MR1 gene and protein expression are enhanced by inhibition of the extracellular signal-regulated kinase ERK.

The MHC class I-related molecule MR1 is ubiquitously expressed, is highly conserved among mammals, and presents bacterial and endogenous antigens in tumor cells. These features indicate that tumor-specific T cells restricted to MR1 may represent ideal candidates for novel cancer-directed T-cell immunotherapy. The very low expression of the MR1 protein at the cell surface is a potential challenge limiting the possible use of MR1-directed immunotherapies. To overcome this challenge, it is important that understanding of the mechanisms regulating MR1 expression is increased, as little is known about this currently. This study identified ERK1/2 as negative regulators of the MR1 gene and protein expression. Inhibition of ERK1/2 in tumor cells or treatment of BRAF-mutant tumor cells with drugs specific for mutated BRAF increased MR1 protein expression and recognition by tumor-reactive and MR1-restricted T cells. The ERK1/2 inhibition of MR1 was mediated by the ELF1 transcription factor, which was required for MR1 gene expression. The effects of ERK1/2 inhibition also occurred in cancer cell lines of different tissue origins, cancer cell lines resistant to drugs that inhibit mutated BRAF, and primary cancer cells, making them potential targets of specific T cells. In contrast to tumor cells, the recognition of healthy cells was very poor or absent after ERK1/2 inhibition. These findings suggest a pharmaceutical approach to increase MR1 protein expression in tumor cells and the subsequent activation of MR1-restricted T cells, and they have potential therapeutic implications.

Promiscuous recognition of MR1 drives self-reactive mucosal-associated invariant T cell responses.

Mucosal-associated invariant T (MAIT) cells use canonical semi-invariant T cell receptors (TCR) to recognize microbial riboflavin precursors displayed by the antigen-presenting molecule MR1. The extent of MAIT TCR crossreactivity toward physiological, microbially unrelated antigens remains underexplored. We describe MAIT TCRs endowed with MR1-dependent reactivity to tumor and healthy cells in the absence of microbial metabolites. MAIT cells bearing TCRs crossreactive toward self are rare but commonly found within healthy donors and display T-helper-like functions in vitro. Experiments with MR1-tetramers loaded with distinct ligands revealed significant crossreactivity among MAIT TCRs both ex vivo and upon in vitro expansion. A canonical MAIT TCR was selected on the basis of extremely promiscuous MR1 recognition. Structural and molecular dynamic analyses associated promiscuity to unique TCRß-chain features that were enriched within self-reactive MAIT cells of healthy individuals. Thus, self-reactive recognition of MR1 represents a functionally relevant indication of MAIT TCR crossreactivity, suggesting a potentially broader role of MAIT cells in immune homeostasis and diseases, beyond microbial immunosurveillance.

The Effects of Friction and Temperature in the Chemical-Mechanical Planarization Process.

Chemical-mechanical planarization (CMP) represents the preferred technology in which both chemical and mechanical interactions are combined to achieve global planarization/polishing of wafer surfaces (wafer patterns from metal with a selective layer, in this paper). CMP is a complex process of material removal process by friction, which interferes with numerous mechanical and chemical parameters. Compared with chemical parameters, mechanical parameters have a greater influence on the material removal rate (MRR). The mechanical parameters manifest by friction force (Ff) and heat generated by friction in the CMP process. The Ff can be estimated by its monitoring in the CMP process, and process temperature is obtained with help of an infrared rays (IR) sensor. Both the Ff and the MRR increase by introducing colloidal silica (SiO2) as an abrasive into the selective layer CMP slurry. The calculated wafer non-uniformity (WNU) was correlated with the friction coefficient (COF). The control of Ff and of the slurry stability is important to maintain a good quality of planarization with optimal results, because Ff participates in mechanical abrasion, and large Ff may generate defects on the wafer surface. Additionally, the temperature generated by the Ff increases as the SiO2 concentration increases. The MRR of the selective layer into the CMP slurry showed a non-linear (Prestonian) behavior, useful not only to improve the planarization level but to improve its non-uniformity due to the various pressure distributions. The evaluation of the Ff allowed the calculation of the friction energy (Ef) to highlight the chemical contribution in selective-layer CMP, from which it derived an empirical model for the material removal amount (MRA) and validated by the CMP results. With the addition of abrasive nanoparticles into the CMP slurry, their concentration increased and the MRA of the selective layer improved; Ff and MRR can be increased due to the number of chemisorbed active abrasive nanoparticles by the selective layer. Therefore, a single abrasive was considered to better understand the effect of SiO2 concentration as an abrasive and of the MRR features depending on abrasive nanoparticle concentration. This paper highlights the correlation between friction and temperature of the SiO2 slurry with CMP results, useful to examine the temperature distribution. All the MRRs depending on Ef after planarization with various SiO2 concentrations had a non-linear characteristic. The obtained results can help in developing a CMP process more effectively.

