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AIMS Primary mediastinal germ cell tumours (PMGCTs) are rare mediastinal neoplasms, and their diagnosis can be challenging, owing to small biopsy samples. The aim of this study was to develop a diagnostic algorithm using immunohistochemical staining, with a focus on novel markers, and molecular analysis of isochromosome 12p [i(12p)]. METHODS AND RESULTS Paraffin-embedded tissues of 32 mediastinal tumours were analysed with immunohistochemical staining for sal-like transcription factor 4 (SALL4), Lin-28 homologue A (LIN28), octamer-binding transcription factor 3/4 (OCT3/4), D2-40, cluster of differentiation 117 (CD117), sex-determining region Y-box 17, sex-determining region Y-box 2 (SOX2), cluster of differentiation 30, the β-subunit of human chorionic gonadotropin (β-hCG), GATA-binding protein 3 (GATA3), forkhead box protein A2 (FOXA2), glypican-3 (GPC3), α-fetoprotein (AFP), terminal deoxynucleotidyl transferase (TdT), nuclear protein of the testis (NUT), and pan-cytokeratin. Quantitative real-time polymerase chain reaction was performed to investigate the i(12p) status. Fifteen seminomas, seven teratomas, one yolk sac tumour, one choriocarcinoma and seven mixed PMGCTs were diagnosed. Each entity had different immunohistochemical staining patterns, which helped to distinguish them: OCT3/4, D2-40, CD117 and TdT for seminoma; OCT3/4 and SOX2 for embryonal carcinoma; FOXA2, GPC3 and AFP for yolk sac tumour; and β-hCG and GATA3 for choriocarcinoma. Mature teratomas stained positively for pan-cytokeratin in epithelial components and focally for SALL4, SOX2, GATA3, D2-40, and FOXA2. Furthermore, a NUT carcinoma mimicking a PMGCT was diagnosed, showing strong nuclear SOX2 staining and speckled nuclear NUT staining. i(12p) was detected in 24 of 27 PMGCTs (89%). CONCLUSION A diagnostic algorithm is of great importance for a reliable diagnosis of PMGCT in, usually small, tissue biopsy samples. Therefore, a combination of three to four antibodies to identify the correct histological subtype is usually necessary, in addition to morphological features. The i(12p) status serves as an additional option to indicate a germ cell origin in selected cases.

Inositol pyrophosphates (PP-InsPs) are highly phosphorylated molecules that have emerged as central nutrient messengers in eukaryotic organisms. They can bind to structurally diverse target proteins to regulate biological functions, such as protein-protein interactions. PP-InsPs are strongly negatively charged and interact with highly basic surface patches in proteins, making their quantitative biochemical analysis challenging. Here, we present the synthesis of biotinylated myo-inositol hexakisphosphates and their application in surface plasmon resonance and grating-coupled interferometry assays, to enable the rapid identification, validation, and kinetic characterization of InsP- and PP-InsP-protein interactions.

Energy dissipation within the respiratory system is a factor promoting ventilator-induced lung injury (VILI). In this animal study, we modulated the expiratory flow, reducing the energy dissipated in the system. However, this reduction happened mostly across the endotracheal tube, and only partly in the respiratory system. Therefore, in healthy lungs, the advantage in energy dissipation does not reduce VILI, but the advantages might be more relevant in diseased lungs under injurious ventilation.

