German sports clubs’ recruitment of executive board members

This paper analyzes the recruitment of executive board members for German sports clubs, an issue of key interest given that volunteers play a dominant role in these sports clubs. Based on a new, micro-level data set, we examine how members of a sports club advance to become executive board members and how board members differ from ordinary members. Based on the results of our study, we examine whether Michels’ “iron law of oligarchy” applies and whether democratic procedures for elections in sports clubs are replaced, even in part, by trust-based oligarchic structures

Prognostic impact of the bone marrow tumor microenvironment, HLA-I and HLA-Ib expression in MDS and CMML progression to sAML

Genetic aberrations and immune escape are fundamental in MDS and CMML initiation and progression to sAML. Therefore, quantitative and spatial immune cell organization, expression of immune checkpoints (ICP), classical human leukocyte antigen class I (HLA-I) and the non-classical HLA-Ib antigens were analyzed in 274 neoplastic and 50 non-neoplastic bone marrow (BM) biopsies using conventional and multiplex immunohistochemistry and correlated to publicly available dataset. Higher numbers of tissue infiltrating lymphocytes (TILs) were found in MDS/CMML (8.8%) compared to sAML (7.5%) and non-neoplastic BM (5.3%). Higher T cell abundance, including the CD8+ T cell subset, inversely correlated with the number of pathogenic mutations and was associated with blast BM counts, ICP expression, spatial T cell distribution and improved patients’ survival in MDS and CMML. In MDS/CMML, higher PD-1/PD-L1/PD-L2 and HLA-I, but lower HLA-G expression correlated with a significantly better patients’ outcome. Moreover, a closer spatial proximity of T cell subpopulations and their proximity to myeloid blasts showed a stronger prognostic impact when compared to TIL numbers. In sAML – the continuum of MDS and CMML – the number of TILs had no impact on prognosis, but higher CD28 and HLA-I expression correlated with a better outcome of sAML patients. This study underlines the independent prognostic value of the tumor microenvironment in MDS/CMML progression to sAML, which shows the most pronounced immune escape. Moreover, new prognostic markers, like HLA-G expression and spatial T cell distribution, were described for the first time, which might also serve as therapeutic targets

Characterization of lymphatic vessels and lymphatic endothelial cells in type 2 diabetes mellitus : structural, morphological and molecular analysis

