targeted doxorubicin (mtDox), and that complementing TDP2 (−/−/−) cells with human TDP2 restores resistance to mtDox. Furthermore, mtDox selectively depletes mitochondrial DNA in TDP2 (−/−/−) cells. Using CRISPR‐engineered human cells expressing only the TDP2(S) isoform, we show that TDP2(S) also protects human cells against mtDox. Finally, lack of TDP2 in the mitochondria reduces the mitochondria transcription levels in two different human cell lines. In addition to identifying a novel TDP2(S) isoform, our report demonstrates the presence and importance of both TDP2 isoforms in the mitochondria.

The highly effective anticancer agent doxorubicin (Dox) is a frontline drug used to treat a number of cancers. While Dox has a high level of activity against cancer cells, its clinical use is often complicated by dose-limiting cardiotoxicity. While this side effect has been linked to the drug's direct activity in the mitochondria of cardiac cells, recent studies have shown that these result primarily from downstream effects of nuclear DNA damage. Our lab has developed a mitochondrially targeted derivative of Dox that enables the selective study of toxicity generated by the presence of Dox in the mitochondria of human cells. We demonstrate that mitochondria-targeted doxorubicin (mtDox) lacks any direct nuclear effects in H9c2 rat cardiomyocytes, and that these cells are able to undergo mitochondrial biogenesis. This recovery response compensates for the mitotoxic effects of Dox and prevents cell death in cardiomyocytes. Furthermore, cardiac toxicity was only observed in Dox but not mtDox treated mice. This study supports the hypothesis that mitochondrial damage is not the main source of the cardiotoxic effects of Dox.