TY - THES A1 - Müller, Verena T1 - Candida albicans-induzierte Genexpression in primären humanen Endothelzellen - Mechanismen der Signaltransduktion und Möglichkeiten der Intervention T1 - Candida albicans-induced gene expression in primary human endothelial cells - mechanisms of signal transduction and possibilities of intervention N2 - Endothelzellen sind ein aktiver Bestandteil der angeborenen Immunabwehr des Menschen gegen mikrobielle Pathogene. Unter ungünstigen Bedingungen kann die Abwehrreaktion sogar zu einer lebensbedrohlichen Sepsis führen. Hier wurde die bislang wenig bekannte Endothelantwort auf den fakultativ humanpathogenen Hefepilz Candida albicans, einem der häufigsten Verursacher von letaler Sepsis beim Menschen, näher untersucht. Mittels Oligonukleotid-Mikroarray-Analyse von HUVEC nach Exposition mit C. albicans konnten 56 hochregulierte Gene identifiziert werden, während 69 Gene herunterreguliert wurden. Ein bedeutender Anteil der regulierten Gene ist an Prozessen der angeborenen Immunantwort beteiligt und dient hauptsächlich der Rekrutierung von Neutrophilen. Weitere Untersuchungen ergaben eine zentrale Rolle des proinflammatorischen NF-kappaB-Weges bei der Regulation des Candida-induzierten Transkriptoms von Endothelzellen. Es konnte gezeigt werden, dass C. albicans diesen Signalweg sequenziell aktiviert. Zusätzlich konnte durch die Expression einer dominant-negativen Mutante einer Signalkomponente des NF-kappaB-Signalwegs die Candida-vermittelte Induktion von kappaB-abhängigen Genen gehemmt werden. Mit einem pharmakologischen Ansatz wurde der p38 MAP Kinase-Signalweg als weiterer bedeutsamer Signalweg identifiziert, der die Expression einzelner Candida-Zielgene wie CXCL8/IL-8 moduliert. Schließlich wurde gezeigt, dass die Candida-induzierte NF-kappaB-Aktivierung im untersuchten endothelialen Zellsystem unabhängig von den Toll-like Rezeptoren TLR2 und TLR4 geschieht, die üblicherweise an der Erkennung mikrobieller Pathogene beteiligt sind. Durch RNA-Interferenz-Experimente konnte jedoch dargelegt werden, dass das Adaptermolekül MyD88 und die Kinase IRAK1, die beide entscheidend an der TLR-vermittelten Signaltransduktion beteiligt sind, essentiell für die Weiterleitung des Signals in Endothelzellen sind. Nachfolgend konnte mit TLR3 zumindest einer der signaltransduzierenden Rezeptoren identifiziert werden. Als erste umfassende Untersuchung der endothelialen Antwort auf Candida albicans erlaubt die vorliegende Arbeit neue Einblicke in die komplexen Signalmuster von Endothelzellen, die dieser klinisch bedeutende Krankheitserreger auslöst. N2 - Endothelial cells (ECs) actively participate in the innate defence against microbial pathogens. Under unfavourable conditions defence reactions can even turn into life-threatening responses resulting in sepsis. Here the so far largely unknown EC reaction patterns to Candida albicans were studied. C. albicans is a facultative human pathogenic fungus and a major cause of lethality in septic patients. Oligonucleotide microarray analysis revealed 56 genes that were transcriptionally up-regulated and 69 that were suppressed upon exposure of ECs to C. albicans. A major portion of these genes is involved in defence mechanisms of the innate immune system with a high representation of genes serving the recruitment of neutrophils to sites of infection. Further examination of candidate signalling cascades established a central role of the proinflammatory NF-kappaB pathway in the regulation of the Candida-modulated transcriptome of ECs. It was shown that the NF-kappaB signalling pathway becomes activated at various levels. In addition, expression of a dominant negative mutant of a NF-kappaB signalling pathway compound blocked the Candida-induced kappaB-dependent gene expression. Using a pharmacological approach the stress-activated p38 mitogen-activated protein (MAP) kinase pathway was identified as a second major regulatory pathway which critically contributes to the regulation of selected Candida target genes such as CXCL8/IL-8. Candida-induced NF-kappaB activation is mediated independently of Toll-like receptors (TLR) 2 and 4 that commonly have been implicated with microbial pattern recognition. Nevertheless, knock-down of the adapter molecule MyD88 and the