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Micro-metastatic growth is considered the main source of early cancer recurrence. Nutritional and microenvironmental components are increasingly recognized to play a significant role in the liver. We explored the predictive potential of preoperative plasma metabolites for postoperative disease recurrence in colorectal cancer liver metastasis (CRCLM) patients. All included patients (n = 71) had undergone R0 liver resection for colorectal cancer liver metastasis in the years between 2012 and 2018. Preoperative blood samples were collected and assessed for 180 metabolites using a preconfigured mass-spectrometry kit (Biocrates Absolute IDQ p180 kit). Postoperative disease-free (DFS) and overall survival (OS) were prospectively recorded. Patients that recurred within 6 months after surgery were defined as "high-risk" and, subsequently, a three-metabolite model was created which can assess DFS in our collective. Multiple lysophosphatidylcholines (lysoPCs) and phosphatidylcholines (PCs) significantly predicted disease recurrence within 6 months (strongest: PC aa C36:1 AUC = 0.83, p = 0.003, PC ae C34:0 AUC = 0.83, p = 0.004 and lysoPC a C18:1 AUC = 0.8, p = 0.006). High-risk patients had a median DFS of 183 days versus 522 days in low-risk population (p = 0.016, HR = 1.98 95% CI 1.16–4.35) with a 6 months recurrence rate of 47.6% versus 4.7%, outperforming routine predictors of oncological outcome. Circulating metabolites identified CRCLM patients at highest risk for 6 months disease recurrence after surgery. Our data also suggests that circulating metabolites might play a significant pathophysiological role in micro-metastatic growth and concomitant early tumor recurrences after liver resection. However, the clinical applicability and performance of this proposed metabolomic concept needs to be independently validated in future studies. [ABSTRACT FROM AUTHOR]

Activation induced deaminase (AID) has two distinct and well defined roles, both relying on its deoxycytidine (dC) deaminating function: one as a DNA mutator and another in DNA demethylation. In chronic lymphocytic leukemia (CLL), AID was previously shown to be an independent negative prognostic factor. While there is substantial impact on DNA mutations, effects of AID on gene expression by promoter demethylation of disease related target genes in leukemia has not been addressed. To shed light on this question, we aimed at determining genome wide methylation changes as well as gene expression changes in response to AID expression in CLL. Although we found minor differences in individual methylation variable positions following AID expression, we could not find recurrent methylation changes of specific target sites or changes in global methylation. [ABSTRACT FROM AUTHOR]

Background: The majority of individuals with acute myeloid leukemia (AML) respond to initial chemotherapy and achieve a complete remission, yet only a minority experience long-term survival because a large proportion of patients eventually relapse with therapy-resistant disease. Relapse therefore represents a central problem in the treatment of AML. Despite this, and in contrast to the extensive knowledge about the molecular events underlying the process of leukemogenesis, information about the mechanisms leading to therapy resistance and relapse is still limited. Purpose and content of review: Recently, a number of studies have aimed to fill this gap and provided valuable information about the clonal composition and evolution of leukemic cell populations during the course of disease, and about genetic, epigenetic, and gene expression changes associated with relapse. In this review, these studies are summarized and discussed, and the data reported in them are compiled in order to provide a resource for the identification of molecular aberrations recurrently acquired at, and thus potentially contributing to, disease recurrence and the associated therapy resistance. This survey indeed uncovered genetic aberrations with known associations with therapy resistance that were newly gained at relapse in a subset of patients. Furthermore, the expression of a number of protein coding and microRNA genes was reported to change between diagnosis and relapse in a statistically significant manner. Conclusions: Together, these findings foster the expectation that future studies on larger and more homogeneous patient cohorts will uncover pathways that are robustly associated with relapse, thus representing potential targets for rationally designed therapies that may improve the treatment of patients with relapsed AML, or even facilitate the prevention of relapse in the first place. [ABSTRACT FROM AUTHOR]

Mutations in KRAS are often associated with resistance to EGFR‐targeting antibody therapy. Using comprehensive systems analyses, GNB5 has been identified as a potential target to overcome therapy resistance targeting the EGFR signaling pathways, whereby the AKT signaling pathway (PI3K) rather than the ERK signaling pathway (RAS) might be dominantly affected. Personalized mathematical modeling and simulations of this signaling pathway/network and respective perturbations are of great utility to customize therapy for patients. [ABSTRACT FROM AUTHOR]

Oxidized phospholipids (oxPLs) are components of oxidized LDL (oxLDL). It is known that oxLDL activates expression of a series of atherogenic genes and their oxPLs contribute to their biological activities. In this study we present the effects of 1-palmitoyl-2-glutaroyl-sn-glycero-3-phosphocholine (PGPC) and 1-palmitoyl-2-(5-oxovaleroyl)-sn-glycero-3-phosphocholine (POVPC) on gene expression in RAW 264.7 macrophages using cDNA microarrays. PGPC affected the regulation of 146 genes, whereas POVPC showed only very minor effects. PGPC preferentially influenced expression of genes related to cell death, angiogenesis, cholesterol efflux, procoagulant mechanisms, atherogenesis, inflammation, and cell cycle. Many of these effects are known from studies with oxLDL or oxidized 1-hexadecanoyl-2-eicosatetra-5′,8′,11′,14′-enoyl-sn-glycero-3-phosphocholine (oxPAPC), containing PGPC in addition to other oxPL species. It is known that POVPC efficiently reacts with proteins by Schiff base formation, whereas PGPC only physically interacts with its biological targets. POVPC seems to affect cell physiology to a great extent on the protein level, whereas PGPC gives rise to both the modulation of protein function and regulation on the transcriptional level. [ABSTRACT FROM AUTHOR]

Maternal lipoproteins have been studied extensively in human pregnancies, but little is known about the role of fetal lipoproteins. The vascularized human placenta interfaces between the mother and fetus to transfer nutrients for sustaining pregnancy. Unlike that of adults, fetal highdensity lipoprotein (HDL), which is in contact with placental vessels, is characterized by a high proportion of apolipoprotein E (apoE). We hypothesize this unique composition of fetal HDL affects key functions of the growing fetal tissues. The aim was to identify genes regulated by apoE-HDL by incubating human placental endothelial cells (HPEC) with either fetal HDL or apoE-rich reconstituted HDL particles (apoE-rHDL). HPEC were exposed to 15 μg/ml fetal HDL, 15 μg/ml apoE-rHDL, or medium for 16 h, respectively. Microarray analysis determined genes regulated by fetal HDL and apoE. Characterization of HDL particles revealed a different hydrodynamic radius for apoE-rHDL (13.70 nm) compared with fetal HDL (18.11 nm). Stepwise gene clustering after microarray experiments identified 79 differentially expressed genes (P< 0.05) when cells were exposed to HDL compared with controls. Among them 16 genes were downregulated, whereas five genes were upregulated by twofold, respectively. When HPEC were incubated with apoE-rHDL 18-fold more genes (1,417, 12% of transcripts) were regulated (P < 0.05) in contrast to HDL. Thereof, 172 genes were downregulated and 376 genes upregulated (twofold). In the common subset of 38 genes regulated by both HDL particles, genes involved in cholesterol biosynthesis and cell protection prevailed. Strikingly, results suggest that HDL has the capability of regulating metallothioneins, which may have an effect on oxidative stress in HPEC. [ABSTRACT FROM AUTHOR]

Abstract: The quest for new cancer biomarkers and targets for therapy requires not only the aggregation and analysis of heterogeneous biomolecular data but also integration of clinical data. In this review we highlight information technology solutions for the integration of biomolecular and clinical data and focus on a solution at the departmental level, i.e., decentralized and medium-scale solution for groups of labs working on a specific topic. Both, hardware and software requirements are described as well as bioinformatics methods and tools for the data analysis. The highlighted IT solutions include storage architecture, high-performance computing, and application servers. Additionally, following computational approaches for data integration are reviewed: data aggregation, integrative data analysis including methodological aspects as well as examples, biomolecular pathways and network reconstruction, and mathematical modelling. Finally, a case study in cancer immunology including the used computational methods is shown, demonstrating how IT solutions for integrating biomolecular and clinical data can help to identify new cancer biomarkers for improving diagnosis and predicting clinical outcome. [ABSTRACT FROM AUTHOR]

Background: Tetracycline regulated ectopic gene expression is a widely used tool to study gene function. However, the tetracycline regulator (tetR) itself has been reported to cause certain phenotypic changes in mammalian cells. We, therefore, asked whether human myeloid U937 cells expressing the tetR in an autoregulated manner would exhibit alterations in gene expression upon removal of tetracycline. Methodology/Principal Findings: Microarray analyses revealed that 172 and 774 unique genes were significantly differentially expressed by at least 2- or 1.5-fold, respectively, when tetR expressing U937 cells were maintained in media with or without the antibiotic. Conclusions/Significance: These alterations in gene expression are likely to contribute to the phenotypic consequences of tetR expression. In addition, they need to be taken into consideration when using the tetR system for the identification of target genes of transcription factors or other genes of interest. [ABSTRACT FROM AUTHOR]