Sulfur mustard (SM) is a banned chemical warfare agent recently used in the Syrian Arab Republic conflict causing erythema and blisters characterized by complicated and delayed wound healing. For medical and legal reasons, the proof of exposure to SM is of high toxicological and forensic relevance. SM reacts with endogenous human serum albumin (HSA adducts) alkylating the thiol group of the cysteine residue C34, thus causing the addition of the hydroxyethylthioethyl (HETE) moiety. Following proteolysis with pronase, the biomarker dipeptide C(-HETE)P is produced. To expand the possibilities for verification of exposure, we herein introduce a novel biomarker produced from that alkylated dipeptide by derivatization with propionic anhydride inducing the selective propionylation of the N-terminus yielding PA-C(-HETE)P. Quantitative derivatization is carried out at room temperature in aqueous buffer within 10 s. The biomarker was found to be stable in the autosampler at 15 °C for at least 24 h, thus documenting its suitability even for larger sets of samples. Selective and sensitive detection is done by micro liquid chromatography-electrospray ionization tandem-mass spectrometry (μLC-ESI MS/MS) operating in the selected reaction monitoring (SRM) mode detecting product ions of the single protonated PA-C(-HETE)P (m/z 379.1) at m/z 116.1, m/z 137.0, and m/z 105.0. The lower limit of detection corresponds to 32 nM SM in plasma in vitro and the limit of identification to 160 nM. The applicability to real exposure scenarios was proven by analyzing samples from the Middle East confirming poisoning with SM. Graphical abstract Deutsche Forschungsgemeinschaft http://dx.doi.org/10.13039/501100001659

Sulfur mustard (SM, bis[2-chloroethyl]-sulfide) is a banned chemical warfare agent that was frequently used in recent years and led to numerous poisoned victims who developed painful erythema and blisters. Post-exposure analysis of SM incorporation can be performed by the detection of human serum albumin (HSA)-derived peptides. HSA alkylated by SM contains a hydroxyethylthioethyl (HETE)-moiety bound to the cysteine residue C34 yielding the dipeptide biomarker C(-HETE)P after pronase-catalyzed proteolysis. We herein present a novel procedure for the selective precolumn nicotinylation of its N-terminus using 1-nicotinoyloxy-succinimide. The reaction was carried out for 2 h at ambient temperature with a yield of 81%. The derivative NA-C(-HETE)P was analyzed by micro liquid chromatography-electrospray ionization tandem-mass spectrometry working in the selected reaction monitoring mode (μLC-ESI MS/MS SRM). The derivative was shown to be stable in the autosampler at 15°C for at least 24 h. The single protonated precursor ion (m/z 428.1) was subjected to collision-induced dissociation yielding product ions at m/z 116.1, m/z 137.0, and m/z 105.0 used for selective monitoring without any plasma-derived interferences. NA-C(-HETE)P showed a mass spectrometric response superior to the non-derivatized dipeptide thus yielding larger peak areas (factor 1.3 ± 0.2). The lower limit of identification corresponded to 80 nM SM spiked to plasma in vitro. The presented procedure was applied to real case plasma samples from 2015 collected in the Middle East confirming SM poisoning.

The great variety of pathological patterns in germ cell tumours, especially in yolk sac tumours but also the possibility of somatic-type malignancies, can complicate daily diagnosis. For the correct diagnosis, knowledge of morphological aspects and additional immunohistochemical staining can be helpful. Also, rare entities like sex cord stromal tumours, tumours of testicular adnexa or mesenchymal tumours of the spermatic cord can be diagnostically challenging.Die große Vielfalt der histologischen Muster von Keimzelltumoren, insbesondere beim Dottersacktumor, aber auch das Phänomen der somatischen Malignität können die tägliche Diagnostik komplizieren. Verschiedene morphologische Aspekte und immunhistochemische Zusatzuntersuchungen können bei der Diagnosefindung hilfreich sein. Auch das Spektrum seltener Entitäten wie Gonadenstromatumoren, testikuläre Adnextumoren und Weichgewebstumoren des Samenstrangs können zu einer diagnostischen Herausforderung werden.