essential downstream kinase of TLR-, IL-1R- and IL-18R-signalling, IRAK1, suggested that recognition and signalling via a TLR apart from TLR2/TLR4 is crucial for Candida-induced gene expression in primary ECs. Finally, RNAi experiments indicate that most likely TLR3 represents this receptor. These data provide the first comprehensive analysis of endothelial gene responses to Candida albicans and present novel insights into the complex signalling patterns triggered by this important pathogen. KW - Angeborene Immunität KW - Endothelzelle KW - Toll-like-Rezeptoren KW - Candida albicans KW - Entzündung KW - innate immunity KW - endothelial cell KW - toll-like receptors KW - Candida albicans KW - inflammation Y1 - 2007 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-26224 ER - TY - JOUR A1 - van Dinther, Maarten A1 - Zhang, Juan A1 - Weidauer, Stella E. A1 - Boschert, Verena A1 - Muth, Eva-Maria A1 - Knappik, Achim A1 - de Gorter, David J. J. A1 - van Kasteren, Puck B. A1 - Frisch, Christian A1 - Müller, Thomas D. A1 - ten Dijke, Peter T1 - Anti-Sclerostin Antibody Inhibits Internalization of Sclerostin and Sclerostin-Mediated Antagonism of Wnt/LRP6 Signaling JF - PLoS ONE N2 - Sclerosteosis is a rare high bone mass disease that is caused by inactivating mutations in the SOST gene. Its gene product, Sclerostin, is a key negative regulator of bone formation and might therefore serve as a target for the anabolic treatment of osteoporosis. The exact molecular mechanism by which Sclerostin exerts its antagonistic effects on Wnt signaling in bone forming osteoblasts remains unclear. Here we show that Wnt3a-induced transcriptional responses and induction of alkaline phosphatase activity, an early marker of osteoblast differentiation, require the Wnt co-receptors LRP5 and LRP6. Unlike Dickkopf1 (DKK1), Sclerostin does not inhibit Wnt-3a-induced phosphorylation of LRP5 at serine 1503 or LRP6 at serine 1490. Affinity labeling of cell surface proteins with \([^{125} I]\) Sclerostin identified LRP6 as the main specific Sclerostin receptor in multiple mesenchymal cell lines. When cells were challenged with Sclerostin fused to recombinant green fluorescent protein (GFP) this was internalized, likely via a Clathrin-dependent process, and subsequently degraded in a temperature and proteasome-dependent manner. Ectopic expression of LRP6 greatly enhanced binding and cellular uptake of Sclerostin-GFP, which was reduced by the addition of an excess of non-GFP-fused Sclerostin. Finally, an anti-Sclerostin antibody inhibited the internalization of Sclerostin-GFP and binding of Sclerostin to LRP6. Moreover, this antibody attenuated the antagonistic activity of Sclerostin on canonical Wnt-induced responses. KW - gene KW - rat model KW - Dickkopf proteins KW - postmenopausal osteoporosis KW - increases bone-formation KW - WNT KW - LRP6 KW - density KW - receptor KW - ligand Y1 - 2013 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-130981 VL - 8 IS - 4 ER - TY - JOUR A1 - Boschert, Verena A1 - Klenk, Nicola A1 - Abt, Alexander A1 - Raman, Sudha Janaki A1 - Fischer, Markus A1 - Brands, Roman C. A1 - Seher, Axel A1 - Linz, Christian A1 - Müller-Richter, Urs D. A. A1 - Bischler, Thorsten A1 - Hartmann, Stefan T1 - The influence of Met receptor level on HGF-induced glycolytic reprogramming in head and neck squamous cell carcinoma JF - International Journal of Molecular Sciences N2 - Head and neck squamous cell carcinoma (HNSCC) is known to overexpress a variety of receptor tyrosine kinases, such as the HGF receptor Met. Like other malignancies, HNSCC involves a mutual interaction between the tumor cells and surrounding tissues and cells. We hypothesized that activation of HGF/Met signaling in HNSCC influences glucose metabolism and therefore substantially changes the tumor microenvironment. To determine the effect of HGF, we submitted three established HNSCC cell lines to mRNA sequencing. Dynamic changes in glucose metabolism were measured in real time by an extracellular flux analyzer. As expected, the cell lines exhibited different levels of Met and responded differently to HGF stimulation. As confirmed by mRNA sequencing, the level of Met expression was associated with the number of upregulated HGF-dependent genes. Overall, Met