Abstract: Adipocyte triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) are intracellular lipases that mobilize triglycerides, the main energy source in mammals. Deletion of genes encoding ATGL (Pnpla2) or HSL (Lipe) in mice results in striking phenotypic differences, suggesting distinct roles for these lipases. The goal of the present study was to identify the biological processes that are modulated in the metabolic tissues of ATGL- and HSL-deficient mice. DNA microarrays were employed to provide full genome coverage concerning the types of genes that are differentially expressed in wild-type and mutant mice. For both mouse models, transcript signatures were identified in white adipose tissue, brown adipose tissue (BAT), skeletal muscle (SM), cardiac muscle (CM), and liver. Genetic ablation of ATGL and HSL alters the transcript levels of a large number of genes in metabolic tissues. The genes affected in the two models are, however, largely different ones. Indeed, only one biological process was modulated in the same way in both mouse models, namely the down-regulation of fatty acid metabolism in BAT. The most pronounced modulation of biological processes was observed in ATGL-/- CM, in which a concerted down-regulation of transcripts associated with oxidative pathways was observed. In HSL-/- mice, in contrast, the most marked changes were seen in SM, namely, alterations in transcript levels reflecting a change of energy source from lipid to carbohydrate. The transcript signatures also provided novel insights into the metabolic derangements that are characteristic of ATGL-/- mice. Our findings suggest that ATGL and HSL differentially modulate biological processes in metabolic tissues. We hypothesize that the intermediary metabolites of the lipolytic pathways are signaling molecules and activators of a wide range of biochemical and cellular processes in mammals. [Copyright &y& Elsevier]

Background: The GOLD.db (Genomics of Lipid-Associated Disorders Database) was developed to address the need for integrating disparate information on the function and properties of genes and their products that are particularly relevant to the biology, diagnosis management, treatment, and prevention of lipid-associated disorders. Description: The GOLD.db http://gold.tugraz.at provides a reference for pathways and information about the relevant genes and proteins in an efficiently organized way. The main focus was to provide biological pathways with image maps and visual pathway information for lipid metabolism and obesity-related research. This database provides also the possibility to map gene expression data individually to each pathway. Gene expression at different experimental conditions can be viewed sequentially in context of the pathway. Related large scale gene expression data sets were provided and can be searched for specific genes to integrate information regarding their expression levels in different studies and conditions. Analytic and data mining tools, reagents, protocols, references, and links to relevant genomic resources were included in the database. Finally, the usability of the database was demonstrated using an example about the regulation of Pten mRNA during adipocyte differentiation in the context of relevant pathways. Conclusions: The GOLD.db will be a valuable tool that allow researchers to efficiently analyze patterns of gene expression and to display them in a variety of useful and informative ways, allowing outside researchers to perform queries pertaining to gene expression results in the context of biological processes and pathways. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented about a gene expression profile associated with relapse of cytogenetically normal acute myeloid leukemia.

Background: Activation of brown adipose tissue (BAT) has gained attention due to its ability to dissipate energy and counteract cardiometabolic diseases (CMDs). Methods: This study investigated the consequences of cold exposure on the BAT and liver proteomes of an established CMD mouse model based on LDL receptor-deficient (LdlrKO) mice fed a high-fat, high-sucrose, high-cholesterol diet for 16 weeks. We analyzed energy metabolism in vivo and performed untargeted proteomics on BAT and liver of LdlrKO mice maintained at 22 °C or 5 °C for 7 days. Results: We identified several dysregulated pathways, miRNAs, and transcription factors in BAT and liver of cold-exposed Ldlrko mice that have not been previously described in this context. Networks of regulatory interactions based on shared downstream targets and analysis of ligand-receptor pairs identified fibrinogen alpha chain (FGA) and fibronectin 1 (FN1) as potential crosstalk factors between BAT and liver in response to cold exposure. Importantly, genetic variations in the genes encoding FGA and FN1 have been associated with cardiometabolic-related phenotypes and traits in humans. Discussion: This study describes the key factors, pathways, and regulatory networks involved in the crosstalk between BAT and the liver in a cold-exposed CMD mouse model. These findings may provide a basis for future studies aimed at testing whether molecular mediators, as well as regulatory and signaling mechanisms involved in tissue adaption upon cold exposure, could represent a target in cardiometabolic disorders. [ABSTRACT FROM AUTHOR]

Experimental evidence suggests sex dependent differences in liver regeneration. Limited evidence is available examining sex differences in post-hepatectomy liver failure (PHLF) and postoperative outcomes. Our aim was to assess the influence of sex on the outcomes after liver resection. The hepatectomy targeted National Surgical Quality Improvement Program (NSQIP) database was assessed for associations between sex and outcomes. A total of 13,401 patients underwent elective hepatic resection between 2014-2017. PHLF was highest among male patients with hepatocellular carcinoma (HCC) (OR = 2.81,95%CI:1.40–5.62). Male sex was independently associated with increased PHLF (OR = 1.47,95%CI:1.15–1.88), major complications (OR = 1.25,95%CI:1.08–1.45), mortality (OR = 1.61,95%CI:1.03–2.50), and if only major resections were assessed (OR = 1.38,95%CI:1.03–1.84). Diagnosis specific subgroup analyses revealed that effects of sex were predominantly HCC associated. This is the largest series investigating the effects of gender on outcomes after hepatic resection. We documented that women undergoing liver resection have significantly lower risk of PHLF. This difference seemed influenced by the striking increase of PHLF in male HCC patients. These hypothesis suggest that sex might play a role in preoperative risk stratification. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs), small non-coding RNAs, have emerged as important, epigenetic regulators of endothelial function. Metabolic disturbances such as diabetes alter miRNA expression. In adults, the miRNA transcriptome as well as endothelial function differ between the sexes. Here, we hypothesized that metabolic disturbances associated with gestational diabetes (GDM) alter miRNA signatures in feto-placental endothelial cells (fpEC), dependent on fetal sex. We isolated human primary fpEC after normal and GDM-complicated pregnancies with male and female neonates and screened for differential miRNA expression using next-generation miRNA sequencing. To test for miRNAs commonly regulated in fpEC of female and male progeny, data were stratified for fetal sex and maternal body mass index (BMI). Analyses were also performed separately for female and male fpEC, again accounting for maternal BMI as covariate. Potential biological pathways regulated by the altered set of miRNAs were determined using mirPath software. Maternal GDM altered 26 miRNA signatures when male and female fpEC were analyzed together. Separate analysis of male versus female fpEC revealed 22 GDM affected miRNAs in the females and only 4 in the males, without overlap. Biological functions potentially modulated by the affected miRNAs related to 'Protein Processing in Endoplasmic Reticulum' and 'Proteoglycans in Cancer'. Maternal GDM alters miRNA signatures in fpEC, and biological functions affected by these miRNAs relate to well-known adverse functional consequences of diabetes on endothelium. GDM effects were highly dependent on fetal sex with miRNA signatures in female fpEC being more susceptible to metabolic derangements of GDM than miRNAs in male fpEC. [ABSTRACT FROM AUTHOR]

Aging is a multifaceted process that unfolds at both the individual and cellular levels, resulting in changes in platelet count and platelet reactivity. These alterations are influenced by shifts in platelet production, as well as by various environmental factors that affect circulating platelets. Aging also triggers functional changes in platelets, including a reduction in RNA content and protein production capacity. Older individuals and RNA-rich immature platelets often exhibit hyperactivity, contributing significantly to pathologic conditions such as cardiovascular diseases, sepsis, and thrombosis. However, the impact of aging on surface receptor expression of circulating platelets, particularly whether these effects vary between immature and mature platelets, remains largely unexplored. Thus, we investigated the expression of certain surface and activation receptors on platelets from young and old mice as well as on immature and mature platelets from mouse models of regenerative thrombocytopenia by flow cytometry. Our findings indicate that aged mice show an upregulated expression of the platelet endothelial cell adhesion molecule-1 (CD31), tetraspanin-29 (CD9), and Toll-like receptor 2 (TLR2) compared to their younger counterparts. Interestingly, when comparing immature and mature platelets in both young and old mice, no differences were observed in mature platelets. However, immature platelets from young mice displayed higher surface expression compared to immature platelets from old mice. Additionally, in mouse models of regenerative thrombocytopenia, the majority of receptors were upregulated in immature platelets. These results suggest that distinct surface receptor expressions are increased on platelets from old mice and immature platelets, which may partially explain their heightened activity and contribute to an increased thrombotic risk. [ABSTRACT FROM AUTHOR]

