Distribution of KIR Genes and Their HLA Ligands in Different Viral Infectious Diseases: Frequency Study in Sicilian Population

Natural killer (NK) cells play a role in defence against viral infections by killing infected cells or by producing cytokines and interacting with adaptive immune cells. Killer immunoglobulin-like receptors (KIRs) regulate the activation of NK cells through their interaction with human leucocyte antigens (HLA). Ninety-six Sicilian patients positive to Human Immunodeficiency Virus-1 (HIV) and ninety-two Sicilian patients positive to SARS-CoV-2 were genotyped for KIRs and their HLA ligands. We also included fifty-six Sicilian patients with chronic hepatitis B (CHB) already recruited in our previous study. The aim of this study was to compare the distribution of KIR–HLA genes/groups of these three different infected populations with healthy Sicilian donors from the literature. We showed that the inhibitory KIR3DL1 gene and the KIR3DL1/HLA-B Bw4 pairing were more prevalent in individual CHB. At the same time, the frequency of HLA-C2 was increased in CHB compared to other groups. In contrast, the HLA-C1 ligand seems to have no contribution to CHB progression whereas it was significantly higher in COVID-19 and HIV-positive than healthy controls. These results suggest that specific KIR–HLA combinations can predict the outcome/susceptibility of these viral infections and allows to plan successful customized therapeutic strategies.     

Effect of Zn2TiO4 and ZnB2O4 additions on the microstructure and dielectric properties of AgNb1 ? xTaxO3 solid solutions

We report the preparation and characterization of AgNb_0.6Ta_0.4O₃ (ANT) based materials. The addition of Zn₂TiO₄ and ZnB₂O₄ influences the sintering temperature, phase composition, and microstructure of ANT ceramics. ANT doped with 1 wt % Zn₂TiO₄ or ZnB₂O₄ has high dielectric permittivity (400–470), low dielectric losses (tanδ ∼ 10⁻³), and a nonlinearity coefficient n _R ≅ 3–9% (at a field strength E = 3 × 10⁶ V/m).     

Different response to acetic acid stress in Saccharomyces cerevisiae wild‐type and l‐ascorbic acid‐producing strains

Biotechnological processes are of increasing significance for industrial production of fine and bulk chemicals, including biofuels. Unfortunately, under operative conditions microorganisms meet multiple stresses, such as non‐optimal pH, temperature, oxygenation and osmotic stress. Moreover, they have to face inhibitory compounds released during the pretreatment of lignocellulosic biomasses, which constitute the preferential substrate for second‐generation processes. Inhibitors include furan derivatives, phenolic compounds and weak organic acids, among which acetic acid is one of the most abundant and detrimental for cells. They impair cellular metabolism and growth, reducing the productivity of the process: therefore, the development of robust cell factories with improved production rates and resistance is of crucial importance. Here we show that a yeast strain engineered to endogenously produce vitamin C exhibits an increased tolerance compared to the parental strain when exposed to acetic acid at moderately toxic concentrations, measured as viability on plates. Starting from this evidence, we investigated more deeply: (a) the nature and levels of reactive oxygen species (ROS); (b) the activation of enzymes that act directly as detoxifiers of reactive oxygen species, such as superoxide dismutase (SOD) and catalase, in parental and engineered strains during acetic acid stress. The data indicate that the engineered strain can better recover from stress by limiting ROS accumulation, independently from SOD activation. The engineered yeast can be proposed as a model for further investigating direct and indirect mechanism(s) by which an antioxidant can rescue cells from organic acid damage; moreover, these studies will possibly provide additional targets for further strain improvements. Copyright © 2013 John Wiley & Sons, Ltd.     

Visualizing and exploring profiles with calling context ring charts

Calling context profiling is an important technique for analyzing the performance of object‐oriented software with complex inter‐procedural control flow. The Calling Context Tree (CCT) is a common data structure that stores dynamic metrics, such as CPU time, separately for each calling context. As CCTs may comprise millions of nodes, there is a need for a condensed visualization that eases the localization of performance bottlenecks. In this article, we discuss Calling Context Ring Charts (CCRCs), a compact visualization for CCTs, where callee methods are represented in ring segments surrounding the caller's ring segment. In order to reveal hot methods, their callers, and callees, the ring segments can be sized according to a chosen dynamic metric. We describe two case studies where CCRCs help us to detect and fix performance problems in applications. A performance evaluation also confirms that our implementation can efficiently handle large CCTs. Copyright © 2010 John Wiley & Sons, Ltd.     

Molecular Mechanism of Action of 2‐Ferrocenyl‐1,1‐diphenylbut‐1‐ene on HL‐60 Leukemia Cells

The aim of this work was to investigate the mechanism of action of 2‐ferrocenyl‐1,1‐diphenylbut‐1‐ene ( 1 ) on HL‐60 human leukemia cells. While inactive against noncancerous cells, 1 provoked a concentration‐dependent decrease in viable tumor cells, primarily via apoptosis, as evidenced by analysis of cell morphology, activation of caspases 3 and 7, increased DNA fragmentation, and externalization of phosphatidylserine. Necrosis was observed only at the highest tested concentration (4 μM ). Compound 1 interfered with the cell cycle, causing an accumulation of cells in the G₁/G₀ phase. Interaction of 1 with dsDNA and ssDNA was observed by differential pulse voltammetry and confirmed by hyperchromicity in the UV/Vis spectra of dsDNA, with an interaction constant of 2×10⁴ M ^?1. Both the organic analogue 1,1,2‐triphenylbut‐1‐ene ( 2 ) and ferrocene were inactive against cancer and noncancer cell lines and did not react with DNA. These results reinforce the idea that the hybrid strategy of conjugating ferrocene to the structure of tamoxifen derivatives is advantageous in finding new substances with antineoplastic activity.     

