Evidence for a Negative Correlation between Human Reactive Enamine-Imine Intermediate Deaminase A (RIDA) Activity and Cell Proliferation Rate: Role of Lysine Succinylation of RIDA

Reactive intermediate deaminase (Rid) proteins are enzymes conserved in all domains of life. UK114, a mammalian member of RidA subfamily, has been firstly identified as a component of liver perchloric acid-soluble proteins (L-PSP). Although still poorly defined, several functions have been attributed to the mammalian protein UK114/RIDA, including the reactive intermediate deamination activity. The expression of UK114/RIDA has been observed in some tumors, arousing interest in this protein as an evaluable tumor marker. However, other studies reported a negative correlation between UK114/RIDA expression, tumor differentiation degree and cell proliferation. This work addressed the question of UK114/RIDA expression in human non-tumor HEK293 cell lines and in some human tumor cell lines. Here we reported that human RIDA (hRIDA) was expressed in all the analyzed cell line and subjected to lysine (K-)succinylation. In HEK293, hRIDA K-succinylation was negatively correlated to the cell proliferation rate and was under the control of SIRT5. Moreover, K-succinylation clearly altered hRIDA quantification by immunoblotting, explaining, at least in part, some discrepancies about RIDA expression reported in previous studies. We found that hRIDA was able to deaminate reactive enamine-imine intermediates and that K-succinylation drastically reduced deaminase activity. As predicted by in silico analysis, the observed reduction of deaminase activity has been related to the drastic alterations of hRIDA structure inferred by K-succinylation. The role of hRIDA and the importance of its K-succinylation in cell metabolism, especially in cancer biology, have been discussed.     

Driving business performance: innovation complementarities and persistence patterns

Complementarities between technological and non-technological innovation are crucial determinants of firm performance. Although innovation complementarity has been extensively tested in the empirical literature, it has not been analysed in conjunction with innovation persistence. This fact is mainly due to the lack of data sets able to provide adequate longitudinal information. The capacities to develop market-oriented behaviour and introduce new organisational innovations, together with technological innovation, are the drivers of a firm’s productivity and profitability. We find that these activities complement technological innovation and that their impact is greater when they persist over time, thus introducing a more general concept of innovation persistence. We present an empirical model based on a large new panel of Italian manufacturing firms covering the period 2000–2012 which enables us to determine the precise impacts of a firm’s innovative attitude, in a broad definition that incorporates non-technological innovation and persistence, on its productivity and profitability.     

Proteins as biomarkers of oxidative/nitrosative stress in diseases: the contribution of redox proteomics

Reactive oxygen species (ROS) and reactive nitrogen species (RNS) contribute to the pathogenesis and/or progression of several human diseases. Proteins are important molecular signposts of oxidative/nitrosative damage. However, it is generally unresolved whether the presence of oxidatively/nitrosatively modified proteins has a causal role or simply reflects secondary epiphenomena. Only direct identification and characterization of the modified protein(s) in a given pathophysiological condition can decipher the potential roles played by ROS/RNS-induced protein modifications. During the last few years, mass spectrometry (MS)-based technologies have contributed in a significant way to foster a better understanding of disease processes. The study of oxidative/nitrosative modifications, investigated by redox proteomics, is contributing to establish a relationship between pathological hallmarks of disease and protein structural and functional abnormalities. MS-based technologies promise a contribution in a new era of molecular medicine, especially in the discovery of diagnostic biomarkers of oxidative/nitrosative stress, enabling early detection of diseases. Indeed, identification and characterization of oxidatively/nitrosatively modified proteins in human diseases has just begun.     

Exposure data and risk indicators for safety performance assessment in Europe

The objective of this paper is the analysis of the state-of-the-art in risk indicators and exposure data for safety performance assessment in Europe, in terms of data availability, collection methodologies and use. More specifically, the concepts of exposure and risk are explored, as well as the theoretical properties of various exposure measures used in road safety research (e.g. vehicle- and person-kilometres of travel, vehicle fleet, road length, driver population, time spent in traffic, etc.). Moreover, the existing methods for collecting disaggregate exposure data for risk estimates at national level are presented and assessed, including survey methods (e.g. travel surveys, traffic counts) and databases (e.g. national registers). A detailed analysis of the availability and quality of existing risk exposure data is also carried out. More specifically, the results of a questionnaire survey in the European countries are presented, with detailed information on exposure measures available, their possible disaggregations (i.e. variables and values), their conformity to standard definitions and the characteristics of their national collection methods. Finally, the potential of international risk comparisons is investigated, mainly through the International Data Files with exposure data (e.g. Eurostat, IRTAD, ECMT, UNECE, IRF, etc.). The results of this review confirm that comparing risk rates at international level may be a complex task, as the availability and quality of exposure estimates in European countries varies significantly. The lack of a common framework for the collection and exploitation of exposure data limits significantly the comparability of the national data. On the other hand, the International Data Files containing exposure data provide useful statistics and estimates in a systematic way and are currently the only sources allowing international comparisons of road safety performance under certain conditions.     

