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Four pinaceae pine resins analyzed in this study: black pine, shore pine, Baltic amber, and rosin demonstrate excellent dielectric properties, outstanding film forming, and ease of processability from ethyl alcohol solutions. Their trap-free nature allows fabrication of virtually hysteresis-free organic field effect transistors operating in a low voltage window with excellent stability under bias stress. Such green constituents represent an excellent choice of materials for applications targeting biocompatibility and biodegradability of electronics and sensors, within the overall effort of sustainable electronics development and environmental friendliness., Competing Interests: The authors declare no conflict of interest., (© 2023 The Authors. Global Challenges published by Wiley‐VCH GmbH.)

Lignin constitutes an impressive resource of high-value low molecular weight compounds. However, robust methods for isolation of the extractable fraction from lignocellulose are yet to be established. In this study, supercritical fluid extraction (SFE) and CO 2 -expanded liquid extraction (CXLE) were employed to extract lignin from softwood and hardwood chips. Ethanol, acetone, and ethyl lactate were investigated as green organic co-solvents in the extractions. Additionally, the effects of temperature, CO 2 percentage and the water content of the co-solvent were investigated using a design of experiment approach employing full factorial designs. Ethyl lactate and acetone provided the highest gravimetric yields. The water content in the extraction mixture had the main impact on the amount of extractable lignin monomers (LMs) and lignin oligomers (LOs) while the type of organic solvent was of minor importance. The most effective extraction was achieved by using a combination of liquid CO 2 /acetone/water (10/72/18, v/v/v) at 60 °C, 350 bar, 30 min and 2 mL min -1 flow rate. The optimized method provided detection of 13 LMs and 6 lignin dimers (LDs) from the hardwood chips. The results demonstrate the potential of supercritical fluids and green solvents in the field of mild and bening lignin extraction from wood., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

The analysis of archeological artifacts, due to the high value of antique objects, is preferably performed by nondestructive, noninvasive, and in situ techniques. At present, the most common in situ protocols used for the analysis of organic materials are spectroscopic approaches. In this work, we tested selected-ion flow tube-mass spectrometry (SIFT-MS), a transportable mass spectrometry system for the characterization and discrimination of natural resins by the analysis of their volatile organic compounds profiles. We chose diterpenoid, triterpenoid, and aromatic resins as reference materials, focusing on the most identified in archeological artifacts. This work aims to create a SIFT-MS database of mass spectra suitable for characterizing archeological samples. The spectral data obtained by SIFT-MS were interpreted with the aid of chromatograms and mass spectra obtained by head space-gas chromatography/mass spectrometry (HS-GC/MS). Finally, principal components analysis (PCA) was used to further underline the differences among the different materials and to investigate the possibility of discriminating different classes of resins based on their SIFT spectra.