stimulation by HGF leads to increased glycolysis, presumably mediated by higher expression of three key enzymes of glycolysis. These effects appear to be stronger in Met\(^{high}\)-expressing HNSCC cells. Collectively, our data support the hypothesized role of HGF/Met signaling in metabolic reprogramming of HNSCC. KW - HNSCC KW - head and neck cancer KW - HGF KW - Met KW - cancer metabolism Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-235995 SN - 1422-0067 VL - 21 IS - 2 ER - TY - JOUR A1 - Hendricks, Anne A1 - Müller, Sophie A1 - Fassnacht, Martin A1 - Germer, Christoph-Thomas A1 - Wiegering, Verena A. A1 - Wiegering, Armin A1 - Reibetanz, Joachim T1 - Impact of lymphadenectomy on the oncologic outcome of patients with adrenocortical carcinoma — a systematic review and meta-analysis JF - Cancers N2 - (1) Background: Locoregional lymphadenectomy (LND) in adrenocortical carcinoma (ACC) may impact oncological outcome, but the findings from individual studies are conflicting. The aim of this systematic review and meta-analysis was to determine the oncological value of LND in ACC by summarizing the available literature. (2) Methods: A systematic search on studies published until December 2020 was performed according to the PRISMA statement. The primary outcome was the impact of lymphadenectomy on overall survival (OS). Two separate meta-analyses were performed for studies including patients with localized ACC (stage I–III) and those including all tumor stages (I–IV). Secondary endpoints included postoperative mortality and length of hospital stay (LOS). (3) Results: 11 publications were identified for inclusion. All studies were retrospective studies, published between 2001–2020, and 5 were included in the meta-analysis. Three studies (N = 807 patients) reported the impact of LND on disease-specific survival in patients with stage I–III ACC and revealed a survival benefit of LND (hazard ratio (HR) = 0.42, 95% confidence interval (95% CI): 0.26–0.68). Based on results of studies including patients with ACC stage I–IV (2 studies, N = 3934 patients), LND was not associated with a survival benefit (HR = 1.00, 95% CI: 0.70–1.42). None of the included studies showed an association between LND and postoperative mortality or LOS. (4) Conclusion: Locoregional lymphadenectomy seems to offer an oncologic benefit in patients undergoing curative-intended surgery for localized ACC (stage I–III). KW - adrenocortical carcinoma KW - adrenal cancer KW - lymphadenectomy KW - lymph node dissection KW - LND KW - LNE KW - review KW - meta-analysis Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-254798 SN - 2072-6694 VL - 14 IS - 2 ER - TY - JOUR A1 - Riedmeier, Maria A1 - Decarolis, Boris A1 - Haubitz, Imme A1 - Müller, Sophie A1 - Uttinger, Konstantin A1 - Börner, Kevin A1 - Reibetanz, Joachim A1 - Wiegering, Armin A1 - Härtel, Christoph A1 - Schlegel, Paul-Gerhardt A1 - Fassnacht, Martin A1 - Wiegering, Verena T1 - Adrenocortical carcinoma in childhood: a systematic review JF - Cancers N2 - Adrenocortical tumors are rare in children. This systematic review summarizes the published evidence on pediatric adrenocortical carcinoma (ACC) to provide a basis for a better understanding of the disease, investigate new molecular biomarkers and therapeutic targets, and define which patients may benefit from a more aggressive therapeutic approach. We included 137 studies with 3680 ACC patients (~65% female) in our analysis. We found no randomized controlled trials, so this review mainly reflects retrospective data. Due to a specific mutation in the TP53 gene in ~80% of Brazilian patients, that cohort was analyzed separately from series from other countries. Hormone analysis was described in 2569 of the 2874 patients (89%). Most patients were diagnosed with localized disease, whereas 23% had metastasis at primary diagnosis. Only 72% of the patients achieved complete resection. In 334 children (23%), recurrent disease was reported: 81% — local recurrence, 19% (n = 65) — distant metastases at relapse. Patients < 4 years old had a different distribution of tumor stages and hormone activity and better overall survival (p < 0.001). Although therapeutic approaches are typically multimodal, no consensus is available on effective standard treatments