Microangiopathy is a major cause of morbidity in type 2 diabetes mellitus (T2DM). While characteristic changes in blood capillary walls and endothelial dysfunction of blood vessels are well studied in type 2 diabetes, examination of lymphatic endothelial cells (LECs) and lymphatic vessels (LVs) is scarcely done. However, complications seen in type 2 diabetes, e.g. increased risk for infections, wound healing defects and obesity, may be related to lymphatic dysfunction. Therefore, we aimed at comprehensively analyzing potential morphological and structural differences of lymphatic endothelial cells and lymphatic vessels in the skin of type 2 diabetes mellitus patients. Further, we wanted to identify gene expression signatures that are deregulated in human dermal lymphatic vessels to define mechanisms that are linked with microvascular complications observed in type 2 diabetes. By immunohistochemistry, basement membranes of lymph vessels were analyzed and blood and lymph vessel densities of diabetic versus normoglycemic skin was evaluated. Further, we identified signs of inflammation, e.g. macrophage infiltration and TNF alpha expression. We compared the gene expression profiles of ex vivo isolated dermal LECs retrieved from normoglycemic versus type 2 diabetic patients using microarrays and subsequent intensive bioinformatical analysis. The up- or downregulated expression of selected candidate genes was confirmed by quantitative real-time PCR and immunofluorescence stainings. Further, we focused on two differentially regulated genes and performed macrophage adhesion, transmigration and chemotatic assays as well as siRNA-mediated knockdown experiments to identify their specific function in lymphatic endothelial cells. Neither prominent alterations in extracellular matrix (ECM) protein deposition, nor morphological BM changes of lymphatic capillaries and collecting LVs were found in the skin of T2DM patients. This excluded the occurrence of diabetic lymphangiopathy comparable to that of blood vessels. However, the evaluation of lymph vessel counts revealed a prominent enhanced lymph vessel density in type 2 diabetic patient's skin. Further, we traced a strong macrophage infiltration in the dermis of type 2 diabetic patients. These macrophages produced vascular endothelial growth factors VEGF-A and VEGF-C, as well as the pro-inflammatory cytokine TNF alpha. Transcriptomal analysis of ex vivo isolated diabetic versus non-diabetic LECs resulted in a list of 180 differently expressed genes. Consistent with earlier studies, we identified several genes that have already been linked to genetic susceptibility for type 2 diabetes, including HP, APOD, HHEX, CD55, ANXA1, LMNA and FABP4. Essentially, we observed multiple changes related to altered LEC proliferation, adhesion and migration. Further, in line with increased TNF alpha abundance, we observed expression changes of CXCL10, VCAM1, CYR61, CXADR, SDC1 and AQP3. TNF alpha treatment of cultured LECs led to deregulated expression of selected genes, recapitulating the array results, indicating that TNF alpha is one major contributor to diabetes-specific gene expression signatures in lymphatic endothelial cells. CXCL10 was confirmed as one important candidate gene only expressed in chronically inflamed lymphatic vessels, contributing to adhesion and transmigration of macrophages and possibly intending to resolute the dermal inflammation. Further, the fatty acid transporter FABP4 was specifically upregulated in LECs and lymphatic vessels in type 2 diabetes in comparison to blood endothelial cells (BECs) and blood vessels. FABP4 was shown to regulate LEC proliferation and permeability in vitro, and pointed out the crucial role of lymphatic vessels in fatty acid transport and metabolism. These data reveal gene sets highlighting the dramatically altered milieu skin lymphatic vessels have to cope with during type 2 diabetes mellitus. Further, we discovered that skin lymphatics show a chronic subacute inflammatory phenotype characterized by macrophage recruitment and de novo lymphangiogenesis. We provide evidence for a paracrine crosstalk, mainly via TNF alpha and CXCL10, fostering macrophage recruitment to LECs as one pathophysiological process that might contribute to persistent inflammation and consecutively, aberrant lymphangiogenesis in the skin.Mikroangiopathie ist eine häufige Komplikation im Verlauf des Typ 2 Diabetes mellitus. Während diabetische Veränderungen der Blutgefäße in der Haut sehr gut charakterisiert sind, ist über mögliche morphologische, strukturelle und molekulare Veränderungen von Lymphgefäßen wenig bekannt. Trotzdem besteht die Vermutung, dass Komplikationen des Diabetes wie erhöhtes Infektionsrisiko, Wundheilungsstörungen