This paper presents a study on the pyrolytic behavior of mixtures of lignocellulosic biomass with hydrocarbon plastics using analytical pyrolysis-GC/MS. Semi-quantitative analysis using chromatographic peak areas was used to investigate the composition of the pyrolysis oils and to highlight the occurrence of synergistic effects. A new method is also proposed to estimate the elemental composition of the pyrolysis oil based on the peak areas and brute formulas of the pyrolysis products. The results indicate that synergistic effects during co-pyrolysis favor secondary pyrolysis of holocellulose and polystyrene oligomers, and hinder radical chain-scission of polyethylene chains. H/C and O/C values of the pyrolysis oils were improved by the addition of plastic, indicating a decrease in the content of oxygenated pyrolysis products. The best performances were observed for the mixture containing 70% fir wood and 30% polyethylene, in which synergistic effects led to both an increase of H/C and a decrease of O/C., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Background: Antigen-specific neuroinflammation and neurodegeneration are characteristic for neuroimmunological diseases. In Parkinson's disease (PD) pathogenesis, α-synuclein is a known culprit. Evidence for α-synuclein-specific T cell responses was recently obtained in PD. Still, a causative link between these α-synuclein responses and dopaminergic neurodegeneration had been lacking. We thus addressed the functional relevance of α-synuclein-specific immune responses in PD in a mouse model., Methods: We utilized a mouse model of PD in which an Adeno-associated Vector 1/2 serotype (AAV1/2) expressing human mutated A53T-α-Synuclein was stereotactically injected into the substantia nigra (SN) of either wildtype C57BL/6 or Recombination-activating gene 1 (RAG1) -/- mice. Brain, spleen, and lymph node tissues from different time points following injection were then analyzed via FACS, cytokine bead assay, immunohistochemistry and RNA-sequencing to determine the role of T cells and inflammation in this model. Bone marrow transfer from either CD4 + /CD8 - , CD4 - /CD8 + , or CD4 + /CD8 + (JHD -/- ) mice into the RAG-1 -/- mice was also employed. In addition to the in vivo studies, a newly developed A53T-α-synuclein-expressing neuronal cell culture/immune cell assay was utilized., Results: AAV-based overexpression of pathogenic human A53T-α-synuclein in dopaminergic neurons of the SN stimulated T cell infiltration. RNA-sequencing of immune cells from PD mouse brains confirmed a pro-inflammatory gene profile. T cell responses were directed against A53T-α-synuclein-peptides in the vicinity of position 53 (68-78) and surrounding the pathogenically relevant S129 (120-134). T cells were required for α-synuclein-induced neurodegeneration in vivo and in vitro, while B cell deficiency did not protect from dopaminergic neurodegeneration., Conclusions: Using T cell and/or B cell deficient mice and a newly developed A53T-α-synuclein-expressing neuronal cell culture/immune cell assay, we confirmed in vivo and in vitro that pathogenic α-synuclein peptide-specific T cell responses can cause dopaminergic neurodegeneration and thereby contribute to PD-like pathology., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Peru Balsam, a resinous substance derived from Myroxylon balsamum var. pereirae , has historically been used as a topical ointment for various skin conditions such as scabies, poorly healing wounds, eczema, and haemorrhoids. The ingredients responsible for these properties are not fully elucidated. We investigated the chemical composition of two Peru Balsam samples, one historical and one modern, using gas chromatography/mass spectrometry to identify the active ingredients responsible for its pharmaceutical properties. Both Peru Balsam specimens investigated had similar compositions, showing the stability of the substance. Components identified are effective against scabies, exhibit antimicrobial activity and aid skin penetration. These properties are consistent with historical uses of Peru Balsam. Several ingredients are also known allergens. This study, combining chemical information with scientific literature related to pharmaceutical properties of natural substances, represents a breakthrough in the elucidation of active ingredients in Peru Balsam.

Modern humans expanded into Eurasia more than 40,000 years ago following their dispersal out of Africa. These Eurasians carried ~2-3% Neanderthal ancestry in their genomes, originating from admixture with Neanderthals that took place sometime between 50,000 and 60,000 years ago, probably in the Middle East. In Europe, the modern human expansion preceded the disappearance of Neanderthals from the fossil record by 3,000-5,000 years. The genetic makeup of the first Europeans who colonized the continent more than 40,000 years ago remains poorly understood since few specimens have been studied. Here, we analyse a genome generated from the skull of a female individual from Zlatý kůň, Czechia. We found that she belonged to a population that appears to have contributed genetically neither to later Europeans nor to Asians. Her genome carries ~3% Neanderthal ancestry, similar to those of other Upper Palaeolithic hunter-gatherers. However, the lengths of the Neanderthal segments are longer than those observed in the currently oldest modern human genome of the ~45,000-year-old Ust'-Ishim individual from Siberia, suggesting that this individual from Zlatý kůň is one of the earliest Eurasian inhabitants following the expansion out of Africa.

Myeloid-derived suppressor cells (MDSCs) represent a major population controlling T cell immune responses. However, little is known about their molecular requirements for homing and T cell interaction to mediate suppression. Here, we investigated the functional role of the homing and collagen IV receptor VLA-1 (α1β1-integrin) on in vitro GM-CSF generated murine MDSCs from wild-type (WT) and CD49a/α1-integrin ( Itga1 -/- ) gene-deficient mice. Here, we found that effector (Teff) but not naive (Tn) CD4 + T cells express VLA-1 and monocytes further up-regulated their expression after culture in GM-CSF when they differentiated into the monocytic subset of resting MDSCs (R-MDSCs). Subsequent activation of R-MDSCs by LPS+IFN-γ (A-MDSCs) showed increased in vitro suppressor potential, which was independent of VLA-1. Surprisingly, VLA-1 deficiency did not influence A-MDSC motility or migration on collagen IV in vitro . However, interaction times of Itga1 -/- A-MDSCs with Teff were shorter than with WT A-MDSCs on collagen IV but not on fibronectin substrate in vitro . After injection, A-MDSCs homed to the splenic red pulp where they co-localized with Teff and showed immediate suppression already after 6 h as shown by inhibition of T cell proliferation and induction of apoptosis. Injection of A-MDSCs from Itga 1 -/- mice showed equivalent homing into the spleen but a reduced suppressive effect. Interaction studies of A-MDSCs with Teff in the subcapsular red pulp with intravital two-photon microscopy revealed also here that MDSC motility and migration parameters were not altered by VLA-1 deficiency, but the interaction times with Teff were reduced. Together, our data point to a new role of VLA-1 adhesion to collagen IV as a prerequisite for extended contact times with Teff required for suppression., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Eckert, Ribechini, Jarick, Strozniak, Potter, Beilhack and Lutz.)

