Dissociation constants of relevant secondary organic aerosol components in the atmosphere.

The presented studies focus on measuring the determination of the acidity constant (pKa) of relevant secondary organic aerosol components. For our research, we selected important oxidation products (mainly carboxylic acids) of the most abundant terpene compounds, such as α-pinene, ß-pinene, ß-caryophyllene, and δ-3-carene. The research covered the synthesis and determination of the acidity constant of selected compounds. We used three methods to measure the acidity constant, i.e., 1H NMR titration, pH-metric titration, Bates-Schwarzenbach spectrophotometric method. Moreover, the pKa values were calculated with Marvin 21.17.0 software to compare the experimentally derived values with those calculated from the chemical structure. pKa values measured with 1H NMR titration ranged from 3.51 ± 0.01 for terebic acid to 5.18 ± 0.06 for ß-norcaryophyllonic acid. Moreover, the data determined by the 1H NMR method revealed a good correlation with the data obtained with the commonly used potentiometric and UV-spectroscopic methods (R2 = 0.92). In contrast, the comparison with in silico results exhibits a relatively low correlation (R2Marvin = 0.66). We found that most of the values calculated with the Marvin Program are lower than experimental values obtained with pH-metric titration with an average difference of 0.44 pKa units. For di- and tricarboxylic acids, we obtained two and three pKa values, respectively. A good correlation with the literature values was observed, for example, Howell and Fisher (1958) used pH-metric titration and measured pKa1 and pKa2 to be 4.48 and 5.48, while our results are 4.24 ± 0.10 and 5.40 ± 0.02, respectively.

Fast Dynamic Siloxane Exchange Mechanism for Reshapable Vitrimer Composites.

To develop siloxane-containing vitrimers with fast dynamic characteristics, different mechanistic pathways have been investigated using a range of catalysts. In particular, one siloxane exchange pathway has been found to show a fast dynamic behavior in a useful temperature range (180-220 °C) for its application in vitrimers. The mechanism is found to involve 1,5,7-triazabicyclo [4.4.0] dec-5-ene (TBD) as an organic catalyst in the presence of hydroxyl groups. Using this new mechanistic approach, vitrimers with ultrafast stress-relaxation characteristics (relaxation times below 10 s) have been prepared with a readily available epoxy resin and siloxane-amine hardener. Subsequently, the low viscosity siloxane-containing vitrimer resin enabled the preparation of glass fiber-reinforced vitrimer composites using an industrially relevant vacuum-assisted resin infusion technique. The resulting composite was successfully thermoformed into a new shape, which makes it possible to envision a second life for such highly engineered materials.

Spatial CRISPR genomics identifies regulators of the tumor microenvironment.

While CRISPR screens are helping uncover genes regulating many cell-intrinsic processes, existing approaches are suboptimal for identifying extracellular gene functions, particularly in the tissue context. Here, we developed an approach for spatial functional genomics called Perturb-map. We applied Perturb-map to knock out dozens of genes in parallel in a mouse model of lung cancer and simultaneously assessed how each knockout influenced tumor growth, histopathology, and immune composition. Moreover, we paired Perturb-map and spatial transcriptomics for unbiased analysis of CRISPR-edited tumors. We found that in Tgfbr2 knockout tumors, the tumor microenvironment (TME) was converted to a fibro-mucinous state, and T cells excluded, concomitant with upregulated TGFß and TGFß-mediated fibroblast activation, indicating that TGFß-receptor loss on cancer cells increased TGFß bioavailability and its immunosuppressive effects on the TME. These studies establish Perturb-map for functional genomics within the tissue at single-cell resolution with spatial architecture preserved and provide insight into how TGFß responsiveness of cancer cells can affect the TME.

Radical Formation in Sugar-Derived Acetals under Solvent-Free Conditions.

The degradation of acetal derivatives of the diethylester of galactarate (GalX) was investigated by electron paramagnetic resonance (EPR) spectroscopy in the context of solvent-free, high-temperature reactions like polycondensations. It was demonstrated that less substituted cyclic acetals are prone to undergo radical degradation at higher temperatures as a result of hydrogen abstraction. The EPR observations were supported by the synthesis of GalX based polyamides via ester-amide exchange-type polycondensations in solvent-free conditions at high temperatures in the presence and in the absence of radical inhibitors. The radical degradation can be offset by the addition of a radical inhibitor. The radical is probably formed on the methylene unit between the oxygen atoms and subsequently undergoes a rearrangement.