The Mitochondrial Amidoxime Reducing Component (mARC): Involvement in Metabolic Reduction of N‐Oxides,Oximes and N‐Hydroxyamidinohydrazones

The mitochondrial amidoxime reducing component (mARC) is a molybdenum‐containing enzyme and capable of reducing N‐hydroxylated structures such as amidoxime prodrugs. In this study, we tested the involvement of mARC in the reduction of N‐oxides (amitriptyline‐N‐oxide, nicotinamide‐N‐oxide), oximes ((E)‐/(Z)‐2,4,6‐trimethylacetophenonoxime) and a N‐hydroxyamidinohydrazone (guanoxabenz). All groups are reduced by mARC proteins, and the enzymes are therefore involved in the interconversion of N‐oxygenated metabolites originating from cytochrome P450s and flavin‐containing monooxygenases. In addition, these structures open up further options for serving as prodrugs. Thus, with respect to these reactions, testing of candidates with N‐oxygenated structures should not solely be carried out in microsomal enzyme sources but as well in mitochondria. However, differences in the reduction of oximes and N‐oxides between the two isoforms, namely mARC1 and mARC2, were detectable; N‐oxides are exclusively reduced by mARC1. We therefore assume differences between the so far unknown 3D structures of the two proteins.     

pH-triggered conduction of amine-functionalized single ZnO wire integrated on a customized nanogap electronic platform

The electrical conductance response of single ZnO microwire functionalized with amine-groups was tested upon an acid pH variation of a solution environment after integration on a customized gold electrode array chip. ZnO microwires were easily synthesized by hydrothermal route and chemically functionalized with aminopropyl groups. Single wires were deposited from the solution and then oriented through dielectrophoresis across eight nanogap gold electrodes on a platform single chip. Therefore, eight functionalized ZnO microwire-gold junctions were formed at the same time, and being integrated on an ad hoc electronic platform, they were ready for testing without any further treatment. Experimental and simulation studies confirmed the high pH-responsive behavior of the amine-modified ZnO-gold junctions, obtaining in a simple and reproducible way a ready-to-use device for pH detection in the acidic range. We also compared this performance to bare ZnO wires on the same electronic platform, showing the superiority in pH response of the amine-functionalized material.     

A potential antitumor drug (arginine deiminase) reengineered for efficient operation under physiological conditions

Arginine deiminase (ADI, EC 3.5.3.6) is a potential antitumor drug for the treatment of arginine‐auxotrophic tumors such as hepatocellular carcinomas (HCCs) and melanomas, and studies on human lymphatic leukemia cell lines have confirmed that ADI has antiangiogenic activity. Recent studies showed that a combination of taxane and ADI‐PEG20, which induces caspase‐independent apoptosis, is more effective than taxane monotherapy for prostate cancer. The main limitation of ADI from Pseudomonas plecoglossicida (PpADI) and of many other ADI enzymes lies in their pH‐dependent activity profile. PpADI has a pH optimum at 6.5 and a pH shift from 6.5 to 7.5 results in an ～80 % activity drop (the pH of human plasma is 7.35 to 7.45). In 2010, we reported a proof of concept for ADI engineering by directed evolution that resulted in variant M2 (K5T/D44E/H404R). M2 has a pH optimum of pH 7.0, a fourfold higher k_cat value than the wild‐type PpADI (pH 7.4, 0.5 M phosphate buffer), and an increased K_m value for substrate arginine. In our latest work, variants M5 (K5T/D38H/D44E/A128T/H404R) and M6 (K5T/D38H/D44E/A128T/E296K/H404R) were generated by directed evolution by employing PBS buffer (pH 7.4), which mimics physiological conditions. The S_0.5 value of parent M3 (K5T/D44E/A128T/H404R) decreased from 2.01 to 1.48 mM (M5) and 0.81 mM (M6). The S_0.5 value of M6 (0.81 mM ) is lower than that of wild‐type PpADI (1.30 mM ); the k_cat values improved from 0.18 s^?1 (wild‐type PpADI) to 17.56 s^?1 (M5, 97.6‐fold) and 11.64 s^?1 (M6, 64.7‐fold).     

Directed Evolution of an Antitumor Drug (Arginine Deiminase PpADI) for Increased Activity at Physiological pH

Arginine deiminase (ADI; EC 3.5.3.6) has been studied as a potential antitumor drug for the treatment of arginine‐auxotrophic tumors, such as hepatocellular carcinomas (HCCs) and melanomas. Studies with human lymphatic leukemia cell lines confirmed that ADI is an antiangiogenic agent for treating leukemia. The main limitation of ADI from Pseudomonas plecoglossicida (PpADI) lies in its pH‐dependent activity profile, its pH optimum is at 6.5. A pH shift from 6.5 to 7.5 results in an approximately 80 % drop in activity. (The pH of human plasma is 7.35 to 7.45.) In order to shift the PpADI pH optimum, a directed‐evolution protocol based on an adapted citrulline‐screening protocol in microtiter‐plate format was developed and validated. A proof of concept for ADI engineering resulted in a pH optimum of pH 7.0 and increased resistance under physiological and slightly alkaline conditions. At pH 7.4, variant M2 (K5T/D44E/H404R) is four times faster than the wild‐type PpADI and retains ～50 % of its activity relative to its pH optimum, compared to ～10 % in the case of the wild‐type PpADI.