Myeloid-derived suppressor cells (MDSC) appear at relatively low frequencies in diseased organs such as tumors or infection sites, but accumulate systemically in the spleen. So far MDSC have been reported in humans and experimental animals such as mice, rats, and nonhuman primates. Therefore, methods to generate MDSC in large amounts in vitro can serve as an additional tool to study their biology. Here, we describe in detail the generation of murine MDSC with GM-CSF from bone marrow (BM). Both subsets of granulocytic (G-MDSC) and monocytic MDSC (M-MDSC) are generated by this cytokine. We provide panels of phenotypic markers to distinguish them from non-suppressive cells and define developmental stages of monocytes developing into M-MDSC by two subsequent steps in vitro.

This work presents a method to characterize high molecular esters in beeswax and resinous substances based on the use of microwave-assisted extraction and flow injection analysis-high resolution mass spectrometry that combines the high efficiency of the extraction procedure with the advantages of high resolution mass spectrometry. This approach allows us to identify archaeological beeswax and plant resinous substances by the characterization of the survived intact high molecular weight components. By this way, several raw materials (beeswax, pine resin and pitch, and resin extracted from Euphorbia tirucalli) were studied and used as reference substances. The procedure was then tested on an adhesive dated 44-42 ka BP recovered from Border Cave (KwaZulu-Natal, Africa), allowing us to detect the high molecular weight species even after almost 50,000 years, and then used to chemically investigate unknown archaeological adhesives from Antinoopolis (Egypt), dated to the 4th-5th century AD. The results allowed us to extend our knowledge on the long-term behavior of beeswax and resinous substances., (Copyright © 2020 Elsevier B.V. All rights reserved.)

A fast non-destructive approach based on the use of portable selected ion flow tube-mass spectrometry (SIFT-MS) was used for the first time to characterize organic materials in archaeological artifacts. The high sensitivity, specificity and selectivity SIFT soft chemical ionization mass spectrometry enabled us to investigate the composition of organic residues collected from ancient Egyptian findings in order to demonstrate the robustness of the techniques with different matrices. In addition, we tested SIFT-MS directly on an archaeological Egyptian amphora to prove its suitability as a completely non-invasive technique. Parallel investigations on all the samples were performed by GC/MS analysis to correlate and confirm the data obtained by SIFT-MS. The possibility of using a portable mass spectrometer on an excavation site or in a museum would be a significant step forward in the non-invasive analysis of organic archaeological materials, enabling archeologists and conservators to obtain real-time information on the molecular composition of organic residues., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Monocytic myeloid-derived suppressor cells (M-MDSCs) and granulocytic MDSCs (G-MDSCs) have been found to be massively induced in TB patients as well in murine Mtb infection models. However, the interaction of mycobacteria with MDSCs and its role in TB infection is not well studied. Here, we investigated the role of Cav-1 for MDSCs infected with Mycobacterium bovis Bacille-Calmette-Guerín (BCG). MDSCs that were generated from murine bone marrow (MDSCs) of wild-type (WT) or Cav1 -/- mice upregulated Cav-1, TLR4 and TLR2 expression after BCG infection on the cell surface. However, Cav-1 deficiency resulted in a selective defect of intracellular TLR2 levels predominantly in the M-MDSC subset. Further analysis indicated no difference in the phagocytosis of BCG by M-MDSCs from WT and Cav1 -/- mice or caveosome formation, but a reduced capacity to up-regulate surface markers, to secrete various cytokines, to induce iNOS and NO production required for suppression of T cell proliferation, whereas Arg-1 was not affected. Among the signaling pathways affected by Cav-1 deficiency, we found lower phosphorylation of the p38 mitogen-activated protein kinase (MAPK). Together, our findings implicate that (i) Cav-1 is dispensable for the internalization of BCG, (ii) vesicular TLR2 signaling in M-MDSCs is a major signaling pathway induced by BCG, (iii) vesicular TLR2 signals are controlled by Cav-1, (iv) vesicular TLR2/Cav-1 signaling is required for T cell suppressor functions., (Copyright © 2019 John, Kotze, Ribechini, Walzl, Du Plessis and Lutz.)