und Veränderungen des Fettstoffwechsels auch die Folge einer Lymphgefäßdysfunktion sind. Das Ziel dieser Arbeit war es, potentielle Veränderungen des Lymphgefäßsystems in der Haut auf morphologischer, zellulärer und molekularer Ebene zu beschreiben. Darüber hinaus war es das Ziel, deregulierte Genexpressionsmuster zu erkennen, um sie mit den oben genannten Komplikationen in Verbindung zu bringen. In dieser Arbeit wird eine umfassende immunhistochemische Analyse der diabetischen Haut, inklusive einer Analyse der Basalmembranen der Gefäße, der Gefäßdichte und von Zeichen einer Entzündung, präsentiert. Zusätzlich wurde mit Hilfe der Genechip Microarray-Technologie und nachfolgender intensiver bioinformatischen Analyse das mRNA-Expressionsprofil der diabetischen im Vergleich zu nicht-diabetischen Lymphendothelzellen, die ex vivo aus Patientenhaut isoliert wurden, analysiert. Die Expression wichtiger Kandidatengene wurde mit Hilfe von quantitativen PCR-Analysen sowie Immunfluoreszenzfärbungen bestätigt. Im weiteren Verlauf haben wir funktionell auf zwei Gene fokussiert. Es wurden Makrophagenadhäsions-, -transmigrations und -chemotaxis-Experimente durchgeführt, um ihre Rolle in der Interaktion mit Makrophagen zu analysieren. In siRNA-mediierten knockdown Studien wurde versucht, die spezifische Funktion dieser Gene in lymphatischen Endothelzellen herauszufinden. Signifikante Veränderungen der Basalmembranen von, sowie erhöhte Expression von Extrazellulärmatrixproteinen rund um diabetische Lymphgefäße konnten nicht gefunden werden, was die Existenz einer sogenannten diabetischen Lymphangiopathie ausschloss. Dennoch zeigte ein Vergleich der Lymphgefäßdichte von diabetischer und nicht-diabetischer Haut eine signifikant erhöhte Dichte der Lymphgefäße beim diabetischen Patienten. Zusätzlich konnte in der diabetischen Haut eine starke Infiltration mit Makrophagen nachgewiesen werden. Diese Makrophagen produzierten vaskuläre Wachstumsfaktoren wie VEGF-A und VEGF-C, sowie das pro-inflammatorische Zytokin TNF alpha. Der transkriptionelle Vergleich des mRNA-Profils von diabetischen und nicht-diabetischen Lymphendothelzellen führte zur Identifikation von 180 differentiell regulierten Genen. Neben Genen, die als mögliche Suszeptibilitätsgene für die Entwicklung des Typ 2 Diabetes gelten, wie z.B. HP, APOD, HHEX, CD55, ANXA1, LMNA und FABP4, wurden Transkripte gefunden, die mit der Proliferation von Lymphendothelzellen, sowie mit der Adhäsion und Migration von inflammatorischen Zellen in Gefäßen assoziiert sind, was mit dem immunhistochemischen Befund korrelierte. Es wurden besonders prominente Expressionsunterschiede von CXCL10, VCAM1, CYR61, CXADR, SDC1 und AQP3 detektiert. Diese konnten durch eine Stimulation von Lymphendothelzellen mit TNF alpha in vitro spezifisch rekapituliert werden. CXCL10 wurde als ein wichtiges Chemokin identifiziert, das wahrscheinlich nur im Rahmen einer Entzündung auf dermalen Lymphgefäßen exprimiert wird und das eine wichtige Rolle bei der Adhäsion und Transmigration von Makrophagen und damit möglicherweise bei der Auflösung einer lokalen Entzündung spielt. Es war auch eine Gensignatur von Lipidtransportern in dLECs dereguliert. Darunter war speziell die Expression von FABP4 nicht nur in diabetischen Lymphendothelzellen und Lymphgefäßen signifikant erhöht, sondern grundsätzlich spezifisch für Lymph- im Vergleich zu Blutendothelzellen. Ein Einfuss von FABP4 auf das Verhalten von Lymphendothelzellen, wie endotheliale Proliferation und Permeabilität wurde gezeigt. Die verstärkte Expression von FABP4 in Lymphendothelzellen hebt damit die essentielle Rolle der Lymphgefäße im Rahmen des Lipidtransports und Fettsäurestoffwechsels hervor. Diese Arbeit beleuchtet die Genexpressionsveränderungen von Lymphendothelzellen der Haut im Rahmen des Typ 2 Diabetes mellitus und zeigt mit welchen metabolischen Veränderungen diese zu kämpfen haben. Die erhöhte Lymphgefäßdichte ist mit einer starken Makrophagendichte assoziiert, und es scheint hier eine enge Kommunikation zwischen diesen Zellen, vor allem mittels der Chemokine CXCL10 und TNF alpha stattzufinden. Umgestellt: Zusammenfassend lässt sich sagen, dass es über einen parakrinen Mechanismus zu einer vermehrten Rekrutierung von Makrophagen in der diabetischen Haut kommt, was zu chronischer Entzündung und in der Konsequenz zu vermehrter Lymphangiogenese führt. Dadurch konnten wir zeigen, dass dermale Lymphkapillaren aktiv an den bekannten Phänomenen der verzögerten Wundheilung und persistierenden Entzündungen im Typ 2 Diabetes beteiligt sind.submitted by Monika HämmerleAbweichender Titel laut Übersetzung der Verfasserin/des VerfassersZsfassung in dt. SpracheWien, Med. Univ., Diss., 2012OeBB(VLID)171396