Purpose Melanocytic nevi in lymph nodes (NNs) are an important histological differential diagnosis of initial sentinel lymph node (SN) metastasis in melanoma. Our aim was to associate NN in SNs with clinicopathologic features and survival rates in 1, 250 patients with SN biopsy for melanoma. Methods To compare patients with present and absent NN, we used Fisher's exact test, Mann–Whitney U test, and multivariate logistic regression models in this retrospective observational study based on a prospectively maintained institutional database. Results NN prevalence in axillary, cervical, and groin SNs was 16.5%, 19.4%, and 9.8%, respectively. NN were observed in combination with all growth patterns of melanoma, but more frequently when the primary was histologically associated with a cutaneous nevus. We observed a decreasing NN prevalence with increasing SN metastasis diameter. Multiple logistic regression determined a significantly increased NN probability for SNs of the neck or axilla, for individuals with ≥ 50 cutaneous nevi, midline primary melanomas, and for individuals who reported non-cutaneous malignancies in their parents. Cancer in parents was also significantly more frequently reported by melanoma patients who had more than 50 cutaneous nevi. In SN-negative patients, NN indicated a tendency for slightly lower melanoma-specific survival. Conclusions We found a highly significant association between NN diagnosis and multiple cutaneous nevi and provided circumstantial evidence that cutaneous nevi in the drainage area of lymph nodes are particularly important. The trend toward lower melanoma-specific survival in SN-negative patients with NN suggests that careful differentiation of SN metastases is important.

Inositol pyrophosphates (PP-InsPs) are highly phosphorylated molecules that have emerged as central nutrient messengers in eukaryotic organisms. They can bind to structurally diverse target proteins to regulate biological functions, such as protein-protein interactions. PP-InsPs are strongly negatively charged and interact with highly basic surface patches in proteins, making their quantitative biochemical analysis challenging. Here, we present the synthesis of biotinylated

Tertiary amides, such as in N-acylated proline or N-methyl glycine residues, react rapidly with nitrate radicals (NO3˙) with absolute rate coefficients in the range of 4–7 × 108 M−1 s−1 in acetonitrile. The major pathway proceeds through oxidative electron transfer (ET) at nitrogen, whereas hydrogen abstraction is only a minor contributor under these conditions. However, steric hindrance at the amide, for example by alkyl side chains at the α-carbon, lowers the rate coefficient by up to 75%, indicating that NO3˙-induced oxidation of amide bonds proceeds through initial formation of a charge transfer complex. Furthermore, the rate of oxidative damage of proline and N-methyl glycine is significantly influenced by its position in a peptide. Thus, neighbouring peptide bonds, particularly in the N-direction, reduce the electron density at the tertiary amide, which slows down the rate of ET by up to one order of magnitude. The results from these model studies suggest that the susceptibility of proline residues in peptides to radical-induced oxidative damage should be considerably reduced, compared with the single amino acid.

An optical, electronic and structural characterisation of three natural dyes potentially interesting for application in organic solar cells, curcumin (C21H20O6), bixin (C25H30O4) and indigo (C16H10N2O2), was performed. X-Ray Diffraction (XRD) measurements, showed that curcumin has a higher degree of crystallinity compared to bixin and indigo. The results from the Pawley unit cell refinements for all dyes are reported. Optical absorption spectra measured by UV-Visible Spectroscopy (UV-Vis) on thermally evaporated films revealed that bixin undergoes chemical degradation upon evaporation, while curcumin and indigo appear to remain unaffected by this process. Combined Ultraviolet Photoemission Spectroscopy (UPS) and Inverse Photoemission Spectroscopy (IPES) spectra measured on the dyes revealed that all of them are hole-conducting materials and allowed for the determination of their electronic bandgaps, and Fermi level position within the gap. UV Photo-Emission Electron Microscopy (PEEM) revealed the workfunction of the dye materials and indicated that indigo has a negative electron affinity. PEEM was also used to study degradation by UV irradiation and showed that they are quite robust to UV exposure.