APOBEC3C, a nucleolar protein induced by genotoxins, is excluded from DNA damage sites.

The human genome contains 11 APOBEC (apolipoprotein B mRNA editing catalytic polypeptide-like) cytidine deaminases classified into four families. These proteins function mainly in innate antiviral immunity and can also restrict endogenous retrotransposable element multiplication. The present study focuses on APOBEC3C (A3C), a member of the APOBEC3 subfamily. Some APOBEC3 proteins use their enzymatic activity on genomic DNA, inducing mutations and DNA damage, while other members facilitate DNA repair. Our results show that A3C is highly expressed in cells treated with DNA-damaging agents. Its expression is regulated by p53. Depletion of A3C slightly decreases proliferation and does not affect DNA repair via homologous recombination or nonhomologous end joining. The A3C interactomes obtained from control cells and cells exposed to the genotoxin etoposide indicated that A3C is a nucleolar protein. This was confirmed by the detection of either endogenous or ectopic A3C in nucleoli. Interestingly, we show that A3C is excluded from areas of DNA breaks in live cells. Our data also indicate that the C-terminal part of A3C is responsible for its nucleolar localization and exclusion from DNA damage sites.

Comparative Study of γ- and e-Radiation-Induced Effects on FBGs Using Different Femtosecond Laser Inscription Methods.

This work presents an extensive, comparative study of the gamma and electron radiation effects on the behaviour of femtosecond laser-inscribed fibre Bragg gratings (FBGs) using the point-by-point and plane-by-plane inscription methods. The FBGs were inscribed in standard telecommunication single mode silica fibre (SMF28) and exposed to a total accumulated radiation dose of 15 kGy for both gamma and electron radiation. The gratings' spectra were measured and analysed before and after the exposure to radiation, with complementary material characterisation using Fourier transform infrared (FTIR) spectroscopy. Changes in the response of the FBGs' temperature coefficients were analysed on exposure to the different types of radiation, and we consider which of the two inscription methods result in gratings that are more robust in such harsh environments. Moreover, we used the FTIR spectroscopy to locate which chemical bonds are responsible for the changes on temperature coefficients and which are related with the optical characteristics of the FBGs.

A Dynamic Motion Analysis of a Six-Wheel Ground Vehicle for Emergency Intervention Actions.

To protect the personnel of the intervention units operating in high-risk areas, it is necessary to introduce (autonomous/semi-autonomous) robotic intervention systems. Previous studies have shown that robotic intervention systems should be as versatile as possible. Here, we focused on the idea of a robotic system composed of two vectors: a carrier vector and an operational vector. The proposed system particularly relates to the carrier vector. A simple analytical model was developed to enable the entire robotic assembly to be autonomous. To validate the analytical-numerical model regarding the kinematics and dynamics of the carrier vector, two of the following applications are presented: intervention for extinguishing a fire and performing measurements for monitoring gamma radiation in a public enclosure. The results show that the chosen carrier vector solution, i.e., the ground vehicle with six-wheel drive, satisfies the requirements related to the mobility of the robotic intervention system. In addition, the conclusions present the elements of the kinematics and dynamics of the robot.

Assessment of NO2 Pollution Level during the COVID-19 Lockdown in a Romanian City.