Neutrophils Mediate Pulmonary Artery Thrombosis In Situ

Pulmonary embolism is a life-threatening condition, which can result in respiratory insufficiency and death. Blood clots occluding branches of the pulmonary artery (PA) are traditionally considered to originate from thrombi in deep veins (usually in legs). However, growing evidence suggests that occlusion of the vessels in the lungs can develop without preceding deep vein thrombosis (DVT). In this work, we used an inferior vena cava (IVC) complete ligation model of DVT in Wistar rats to explore the possibility and mechanisms of PA thrombosis under the conditions where all routes of thrombotic mass migration from peripheral veins are blocked. We demonstrate that rats both with normal and reduced neutrophil counts developed thrombi in the IVC, although, neutropenia caused a substantial decrease in thrombus size and a shift from fresh fibrin toward mature fibrin and connective tissue inside the thrombus. Massive fibrin deposition was found in the PA branches in the majority of DVT rats with normal neutrophil counts, but in none of the neutropenic animals. Neutrophil ablation also abolished macroscopic signs of lung damage. Altogether, the results demonstrate that thrombi in the lung vasculature can form in situ by mechanisms that require local neutrophil recruitment taking place in the DVT setting.     

State-of-the-Art on Wound Vitality Evaluation: A Systematic Review

The vitality demonstration refers to determining if an injury has been caused ante- or post-mortem, while wound age means to evaluate how long a subject has survived after the infliction of an injury. Histology alone is not enough to prove the vitality of a lesion. Recently, immunohistochemistry, biochemistry, and molecular biology have been introduced in the field of lesions vitality and age demonstration. The study was conducted according to the preferred reporting items for systematic review (PRISMA) protocol. The search terms were “wound”, “lesion”, “vitality”, “evaluation”, “immunohistochemistry”, “proteins”, “electrolytes”, “mRNAs”, and “miRNAs” in the title, abstract, and keywords. This evaluation left 137 scientific papers. This review aimed to collect all the knowledge on vital wound demonstration and provide a temporal distribution of the methods currently available, in order to determine the age of lesions, thus helping forensic pathologists in finding a way through the tangled jungle of wound vitality evaluation.     

Engineered Sleeping Beauty Transposon as Efficient System to Optimize Chimp Adenoviral Production

Sleeping Beauty (SB) is the first DNA transposon employed for efficient transposition in vertebrate cells, opening new applications for genetic engineering and gene therapies. A transposon-based gene delivery system holds the favourable features of non-viral vectors and an attractive safety profile. Here, we employed SB to engineer HEK293 cells for optimizing the production of a chimpanzee Adenovector (chAd) belonging to the Human Mastadenovirus C species. To date, chAd vectors are employed in several clinical settings for infectious diseases, last but not least COVID-19. A robust, efficient and quick viral vector production could advance the clinical application of chAd vectors. To this aim, we firstly swapped the hAd5 E1 with chAd-C E1 gene by using the CRISPR/Cas9 system. We demonstrated that in the absence of human Ad5 E1, chimp Ad-C E1 gene did not support HEK293 survival. To improve chAd-C vector production, we engineered HEK293 cells to stably express the chAd-C precursor terminal protein (ch.pTP), which plays a crucial role in chimpanzee Adenoviral DNA replication. The results indicate that exogenous ch.pTP expression significantly ameliorate the packaging and amplification of recombinant chAd-C vectors thus, the engineered HEK293ch.pTP cells could represent a superior packaging cell line for the production of these vectors.     

Parallel enzymatic and non-enzymatic formation of zinc protoporphyrin IX in pork

Zinc protoporphyrin is formed in pork homogenates in both enzymatic and non-enzymatic reactions. Ferrochelatase is active in formation of the highly fluorescent pigment known from Parma ham as demonstrated by inhibition with N-methylmesoporphyrin and by thermal inactivation. A non-enzymatic transmetallisation reaction, exchange of iron in myoglobin by zinc(II), is demonstrated by Pb(II) inhibition of zinc protoporphyrin formation at low Pb(II) concentrations, but promoted at higher Pb(II) concentrations. The non-enzymatic reaction is characterised as a slow bimolecular reaction between protoporphyrin IX and zinc(II) with a second-order rate constant of 0.63 l mol⁻¹ s⁻¹ at 35 °C and a high energy of activation of 98 kJ mol⁻¹ for acetone:water (3:1, v/v) as solvent. Zinc protoporphyrin formation is concluded to be thermodynamically controlled with a formation constant of 4 × 10⁵ M (35 °C, acetone:water (3:1)). An efficient inhibition of formation of zinc protoporphyrin by nitrite is related to myoglobin as substrate and involves both enzymatic and non-enzymatic reactions.