Epithelial ovarian cancer (OC) is the deadliest gynecological malignancy worldwide. Blocking angiogenesis with bevacizumab, an antibody targeting vascular endothelial growth factor (VEGF), shows efficacy in different lines of OC therapy. This study investigates the clinical impact of tumoral expression of angiogenesis-related genes and their association with bevacizumab response in OC in retrospective analysis of three independent cohorts. mRNA expression of seven angiogenic genes (VEGF , VEGFR2 , PDGFA , PDGFB , PDGFRA , PDGFRB , KIT) was quantified in an inception OC cohort (n = 195) and a transcriptional tumor angiogenesis score from 0 to 3 was established and linked to progression-free survival (PFS) and overall survival (OS). This score was corroborated in an independent publicly available cohort from The Cancer Genome Atlas (TCGA, n = 582) and prediction of therapeutic efficacy of bevacizumab by the angiogenesis score was analyzed in the Gene Expression Omnibus (GEO) dataset GSE140082 (n = 380) from the ICON7-trial. The tumor angiogenesis score prognosticated PFS and OS in patients with OC from the inception cohort (p < 0.001, respectively). Tumoral PDGFA expression (PFS: HR 2.46, p = 0.005; OS: HR 2.26, p = 0.011) and a high tumoral transcriptional angiogenesis score (PFS: HR 1.41, p = 0.018) were identified as independent predictors of clinical outcome. The transcriptional angiogenesis score exhibited a significant though smaller effect size on PFS in the TCGA cohort. However, in the ICON7-trial, the angiogenesis score was not associated with benefit of bevacizumab treatment. Our study indicates that tumoral expression of angiogenic genes is unfavorable in OC. The established score could be used to identify patients who respond to targeted angiogenic therapies, a concept that warrants prospective controlled clinical trials. • Blockade of angiogenesis, i.e. bevacizumab treatment is a therapeutic pillar in OC. • Identification of patients who benefit from anti-angiogenic therapy is expected to improve clinical care. • An angiogenic tumor phenotype defined by high expression of tumoral angiogenic genes predicts poor outcome of OC patients. • The transcriptional angiogenesis does not predict treatment response to bevacizumab in a subset of ICON-7 patients. [ABSTRACT FROM AUTHOR]

Cancer associated fibroblasts (CAF) are known to orchestrate multiple components of the tumor microenvironment, whereas the influence of the whole stromal-fibroblast compartment is less understood. Here, an extended stromal fibroblast signature was investigated to define its impact on immune cell infiltration. The lung cancer adenocarcinoma (LUAD) data set of the cancer genome atlas (TCGA) was used to test whole stroma signatures and cancer-associated fibroblast signatures for their impact on prognosis. 3D cell cultures of the NSCLC cancer cell line A549 together with the fibroblast cell line SV80 were used in combination with infiltrating peripheral blood mononuclear cells (PBMC) for in-vitro investigations. Immune cell infiltration was assessed via flow cytometry, chemokines were analyzed by immunoassays and RNA microarrays. Results were confirmed in specimens from NSCLC patients by flow cytometry or immunohistochemistry as well as in the TCGA data set. The TCGA analyses correlated the whole stromal-fibroblast signature with an improved outcome, whereas no effect was found for the CAF signatures. In 3D microtumors, the presence of fibroblasts induced infiltration of B cells and CD69+CD4+ T cells, which was linked to an increased expression of CCL13 and CXCL16. The stroma/lymphocyte interaction was confirmed in NSCLC patients, as stroma-rich tumors displayed an elevated B cell count and survival in the local cohort and the TCGA data set. A whole stromal fibroblast signature was associated with an improved clinical outcome in lung adenocarcinoma and in vitro and in vivo experiments suggest that this signature increases B and T cell recruitment via induction of chemokines. [ABSTRACT FROM AUTHOR]

Background: The transcription factor nuclear factor erythroid 2-related factor 2 (NFE2L2; previously known as NRF2) is a crucial regulator of the intracellular antioxidant response. It controls the expression of genes involved in the detoxification and elimination of reactive oxidants and electrophilic agents. The role of NFE2L2 in cancer is subject of controversial discussion, as it has been reported to have both pro-and anti-tumourigenic functions. To shed some light on this paradox, we analysed the NFE2L2 mRNA expression levels in breast cancer and its association with clinicopathological features and survival. Methods: We retrospectively evaluated the NFE2L2 mRNA expression levels in tumour tissue of two independent breast cancer patient cohorts. In the training set we analysed data from the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC). In the test set we measured the NFE2L2 mRNA expression levels in 176 breast tumour tissues by quantitative real-time reverse transcription PCR (qRT-PCR). Group differences were analysed using Mann-Whitney U-test, and associations between NFE2L2 mRNA expression levels and clinicopathological features were examined by means of univariate and multivariate survival analyses. Furthermore, we compared NFE2L2 mRNA expression levels between tumour and normal breast tissue samples by means of 108 paired samples from the The Cancer Genome Atlas (TCGA) dataset. Results: In the training set we identified an independent predictive value for high NFE2L2 mRNA expression levels [HRdisease specific death 0.8 (0.6-1.0), P = 0.041; HRdeath 0.8 (0.6-1.0), P = 0.023] especially in the subgroup of oestrogen receptor (ER) positive tumours [HRdisease specific death 0.6 (0.4-0.9), P = 0.008; HRdeath 0.6 (0.4-0.8), P = 0.001]. Similarly, we found this association also in the test set [HRrelapse 0.4 (0.2-0.9), P = 0.031] and again, more pronounced in patients with ER positive tumours [HRrelapse 0.2 (0.1-0.7), P = 0.012]. In addition, we observed generally lower NFE2L2 expression levels in tumour tissues than in normal breast tissues. Conclusion: We concluded that reduced NFE2L2 mRNA expression in tumour tissues is an independent predictor of shortened survival in breast cancer patients. [ABSTRACT FROM AUTHOR]

Early on in human immunodeficiency virus (HIV) type 1 infection, gut T-helper (Th) 17 cells are massively depleted leading eventually to compromised intestinal barrier function and excessive immune activation. In contrast, the functional Th17 cell compartment of the gut is well-maintained in nonpathogenic simian immunodeficiency virus infection as well as HIV-1 long-term nonprogressors. Here, we show that dendritic cells (DCs) loaded with HIV-1 bearing high surface complement levels after incubation in plasma from HIV-infected individuals secreted significantly higher concentrations of Th17-polarizing cytokines than DCs exposed to nonopsonized HIV-1. The enhanced Th17-polarizing capacity of in vitro-generated and BDCA-1+ DCs directly isolated from blood was linked to activation of ERK. In addition, C3a produced from DCs exposed to complement- opsonized HIV was associated with the higher Th17 polarization. Our in vitro and ex vivo data, therefore, indicate that complement opsonization of HIV-1 strengthens DC-mediated antiviral immune functions by simultaneously triggering Th17 expansion and intrinsic C3 formation via DC activation. [ABSTRACT FROM AUTHOR]

Acute skin rejection in vascularized composite allotransplantation (VCA) is the major obstacle for wider adoption in clinical practice. This study utilized computational modeling to identify biomarkers for diagnosis and targets for treatment of skin rejection. Protein levels of 14 inflammatory mediators in skin and muscle biopsies from syngeneic grafts [n = 10], allogeneic transplants without immunosuppression [n = 10] and allografts treated with tacrolimus [n = 10] were assessed by multiplexed analysis technology. Hierarchical Clustering Analysis, Principal Component Analysis, Random Forest Classification and Multinomial Logistic Regression models were used to segregate experimental groups. Based on Random Forest Classification, Multinomial Logistic Regression and Hierarchical Clustering Analysis models, IL-4, TNF-α and IL-12p70 were the best predictors of skin rejection and identified rejection well in advance of histopathological alterations. TNF-α and IL-12p70 were the best predictors of muscle rejection and also preceded histopathological alterations. Principal Component Analysis identified IL-1α, IL-18, IL-1β, and IL-4 as principal drivers of transplant rejection. Thus, inflammatory patterns associated with rejection are specific for the individual tissue and may be superior for early detection and targeted treatment of rejection. [ABSTRACT FROM AUTHOR]