This study investigates changes in pollution associated with the lockdown period caused by the COVID-19 pandemic in Galati (45.43° N, 28.03° E), a Romanian city located in the southeast of Romania. The study is focused on nitrogen dioxide (NO2), a trace gas which can be related to emissions from industrial activities, heating, and transportation. The investigation is based on in situ observations from local Air Quality Monitoring Stations (AQMS) and mobile remote sensing observations by Differential Optical Absorption Spectroscopy (DOAS) technique. We also show results of the NO2 vertical column measured by TROPOMI (TROPOspheric Monitoring Instrument), a space instrument onboard of satellite mission Sentinel-5P, to complement local ground-based measurements. For in situ observations, the lockdown interval (23 March 2020-15 May 2020) was separated from normal periods. The decrease in local NO2 concentration during lockdown, measured in situ, is rather small, of about 10-40% at the most, is observed only at some stations, and is better seen during workdays than during weekends. We conclude that the decrease in NO2 content over Galati city during lockdown is relatively small and may be attributed to the reduction in local traffic, a consequence of special measures and restrictions imposed during the COVID-19 lockdown by the Romanian authorities.

ASH2L drives proliferation and sensitivity to bleomycin and other genotoxins in Hodgkin's lymphoma and testicular cancer cells.

It is of clinical importance to identify biomarkers predicting the efficacy of DNA damaging drugs (genotoxins) so that nonresponders are not unduly exposed to the deleterious effects of otherwise inefficient drugs. Here, we initially focused on the bleomycin genotoxin because of the limited information about the genes implicated in the sensitivity or resistance to this compound. Using a whole-genome CRISPR/Cas9 gene knockout approach, we identified ASH2L, a core component of the H3K4 methyl transferase complex, as a protein required for bleomycin sensitivity in L1236 Hodgkin lymphoma. Knocking down ASH2L in these cells and in the NT2D1 testicular cancer cell line rendered them resistant to bleomycin, etoposide, and cisplatin but did not affect their sensitivity toward ATM or ATR inhibitors. ASH2L knockdown decreased cell proliferation and facilitated DNA repair via homologous recombination and nonhomologous end-joining mechanisms. Data from the Tumor Cancer Genome Atlas indicate that patients with testicular cancer carrying alterations in the ASH2L gene are more likely to relapse than patients with unaltered ASH2L genes. The cell models we have used are derived from cancers currently treated either partially (Hodgkin's lymphoma), or entirely (testicular cancer) with genotoxins. For such cancers, ASH2L levels could be used as a biomarker to predict the response to genotoxins. In situations where tumors are expressing low levels of ASH2L, which may allow them to resist genotoxic treatment, the use of ATR or ATM inhibitors may be more efficacious as our data indicate that ASH2L knockdown does not affect sensitivity to these inhibitors.

Role of Meteorological Parameters in the Diurnal and Seasonal Variation of NO2 in a Romanian Urban Environment.

The main purpose of this study was to investigate whether meteorological parameters (temperature, relative humidity, direct radiation) play an important role in modifying the NO2 concentration in an urban environment. The diurnal and seasonal variation recorded at a NO2 traffic station was analyzed, based on data collected in situ in a Romanian city, Braila (45.26° N, 27.95° E), during 2009-2014. The NO2 atmospheric content close to the ground had, in general, a summer minimum and a late autumn/winter maximum for most years. Two diurnal peaks were observed, regardless of the season, which were more evident during cold months. Traffic is an important contributor to the NO2 atmospheric pollution during daytime hours. The variability of in situ measurements of NO2 concentration compared relatively well with space-based observations of the NO2 vertical column by the Ozone Monitoring Instrument (OMI) satellite for most of the period under scrutiny. Data for daytime and nighttime (when the traffic is reduced) were analyzed separately, in the attempt to isolate meteorological effects. Meteorological parameters are not fully independent and we used partial correlation analysis to check whether the relationships with one parameter may be induced by another. The correlation between NO2 and temperature was not coherent. Relative humidity and solar radiation seemed to play a role in shaping the NO2 concentration, regardless of the time of day, and these relationships were only partially interconnected.

A New Setup for Real-Time Investigations of Optical Fiber Sensors Subjected to Gamma-Rays: Case Study on Long Period Gratings.