A microRNA expression and regulatory element activity atlas of the mouse immune system.

To better define the control of immune system regulation, we generated an atlas of microRNA (miRNA) expression from 63 mouse immune cell populations and connected these signatures with assay for transposase-accessible chromatin using sequencing (ATAC-seq), chromatin immunoprecipitation followed by sequencing (ChIP-seq) and nascent RNA profiles to establish a map of miRNA promoter and enhancer usage in immune cells. miRNA complexity was relatively low, with >90% of the miRNA compartment of each population comprising <75 miRNAs; however, each cell type had a unique miRNA signature. Integration of miRNA expression with chromatin accessibility revealed putative regulatory elements for differentially expressed miRNAs, including miR-21a, miR-146a and miR-223. The integrated maps suggest that many miRNAs utilize multiple promoters to reach high abundance and identified dominant and divergent miRNA regulatory elements between lineages and during development that may be used by clustered miRNAs, such as miR-99a/let-7c/miR-125b, to achieve distinct expression. These studies, with web-accessible data, help delineate the cis-regulatory elements controlling miRNA signatures of the immune system.

Post-Traumatic Growth among Patients after Living and Cadaveric Donor Kidney Transplantation: The Role of Resilience and Alexithymia.

The aim of this study was to determine the role of resilience and alexithymia in the post-traumatic growth as a response to extreme stress in patients after kidney transplantation and to determine whether there are differences in the level of posttraumatic growth in patients after living and cadaveric donor kidney transplantation. The relationships between these variables were also evaluated. The questionnaire survey of 91 kidney recipients took place in 2018 and 2019. The following tools were used: authorial post-transplant questionnaire for recipients and validated questionnaires, Post Traumatic Growth Inventory (PTGI-R), Resilience Coping Scale Questionnaire, and Toronto Alexithymia Scale Questionnaire (TAS20). The results obtained showed significant differences between the group of kidney recipients from living donors and recipients from cadaveric donors, in terms of overall post-traumatic growth, as well as changes in self-perception and a greater appreciation for life. Post-traumatic growth in both groups was related to the level of resilience and the level of alexithymia. Resilience is an accurate predictor of posttraumatic growth in general and for each of the groups of recipients separately.

A Critical Role for Fas-Mediated Off-Target Tumor Killing in T-cell Immunotherapy.

T cell-based therapies have induced cancer remissions, though most tumors ultimately progress, reflecting inherent or acquired resistance including antigen escape. Better understanding of how T cells eliminate tumors will help decipher resistance mechanisms. We used a CRISPR/Cas9 screen and identified a necessary role for Fas-FasL in antigen-specific T-cell killing. We also found that Fas-FasL mediated off-target "bystander" killing of antigen-negative tumor cells. This localized bystander cytotoxicity enhanced clearance of antigen-heterogeneous tumors in vivo, a finding that has not been shown previously. Fas-mediated on-target and bystander killing was reproduced in chimeric antigen receptor (CAR-T) and bispecific antibody T-cell models and was augmented by inhibiting regulators of Fas signaling. Tumoral FAS expression alone predicted survival of CAR-T-treated patients in a large clinical trial (NCT02348216). These data suggest strategies to prevent immune escape by targeting both the antigen expression of most tumor cells and the geography of antigen-loss variants. SIGNIFICANCE: This study demonstrates the first report of in vivo Fas-dependent bystander killing of antigen-negative tumors by T cells, a phenomenon that may be contributing to the high response rates of antigen-directed immunotherapies despite tumoral heterogeneity. Small molecules that target the Fas pathway may potentiate this mechanism to prevent cancer relapse.This article is highlighted in the In This Issue feature, p. 521.

Toxicity of selected airborne nitrophenols on eukaryotic cell membrane models.

