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Limited number of tumor types have been examined for Orthopedia Homeobox (OTP) expression. In pulmonary carcinoids, loss of expression is a strong indicator of poor prognosis. Here, we investigated OTP expression in 37 different tumor types, and the association between OTP expression and DNA methylation levels in lung neuroendocrine neoplasms. We analyzed publicly available multi-omics data (whole-exome-, whole-genome-, RNA sequencing and Epic 850K-methylation array) of 58 typical carcinoids, 27 atypical carcinoids, 69 large cell neuroendocrine carcinoma and 51 small cell lung cancer patients and TCGA (The Cancer Genome Atlas) data of 33 tumor types. 850K-methylation analysis was cross-validated using targeted pyrosequencing on 35 carcinoids. We report bimodality of OTP expression in carcinoids (OTPhigh vs OTPlow group, likelihood-ratio test P = 1.5 × 10−2), with the OTPhigh group specific to pulmonary carcinoids while absent from all other cohorts analyzed. Significantly different DNA methylation levels were observed between OTPhigh and OTPlow carcinoids in 12/34 OTP infinium probes (FDR < 0.05 and β-value effect size >.2). OTPlow carcinoids harbor high DNA methylation levels as compared to OTPhigh carcinoids. OTPlow carcinoids showed a significantly worse overall survival (log-rank test P =.0052). Gene set enrichment analysis for somatically mutated genes associated with hallmarks of cancer showed robust enrichment of three hallmarks in the OTPlow group, that is, sustaining proliferative signaling, evading growth suppressor and genome instability and mutation. Together our data suggest that high OTP expression is a unique feature of pulmonary carcinoids with a favorable prognosis and that in poor prognostic patients, OTP expression is lost, most likely due to changes in DNA methylation levels.

The in situ Grignard Metalation Method (iGMM) is a straightforward one-pot strategy to synthesize alkaline-earth metal amides in multi-gram scale with high yields via addition of bromoethane to an ethereal suspension of a primary or secondary amine and magnesium (Part I) or calcium (Part II). This method is highly advantageous because no activation of calcium is required prior to the reaction. Contrary to the magnesium-based iGMM, there are some limitations, the most conspicuous one is the large influence of steric factors. The preparation of Ca(hmds)

Sphingosine 1-phosphate (S1P) signaling influences numerous cell biological mechanisms such as differentiation, proliferation, survival, migration, and angiogenesis. Intriguingly, our current knowledge is based solely on the role of S1P with an 18-carbon long-chain base length, S1P d18:1. Depending on the composition of the first and rate-limiting enzyme of the sphingolipid de novo metabolism, the serine palmitoyltransferase, other chain lengths have been described in vivo. While cells are also able to produce S1P d20:1, its abundance and function remains elusive so far. Our experiments are highlighting the role of S1P d20:1 in the mouse central nervous system (CNS) and human glioblastoma. We show here that S1P d20:1 and its precursors are detectable in both healthy mouse CNS-tissue and human glioblastoma. On the functional level, we focused our work on one particular, well-characterized pathway, the induction of cyclooxygenase (COX)-2 expression via the S1P receptor 2 (S1P2 ). Intriguingly, S1P d20:1 only fairly induces COX-2 expression and can block the S1P d18:1-induced COX-2 expression mediated via S1P2 activation in the human glioblastoma cell line LN229. This data indicates that S1P d20:1 might act as an endogenous modulator of S1P signaling via a partial agonism at the S1P2 receptor. While our findings might stimulate further research on the relevance of long-chain base lengths in sphingolipid signaling, the metabolism of S1P d20:1 has to be considered as an integral part of S1P signaling pathways in vivo.

This is a protocol for a Cochrane Review (Intervention). The objectives are as follows: To assess the efficacy and safety of gastrografin in relieving manifestations of meconium obstruction in newborn infants.

The N‐Myc Downstream‐Regulated Gene 4 (NDRG4), a prominent biomarker for colorectal cancer (CRC), is specifically expressed by enteric neurons. Considering that nerves are important members of the tumor microenvironment, we here establish different Ndrg4 knockout (Ndrg4 −/−) CRC models and an indirect co‐culture of primary enteric nervous system (ENS) cells and intestinal organoids to identify whether the ENS, via NDRG4, affects intestinal tumorigenesis. Linking immunostainings and gastrointestinal motility (GI) assays, we show that the absence of Ndrg4 does not trigger any functional or morphological GI abnormalities. However, combining in vivo, in vitro, and quantitative proteomics data, we uncover that Ndrg4 knockdown is associated with enlarged intestinal adenoma development and that organoid growth is boosted by the Ndrg4 −/− ENS cell secretome, which is enriched for Nidogen‐1 (Nid1) and Fibulin‐2 (Fbln2). Moreover, NID1 and FBLN2 are expressed in enteric neurons, enhance migration capacities of CRC cells, and are enriched in human CRC secretomes. Hence, we provide evidence that the ENS, via loss of Ndrg4, is involved in colorectal pathogenesis and that ENS‐derived Nidogen‐1 and Fibulin‐2 enhance colorectal carcinogenesis., Loss of enteric neuronal N‐Myc Downstream‐Regulated Gene 4 (Ndrg4) increases the release of the extracellular matrix proteins, Nidogen‐1 and Fibulin‐2, which accelerates intestinal growth in vitro and promotes colorectal cancer progression in vivo.