Olive mill wastewaters (OMWs) are important by-products of olive oil production. The wide availability and the forbidding economic costs of OMWs disposal have generated interest in their possible exploitation as a sustainable source of polyphenols. The development and optimisation of improved analytical methods for the detailed characterisation of polyphenol molecular profiles after production and during storage conditions are thus required. A new analytical method based on reverse-phase HPLC separation of polyphenols in OMWs on an embedded-polar group (EPG) stationary phase is proposed as an alternative to conventional C18 columns. The procedure was used for the quantitation of 11 polyphenols, using diode-array detection (DAD), and achieving quantitation limits equal to or lower than 0.1 µg g -1 . The same chromatographic set-up, coupled with high resolution tandem mass spectrometry (HPLC/ESI-Q-ToF), allowed us to identify hydroxytyrosyl esters of elenolic acid, whose relative abundance is proposed for monitoring the ageing of OMWs during storage.

Bone is one of the main sample types used for building chronologies in archaeology. It is also used in other research areas such as palaeodiet and palaeoenvironmental studies. However, for results to be accurate, samples must be free of exogenous carbon. Contamination can originate from a wide range of sources in the post-depositional environment but may also occur during excavation and post excavation activities (i.e. with the application of conservation materials) or during laboratory handling. Efficient procedures to remove contamination are therefore crucial prior to radiocarbon or stable isotope measurements. This work describes the development of an innovative sample pretreatment for bones, based on using supercritical CO2, which shows unique solvation properties. The effectiveness of supercritical fluid extraction (SFE) to remove conservation materials was compared with that obtained when applying a routine extraction based on the use of organic solvents (methanol, acetone and chloroform). The chemical composition of the bone samples before and after the two pre-treatments was then investigated using analytical pyrolysis-based techniques: EGA-MS (Evolved Gas Analysis-Mass Spectrometry) and Py-GC/MS (Pyrolysis-Gas Chromatography coupled with Mass Spectrometry). Collagen samples extracted from the same bones, prepared with the two cleaning protocols, were also radiocarbon dated by Accelerator Mass Spectrometry (AMS). The results of this study show that SFE is an efficient alternative method because it was as effective as the established treatment protocol. It removes contaminants such as conservation materials from bone samples with a minimum of handling and can be used routinely in radiocarbon dating laboratories. This work also demonstrates that analytical pyrolysis is not only a very efficient method to identify contaminants in bones but also to assess the effectiveness of the pretreatment prior to the radiocarbon measurement of the samples.

Tuberculosis patients and mice infected with live Mycobacterium tuberculosis (Mtb) accumulate high numbers of myeloid-derived suppressor cells (MDSCs). Here, we hypothesized that also dead Mtb vaccines may induce MDSCs that could impair the efficacy of vaccination. We found that repeated injections of Mtb vaccines (heat-killed Mtb in Incomplete Freund's Adjuvant, like Montanide) but not single or control vaccines without Mtb strongly expanded CD11b+ myeloid cells in the spleen, that suppressed T cell proliferation and killing ex vivo. Dead Mtb vaccination induced the generation of CD11b+ Ly-6Chigh CD115+ iNOS/Nos2+ monocytic MDSCs (M-MDSCs) upon application of inflammatory or microbial activation signals. In vivo these M-MDSCs positioned strategically in the spleen by infiltrating the splenic bridging channels and white pulp areas. Notably, within 6 to 24 hours in a Nos2-dependent fashion they produced NO to rapidly kill conventional and plasmacytoid dendritic cells (cDCs, pDCs) while, surprisingly, sparing T cells in vivo. Thus, we demonstrate that Mtb vaccine induced M-MDSCs to not directly suppress T cell in vivo but, instead, M-MDSCs directly target DC subpopulations thereby indirectly suppressing effector T cell responses. Collectively, we demonstrate that Mtb booster vaccines induce M-MDSCs in the spleen that can be activated to kill DCs cautioning to thoroughly investigate MDSC formation in individuals after Mtb vaccination in clinical trials.