Background: STAT1 has been attributed a function as tumor suppressor. However, in breast cancer data from microarray analysis indicated a predictive value of high mRNA expression levels of STAT1 and STAT1 target genes belonging to the interferon-related signature for a poor response to therapy. To clarify this issue we have determined STAT1 expression levels and activation by different methods, and investigated their association with tumor infiltration by immune cells. Additionally, we evaluated the interrelationship of these parameters and their significance for predicting disease outcome. Methods: Expression of STAT1, its target genes SOCS1, IRF1, CXCL9, CXCL10, CXCL11, IFIT1, IFITM1, MX1 and genes characteristic for immune cell infiltration (CD68, CD163, PD-L1, PD-L2, PD-1, CD45, IFN-γ, FOXP3) was determined by RT-PCR in two independent cohorts comprising 132 breast cancer patients. For a subset of patients, protein levels of total as well as serine and tyrosine-phosphorylated STAT1 were ascertained by immunohistochemistry or immunoblotting and protein levels of CXCL10 by ELISA. Results: mRNA expression levels of STAT1 and STAT1 target genes, as well as protein levels of total and serine-phosphorylated STAT1 correlated with each other in neoplastic tissue. However, there was no association between tumor levels of STAT1 mRNA and tyrosine-phosphorylated STAT1 and between CXCL10 serum levels and CXCL10 expression in the tumor. Tumors with increased STAT1 mRNA amounts exhibited elevated expression of genes characteristic for tumor-associated macrophages and immunosuppressive T lymphocytes. Survival analysis revealed an association of high STAT1 mRNA levels and bad prognosis in both cohorts. A similar prognostically relevant correlation with unfavorable outcome was evident for CXCL10, MX1, CD68, CD163, IFN-?, and PD-L2 expression in at least one collective. By contrast, activation of STAT1 as assessed by the level of STAT1-Y701 phosphorylation was linked to positive outcome. In multivariate Cox regression, the predictive power of STAT1 mRNA expression was lost when including expression of CXCL10, MX1 and CD68 as confounders. Conclusions: Our study confirms distinct prognostic relevance of STAT1 expression levels and STAT1 tyrosine phosphorylation in breast cancer patients and identifies an association of high STAT1 levels with elevated expression of STAT1 target genes and markers for infiltrating immune cells. [ABSTRACT FROM AUTHOR]

Background: STAT1 has been attributed a function as tumor suppressor. However, in breast cancer data from microarray analysis indicated a predictive value of high mRNA expression levels of STAT1 and STAT1 target genes belonging to the interferon-related signature for a poor response to therapy. To clarify this issue we have determined STAT1 expression levels and activation by different methods, and investigated their association with tumor infiltration by immune cells. Additionally, we evaluated the interrelationship of these parameters and their significance for predicting disease outcome. Methods: Expression of STAT1, its target genes SOCS1, IRF1, CXCL9, CXCL10, CXCL11, IFIT1, IFITM1, MX1 and genes characteristic for immune cell infiltration (CD68, CD163, PD-L1, PD-L2, PD-1, CD45, IFN-?, FOXP3) was determined by RT-PCR in two independent cohorts comprising 132 breast cancer patients. For a subset of patients, protein levels of total as well as serine and tyrosine-phosphorylated STAT1 were ascertained by immunohistochemistry or immunoblotting and protein levels of CXCL10 by ELISA. Results: mRNA expression levels of STAT1 and STAT1 target genes, as well as protein levels of total and serine-phosphorylated STAT1 correlated with each other in neoplastic tissue. However, there was no association between tumor levels of STAT1 mRNA and tyrosinephosphorylated STAT1 and between CXCL10 serum levels and CXCL10 expression in the tumor. Tumors with increased STAT1 mRNA amounts exhibited elevated expression of genes characteristic for tumor-associated macrophages and immunosuppressive T lymphocytes. Survival analysis revealed an association of high STAT1 mRNA levels and bad prognosis in both cohorts. A similar prognostically relevant correlation with unfavorable outcome was evident for CXCL10, MX1, CD68, CD163, IFN-?, and PD-L2 expression in at least one collective. By contrast, activation of STAT1 as assessed by the level of STAT1- Y701 phosphorylation was linked to positive outcome. In multivariate Cox regression, the predictive power of STAT1 mRNA expression was lost when including expression of CXCL10, MX1 and CD68 as confounders. Conclusions: Our study confirms distinct prognostic relevance of STAT1 expression levels and STAT1 tyrosine phosphorylation in breast cancer patients and identifies an association of high STAT1 levels with elevated expression of STAT1 target genes and markers for infiltrating immune cells. [ABSTRACT FROM AUTHOR]

Molecular mechanisms underlying sexual dimorphism in mammals, fetal sex influences on intrauterine development, and the sex-biased susceptibility for selected diseases in adulthood are novel areas of current research. As importantly, two decades of multifaceted research has established that susceptibility to many adult disorders originates in utero, commonly secondary to the effects of placental dysfunction. We hypothesized that fetal sex influences gene expression and produces functional differences in human placentas. We thus extended previous studies on sexual dimorphism in mammals, which used RNA isolated from whole tissues, to investigate the effects of sex on four cell-phenotypes within a single key tissue, human placental villi. The cells studied included cytotrophoblasts, syncytiotrophoblast, arterial and venous endothelial cells. The cells were isolated from placentas of male or female fetuses and subjected to microarray analysis. We found that fetal sex differentially affected gene expression in a cell-phenotype dependent manner among all four cell-phenotypes. The markedly enriched pathways in males were identified to be signaling pathways for graft-versus-host disease as well as the immune and inflammatory systems that parallel the reported poorer outcome of male fetuses. Our study is the first to compare global gene expression by microarray analysis in purified, characterized, somatic cells from a single human tissue, i.e. placental villi. Importantly, our findings demonstrate that there are cell-phenotype specific, and tissue-specific, sex-biased responses in the human placenta, suggesting fetal sex should be considered as an independent variable in gene expression analysis of human placental villi. [ABSTRACT FROM AUTHOR]

Background: Acute myeloid leukemia (AML) is a hematopoietic malignancy with a dismal outcome in the majority of cases. A detailed understanding of the genetic alterations and gene expression changes that contribute to its pathogenesis is important to improve prognostication, disease monitoring, and therapy. In this context, leukemia-associated misexpression of microRNAs (miRNAs) has been studied, but no coherent picture has emerged yet, thus warranting further investigations.Methods: The expression of 636 human miRNAs was compared between samples from 52 patients with AML and 13 healthy individuals by highly specific locked nucleic acid (LNA) based microarray technology. The levels of individual mature miRNAs and of primary miRNAs (pri-miRs) were determined by quantitative reverse transcriptase (qRT) PCR. Transfections and infections of human cell lines were performed using standard procedures.Results: 64 miRNAs were significantly differentially expressed between AML and controls. Further studies on the clustered miRNAs 221 and 222, already known to act as oncogenes in other tumor types, revealed a deficiency of human myeloid cell lines to process vector derived precursor transcripts. Moreover, endogenous pri-miR-221/222 was overexpressed to a substantially higher extent than its mature products in most primary AML samples, indicating that its transcription was enhanced, but processing was rate limiting, in these cells. Comparison of samples from the times of diagnosis, remission, and relapse of AML demonstrated that pri-miR-221/222 levels faithfully reflected the stage of disease.Conclusions: Expression of some miRNAs is strongly regulated at the posttranscriptional level in AML. Pri-miR-221/222 represents a novel molecular marker and putative oncogene in this disease. [ABSTRACT FROM AUTHOR]

Background: Post-hepatectomy liver failure (PHLF) represents the major determinant for death after liver resection. Early recognition is essential. Perioperative lactate dynamics for risk assessment of PHLF and associated morbidity were evaluated. Methods: This was a multicentre observational study of patients undergoing hepatectomy with validation in international highvolume units. Receiver operating characteristics analysis and cut-off calculation for the predictive value of lactate for clinically relevant International Study Group of Liver Surgery grade B/C PHLF (clinically relevant PHLF (CR-PHLF)) were performed. Lactate and other perioperative factors were assessed in a multivariable CR-PHLF regression model. Results: The exploratory cohort comprised 509 patients. CR-PHLF, death, overall morbidity and severe morbidity occurred in 7.7, 3.3, 40.9 and 29.3 per cent of patients respectively. The areas under the curve (AUCs) regarding CR-PHLF were 0.829 (95 per cent c.i. 0.770 to 0.888) for maximum lactate within 24 h (Lactate_Max) and 0.870 (95 per cent c.i. 0.818 to 0.922) for postoperative day 1 levels (Lactate_POD1). The respective AUCs in the validation cohort (482 patients) were 0.812 and 0.751 and optimal Lactate_Max cut-offs were identical in both cohorts. Exploration cohort patients with Lactate_Max 50 mg/dl or greater more often developed CR-PHLF (50.0 per cent) than those with Lactate_Max between 20 and 49.9 mg/dl (7.4 per cent) or less than 20 mg/dl (0.5 per cent; P<0.001). This also applied to death (18.4, 2.7 and 1.4 per cent), severe morbidity (71.1, 35.7 and 14.1 per cent) and associated complications such as acute kidney injury (26.3, 3.1 and 2.3 per cent) and haemorrhage (15.8, 3.1 and 1.4 per cent). These results were confirmed in the validation group. Combining Lactate_Max with Lactate_POD1 further increased AUC (DAUC=0.053) utilizing lactate dynamics for risk assessment. Lactate_Max, major resections, age, cirrhosis and chronic kidney disease were independent risk factors for CR-PHLF. A freely available calculator facilitates clinical risk stratification (www.liver-calculator.com). Conclusion: Early postoperative lactate values are powerful, readily available markers for CR-PHLF and associated complications after hepatectomy with potential for guiding postoperative care. [ABSTRACT FROM AUTHOR]