for advanced ACC. Thus, knowledge regarding pediatric ACC is still scarce and international prospective studies are needed to implement standardized clinical stratifications and risk-adapted therapeutic strategies. KW - pediatric adrenocortical cancer KW - pediatric adrenocortical adenoma KW - pediatric adrenocortical tumor KW - prognostic factors KW - therapy Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-248507 SN - 2072-6694 VL - 13 IS - 21 ER - TY - JOUR A1 - Boschert, Verena A1 - Teusch, Jonas A1 - Aljasem, Anwar A1 - Schmucker, Philipp A1 - Klenk, Nicola A1 - Straub, Anton A1 - Bittrich, Max A1 - Seher, Axel A1 - Linz, Christian A1 - Müller-Richter, Urs D. A. A1 - Hartmann, Stefan T1 - HGF-induced PD-L1 expression in head and neck cancer: preclinical and clinical findings JF - International Journal of Molecular Sciences N2 - Head and neck squamous cell carcinoma (HNSCC) is a widespread disease with a low survival rate and a high risk of recurrence. Nowadays, immune checkpoint inhibitor (ICI) treatment is approved for HNSCC as a first-line treatment in recurrent and metastatic disease. ICI treatment yields a clear survival benefit, but overall response rates are still unsatisfactory. As shown in different cancer models, hepatocyte growth factor/mesenchymal–epithelial transition (HGF/Met) signaling contributes to an immunosuppressive microenvironment. Therefore, we investigated the relationship between HGF and programmed cell death protein 1 (PD-L1) expression in HNSCC cell lines. The preclinical data show a robust PD-L1 induction upon HGF stimulation. Further analysis revealed that the HGF-mediated upregulation of PD-L1 is MAP kinase-dependent. We then hypothesized that serum levels of HGF and soluble programmed cell death protein 1 (sPD-L1) could be potential markers of ICI treatment failure. Thus, we determined serum levels of these proteins in 20 HNSCC patients before ICI treatment and correlated them with treatment outcomes. Importantly, the clinical data showed a positive correlation of both serum proteins (HGF and sPD-L1) in HNSCC patient’s sera. Moreover, the serum concentration of sPD-L1 was significantly higher in ICI non-responsive patients. Our findings indicate a potential role for sPD-L1 as a prognostic marker for ICI treatment in HNSCC. KW - HNSCC KW - head and neck cancer KW - HGF KW - Met KW - PD-L1 KW - immune therapy Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-236220 SN - 1422-0067 VL - 21 IS - 20 ER - TY - JOUR A1 - Boschert, Verena A1 - Teusch, Jonas A1 - Müller-Richter, Urs D. A. A1 - Brands, Roman C. A1 - Hartmann, Stefan T1 - PKM2 modulation in head and neck squamous cell carcinoma JF - International Journal of Molecular Sciences N2 - The enzyme pyruvate kinase M2 (PKM2) plays a major role in the switch of tumor cells from oxidative phosphorylation to aerobic glycolysis, one of the hallmarks of cancer. Different allosteric inhibitors or activators and several posttranslational modifications regulate its activity. Head and neck squamous cell carcinoma (HNSCC) is a common disease with a high rate of recurrence. To find out more about PKM2 and its modulation in HNSCC, we examined a panel of HNSCC cells using real-time cell metabolic analysis and Western blotting with an emphasis on phosphorylation variant Tyr105 and two reagents known to impair PKM2 activity. Our results show that in HNSCC, PKM2 is commonly phosphorylated at Tyrosine 105. Its levels depended on tyrosine kinase activity, emphasizing the importance of growth factors such as EGF (epidermal growth factor) on HNSCC metabolism. Furthermore, its correlation with the expression of CD44 indicates a role in cancer stemness. Cells generally reacted with higher glycolysis to PKM2 activator DASA-58 and lower glycolysis to PKM2 inhibitor Compound 3k, but some were more susceptible to activation and others to inhibition. Our findings emphasize the need to further investigate the role of PKM2 in HNSCC, as it could aid understanding and treatment of the disease. KW - HNSCC KW - head and neck cancer KW - cancer metabolism KW - glycolysis KW - PKM2 KW - Warburg effect KW - CD44 KW - Compound 3k KW - DASA-58 KW - AMPK KW - TXNIP Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-284458 SN - 1422-0067 VL - 23 IS - 2 ER -