In this work, we present a new setup for real-time investigations of optical fibers and optical fiber sensors while being subjected to gamma-rays. The investigation of the radiation effects on novel or well-assessed sensing devices has attracted a lot of interest, however, the facilities required to do this (when available) are barely accessible to the device to be characterized. In order to reduce the limitations of these types of experiments and ensure a highly controlled environment, we implemented a configuration that permits the on-line testing of optical components inside a Co-60 gamma chamber research irradiator. To show the advantages of this new approach, we present a case study that compares an arc-induced optical fiber long period grating (LPG) irradiated in a gamma chamber with the same type of grating irradiated with gamma-rays from a Co-60 industrial irradiator. In order to better understand the effects of radiation on such components and their behavior in radiation environments, we focus on the homogeneity of the radiation field and parameter customizability as well as the high reproducibility of the experiments.

Evolution of SO2 and NOx Emissions from Several Large Combustion Plants in Europe during 2005-2015.

The aim of this paper is to investigate the evolution of SO2 and NOx emissions of ten very large combustion plants (LCPs >500 MW) located in the European Union (EU) during 2005-2015. The evolution of NOx and SO2 emissions were analyzed against the EU Directives in force during 2005-2015. The investigation was performed using space-borne observations and estimated emissions collected from the EEA (European Environment Agency) inventory of air pollutant emissions. The power plants were chosen according to their capacity and emissions, located in various parts of Europe, to give an overall picture of atmospheric pollution with NOx and SO2 associated with the activity of very large LCPs in Europe. Satellite observations from OMI (Ozone Monitoring Instrument) are compared with calculated emissions in order to assess whether satellite observations can be used to monitor air quality, as a standard procedure, by governmental or nongovernmental institutions. Our results show that both space observations and estimated emissions of NOx and SO2 atmospheric content have a descending trend until 2010, complying with the EU Directives. The financial and economic crisis during 2007-2009 played an important role in reducing emissions.

Quantitative assessment of morphological changes of dental pulp components of teeth affected by occlusal trauma.

AIM: In the general economy of the stomatognathic system functionality, the occlusal function has an important role, meaning both the dental-dental reports and the dynamic reports interarch-interarch. Because of the interrelationships and inter-dependency that govern the functioning of the biological systems, a pathological change affecting a component of the stomatognathic system produces impaired functioning of the others. The aim of the present study is to assess the morphological changes occurred in the dental pulp components of teeth affected by occlusal trauma. MATERIALS AND METHODS: Fragments of dental pulp coming from 45 patients with occlusal trauma were processed using classical histological techniques (formalin fixation and paraffin embedment) and stained with Hematoxylin-Eosin (HE), Masson's trichrome and anti-CD34 antibody immunostaining, in order to highlight the peripheral zone and central connective tissue of dental pulp morphological changes. A set of parameters namely thickness of peripheral zone components, calcifications, fibrosis and vascular density in the dental pulp were assessed individually and based on three topographical criteria namely the affected tooth, the position on the dental arches and the position according to sagittal plane. RESULTS: There was no correlation between morphological changes of dental pulp and the topography of teeth with occlusal trauma. The size of the peripheral area of the dental pulp and that of its components evolved in the same sense, whether it was growth or decrease. Decrease of the peripheral area was associated with the increase of collagen fibers density, calcium deposits and density of the capillary network. The direct correlation between the amount of collagen fibers and vascular density seems somewhat paradoxical but it can be explained by the reemergence of chronic inflammatory events located in the dental pulp. CONCLUSIONS: It seems that dental pulp morphological changes are not influenced by the teeth with occlusal trauma topography. With one exception (the components of peripheral zone), most of the correlations between the dental pulp morphological changes were only suggested but not validated statistically, which requires further studies on larger groups together with the introduction of inflammatory cell population studies.

Morphological changes of gums in occlusal trauma.

AIM: Occlusal trauma causes major modifications of the coverage periodontium, which in turn reflect on the dental unit. The aim of the present study is to evaluate some of the morphological modifications occurred in the marginal periodontium surrounding teeth affected by occlusal trauma. MATERIALS AND METHODS: Fragments of marginal periodontium coming from 51 patients with occlusal trauma were processed using classical histological techniques (formalin fixation and paraffin embedment) and stained with Hematoxylin-Eosin (HE), Masson's trichrome and anti-CD34 antibody immunostaining, in order to highlight the epithelial and connective tissue changes of gingival mucosa. A set of epithelial and connective tissue morphological parameters were assessed individually and based on three topographical criteria concerning the affected tooth. RESULTS: The epithelium and especially its superficial compartment presented changes depending on the tooth type and the dental arch. Epithelial thickness had the tendency to decrease as the fibrosis in both corium compartments and vascular density in the deep corium compartment were increasing. Leukoplakia present around the affected teeth but not always was related with the tooth type and was more obvious as the superficial compartment of the epithelium was thicker and as fibrosis was more reduced in the papillary compartment of the corium. Vascular density reduced when fibrosis process increased in the corium. CONCLUSIONS: Lesions determined by occlusal trauma and their topography can and are influencing locally the different structures of the surrounding periodontium.