The Grignard reaction offers a straight forward atom-economic synthesis of organomagnesium halides, which undergo redistribution reactions (Schlenk equilibrium) yielding diorganylmagnesium and magnesium dihalides. The homologous organocalcium complexes (heavy Grignard reagents) gained interest only quite recently owing to several reasons. The discrepancy between the inertness of this heavy alkaline earth metal and the enormous reactivity of its organometallics hampered a vast and timely development after the first investigation more than 100 years ago. In this overview the synthesis of organocalcium reagents is described as is the durability in ethereal solvents. Aryl-, alkenyl-, and alkylcalcium halides are prepared by direct synthesis. Characteristic structural features and NMR parameters are discussed. Ligand redistribution reactions can be performed by addition of potassium tert-butanolate to ethereal solutions of arylcalcium iodides yielding soluble diarylcalcium, whereas sparingly soluble potassium iodide and calcium bis(tert-butanolate) precipitate. Furthermore, reactivity studies with respect to metalation and addition to unsaturated organic compounds and metal-based Lewis acids, leading to the formation of heterobimetallic complexes, are presented.

Background Copeptin, also termed C-terminal pre-pro-vasopressin or CTproAVP, mirrors endogenous vasopressin (anti-diuretic hormone, ADH) activity and might thereby serve as a biomarker reflecting the biological stress level. We therefore hypothesized that copeptin plasma concentrations are associated with disease severity in critically ill patients and could predict mortality. Methods We analyzed plasma copeptin levels in a prospective, single-center, observational study comprising 218 critically ill patients at admission to the medical intensive care unit (ICU). Mortality was assessed during a 2-year observational follow-up period. Results Copeptin plasma levels were significantly elevated in critically ill patients (n = 218) at ICU admission, as compared with 66 healthy controls. Neither sepsis as the cause of critical illness nor pre-existing metabolic disorders (type 2 diabetes, obesity) were found to influence copeptin levels. On the contrary, plasma copeptin was closely associated with disease severity (eg APACHE-II score) and correlated with biomarkers of inflammation, renal failure, metabolism, vascular tone, and tissue perfusion. Elevated copeptin levels at ICU admission predicted short-term and long-term mortality. Conclusions Copeptin plasma concentrations are significantly elevated in critically ill patients, correlate with disease severity and predict ICU and long-term outcome. Thus, copeptin could be a promising tool for prognostication and management of critically ill patients.

Next-generation transcriptome sequencing is increasingly integrated with mass spectrometry to enhance MS-based protein and peptide identification. Recently, a breakthrough in transcriptome analysis was achieved with the development of ribosome profiling (ribo-seq). This technology is based on the deep sequencing of ribosome-protected mRNA fragments, thereby enabling the direct observation of in vivo protein synthesis at the transcript level. In order to explore the impact of a ribo-seq-derived protein sequence search space on MS/MS spectrum identification, we performed a comprehensive proteome study on a human cancer cell line, using both shotgun and N-terminal proteomics, next to ribosome profiling, which was used to delineate (alternative) translational reading-frames. By including protein-level evidence of sample-specific genetic variation and alternative translation, this strategy improved the identification score of 69 proteins and identified 22 new proteins in the shotgun experiment. Furthermore, we discovered 18 new alternative translation start sites in the N-terminal proteomics data and observed a correlation between the quantitative measures of ribo-seq and shotgun proteomics with a Pearson correlation coefficient ranging from 0.483 to 0.664. Overall, this study demonstrated the benefits of ribosome profiling for MS-based protein and peptide identification and we believe this approach could develop into a common practice for next-generation proteomics.

Background & Aims Serum concentrations of miR-122 were proposed as a marker for various inflammatory diseases, but the mechanisms driving alterations in miR-122 serum levels are unknown. Methods We analysed miR-122 serum levels and hepatic miR-122 expression in mice after hepatic ischaemia and reperfusion (I/R) injury. These data were compared with data from mice after caecal pole ligation and puncture (CLP) procedure. To translate these data into the human, we analysed miR-122 serum concentrations in a cohort of 223 patients with critical illness and 57 patients with cirrhosis. Results We detected strongly elevated levels of miR-122 in mice after hepatic I/R injury. miR-122-concentrations correlated with the degree of liver damage according to AST/ALT and were associated with the presence of hepatic cell death detected by TUNEL staining. miR-122 levels were elevated in the cellular supernatants in an in vitro model of hepatocyte injury, supporting the hypothesis that the passive release of miR-122 represents a surrogate for hepatocyte death in liver injury. Moreover, miR-122 levels were almost normal in patients with cirrhosis without ongoing liver damage, but were elevated when liver injury was present. In contrast to previous assumptions, miR-122-concentrations were independent of the presence of infection/sepsis in mice or human patients. miR-122 levels did not correlate with disease severity or mortality in critically ill patients. In contrast, serum miR-122 levels strictly correlated with the presence of hepatic injury in these patients. Conclusion In mice and humans, miR-122 levels represent an independent and potent marker of ongoing liver injury and hepatic cell death regardless of the underlying disease.