Single Cell Analysis of Cultivated Fibroblasts from Chronic Pancreatitis and Pancreatic Cancer Patients

Cancer-associated fibroblasts (CAFs) play a major role in the progression and drug resistance of pancreatic cancer. Recent studies suggest that CAFs exhibit functional heterogeneity and distinct transcriptomic signatures in pancreatic cancer. Pancreatic fibroblasts also form an integral component in pancreatic diseases such as chronic pancreatitis named disease-associated fibroblasts (DAFs). However, intra-tumoral heterogeneity of CAFs in pancreatic cancer patients and their pivotal role in cancer-related mechanisms have not been fully elucidated. Further, it has not been elucidated whether CAF subtypes identified in pancreatic cancer also exist in chronic pancreatitis. In this study, we used primary isolated fibroblasts from pancreatic cancer and chronic pancreatitis patients using the outgrowth method. Single-cell RNA sequencing (scRNA-seq) was performed, and bioinformatics analysis identified highly variable genes, including factors associated with overall survival of pancreatic cancer patients. The majority of highly variable genes are involved in the cell cycle. Instead of previously classified myofibroblastic (myCAFs), inflammatory (iCAFs), and antigen-presenting (ap) CAFs, we identified a myCAFs-like subtype in all cases. Most interestingly, after cell cycle regression, we observed 135 highly variable genes commonly identified in chronic pancreatitis and pancreatic cancer patients. This study is the first to conduct scRNAseq and bioinformatics analyses to compare CAFs/DAFs from both chronic pancreatitis and pancreatic cancer patients. Further studies are required to select and identify stromal factors in DAFs from chronic pancreatitis cases, which are commonly expressed also in CAFs potentially contributing to pancreatic cancer development

Periostin is associated with prognosis and immune cell infiltration in pancreatic adenocarcinoma based on integrated bioinformatics analysis

Background:Pancreatic cancer is one of the most aggressive human malignancies.Previous research has shown that periostin (POSTN) promotes pancreatic cancer cellproliferation, migration, and invasion. Further, POSTN is involved in tumor microenvi-ronment remodeling during tumor progression. However, the relationship betweenPOSTN expression, immune cell infiltration, and the efficacy of immunotherapy inpancreatic cancer is unclear.Methods:We conducted a comprehensive evaluation of POSTN differential expres-sion, examining mRNA and protein levels. To gather data, we utilized various data-bases including gene expression profiling interactive analysis 2 (GEPIA2), geneexpression omnibus (GEO), and the human protein atlas (HPA). To investigate thecorrelation betweenPOSTNexpression and clinical characteristics, we analyzed datafrom the Kaplan–Meier plotter database and clinical data sourced from the cancergenome atlas (TCGA). Furthermore, we performed gene ontology (GO) analysis,Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis, and gene setenrichment analysis (GSEA). Additionally, we explored the relationship betweenPOSTNexpression and immune cell infiltration, as well as the immunophenoscore(IPS), by leveraging the cancer immunome atlas (TCIA) database. Lastly, we examinedthe tumor mutational burden (TMB) in pancreatic cancer in relation toPOSTNexpression.Results:When compared with healthy pancreatic tissues, pancreatic cancer tissuesdisplayed significantly higher levels of POSTN, which was indicative of a worse prog-nosis.POSTNexpression was closely associated with extracellular matrix (ECM) orga-nization, ECM-receptor interaction, and focal adhesion by GO, KEGG pathway, and GSEA analyses. Higher expression ofPOSTNwas associated with increased infiltra-tion of M2 macrophages. Additionally, increased IPS was linked to lowerPOSTNexpression. IPS scores for CTLA4, PD-1/PDL1, and CTLA4/PD-1/PDL1 immunecheckpoint inhibitors were also higher in thePOSTN-low expression group, suggest-ing that lower expression ofPOSTNis associated with a better outcome with check-point inhibitor treatment.Conclusion:POSTN is related to pancreatic cancer prognosis, and may influenceimmune cell infiltration. High expression ofPOSTNis predicted to correlate withlower sensitivity to immunotherapy with checkpoint inhibitors in pancreaticcancer

Palatal wound healing using a xenogeneic collagen matrix - histological outcomes of a randomized controlled clinical trial