Aims Malignant germ cell tumours (GCTs) of the testis are rare neoplasms, but the most common solid malignancies in young men. World Health Organization guidelines divide GCTs into five types, for which numerous immunohistochemical markers allow exact histological subtyping in the majority of cases. In contrast, a germ cell origin is often hard to prove in metastatic GCTs that have developed so-called somatic malignant transformation. A high percentage, up to 89%, of GCTs are characterised by the appearance of isochromosome 12p [i(12p)]. Fluorescence in-situ hybridisation has been the most common diagnostic method for the detection of i(12p) so far, but has the disadvantages of being time-consuming, demanding, and not being a stand-alone method. The aim of the present study was to establish a quantitative real-time polymerase chain reaction assay as an independent method for detecting i(12p) and regional amplifications of the short arm of chromosome 12 by using DNA extracted from formalin-fixed paraffin-embedded tissue. Methods and results A cut-off value to distinguish between the presence and absence of i(12p) was established in a control set consisting of 36 tumour-free samples. In a training set of 149 GCT samples, i(12p) was detectable in 133 tumours (89%), but not in 16 tumours (11%). In a test set containing 27 primary and metastatic GCTs, all 16 tumours with metastatic spread and/or somatic malignant transformation were successfully identified by the detection of i(12p). Conclusion In summary, the qPCR assay presented here can help to identify, further characterise and assign a large proportion of histologically inconclusive malignancies to a GCT origin.

Organophosphorus (OP) nerve agents represent a class of highly toxic chemical warfare agents banned by the Chemical Weapons Convention. Nevertheless, in the past few years they have been used repeatedly for warfare, assassination and attempted murder. In addition, the chemically related OP pesticides were frequently used for suicide and may be deployed for terroristic attacks. Therefore, sensitive and selective bioanalytical methods are indispensable to investigate biological specimens as pieces of evidence to prove poisoning. OP agents form long-lived covalent reaction products (adducts) with endogenous proteins like human serum albumin (HSA) and butyrylcholinesterase (BChE). The adducted nonapeptide (NP) obtained by proteolysis of the BChE-adduct is one of the most sensitive and important biomarkers. We herein present a novel class of NP-adducts propionylated at its N-terminal phenylalanine residue (F195). The biomarker derivative is produced by addition of propionic anhydride to the NP-adduct inducing its quantitative conversion in aqueous buffer within 5 min at room temperature. Afterwards the mixture is directly analyzed by micro-liquid chromatography-electrospray ionization tandem-mass spectrometry (µLC-ESI MS/MS). The sensitivity of the method is comparable to that of the non-derivatized NP-adduct. These characteristics make the method highly beneficial for forensic analysis especially in cases in which the OP agent does not form adducts with HSA that are typically targeted as a second biomarker of exposure. This novel procedure was successfully applied to nerve agent-spiked samples sent by the Organisation for the Prohibition of Chemical Weapons (OPCW) as well as to plasma samples of real cases of pesticide poisoning.

Apart from the well-known sulfur mustard (SM), additional sulfur-containing blistering chemical warfare agents exist. Sesquimustard (Q) is one of them and five times more blistering than SM. It is a common impurity in mustard mixtures and regularly found in old munitions but can also be used in pure form. Compared to the extensive literature on SM, very little experimental data is available on Q and no protein biomarkers of exposure have been reported. We herein report for the first time the adduct of Q with the nucleophilic Cys34 residue of human serum albumin (HSA) formed in vitro and introduce two novel bioanalytical procedures for detection. After proteolysis of this HSA adduct catalyzed either by pronase or by proteinase K, two biomarkers were identified by high-resolution tandem mass spectrometry (MS/HR MS), namely a dipeptide and a tripeptide, both alkylated at their Cys residue, which we refer to as HETETE-CP and HETETE-CPF. HETETE represents the Q-derived thio-alkyl moiety bearing a terminal hydroxyl group: “hydroxyethylthioethylthioethyl.” Targeting both peptide markers from plasma, a micro liquid chromatography–electrospray ionization tandem mass spectrometry method working in the selected reaction monitoring mode (μLC-ESI MS/MS SRM) was developed and validated as well suited for the verification of exposure to Q. Fulfilling the quality criteria defined by the Organisation for the Prohibition of Chemical Weapons, the novel methods enable the detection of exposure to Q alone or in mixtures with SM. We further report on the relative reactivity of Q compared to SM. Based on experiments making use of partially deuterated Q as the alkylating agent, we rule out a major role for six-membered ring sulfonium ions as relevant reactive species in the alkylation of Cys34. Furthermore, the results of molecular dynamics simulations are indicative that the protein environment around Cys34 allows adduct formation with elongated but not bulky molecules such as Q, and identify important hydrogen bonding interactions and hydrophobic contacts. Graphical abstract Electronic supplementary material The online version of this article (10.1007/s00216-020-02917-w) contains supplementary material, which is available to authorized users.