A skullcap found in the Salkhit Valley in northeast Mongolia is, to our knowledge, the only Pleistocene hominin fossil found in the country. It was initially described as an individual with possible archaic affinities, but its ancestry has been debated since the discovery. Here, we determine the age of the Salkhit skull by compound-specific radiocarbon dating of hydroxyproline to 34,950-33,900 Cal. BP (at 95% probability), placing the Salkhit individual in the Early Upper Paleolithic period. We reconstruct the complete mitochondrial genome (mtDNA) of the specimen. It falls within a group of modern human mtDNAs (haplogroup N) that is widespread in Eurasia today. The results now place the specimen into its proper chronometric and biological context and allow us to begin integrating it with other evidence for the human occupation of this region during the Paleolithic, as well as wider Pleistocene sequences across Eurasia.

A combined study of crystallinity index (CI), degree of polymerisation (DP) and thermal stability of cellulose was carried out for monitoring the effect of ball-milling. DP and CI are two fundamental quantities that describe the physico-chemical behaviour of cellulose. Milling is a common strategy to reduce cellulose crystallinity. In this work, four different commercially available celluloses were milled at 30, 60 and 120 min, and the changes in DP and CI were monitored using spectroscopic, diffraction and chromatographic techniques. Evolved gas analysis-mass spectrometry (EGA-MS) was also used to evaluate the changes in apparent activation energy (E a ) of the pyrolysis reaction upon different milling times by using model-free isoconversional methods. The results showed substantial decrease in CI values and moderate changes in DP after two-hours of ball-milling. E a were found in the range 110-140 kJ/mol, and were reduced by 10% on average after two hours of ball-milling., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Reactive pyrolysis is a technique that provides mechanistic information by performing pyrolysis of the substrate in a sealed glass capsule at elevated temperature and pressure for relatively long time. This technique has already shown great potential for the analysis of biomass, favouring the formation of only the most thermostable compounds. In this work, both fast and reactive pyrolysis with on-line gas chromatography-mass spectrometry analysis (Py-GC/MS) are used to study fructose, inulin and Jerusalem artichoke tubers (Heliantus tuberosus). Interesting differences were found between the two systems, and became even more evident as the reaction time was increased. The most striking result was the formation of di-fructose dianhydrides (DFAs), a class of compounds with interesting biological activities. DFAs were obtained in high yields from reactive pyrolysis, but not from fast pyrolysis. Hypotheses on the pyrolysis mechanisms were made based upon the composition of the pyrolysates. This work describes for the first time the behaviour of fructans under reactive pyrolysis., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The molecular characterization of organic materials in samples from artworks and historical objects traditionally entailed qualitative and quantitative analyses by HPLC and GC. Today innovative approaches based on analytical pyrolysis enable samples to be analysed without any chemical pre-treatment. Pyrolysis, which is often considered as a screening technique, shows previously unexplored potential thanks to recent instrumental developments. Organic materials that are macromolecular in nature, or undergo polymerization upon curing and ageing can now be better investigated. Most constituents of paint layers and archaeological organic substances contain major insoluble and chemically non-hydrolysable fractions that are inaccessible to GC or HPLC. To date, molecular scientific investigations of the organic constituents of artworks and historical objects have mostly focused on the minor constituents of the sample. This review presents recent advances in the qualitative and semi-quantitative analyses of organic materials in heritage objects based on analytical pyrolysis coupled with mass spectrometry., (© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The Mediterranean buckthorn, Rhamnus alaternus L., is a plant used in traditional medicine in Mediterranean countries. We aimed at characterizing its phenolic compounds and explore potential antihyperlipidemic activity of this plant. The profile of phenolic compounds in R. alaternus leaf crude methanolic extract (CME) and its liquid-liquid extraction-derived fractions were analyzed by high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC/ESI-MS 2 ). Effects of CME on: circulating lipids in rats with Triton WR-1339-induced hyperlipidemia, intracellular lipid accumulation and expression of genes of fatty acid metabolism in human hepatoma HepG2 cells, and adipogenesis in the 3T3-L1 murine adipocyte cell model were assessed. The HPLC/ESI-MS 2 analytical profile revealed a total of fifteen compounds, of which eleven were identified. Oral CME administration decreased blood levels of cholesterol and triacylglycerols in hyperlipidemic rats (by 60% and 70%, respectively, at 200 mg CME/kg). In HepG2 cells, CME exposure dose-dependently decreased intracellular lipids and up-regulated gene expression of carnitine palmitoyltransferase 1 involved in fatty acid oxidation. In the 3T3-L1 model, CME favored preadipocyte proliferation and adipogenesis, pointing to positive effects on adipose tissue expandability. These results suggest novel uses of R. alaternus by showing that its leaves are rich in flavonoids and flavonoid derivatives with an antihyperlipidemic effect in vivo and in hepatic cells., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Cereals were very important in ancient diets, however evidence from archaeological sites of the vessels used for processing or storing cereals is comparatively rare. Micro-organisms, as well as chemical-physical effects can easily degrade cereals during the burial period. This can lead to a complete cereal decay and to serious difficulties in estimating the intensity of use of the cereals by ancient populations. Here, we present a novel biomarker approach entailing the detection of secondary lipid metabolites produced by ergot fungi (genus Claviceps), which are common cereal pests. The aim was to identify the original presence of Gramineae and to indirectly establish if vessels were used for cereal storage/processing. The fatty acid and TAG-estolide profiles of the remains from more than 30 archaeological vessels were investigated by gas chromatography/mass spectrometry (GC/MS) and high performance liquid chromatography/high resolution mass spectrometry (HPLC/ESI-Q-ToF). The detection of lipids derived from ergot in archaeological and historic contexts rests on its complex chemistry, providing a unique and relatively recalcitrant chemical signature for cereals. This research demonstrated that the combination of our innovative biomarker approach along with environmental and archaeological evidence can provide unprecedented insights into the incidence of cereals and related processing activities in ancient societies.