AIM The aim of this study was to test whether or not a collagen matrix can improve early wound healing compared to spontaneous healing based on histological and immunohistologic analyses. METHODS In 20 volunteers, 6 mm punch biopsies were harvested at the palate. A xenogeneic collagen matrix (XCM) was sutured in one site; the other one was left untreated (control). Biopsies with a diameter of 8 mm were subsequently obtained at 4, 8, 15 and 29 days and histological and immunohistologic analyses were performed. RESULTS At day 4, wound bed keratinization amounted to 12.4 ± 7.5% (control) and 18.0 ± 10.2% (XCM). This increased up to day 8 (19.7 ± 25.5% control; 29.1 ± 8.0% XCM) and reached complete keratinization at day 15 in both groups. The quantitative analyses of the superficial compartment measured an increase in the amount of granulation tissue (32-88% control; 14-41% XCM) from day 4 to day 8. Angiogenesis was first detected at 8 days. At day 29, the amount of connective tissue in all compartments reached values similar to the native tissue at baseline. CONCLUSIONS The application of a XCM as a wound dressing on palatal wounds might be beneficial in the early stages of wound healing. Further research with a larger sample size is needed to confirm these results

Loss of the abundant nuclear non-coding RNA MALAT1 is compatible with life and development

The metastasis-associated lung adenocarcinoma transcript 1, MALAT1, is a long non-coding RNA (lncRNA) that has been discovered as a marker for lung cancer metastasis. It is highly abundant, its expression is strongly regulated in many tumor entities including lung adenocarcinoma and hepatocellular carcinoma as well as physiological processes, and it is associated with many RNA binding proteins and highly conserved throughout evolution. The nuclear transcript MALAT-1 has been functionally associated with gene regulation and alternative splicing and its regulation has been shown to impact proliferation, apoptosis, migration and invasion. Here, we have developed a human and a mouse knockout system to study the loss-of-function phenotypes of this important ncRNA. In human tumor cells, MALAT1 expression was abrogated using Zinc Finger Nucleases. Unexpectedly, the quantitative loss of MALAT1 did neither affect proliferation nor cell cycle progression nor nuclear architecture in human lung or liver cancer cells. Moreover, genetic loss of Malat1 in a knockout mouse model did not give rise to any obvious phenotype or histological abnormalities in Malat1-null compared with wild-type animals. Thus, loss of the abundant nuclear long ncRNA MALAT1 is compatible with cell viability and normal development

Posttranscriptional destabilization of the liver-specific long noncoding RNA HULC by the IGF2 mRNA-binding protein 1 (IGF2BP1).

Selected long noncoding RNAs (lncRNAs) have been shown to play important roles in carcinogenesis. Although the cellular functions of these transcripts can be diverse, many lncRNAs regulate gene expression. In contrast, factors that control the expression of lncRNAs remain largely unknown. Here we investigated the impact of RNA binding proteins on the expression of the liver cancer-associated lncRNA HULC (highly up-regulated in liver cancer). First, we validated the strong up-regulation of HULC in human hepatocellular carcinoma. To elucidate posttranscriptional regulatory mechanisms governing HULC expression, we applied an RNA affinity purification approach to identify specific protein interaction partners and potential regulators. This method identified the family of IGF2BPs (IGF2 mRNA-binding proteins) as specific binding partners of HULC. Depletion of IGF2BP1, also known as IMP1, but not of IGF2BP2 or IGF2BP3, led to an increased HULC half-life and higher steady-state expression levels, indicating a posttranscriptional regulatory mechanism. Importantly, HULC represents the first IGF2BP substrate that is destabilized. To elucidate the mechanism by which IGF2BP1 destabilizes HULC, the CNOT1 protein was identified as a novel interaction partner of IGF2BP1. CNOT1 is the scaffold of the human CCR4-NOT deadenylase complex, a major component of the cytoplasmic RNA decay machinery. Indeed, depletion of CNOT1 increased HULC half-life and expression. Thus, IGF2BP1 acts as an adaptor protein that recruits the CCR4-NOT complex and thereby initiates the degradation of the lncRNA HULC. Conclusion: Our findings provide important insights into the regulation of lncRNA expression and identify a novel function for IGF2BP1 in RNA metabolism. (Hepatology 2013)