NFκB regulates p21 expression and controls DNA damage-induced leukemic differentiation.

DNA damage exposure is a major modifier of cell fate in both normal and cancer tissues. In response to DNA damage, myeloid leukemia cells activate a poorly understood terminal differentiation process. Here, we show that the NFκB pathway directly activates expression of the proliferation inhibitor p21 in response to DNA damage in myeloid leukemia cells. In order to understand the role of this unexpected regulatory event, we ablated the NFκB binding site we identified in the p21 promoter, using CRISPR/Cas9-mediated genome editing. We found that NFκB-mediated p21 activation controls DNA damage-induced myeloid differentiation. Our results uncover a p53-independent pathway for p21 activation involved in controlling hematopoietic cell fate.

Assessing tumor angiogenesis in colorectal cancer by quantitative contrast-enhanced endoscopic ultrasound and molecular and immunohistochemical analysis.

BACKGROUND AND OBJECTIVES: Data on contrast-enhanced endoscopic ultrasound (CE-EUS) for colorectal cancer (CRC) evaluation are scarce. Therefore, we aimed to assess the vascular perfusion pattern in CRC by quantitative CE-EUS and compare it to immunohistochemical and genetic markers of angiogenesis. PATIENTS AND METHODS: We performed a retrospective analysis of CE-EUS examinations of 42 CRC patients, before any therapy. CE-EUS movies were processed using a dedicated software. Ten parameters were automatically generated from the time-intensity curve (TIC) analysis: peak enhancement (PE), rise time (RT), mean transit time, time to peak (TTP), wash-in area under the curve (WiAUC), wash-in rate (WiR), wash-in perfusion index (WiPI), wash-out AUC (WoAUC), and wash-in and wash-out AUC (WiWoAUC). The expression levels of the vascular endothelial growth factor receptor 1 (VEGFR1) and VEGFR2 genes were assessed from biopsy samples harvested during colonoscopy. Microvascular density and vascular area were calculated after CD31 and CD105 immunostaining. RESULTS: Forty-two CE-EUS video sequences were analyzed. We found positive correlations between the parameters PE, WiAUC, WiR, WiPI, WoAUC, WiWoAUC, and N staging (Spearman r = 0.437, r = 0.336, r = 0.462, r = 0.437, r = 0.358, and r = 0.378, respectively, P < 0.05), and also between RT and TTP and CD31 vascular area (r = 0.415, and r = 0.421, respectively, P < 0.05). VEGFR1 and VEGFR2 expression did not correlate with any of the TIC parameters. CONCLUSIONS: CE-EUS with TIC analysis enables minimally invasive assessment of CRC angiogenesis and may provide information regarding the lymph nodes invasion. However, further studies are needed for defining its role in the evaluation of CRC patients.

OMI and Ground-Based In-Situ Tropospheric Nitrogen Dioxide Observations over Several Important European Cities during 2005-2014.

In this work we present the evolution of tropospheric nitrogen dioxide (NO2) content over several important European cities during 2005-2014 using space observations and ground-based in-situ measurements. The NO2 content was derived using the daily observations provided by the Ozone Monitoring Instrument (OMI), while the NO2 volume mixing ratio measurements were obtained from the European Environment Agency (EEA) air quality monitoring stations database. The European cities selected are: Athens (37.98° N, 23.72° E), Berlin (52.51° N, 13.41° E), Bucharest (44.43° N, 26.10° E), Madrid (40.38° N, 3.71° W), Lisbon (38.71° N, 9.13° W), Paris (48.85° N, 2.35° E), Rome (41.9° N, 12.50° E), and Rotterdam (51.91° N, 4.46° E). We show that OMI NO2 tropospheric column data can be used to assess the evolution of NO2 over important European cities. According to the statistical analysis, using the seasonal variation, we found good correlations (R > 0.50) between OMI and ground-based in-situ observations for all of the cities presented in this work. Highest correlation coefficients (R > 0.80) between ground-based monitoring stations and OMI observations were calculated for the cities of Berlin, Madrid, and Rome. Both types of observations, in-situ and remote sensing, show an NO2 negative trend for all of locations presented in this study.