Nitroaromatics belong to the group of toxic components of aerosol particles and atmospheric hydrometeors that enter the atmosphere through biomass burning and fuel combustion. In the present work, we report on the cytotoxic effects of a 2-, 3- and 4-nitrophenol mixture on a model eukaryotic-like cell membrane and compared it with in vitro cellular models BEAS-2B (immortalized bronchial epithelial cells) and A549 (cancerous alveolar epithelial cells). A selected model biomembrane comprised of DMPC (1,2-dimyristoyl-sn-glycero-3-phosphocholine), DPPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine) and POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) was studied. The electrochemical-based method, combined with atomic force microscopy (AFM) and phase-contrast microscopy imaging, allowed to get insights into the mechanism of cellular function disruption caused by airborne nitrophenols. The efficacy of the method is supported by the data obtained from in vitro experiments performed on cell models. The nitrophenol mixture exhibited cytotoxic effects at concentrations above 100 µg mL-1, as demonstrated by phase-contrast microscopy in real lung cell lines. Electrochemical impedance spectroscopy (EIS) revealed the formation of membrane defects at a nitrophenol concentration of 200 µg mL-1. AFM imaging confirmed the model membrane disintegration and phospholipids rearrangement in the presence of nitrophenols. These observations indicate that particle-bound nitrophenols induce substantial changes in cell membranes and make them more permeable to aerosol, resulting in major cellular damage in the lungs when inhaled. The study provides initial evidence of cellular membrane damage induced by three important nitrated phenols present in the environment.

CSF-1 controls cerebellar microglia and is required for motor function and social interaction.

Microglia, the brain resident macrophages, critically shape forebrain neuronal circuits. However, their precise function in the cerebellum is unknown. Here we show that human and mouse cerebellar microglia express a unique molecular program distinct from forebrain microglia. Cerebellar microglial identity was driven by the CSF-1R ligand CSF-1, independently of the alternate CSF-1R ligand, IL-34. Accordingly, CSF-1 depletion from Nestin+ cells led to severe depletion and transcriptional alterations of cerebellar microglia, while microglia in the forebrain remained intact. Strikingly, CSF-1 deficiency and alteration of cerebellar microglia were associated with reduced Purkinje cells, altered neuronal function, and defects in motor learning and social novelty interactions. These findings reveal a novel CSF-1-CSF-1R signaling-mediated mechanism that contributes to motor function and social behavior.

Towards High-performance Materials Based on Carbohydrate-Derived Polyamide Blends.

A bio-derived monomer called 2,3:4,5-di-O-isopropylidene-galactarate acid/ester (GalXMe) has great potential in polymer production. The unique properties of this molecule, such as its rigidity and bulkiness, contribute to the good thermal properties and appealing transparency of the material. The main problem, however, is that like other biobased materials, the polymers derived thereof are very brittle. In this study, we report on the melt blending of GalXMe polyamides (PAs) with different commercial PA grades using extrusion as well as blend characterization. Biobased PA blends showed limited to no miscibility with other polyamides. However, their incorporation resulted in strong materials with high Young moduli. The increase in modulus of the prepared GalXMe blends with commercial PAs ranged from up to 75% for blends with aliphatic polyamide composed of 1,6-diaminohexane and 1,12-dodecanedioic acid PA(6,12) to up to 82% for blends with cycloaliphatic polyamide composed of 4,4'-methylenebis(cyclohexylamine) and 1,12-dodecanedioic acid PA(PACM,12). Investigation into the mechanism of blending revealed that for some polyamides a transamidation reaction improved the blend compatibility. The thermal stability of the biobased PAs depended on which diamine was used. Polymers with aliphatic/aromatic or alicyclic diamines showed no degradation, whereas with fully aromatic diamines such as p-phenylenediamine, some degradation processes were observed under extrusion conditions (260/270 °C).

The cis-Regulatory Atlas of the Mouse Immune System.

A complete chart of cis-regulatory elements and their dynamic activity is necessary to understand the transcriptional basis of differentiation and function of an organ system. We generated matched epigenome and transcriptome measurements in 86 primary cell types that span the mouse immune system and its differentiation cascades. This breadth of data enable variance components analysis that suggests that genes fall into two distinct classes, controlled by either enhancer- or promoter-driven logic, and multiple regression that connects genes to the enhancers that regulate them. Relating transcription factor (TF) expression to the genome-wide accessibility of their binding motifs classifies them as predominantly openers or closers of local chromatin accessibility, pinpointing specific cis-regulatory elements where binding of given TFs is likely functionally relevant, validated by chromatin immunoprecipitation sequencing (ChIP-seq). Overall, this cis-regulatory atlas provides a trove of information on transcriptional regulation through immune differentiation and a foundational scaffold to define key regulatory events throughout the immunological genome.

Protein Barcodes Enable High-Dimensional Single-Cell CRISPR Screens.