In vascularized composite allotransplantation (VCA), invasive tissue biopsies remain the gold standard in diagnosing rejection carrying significant morbidity. We aimed to show feasibility of tape‐stripping for noninvasive immune monitoring in VCA. Tape‐stripping was performed on allografts and native skin of upper extremity transplant recipients. Healthy nontransplanted individuals served as controls. The technique was also used in swine on naïve skin in nontransplanted animals, native skin of treated, transplanted swine, nonrejecting VCAs, and rejecting VCAs. Extracted protein was analyzed for differences in cytokine expression using Luminex technology. Significantly decreased levels of INFγ and IL‐1Ra were seen between human allograft samples and native skin. In swine, rejecting grafts had increased IL‐1Ra compared to naïve and native skin, decreased levels of GM‐CSF compared to native skin, and decreased IL‐10 compared to nonrejecting grafts. Unsupervised hierarchical clustering revealed rejecting grafts separated from the nonrejecting (P = 0.021). Variable importance in projection scores identified GM‐CSF, IL‐1Ra, and IL‐2 as the most important profiles for group discrimination. Differences in cytokine expression are detectable in human VCA patient native skin and VCA graft skin using a noninvasive tape‐stripping method. Swine studies suggest that differences in cytokines between rejecting and nonrejecting grafts are discernable. [ABSTRACT FROM AUTHOR]

Summary MicroRNAs have been shown to be able to modulate the tumor microenvironment and the immune response and hence could be interesting biomarkers and therapeutic targets in immuno-oncology; however, dedicated analysis tools are missing. Here, we present a user-friendly web platform MIO and a Python toolkit miopy integrating various methods for visualization and analysis of provided or custom bulk microRNA and gene expression data. We include regularized regression and survival analysis and provide information of 40 microRNA target prediction tools as well as a collection of curated immune related gene and microRNA signatures and processed TCGA data including estimations of infiltrated immune cells and the immunophenoscore. The integration of several machine learning methods enables the selection of prognostic and predictive microRNAs and gene interaction network biomarkers. Availability and implementation https://mio.icbi.at , https://github.com/icbi-lab/mio and https://github.com/icbi-lab/miopy. Supplementary information Supplementary data are available at Bioinformatics online. [ABSTRACT FROM AUTHOR]

In the article, the authors present their study on the clinical effects of breast cancer 2 messenger ribonucleic acid (BRCA2 mRNA) expression in high-grade serous ovarian cancer (HGSOC) using dataset from The Cancer Genome Atlas (TCGA) project. Topics include the percentage of epithelial OCs that are hereditary, and the use of tests like the non-parametric Mann-Whitney U test and univariate Kaplan-Meier analyses in the study.

Summary: Between 2000 and 2014, five patients received bilateral hand (n = 3), bilateral forearm (n = 1), and unilateral hand (n = 1) transplants at the Innsbruck Medical University Hospital. We provide a comprehensive report of the long‐term results at 20 years. During the 6–20 years follow‐up, 43 rejection episodes were recorded in total. Of these, 27.9% were antibody‐related with serum donor‐specific alloantibodies (DSA) and skin‐infiltrating B‐cells. The cell phenotype in rejecting skin biopsies changed and C4d‐staining increased with time post‐transplantation. In the long‐term, a change in hand appearance was observed. The functional outcome was highly depending on the level of amputation. The number and severity of rejections did not correlate with hand function, but negatively impacted on the patients´ well‐being and quality of life. Patient satisfaction significantly correlated with upper limb function. One hand allograft eventually developed severe allograft vasculopathy and was amputated at 7 years. The patient later died due to progressive gastric cancer. The other four patients are currently rejection‐free with moderate levels of immunosuppression. Hand transplantation remains a therapeutic option for carefully selected patients. A stable immunologic situation with optimized and individually adopted immunosuppression favors good compliance and patient satisfaction and may prevent development of DSA. [ABSTRACT FROM AUTHOR]

Background: In metastatic breast cancer (MBC) patients, no biomarker predicting benefit to a bevacizumab-containing therapy has been established yet. MicroRNAs (miRNAs) are involved in angiogenesis and treatment resistance and therefore could be of predictive value. Methods: Profiling of 754 miRNAs was performed in tumor samples of 58 MBC patients treated with a bevacizumab-containing first-line regimen (learning set). Based on progression-free survival (PFS), patients were divided into responders (R) and non-responders (NR). Differentially expressed miRNAs between R and NR were analyzed in a cohort of 57 patients treated with first-line chemotherapy without bevacizumab (control set), to exclude miRNAs providing prognostic information. MiRNA candidates significantly associated with PFS in multivariate analysis were further validated in tumor samples of 203 patients treated within the phase III trial TANIA randomizing between chemotherapy either alone or with bevacizumab after progression on first-line bevacizumab. Results: Low expression of miR-20a-5p (multivariate p = 0.035) and miR-21-5p (multivariate p = 0.004) were significantly associated with longer PFS in the learning set, but not in the control set. In samples from the TANIA trial, low expression of miR-20a-5p was also significantly associated with longer PFS (hazard ration (HR) 0.60; 95%-CI 0.37–0.89; p = 0.012) and longer overall survival (OS; HR 0.54; 95%-CI 0.32–0.83; p = 0.007) in the bevacizumab arm but not in the chemotherapy-only arm (PFS: HR 0.73, p = 0.119; OS: HR 1.01; p = 0.964). For miR-21-5p no significant association with PFS or OS in both treatment arms was observed. Conclusion: MiR-20a-5p expression in breast cancer tissue was predictive for a greater benefit from bevacizumab-containing therapy in two independent cohorts. [ABSTRACT FROM AUTHOR]

Endometrial cancer (EC) is the most common gynaecologic tumour in the Western world. Previous studies have implicated an imbalance of oestrogens and progestogens in the development of most ECs, while the role of low-grade tissue inflammation remains largely unexplored. We investigated the impact of tumour necrosis factor alpha (TNFα), a central mediator of inflammation and spermatogenesis-associated protein 2 (SPATA2), a regulator of TNF receptor signalling, on clinical outcomes in EC. We evaluated TNFA and SPATA2 transcript levels in 239 EC patients and 25 non-malignant control tissues. Findings were validated in a cohort of 332 EC patients from The Cancer Genome Atlas (TCGA). Expression of TNFA and SPATA2 was increased in EC when compared with control tissues (P < 0.001). TNFA expression correlated with SPATA2 expression in non-malignant (P = 0.003, r S = 0.568) and EC tissue (P = 0.005, r S = 0.179). High TNFA and SPATA2 expression were associated with poor recurrence-free survival (RFS; P = 0.049 and P = 0.018) and disease-specific (P = 0.034 and P = 0.002) survival. Increased SPATA2 expression was also associated with decreased overall survival (OS; P = 0.013). In multivariate analysis, both TNFA and SPATA2 were predictors of clinical outcome. The impact of SPATA2 on RFS and OS could be validated in the TCGA cohort. Our study demonstrates that ECs exhibit a TNF signature which predicts clinical outcome. These findings indicate that TNF signalling modulates the course of EC, which could be therapeutically utilized in the future. [ABSTRACT FROM AUTHOR]