Correlations between endothelial cell markers CD31, CD34 and CD105 in colorectal carcinoma.

PURPOSE: Colorectal carcinoma is an important cause of mortality worldwide. The fact that tumor growth is dependent on angiogenesis has supported researches for new prognostic parameters and the development of novel therapeutic strategies. Accordingly, we sought to evaluate angiogenesis quantitatively by assessing microvessel density in colorectal cancer. MATERIALS AND METHODS: The blood vessels stained with CD31, CD34 and CD105 were counted, and we reported their number per square millimeter in order to obtain microvascular density (MVD). Then, we aimed at comparing the performance of three endothelial cell markers (CD31, CD34, and CD105) on formalin-fixed tissues from 58 patients diagnosed with colorectal cancer. RESULTS: Following the comparison of the average effective vessels marked with the three markers, Student's t-test showed that the mean number of blood vessels marked with CD34 is higher than the blood vessels marked with CD31 and CD105. A significant difference that has been registered between the three levels of the T stage was found in the patients in our study, in terms of value marker CD105, ANOVA p=0.049, which returns to a value <0.05. Quick time decreases the pT stage, the observed differences being close to statistical significance. However, the result of ANOVA test does not allow us to say that differences can be generalized and not just a particular result, valid only for the study group, p=0.061 >0.05. There is a significant difference between patients with stage T, in terms of value: hemoglobin (ANOVA p<0.001), hematocrit (ANOVA p<0.001), mean corpuscular volume (MCV) (ANOVA p<0.001), mean corpuscular hemoglobin (MCH) (ANOVA p=0.002 <0.01 - significant difference with 99% confidence). By calculating the Pearson's correlation coefficient for the relationship CD31-CD105, we obtained a value r=0.440, which corresponds to p=0.0013 <0.05, indicating a statistically noteworthy direct correlation between the two factors. CONCLUSIONS: CD31 marker increases simultaneously with the CD105, in the cases analyzed throughout the present study. The ability of tumors to maintain a high vascular blood density in their inner portions may represent a reliable parameter to evaluate tumor angiogenesis and a finding relevant for future development of therapeutic angiogenesis strategies.

Synthesis and Structural Investigation of New Bio-Relevant Complexes of Lanthanides with 5-Hydroxyflavone: DNA Binding and Protein Interaction Studies.

In the present work, we attempted to develop new metal coordination complexes of the natural flavonoid 5-hydroxyflavone with Sm(III), Eu(III), Gd(III), Tb(III). The resultant hydroxo complexes have been characterized by a variety of spectroscopic techniques, including fluorescence, FT-IR, UV-Vis, EPR and mass spectral studies. The general chemical formula of the complexes is [Ln(C15H9O3)3(OH)2(H2O)x]·nH2O, where Ln is the lanthanide cation and x = 0 for Sm(III), x = 1 for Eu(III), Gd(III), Tb(III) and n = 0 for Sm(III), Gd(III), Tb(III), n = 1 for Eu(III), respectively. The proposed structures of the complexes were optimized by DFT calculations. Theoretical calculations and experimental determinations sustain the proposed structures of the hydroxo complexes, with two molecules of 5-hydroxyflavone acting as monoanionic bidentate chelate ligands. The interaction of the complexes with calf thymus DNA has been explored by fluorescence titration and UV-Vis absorption binding studies, and revealed that the synthesized complexes interact with DNA with binding constants (Kb) ~ 104. Human serum albumin (HSA) and transferrin (Tf) binding studies have also been performed by fluorescence titration techniques (fluorescence quenching studies, synchronous fluorescence spectra). The apparent association constants (Ka) and thermodynamic parameters have been calculated from the fluorescence quenching experiment at 299 K, 308 K, and 318 K. The quenching curves indicate that the complexes bind to HSA with smaller affinity than the ligand, but to Tf with higher binding affinities than the ligand.