CRISPR pools are being widely employed to identify gene functions. However, current technology, which utilizes DNA as barcodes, permits limited phenotyping and bulk-cell resolution. To enable novel screening capabilities, we developed a barcoding system operating at the protein level. We synthesized modules encoding triplet combinations of linear epitopes to generate >100 unique protein barcodes (Pro-Codes). Pro-Code-expressing vectors were introduced into cells and analyzed by CyTOF mass cytometry. Using just 14 antibodies, we detected 364 Pro-Code populations; establishing the largest set of protein-based reporters. By pairing each Pro-Code with a different CRISPR, we simultaneously analyzed multiple phenotypic markers, including phospho-signaling, on dozens of knockouts. Pro-Code/CRISPR screens found two interferon-stimulated genes, the immunoproteasome component Psmb8 and a chaperone Rtp4, are important for antigen-dependent immune editing of cancer cells and identified Socs1 as a negative regulator of Pd-l1. The Pro-Code technology enables simultaneous high-dimensional protein-level phenotyping of 100s of genes with single-cell resolution.

Solvent-Free Method for the Copolymerization of Labile Sugar-Derived Building Blocks into Polyamides.

This research focuses on the preparation of biobased copolyamides containing biacetalized galactaric acid (GalX), namely, 2,3:4,5-di-O-isopropylidene-galactaric acid (GalXMe) and 2,3:4,5-di-O-methylene-galactaric acid (GalXH), in bulk by melt polycondensation of salt monomers. In order to allow the incorporation of temperature-sensitive sugar-derived building blocks into copolyamides at temperatures below the degradation temperature of the monomers and below their melting temperatures, a clever selection of salt monomers is required, such that the sugar-derived salt monomer dissolves in the other salt monomers. The polymerization was investigated by temperature dependent FT-IR and optical microscopy. The structure of the obtained copolyamides was elucidated by NMR and matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) techniques. The positive outcome of this modified polycondensation method depends on the solubility of sugar-derived polyamide salts in polyamide salts of comonomers and the difference between their melting temperatures, however does not depend on the melting temperature of the used sugar-derived monomer. A variety of comonomers was screened in order to establish the underlying mechanisms of the process.

SIRT6 haploinsufficiency induces BRAFV600E melanoma cell resistance to MAPK inhibitors via IGF signalling.

While multiple mechanisms of BRAFV600-mutant melanoma resistance to targeted MAPK signaling inhibitors (MAPKi) have been reported, the epigenetic regulation of this process remains undetermined. Here, using a CRISPR-Cas9 screen targeting chromatin regulators, we discover that haploinsufficiency of the histone deacetylase SIRT6 allows melanoma cell persistence in the presence of MAPKi. Haploinsufficiency, but not complete loss of SIRT6 promotes IGFBP2 expression via increased chromatin accessibility, H3K56 acetylation at the IGFBP2 locus, and consequent activation of the IGF-1 receptor (IGF-1R) and downstream AKT signaling. Combining a clinically applicable IGF-1Ri with BRAFi overcomes resistance of SIRT6 haploinsufficient melanoma cells in vitro and in vivo. Using matched melanoma samples derived from patients receiving dabrafenib + trametinib, we identify IGFBP2 as a potential biomarker for MAPKi resistance. Our study has not only identified an epigenetic mechanism of drug resistance, but also provides insights into a combinatorial therapy that may overcome resistance to standard-of-care therapy for BRAFV600-mutant melanoma patients.

Structure-Property Relations in New Cyclic Galactaric Acid Derived Monomers and Polymers Therefrom: Possibilities and Challenges.

In order to fully exploit the potential of carbohydrate-based monomers, different (and some new) functionalities are introduced on galactaric acid via acetalization, and subsequently, partially-biobased polyamides are prepared therefrom via polycondensation in the melt. Compared to nonsubstituted linear monomer, faster advancement of the reaction is observed for the different biacetal derivatives of galactaric acid. This kinetic observation is of great significance since it allows conducting a polymerization reaction at lower temperatures than normally expected for polyamides, which allows overcoming typical challenges (e.g., thermal degradation) encountered upon polymerization of carbohydrate-derived monomers in the melt. The polymers derived from the modified galactaric acid monomers vary in terms of glass transition temperature, thermal stability, hydrophilicity, and functionality.

Assessment of knowledge about the effects of UV radiation on health behaviors associated with sunbathing in gymnasium students.