Aims As many current approaches for heart regeneration exert unfavourable side effects, the induction of endogenous repair mechanisms in ischaemic heart disease is of particular interest. Recently, exosomes carrying angiogenic miRNAs have been described to improve heart function. However, it remains challenging to stimulate specific release of reparative exosomes in ischaemic myocardium. In the present study, we sought to test the hypothesis that the physical stimulus of shock wave therapy (SWT) causes the release of exosomes. We aimed to substantiate the pro-angiogenic impact of the released factors, to identify the nature of their cargo, and to test their efficacy in vivo supporting regeneration and recovery after myocardial ischaemia. Methods and results Mechanical stimulation of ischaemic muscle via SWT caused extracellular vesicle (EV) release from endothelial cells both in vitro and in vivo. Characterization of EVs via electron microscopy, nanoparticle tracking analysis and flow cytometry revealed specific exosome morphology and size with the presence of exosome markers CD9, CD81, and CD63. Exosomes exhibited angiogenic properties activating protein kinase b (Akt) and extracellular-signal regulated kinase (ERK) resulting in enhanced endothelial tube formation and proliferation. A miRNA array and transcriptome analysis via next-generation sequencing were performed to specify exosome content. miR-19a-3p was identified as responsible cargo, antimir-19a-3p antagonized angiogenic exosome effects. Exosomes and target miRNA were injected intramyocardially in mice after left anterior descending artery ligation. Exosomes resulted in improved vascularization, decreased myocardial fibrosis, and increased left ventricular ejection fraction as shown by transthoracic echocardiography. Conclusion The mechanical stimulus of SWT causes release of angiogenic exosomes. miR-19a-3p is the vesicular cargo responsible for the observed effects. Released exosomes induce angiogenesis, decrease myocardial fibrosis, and improve left ventricular function after myocardial ischaemia. Exosome release via SWT could develop an innovative approach for the regeneration of ischaemic myocardium. [ABSTRACT FROM AUTHOR]

Open in new tab Download slide Open in new tab Download slide OBJECTIVES Ischaemia and subsequent reperfusion during heart transplantation inevitably result in donor organ injury. Toll-like receptor (TLR)-3 is a pattern recognition receptor activated by viral and endogenous RNA released by injured cells. We hypothesized that ischaemia/reperfusion injury (IRI) leads to RNA release with subsequent TLR3 activation in transplanted hearts. METHODS Human endothelial cells were subjected to IRI and treated with TLR3 agonist polyinosinic–polycytidylic acid or a TLR3/double-stranded RNA complex inhibitor. TLR3 activation was analysed using reporter cells. Gene expression profiles were evaluated via next-generation sequencing. Neutrophil adhesion was assessed in vitro. Syngeneic heart transplantation of wild-type or Tlr3−/− mice was performed following 9 h of cold ischaemia. Hearts were analysed for inflammatory gene expression, cardiac damage, apoptosis and infiltrating leucocytes. RESULTS IRI resulted in RNA release with subsequent activation of TLR3. Treatment with a TLR3 inhibitor abrogated the inflammatory response upon IRI. In parallel, TLR3 stimulation caused activation of the inflammasome. Endothelial IRI resulted in TLR3-dependent adhesion of neutrophils. Tlr3 −/− animals showed reduced intragraft and splenic messenger ribonucleic acid (mRNA) expression of proinflammatory cytokines, resulting in decreased myocardial damage, apoptosis and infiltrating cells. Tlr3 deficiency protected from cardiac damage, apoptosis and leucocyte infiltration after cardiac transplantation. CONCLUSIONS We uncover the release of RNA by injured cells with subsequent activation of TLR3 as a crucial pathomechanism of IRI. Our data indicate that TLR3 represents a novel target in the prevention of IRI in solid organ transplantation. [ABSTRACT FROM AUTHOR]

Function and dysfunction of endothelial cells are regulated by a multitude of factors. Endothelial cell research often requires in vitro cell culture experiments. Hence, various culture media specifically designed to promote endothelial cell growth are available. These strikingly differ in their composition: complex media contain endothelial cell growth supplement (ECGS), an extract produced of bovine brain with undefined amounts of biologically active compounds, whilst defined media contain selected growth factors in defined concentrations. We here compared the effect of seven purchasable endothelial cell culture media on colony outgrowth, proliferation, viability, in vitro angiogenesis and phenotype of mature primary human endothelial cells using feto-placental endothelial cells isolated from chorionic arteries (fpEC). The effect of media on colony outgrowth was additionally tested on umbilical cord blood-derived endothelial progenitor cells (ECFCs). Outgrowth, purity, proliferation and viability differed between media. Outgrowth of fpEC and ECFCs was best in a defined medium containing EGF, FGF2 and VEGF. By contrast, established fpEC isolations proliferated best in complex media containing ECGS, heparin and ascorbic acid. Also viability of cells was higher in complex media. In vitro angiogenesis was most intense in a defined medium containing the highest number of individual growth factors. FACS analysis of surface markers for endothelial cell subtypes revealed that endothelial phenotype of fpEC was unaffected by media composition. Our data demonstrate the fundamental effect of endothelial cell culture media on primary cell isolation success and behaviour. Whether the composition of supplements is suitable also for individual experiments needs to be tested specifically. [ABSTRACT FROM AUTHOR]

Background: Atherosclerosis is the leading cause of death worldwide. The disease development is by and large driven by old age and lifestyle factors, such as diet, physical activity, and smoking. In the present study, we have investigated the effect of exercise and diet on the development of atherosclerosis in young and aged mice.Objective: This study aimed at comparing multiple age-dependent factors that may influence atherosclerosis in a transgenic mouse model.Methods: Young (14 weeks) and aged (49-52 weeks) C57BL/6 wild-type (WT) and atherosclerosis-prone ApoE-/- mice were subjected to physical endurance exercise on a treadmill, with or without a high-fat diet. Five weeks later, the frequencies of regulatory T cells (TREGs) in lymph nodes were assessed by flow cytometry, plasmatic cytokines (interleukin [IL]-1β, IL-6, IL-10, IL-17, interferon-γ, tumor necrosis factor-α, and transforming growth factor [TGF]-β1) levels were determined by Luminex assay. Lipids (cholesterol and triglycerides) and anti-heat shock protein 60 (HSP60) autoantibodies were measured by ELISA. Aortic lesion sizes were assessed by en face imaging. Microarray analysis and qPCR of skeletal muscle gene expression were also performed.Results: Exercise leads to a reduction of aortic lesions in young ApoE-/- and aged WT mice independent of diet. In most groups, this reduction was followed by an increased proportion of TREGs and TGF-β1 levels. Moreover, gene expression analysis showed that exercise seems to affect the AMPK signaling pathway. In particular, PGC-1α1 mRNA was induced in aged WT mice, whereas it was reduced in young ApoE-/- mice. In addition, GSEA analysis showed a marked reduction in the insulin signaling pathway in aged ApoE-/- mice.Conclusion: Practicing endurance exercise seems to be enough for reducing early aortic lesion formation, independent of diet. However, this was only true in mice with smaller aortic lesions, since mice with large, advanced, complicated atherosclerotic plaques did not show any reduction in lesion size with exercise training. [ABSTRACT FROM AUTHOR]

Background: In metastatic colorectal cancer (mCRC), the localization of the primary tumour has been shown to be of prognostic as well as predictive relevance.Methods: With the aim to investigate clinical and molecular disease characteristics with respect to sidedness in a real-world cohort, we analyzed 161 mCRC patients included in the KRAS Registry of the Arbeitsgemeinschaft Medikamentöse Tumortherapie (AGMT) between January 2006 and October 2013.Results: Right-sided mCRC displayed a worse median overall survival (OS) in comparison to left-sided disease (18.1 months [95%-CI: 14.3-40.7] versus 32.3 months [95%-CI: 25.5-38.6]; HR: 1.63 [95%-CI: 1.13-2.84]; p = 0.013). The choice of the biological agent in front-line therapy had a statistically significant impact on median OS in patients with right-sided tumours (anti-epidermal growth factor receptor (EGFR): 10.6 months [95%-CI: 5.2-NA]; anti-vascular endothelial growth factor (VEGF): 26.2 months [95%-CI: 17.9-NA]; HR: 2.69 [95%-CI: 1.30-12.28]; p = 0.015) but not in patients with left-sided tumours (anti-EGFR: 37.0 months [95%-CI: 20.2-56.6]; anti-VEGF: 32.3 months [95%-CI: 23.6-41.1]; HR: 0.97 [95%-CI: 0.56-1.66]; p = 0.905). When evaluating molecular characteristics of tumour samples, we found a clinically meaningful trend towards an inferior OS in TP53 mutant mCRC treated with anti-EGFR based therapy compared to anti-VEGF based therapy (17.1 months [95%-CI: 8.7-NA] versus 38.3 months [95%-CI: 23.6-48.0], HR = 1.95 [95%-CI: 0.95-5.88]; p = 0.066), which was not significantly dependent on sidedness. This was not the case in patients with TP53 wild-type tumours. Therefore we evaluated the combined impact of sidedness and TP53 mutation status in the anti-EGFR treated cohort and patients with left-sided/TP53 wild-type mCRC showed the longest median OS (38.9 months) of all groups (right-sided/TP53 mutant: 12.1 months; right-sided/TP53 wild-type: 8.9 months; left-sided/TP53 mutant: 18.4 months; p = 0.020).Conclusions: TP53 mutation and right-sidedness are associated with shorter OS in patients treated with anti-EGFR based therapy but not with anti-VEGF based therapy. The confirmation of the predictive value of TP53 mutation status in a larger cohort is warranted. [ABSTRACT FROM AUTHOR]