For the first time, the exploitation of hazelnut shells for the combined production of levulinic acid (LA) and hydrochar was investigated. The optimization of the catalytic hydrothermal treatment was performed both in autoclave and microwave reactor, approaching a maximum LA yield of ∼9-12wt%. Hydrochars recovered with high yield (∼43-47wt%) were characterized by different techniques, including elemental and proximate analysis, heating value, FT-IR, XPS, XRD, SEM-EDX, and SAA. Their "lignite-like" energetic properties make them suitable for the energy recovery within the same biorefinery plant for LA production, thus partially offsetting the cost of the entire process. Alternatively, since the synthesized hydrochars maintain high levels of oxygenated groups, they could be smartly exploited as natural sorbents for environmental applications. The proposed integrated approach makes possible to fully exploit this waste, smartly closing its biorefinery cycle in a sustainable development perspective., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Thymus-derived natural Foxp3 + CD4 + regulatory T cells (nTregs) play a key role in maintaining immune tolerance and preventing autoimmune disease. Several studies indicate that dendritic cells (DCs) are critically involved in the maintenance and proliferation of nTregs. However, the mechanisms how DCs manage to keep the peripheral pool at constant levels remain poorly understood. Here, we describe that the NF-κB/Rel family transcription factor RelB controls the frequencies of steady-state migratory DCs (ssmDCs) in peripheral lymph nodes and their numbers control peripheral nTreg homeostasis. DC-specific RelB depletion was investigated in CD11c-Cre × RelB fl/fl mice (RelB DCko ), which showed normal frequencies of resident DCs in lymph nodes and spleen while the subsets of CD103 - Langerin - dermal DCs (dDCs) and Langerhans cells but not CD103 + Langerin + dDC of the ssmDCs in skin-draining lymph nodes were increased. Enhanced frequencies and proliferation rates were also observed for nTregs and a small population of CD4 + CD44 high CD25 low memory-like T cells (Tml). Interestingly, only the Tml but not DCs showed an increase in IL-2-producing capacity in lymph nodes of RelB DCko mice. Blocking of IL-2 in vivo reduced the frequency of nTregs but increased the Tml frequencies, followed by a recovery of nTregs. Taken together, by employing RelB DCko mice with increased frequencies of ssmDCs our data indicate a critical role for specific ssmDC subsets for the peripheral nTreg and IL-2 + Tml frequencies during homeostasis.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) controls proliferation and survival of myeloid cells including monocytes. Here, we describe a time-dependent licensing process driven by GM-CSF in murine Ly6C high and human CD14 + monocytes that disables their inflammatory functions and promotes their conversion into suppressor cells. This 2-step licensing of monocytes requires activation of the AKT/mTOR/mTORC1 signaling cascade by GM-CSF followed by signaling through the interferon-γ receptor (IFN-γR)/interferon regulatory factor-1 (IRF-1) pathway. Only licensing-dependent adaptations in Toll-like receptor/inflammasome, IFN-γR, and phosphatidylinositol 3-kinase/AKT/mTOR signaling lead to stabilized expression of inducible nitric oxide synthase by mouse and indoleamine 2,3-dioxygenase (IDO) by human monocytes, which accounts for their suppressor activity. This study suggests various myeloid cells with characteristics similar to those described for monocytic myeloid-derived suppressor cells, Mreg, or suppressor macrophages may arise from licensed monocytes. Markers of GM-CSF-driven monocyte licensing, including p -Akt, p -mTOR, and p -S6, distinguish inflammatory monocytes from potentially suppressive monocytes in peripheral blood of patients with high-grade glioma., Competing Interests: Conflict-of-interest disclosure: The authors declare no competing financial interests.