INTRODUCTION: The health behaviors or health-related behaviors is behavior (or activity) that are part of everyday life, affecting the health of the individual. An example of the behavior of health is also sunbathing, or exposing the body to excessive solar radiation dosage. It may be positive and negative effects on health. AIM: Evaluation of knowledge about gymnasium students. The impact of UV radiation on health and health behaviors associated with sunbathing. MATERIAL AND METHODS: The study was a diagnostic survey, with author's questionnaire. The sample was comprised students from classes II and III. A total of 312 questionnaires were collected among 181 girls and 131 boys. Used purposeful sampling. Results were considered statistically significant at p < 0.05. RESULTS: There is a statistically significant relationship between the stated sunbathing to get a sun tan, and sex of the respondent (p = 0.0002). Definitely more girls (77.35%) admit that tans in the sun to get a tan as compared to boys (58.02%). CONCLUSION: It is recommended that further research aimed at checking the causes and incidence of sunburn among young people. Consideration should be given to create and implement the appropriate health programs taking about tanning that could be implemented under the School Health Promotion Program, or to supplement this knowledge on subjects such as Biology or Nature.

Mapping the effects of drugs on the immune system.

Understanding how drugs affect the immune system has consequences for treating disease and minimizing unwanted side effects. Here we present an integrative computational approach for predicting interactions between drugs and immune cells in a system-wide manner. The approach matches gene sets between transcriptional signatures to determine their similarity. We apply the method to model the interactions between 1,309 drugs and 221 immune cell types and predict 69,995 interactions. The resulting immune-cell pharmacology map is used to predict how five drugs influence four immune cell types in humans and mice. To validate the predictions, we analyzed patient records and examined cell population changes from in vivo experiments. Our method offers a tool for screening thousands of interactions to identify relationships between drugs and the immune system.

Rational design of small molecules targeting the C2 domain of coagulation factor VIII.

The C domains of coagulation factors V (FV) and VIII (FVIII) are structurally conserved domains and share a common and essential function in membrane binding. In vivo regulation of thrombin formation strongly depends on the expression and regulation of the cofactor activities of FVIII and FV. With this study, we explored the possibility of inhibition of thrombin formation in full blood with small druglike molecules. Such compounds may serve as lead molecules for the development of a new type of orally available coagulation inhibitors that act by blocking the interaction between the C domains of FVIII and the membrane surface. We identified 9 novel molecules that are able to inhibit binding of the FVIII C2 domain to a model membrane by application of a combined ligand-based and target structure-based virtual screening approach that took into account the knowledge of a set of previously identified low-molecular-weight FVIII binders that were, however, not active in full blood. The half-maximal inhibitory concentration values of our newly identified compounds varied from 2.1 to 19.9 µM, of which 7 of 9 molecules did not appreciably inhibit FV membrane binding and were thus specific for FVIII. The most active bioactive compound showed activity in both plasma and in full blood.

Limited promiscuity of HLA-DRB1 presented peptides derived of blood coagulation factor VIII.

The formation of inhibitory antibodies directed against coagulation factor VIII (FVIII) is a severe complication in the treatment of hemophilia A patients. The induction of anti-FVIII antibodies is a CD4(+) T cell-dependent process. Activation of FVIII-specific CD4(+) T cells is dependent on the presentation of FVIII-derived peptides on MHC class II by antigen-presenting cells. Previously, we have shown that FVIII-pulsed human monocyte-derived dendritic cells can present peptides from several FVIII domains. In this study we show that FVIII peptides are presented on immature as well as mature dendritic cells. In immature dendritic cells half of the FVIII-loaded MHC class II molecules are retained within the cell, whereas in LPS-matured dendritic cells the majority of MHC class II/peptide complexes is present on the plasma membrane. Time-course studies revealed that presentation of FVIII-derived peptides was optimal between 12 and 24 hours after maturation but persisted for at least 96 hours. We also show that macrophages are able to internalize FVIII as efficiently as dendritic cells, however FVIII was presented on MHC class II with a lower efficiency and with different epitopes compared to dendritic cells. In total, 48 FVIII core-peptides were identified using a DCs derived of 8 different donors. Five HLA-promiscuous FVIII peptide regions were found - these were presented by at least 4 out of 8 donors. The remaining 42 peptide core regions in FVIII were presented by DCs derived from a single (30 peptides) or two to three donors (12 peptides). Overall, our findings show that a broad repertoire of FVIII peptides can be presented on HLA-DR.