Salt-affected farmlands are increasingly burdened by chlorides, carbonates, and sulfates of sodium, calcium, and magnesium. Intriguingly, the underlying physiological processes are studied almost always under NaCl stress. Two faba bean cultivars were subjected to low- and high-salt treatments of NaCl, Na2 SO4 , and KCl. Assimilation rate and leaf water vapor conductance were reduced to approximately 25-30% without biomass reduction after 7 days salt stress, but this did not cause severe carbon shortage. The equimolar treatments of Na+ , K+ , and Cl- showed comparable accumulation patterns in leaves and roots, except for SO4 2- which did not accumulate. To gain a detailed understanding of the effects caused by the tested ion combinations, we performed nontargeted gas chromatography-mass spectrometry-based metabolite profiling. Metabolic responses to various salts were in part highly linearly correlated, but only a few metabolite responses were common to all salts and in both cultivars. At high salt concentrations, only myo-inositol, allantoin, and glycerophosphoglycerol were highly significantly increased in roots under all tested conditions. We discovered several metabolic responses that were preferentially associated with the presence of Na+ , K+ , or Cl- . For example, increases of leaf proline and decreases of leaf fumaric acid and malic acid were apparently associated with Cl- accumulation.

Zusammenfassung Hintergrund Auf einer neonatologischen Intensivstation am Universitätsklinikum Leipzig wurden im Jahr 2016 vermehrt Kolonisationen mit Methicillin-sensiblen Staphylococcus aureus Stämmen festgestellt. Typisierungsergebnisse zeigten einen prädominierenden spa-Typ t091. Aufgrund der nosokomialen Häufungen wurden verschiedene Maßnahmen zur Infektionsprävention etabliert, z. B. intensive Barrieremaßnahmen, Einzelzimmer, Kohortierung, Umgebungsuntersuchungen auf S. aureus, Schulungen. Ein Teil dieses Maßnahmenbündels war die Durchführung eines Mitarbeiterscreenings, um kolonisierte Mitarbeiter als mögliches Reservoir zu sanieren. Methodik Mitarbeiter mit S. aureus-Nachweis führten in der eigenen Häuslichkeit eine 5-tägige Sanierung durch. Hierfür wurden Produkte mit dem Wirkstoff Octenidin verwendet. Kontrollabstriche erfolgten frühestens 48 Stunden nach Beendigung des Sanierungsintervalls (3 im Abstand von mind. 24 Stunden). Mitarbeiter mit 2 erfolglosen Sanierungsversuchen erhielten anstelle von Octenidin-, Mupirocin-haltige Nasensalbe. Ergebnisse Bei 128 untersuchten Mitarbeitern wurde in 43 Fällen (33,6%) S. aureus nachgewiesen, der in 9 Fällen (20,9%; 9/43) dem Typ t091 entsprach. Unter den S. aureus-positiven Mitarbeitern waren 2 MRSA-Nachweise (4,7%; 2/43) als Zufallsbefund zu verzeichnen. Die Erfolgsquote nach dem ersten Sanierungsversuch von t091 und MRSA lag bei 0%. In 3 Fällen lagen nach dem zweiten Versuch 3 negative Kontrollabstriche vor und 8 Mitarbeiter konnten erst unter Einsatz von Mupirocin-haltiger Nasensalbe saniert werden. Schlussfolgerungen Ursachen für die erschwerte Sanierung können vielfältig sein: Schwer umzusetzende Sanierung im eigenen Haushalt, Vorhandensein von dekolonisierungshemmenden Faktoren, nicht konsequente Durchführung aller Sanierungsmaßnahmen und eine mögliche Rekolonisation während der Arbeitszeit oder durch Familienangehörige. Weiterhin ist zu diskutieren, ob der Erfolg der Sanierung bei Verwendung von Mupirocin-Nasensalbe in der besseren Wirksamkeit von Mupirocin im Vergleich zu Octenidin begründet ist oder ob die unterschiedliche Konsistenz der Präparate ausschlaggebend ist.