PPARγ agonists [thiazolidinediones (TZDs)] are known to exert anti-fibrotic effects in the kidney. In addition, we previously demonstrated that sphingosine kinase 1 (SK-1) and intracellular sphingosine-1-phosphate (S1P), by reducing the expression of connective tissue growth factor (CTGF), have a protective role in the fibrotic process.

Background Extracellular release of high mobility group box 1 (HMGB1) acts as a danger-associated molecular pattern, thereby "alarming" the immune system and promoting systemic inflammation. We investigated plasma HMGB1 concentrations as a potential diagnostic and prognostic biomarker in critical illness. Methods Our study included 218 critically ill patients (145 with sepsis, 73 without sepsis), of whom blood samples were obtained at the time-point of admission to the medical intensive care unit (ICU). Results High mobility group box 1 levels were significantly elevated in critically ill patients (n = 218) compared with healthy controls (n = 66). Elevated HMGB1 plasma levels were independent from the presence of sepsis. Moreover, HMGB1 was not associated with disease severity, organ failure, or mortality in the ICU. We observed a trend toward lower HMGB1 levels in ICU patients with pre-existing obesity, type 2 diabetes and end-stage renal disease patients on chronic hemodialysis. Conclusion In conclusion, our study did not reveal significant associations between HMGB1 levels at ICU admission and clinical outcomes in critically ill patients. Due to the pathogenic role of HMGB1 in the late phases of experimental sepsis, future studies might assess the potential value of HMGB1 by measuring its plasma concentrations at later time points during the course of critical illness.

Background: Critical illness in cirrhotic patients is associated with a poor prognosis and increased susceptibility to infections. Monocyte HLA-DR expression is decreased in cirrhotic patients, but its prognostic value has not been investigated prospectively. Methods: Thirty-eight critically ill patients with decompensated liver cirrhosis were included in this prospective study. On admission to the intensive care unit (ICU), inflammatory parameters (C-reactive protein, procalcitonin and lipopolysaccharide-binding protein), interleukin (IL)-10, interferon (IFN)-γ serum levels, tumour necrosis factor (TNF)-αex vivo stimulation (whole blood assay) and HLA-DR expression on monocytes (FACS analysis) were determined. Immune parameters were furthermore measured every third day until discharge from the ICU or death of the patients. Results: Intensive care unit mortality of the cirrhotic patients was 34.2%. During admission, TNF ex vivo, IFN-γ and HLA-DR expression were lower in non-survivors (all P

Background Inflammation is a major factor for the progression of chronic liver diseases. Interactions between urokinase plasminogen activator (uPA) and its receptor (uPAR) have been functionally linked to hepatic inflammation and fibrosis in mice. High serum concentrations of soluble uPAR (suPAR) are suggested to reflect activated immune cells. Aims We evaluated suPAR serum levels as a diagnostic and prognostic biomarker in patients with chronic liver diseases. Methods Prospective, cross-sectional cohort study of 159 patients with chronic liver diseases (61 without, 98 with established cirrhosis) and 43 healthy controls. Transplant-free survival was monitored for up to 3 years. Results Soluble urokinase plasminogen activator serum concentrations were significantly elevated in patients with chronic liver diseases compared with controls. Cirrhotic patients displayed higher levels than non-cirrhotics, closely depending on stage of fibrosis or cirrhosis. suPAR levels had high diagnostic power to identify established cirrhosis in chronic liver diseases. Circulating suPAR closely correlated with liver function, fibrosis markers, but also with systemic inflammation and renal function. A distinct suPAR elevation was noticed in patients with alcoholic aetiology of liver disease. suPAR identified alcoholic origin more precisely compared with classical indicators of alcoholism (mean corpuscular volume, gamma glutamyl transpeptidase). Strikingly, elevated suPAR levels were identified as a strong predictor of mortality or need for transplantation. suPAR levels >9 ng/ml indicated adverse prognosis (sensitivity: 70.7%, specificity: 77.8%, relative risk: 8.5; 95% confidence interval: 3.5–20.3). Conclusions Serum suPAR is a potential novel biomarker for the diagnosis of cirrhosis, identification of alcoholic origin and for determining prognosis in patients with chronic liver disease.