Obesity is a well-known risk factor for the development of cancer, but its influence on the course of disease is still controversial. We investigated the influence of body mass index (BMI) on overall survival (OS) in 502 patients with indolent non-Hodgkin's lymphoma or mantle cell lymphoma in a subgroup analysis of the StiL (Study Group Indolent Lymphomas) NHL1 trial. We defined a cut-off of 22.55 kg/m2 by ROC calculation and Youden Index analysis and stratified patients into "low BMI" and "high BMI". Five-year OS was significantly longer in the high BMI group (82.2%) when compared to that of the low BMI group (66.2%) (HR 0.597; 95%CI 0.370-0.963; p = 0.034). BMI was also an independent prognostic factor for OS in multivariate analysis (HR 0.541; 95%CI 0.332-0.883; p = 0.014). Of note, patients had a significantly lower BMI in the presence than patients in the absence of B-symptoms (p = 0.025). BMI significantly impacts on OS in indolent non-Hodgkin's lymphoma and mantle cell lymphoma, which may be influenced by the effect of B-symptoms on BMI. [ABSTRACT FROM AUTHOR]

Summary: GPViz is a versatile Java-based software for dynamic gene-centered visualization of genomic regions and/or variants. User-defined data can be loaded in common formats as resulting from analysis workflows used in sequencing applications and studied in the context of the gene, the corresponding transcript isoforms, proteins and their domains or other protein features. Both the genomic regions and variants can be also defined interactively. Various gene filter options are provided to enable an intersection of variants, genomic regions and affected protein features. Finally, by using GPViz, we identified differentially expressed exons, which could indicate alternative splicing events, and found somatic variants in different cancer types affecting metabolic proteins. GPViz is freely available at http://icbi.at/gpviz (released under GNU general public license), is based on Java 7 and can be used as a stand-alone or Web Start application.Availability: http://icbi.at/gpvizContact: hubert.hackl@i-med.ac.atSupplementary information: Supplementary data are available at Bioinformatics online. [ABSTRACT FROM AUTHOR]

Tumor infiltrating lymphocytes (TILs) represent a strong independent predictor of relapse and overall survival in colorectal cancer (CRC). However, it appears that a majority of CRCs, i.e., microsatellite stable (MSS) tumors, are refractory to immune checkpoint blockers. The results of recent comprehensive analyses of genomic data provide possible answers. [ABSTRACT FROM AUTHOR]

Background: TPA Induced Sequence 7 acts as a transcriptional co-regulator controlling the expression of genes involved in differentiation of various cell types, including skeletal myoblasts. We and others have shown that TIS7 regulates adult myogenesis through MyoD, one of the essential myogenic regulatory factors. Results: Here, we present data identifying ICln as the specific, novel protein downstream of TIS7 controlling myogenesis. We show that TIS7/ICln epigenetically regulate myoD expression controlling protein methyl transferase activity. In particular, ICln regulates MyoD expression via its interaction with PRMT5 by an epigenetic modification that utilizes symmetrical di-methylation of histone H3 on arginine 8. We provide multiple evidences that TIS7 directly binds DNA, which is a functional feature necessary for its role in transcriptional regulation. Conclusion: We present here a molecular insight into TIS7-specific control of MyoD gene expression and thereby skeletal muscle differentiation. [ABSTRACT FROM AUTHOR]

Filamentous green algae of the genus Zygnema (Zygnematophyceae, Streptophyta) are key components of polar hydro-terrestrial mats where they face various stressors including UV irradiation, freezing, desiccation and osmotic stress. Their vegetative cells can develop into pre-akinetes, i.e. reserve-rich, mature cells. We investigated lipid accumulation and fatty acid (FA) composition upon pre-akinete formation in an Arctic and an Antarctic Zygnema strain using transmission electron microscopy and gas chromatography coupled with mass spectrometry. Pre-akinetes formed after 9 weeks of cultivation in nitrogen-free medium, which was accompanied by massive accumulation of lipid bodies. The composition of FAs was similar in both strains, and a-linolenic acid (C18:3) dominated in young vegetative cells. Pre-akinete formation coincided with a significant change in FA composition. Oleic (C18:1) and linoleic (C18:2) acid increased the most (up to 17- and 8-fold, respectively). Small amounts of long-chain polyunsaturated FAs were also detected, e.g. arachidonic (C20:4) and eicosapentaenoic (C20:5) acid. Pre-akinetes exposed to desiccation at 86% relative humidity were able to recover maximum quantum yield of photosystem II, but desiccation had no major effect on FA composition. The results are discussed with regard to the capability of Zygnema spp. to thrive in extreme conditions. [ABSTRACT FROM AUTHOR]

Multiple system atrophy (MSA) is a fatal rapidly progressive α-synucleinopathy, characterized by α-synuclein accumulation in oligodendrocytes. It is accepted that the pathological α-synuclein accumulation in the brain of MSA patients plays a leading role in the disease process, but little is known about the events in the early stages of the disease. In this study we aimed to define potential roles of the miRNA-mRNA regulatory network in the early pre-motor stages of the disease, i.e., downstream of α-synuclein accumulation in oligodendroglia, as assessed in a transgenic mouse model of MSA. We investigated the expression patterns of miRNAs and their mRNA targets in substantia nigra (SN) and striatum, two brain regions that undergo neurodegeneration at a later stage in the MSA model, by microarray and RNA-seq analysis, respectively. Analysis was performed at a time point when α-synuclein accumulation was already present in oligodendrocytes at neuropathological examination, but no neuronal loss nor deficits of motor function had yet occurred. Our data provide a first evidence for the leading role of gene dysregulation associated with deficits in immune and inflammatory responses in the very early, non-symptomatic disease stages of MSA. While dysfunctional homeostasis and oxidative stress were prominent in SN in the early stages of MSA, in striatum differential gene expression in the non-symptomatic phase was linked to oligodendroglial dysfunction, disturbed protein handling, lipid metabolism, transmembrane transport and altered cell death control, respectively. A large number of putative miRNA-mRNAs interaction partners were identified in relation to the control of these processes in the MSA model. Our results support the role of early changes in the miRNA-mRNA regulatory network in the pathogenesis of MSA preceding the clinical onset of the disease. The findings thus contribute to understanding the disease process and are likely to pave the way towards identifying disease biomarkers for early diagnosis of MSA. [ABSTRACT FROM AUTHOR]

Aims: Invasive fungal infections have significantly increased over the past decades in immunocompromised individuals and high-risk patients. Amphotericin B (AmB) exerts a powerful and broad activity against a vast array of fungi and has a remarkably low rate of microbial resistance. However, most isolates of Aspergillus terreus developed an intrinsic resistance against AmB, and during this study, we characterized the mode of action of this polyene antifungal drug in more detail in resistant (ATR) and rare susceptible (ATS) clinical isolates of A. terreus. Results: We illustrate that AmB treatment changes cellular redox status and promotes the generation of high levels of reactive oxygen species (ROS) in ATS. In contrast, ATR isolates were able to cope better with AmB-induced oxidative stress. Innovation: Most importantly, we demonstrate in this study that coapplication of anti- and pro-oxidants significantly affects AmB efficacy in an antithetic manner-antioxidants and ROS-scavenging agents increase AmB tolerance in susceptible strains, while pro-oxidants render formerly resistant isolates considerably susceptible to the antifungal drug also in vivo in a Galleria animal model. Conclusion: Thereby, our study provides novel therapeutic options to treat formerly resistant fungal strains by a combination of AmB and pro-oxidant compounds. Antioxid. Redox Signal. 23, 1424-1438. [ABSTRACT FROM AUTHOR]

DCs express intrinsic cellular defense mechanisms to specifically inhibit HIV-1 replication. Thus, DCs are productively infected only at very low levels with HIV-1, and this non-permissiveness of DCs is suggested to go along with viral evasion. We now illustrate that complement-opsonized HIV-1 (HIV-C) efficiently bypasses SAMHD1 restriction and productively infects DCs including BDCA-1 DCs. Efficient DC infection by HIV-C was also observed using single-cycle HIV-C, and correlated with a remarkable elevated SAMHD1 T592 phosphorylation but not SAMHD1 degradation. If SAMHD1 phosphorylation was blocked using a CDK2-inhibitor HIV-C-induced DC infection was also significantly abrogated. Additionally, we found a higher maturation and co-stimulatory potential, aberrant type I interferon expression and signaling as well as a stronger induction of cellular immune responses in HIV-C-treated DCs. Collectively, our data highlight a novel protective mechanism mediated by complement opsonization of HIV to effectively promote DC immune functions, which might be in the future exploited to tackle HIV infection. [ABSTRACT FROM AUTHOR]