177Lu-PSMA radioligand therapy is a promising new option for patients with metastasized castration-resistant prostate cancer, and the spectrum of adverse events with this treatment has to be evaluated. Here, we describe the case of a patient with M1c disease (metastasis to the mediastinum, lungs, bones, and liver) who presented with elevated liver enzyme levels after receiving 177Lu-PSMA radioligand therapy for castration-resistant prostate cancer. Pretreatment 68Ga-PSMA PET/CT showed at least 4 liver lesions with low uptake. Overall, the liver uptake was inhomogeneous. Liver biopsy was performed subsequently.

In 2016, an increased rate of methicillin-susceptibleStaff members showing positive on screening carried out a 5-day decolonization program at home. Decolonization products containing octenidine as active ingredient were used first. At the earliest, 48 h after completing the procedure, the success of the intervention was tested (3 buccal and nasal swabs were taken on consecutive days). If 2 attempts at decolonization were not successful, staff members were provided with a mupirocin-containing nasal ointment instead of octenidine.Of 128 employees examined, 43 (33.6%) were identified as carriers ofVarious reasons might explain the difficulties of decolonization such as the challenge of managing decolonization at home, inhibitory factors as well as inconsistent performance of decolonization measures. Additionally, differences between the preparations for the nasal decontamination may be considered.Auf einer neonatologischen Intensivstation am Universitätsklinikum Leipzig wurden im Jahr 2016 vermehrt Kolonisationen mit Methicillin-sensiblenMitarbeiter mitBei 128 untersuchten Mitarbeitern wurde in 43 Fällen (33,6%)Ursachen für die erschwerte Sanierung können vielfältig sein: Schwer umzusetzende Sanierung im eigenen Haushalt, Vorhandensein von dekolonisierungshemmenden Faktoren, nicht konsequente Durchführung aller Sanierungsmaßnahmen und eine mögliche Rekolonisation während der Arbeitszeit oder durch Familienangehörige. Weiterhin ist zu diskutieren, ob der Erfolg der Sanierung bei Verwendung von Mupirocin-Nasensalbe in der besseren Wirksamkeit von Mupirocin im Vergleich zu Octenidin begründet ist oder ob die unterschiedliche Konsistenz der Präparate ausschlaggebend ist.

Salt stress reduces the growth of salt-sensitive plants such as maize. The cultivation of salt-resistant maize varieties might therefore help to reduce yield losses. For the elucidation of the underlying physiological and biochemical processes of a resistant hybrid, we used a gas chromatography mass spectrometry approach and analyzed five different salt stress levels. By comparing a salt-sensitive and a salt-resistant maize hybrid, we were able to identify an accumulation of sugars such as glucose, fructose, and sucrose in leaves as a salt-resistance adaption of the salt-sensitive hybrid. Although, both hybrids showed a strong decrease of the metabolite concentration in the tricarboxylic acid cycle. These decreases resulted in the same reduced catabolism for the salt-sensitive and even the salt-resistant maize hybrid. Surprisingly, the change of root metabolism was negligible under salt stress. Moreover, the salt-resistance mechanisms were the most effective at low salt-stress levels in the leaves of the salt-sensitive maize.