A novel analytical approach based on pyrolysis-gas chromatography coupled with mass spectrometry of carbohydrates with in situ silylation using hexamethyldisilazane is presented in this work for the first time. A micro reaction sampler was used to simultaneously achieve the pyrolyis reaction and facilitate the derivatization of pyrolysis products, by enabling the materials to react with the derivatizing agent in a sealed capsule at high temperature and pressure for long periods of time. This drastically increased the complete silylation of the pyrolysis products and the chromatographic resolution, resulting in less complex pyrograms and increased sensitivity toward the most stable compounds. Different results were obtained for glucose and cellulose in terms of predominant pyrolytic pathways. The formation of anhydrosugars was the preferential pyrolytic reaction for glucose, while the formation of cyclopentenones and small molecules was predominant for the pyrolysis of cellulose. Steric hindrance effects of polysaccharide chains on the efficiency of the derivatizing agent were hypothesized in order to explain the different results. A good reproducibility was found, with relative standard deviations not greater than 10%. Semiquantitative calculations showed that the partial silylation of anhydrosugars was almost completely overcome after 10 min of reactive pyrolysis. This work discloses a powerful and potentially widely applicable analytical method for the investigations of organic materials under controlled pyrolytic conditions, with the advantage of increasing the effectiveness of in situ derivatization.

Donor CD4(+)Foxp3(+) regulatory T cells (T reg cells) suppress graft-versus-host disease (GvHD) after allogeneic hematopoietic stem cell transplantation (HCT [allo-HCT]). Current clinical study protocols rely on the ex vivo expansion of donor T reg cells and their infusion in high numbers. In this study, we present a novel strategy for inhibiting GvHD that is based on the in vivo expansion of recipient T reg cells before allo-HCT, exploiting the crucial role of tumor necrosis factor receptor 2 (TNFR2) in T reg cell biology. Expanding radiation-resistant host T reg cells in recipient mice using a mouse TNFR2-selective agonist before allo-HCT significantly prolonged survival and reduced GvHD severity in a TNFR2- and T reg cell-dependent manner. The beneficial effects of transplanted T cells against leukemia cells and infectious pathogens remained unaffected. A corresponding human TNFR2-specific agonist expanded human T reg cells in vitro. These observations indicate the potential of our strategy to protect allo-HCT patients from acute GvHD by expanding T reg cells via selective TNFR2 activation in vivo., (© 2016 Chopra et al.)

Neutralization or deletion of tumor necrosis factor (TNF) causes loss of control of intracellular pathogens in mice and humans, but the underlying mechanisms are incompletely understood. Here, we found that TNF antagonized alternative activation of macrophages and dendritic cells by IL-4. TNF inhibited IL-4-induced arginase 1 (Arg1) expression by decreasing histone acetylation, without affecting STAT6 phosphorylation and nuclear translocation. In Leishmania major-infected C57BL/6 wild-type mice, type 2 nitric oxide (NO) synthase (NOS2) was detected in inflammatory dendritic cells or macrophages, some of which co-expressed Arg1. In TNF-deficient mice, Arg1 was hyperexpressed, causing an impaired production of NO in situ. A similar phenotype was seen in L. major-infected BALB/c mice. Arg1 deletion in hematopoietic cells protected these mice from an otherwise lethal disease, although their disease-mediating T cell response (Th2, Treg) was maintained. Thus, deletion or TNF-mediated restriction of Arg1 unleashes the production of NO by NOS2, which is critical for pathogen control., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)