Background: The transcription factor Ecotropic Virus Integration site 1 (EVI1) regulates cellular proliferation, differentiation, and apoptosis, and its overexpression contributes to an aggressive course of disease in myeloid leukemias and other malignancies. Notwithstanding, knowledge about the target genes mediating its biological and pathological functions remains limited. We therefore aimed to identify and characterize novel EVI1 target genes in human myeloid cells. Methods: U937T_EVI1, a human myeloid cell line expressing EVI1 in a tetracycline regulable manner, was subjected to gene expression profiling. qRT-PCR was used to confirm the regulation of membrane-spanning-4-domains subfamily-A member-3 (MS4A3) by EVI1. Reporter constructs containing various parts of the MS4A3 upstream region were employed in luciferase assays, and binding of EVI1 to the MS4A3 promoter was investigated by chromatin immunoprecipitation. U937 derivative cell lines experimentally expressing EVI1 and/or MS4A3 were generated by retroviral transduction, and tested for their tumorigenicity by subcutaneous injection into severe combined immunodeficient mice. Results: Gene expression microarray analysis identified 27 unique genes that were up-regulated, and 29 unique genes that were down-regulated, in response to EVI1 induction in the human myeloid cell line U937T. The most strongly repressed gene was MS4A3, and its down-regulation by EVI1 was confirmed by qRT-PCR in additional, independent experimental model systems. MS4A3 mRNA levels were also negatively correlated with those of EVI1 in several published AML data sets. Reporter gene assays and chromatin immunoprecipitation showed that EVI1 regulated MS4A3 via direct binding to a promoter proximal region. Experimental re-expression of MS4A3 in an EVI1 overexpressing cell line counteracted the tumor promoting effect of EVI1 in a murine xenograft model by increasing the rate of apoptosis. Conclusions: Our data reveal MS4A3 as a novel direct target of EVI1 in human myeloid cells, and show that its repression plays a role in EVI1 mediated tumor aggressiveness. [ABSTRACT FROM AUTHOR]

Advances in microsurgical techniques and immunosuppressive medication have rendered transplantation of vascularized composite allografts possible, when autologous tissue is neither available nor sufficient for reconstruction. However, skin rejection and side effects of long-term immunosuppression still remain a major hurdle for wide adoption of this excellent reconstructive technique. Histopathologic changes during acute skin rejection in vascular composite allotransplantation often mimic inflammatory skin disorders and are hard to distinguish. Hence, the identification of diagnostic and therapeutic markers specific for skin rejection is of particular clinical need. Here we present novel markers allowing for early differentiation between rejection in hind limb allotransplantation and contact hypersensitivity. Assessment of Ccl7, Il18, and Il1b expression is most indicative of distinguishing skin rejection from skin inflammatory disorders. Gene expression levels varied significantly across skin types and regions, indicating localization specific mechanism of leukocyte migration and infiltration. Expression of Il12b, Il17a, and Il1b gene expression levels differed significantly between rejection and inflammation, independent of the skin type. In synopsis of the RNA expression profile and previously assessed protein expression, the Il1 family appears as a promising option for accurate skin rejection diagnosis and, as a following step, for development of novel rejection treatments. [ABSTRACT FROM AUTHOR]

The International Prognostic Index ( IPI) has been used for decades in diffuse large B-cell lymphoma ( DLBCL). A retrospective cancer registry analysis in North America showed significantly improved results when an enhanced IPI, the National Comprehensive Cancer Network ( NCCN)- IPI was applied. This novel score puts more weight on age and high levels of lactate dehydrogenase ( LDH). Nevertheless, it remains unclear if these results can be extrapolated to the general population. This retrospective bi-centre analysis included 499 unselected DLBCL patients who were treated with rituximab and anthracycline-based chemoimmunotherapy between 2004 and 2013. In our cohort, the NCCN- IPI was more accurate in identifying patients at low or high risk, despite older age, and more patients with increased LDH. Nevertheless, a modified scoring of the risk factors was required to more accurately identify elderly patients with a very favourable diagnosis, suggesting an impaired value of the original NCCN- IPI in the elderly. Serum β2-microglobulin and albumin were retained as independent prognostic factors for survival in a multivariate analysis. Our data confirm, for the first time, the superior prognostic power of the NCCN- IPI in an unselected, middle-European cohort. We furthermore propose a modified NCCN- IPI for more accurate prognostication in the elderly. Albumin and β2-microglobulin levels are likely to add significant information to the NCCN- IPI. [ABSTRACT FROM AUTHOR]

Background: STAT1 has been attributed a function as tumor suppressor. However, in breast cancer data from microarray analysis indicated a predictive value of high mRNA expression levels of STAT1 and STAT1 target genes belonging to the interferon-related signature for a poor response to therapy. To clarify this issue we have determined STAT1 expression levels and activation by different methods, and investigated their association with tumor infiltration by immune cells. Additionally, we evaluated the interrelationship of these parameters and their significance for predicting disease outcome.Methods: Expression of STAT1, its target genes SOCS1, IRF1, CXCL9, CXCL10, CXCL11, IFIT1, IFITM1, MX1 and genes characteristic for immune cell infiltration (CD68, CD163, PD-L1, PD-L2, PD-1, CD45, IFN-γ, FOXP3) was determined by RT-PCR in two independent cohorts comprising 132 breast cancer patients. For a subset of patients, protein levels of total as well as serine and tyrosine-phosphorylated STAT1 were ascertained by immunohistochemistry or immunoblotting and protein levels of CXCL10 by ELISA.Results: mRNA expression levels of STAT1 and STAT1 target genes, as well as protein levels of total and serine-phosphorylated STAT1 correlated with each other in neoplastic tissue. However, there was no association between tumor levels of STAT1 mRNA and tyrosine-phosphorylated STAT1 and between CXCL10 serum levels and CXCL10 expression in the tumor. Tumors with increased STAT1 mRNA amounts exhibited elevated expression of genes characteristic for tumor-associated macrophages and immunosuppressive T lymphocytes. Survival analysis revealed an association of high STAT1 mRNA levels and bad prognosis in both cohorts. A similar prognostically relevant correlation with unfavorable outcome was evident for CXCL10, MX1, CD68, CD163, IFN-γ, and PD-L2 expression in at least one collective. By contrast, activation of STAT1 as assessed by the level of STAT1-Y701 phosphorylation was linked to positive outcome. In multivariate Cox regression, the predictive power of STAT1 mRNA expression was lost when including expression of CXCL10, MX1 and CD68 as confounders.Conclusions: Our study confirms distinct prognostic relevance of STAT1 expression levels and STAT1 tyrosine phosphorylation in breast cancer patients and identifies an association of high STAT1 levels with elevated expression of STAT1 target genes and markers for infiltrating immune cells. [ABSTRACT FROM AUTHOR]

Aims While chronic alterations in cardiac triacylglycerol (TAG) metabolism and accumulation are associated with cardiomyopathy, it is unclear whether TAG catabolizing enzymes such as adipose triglyceride lipase (ATGL) play a role in acquired cardiomyopathies. Importantly, germline deletion of ATGL leads to marked cardiac steatosis and heart failure in part through reducing peroxisome proliferator-activated receptor α (PPARα) activity and subsequent fatty acid oxidation (FAO). However, whether ATGL deficiency specifically in adult cardiomyocytes contributes to impaired PPARα activity, cardiac function, and metabolism is not known. Methods and results To study the effects of acquired cardiac ATGL deficiency on cardiac PPARα activity, function, and metabolism, we generated adult mice with tamoxifen-inducible cardiomyocyte-specific ATGL deficiency (icAtglKO). Within 4–6 weeks following ATGL ablation, icAtglKO mice had markedly increased myocardial TAG accumulation, fibrotic remodelling, and pathological hypertrophy. Echocardiographic analysis of hearts in vivo revealed that contractile function was moderately reduced in icAtglKO mice. Analysis of energy metabolism in ex vivo perfused working hearts showed diminished FAO rates which was not paralleled by markedly impaired PPARα target gene expression. Conclusions This study shows that acquired cardiomyocyte-specific ATGL deficiency in adult mice is sufficient to promote fibrotic and hypertrophic cardiomyopathy and impair myocardial FAO in the absence of markedly reduced PPARα signalling. [ABSTRACT FROM PUBLISHER]