Inga Bekes

VEGF induces ascites in ovarian cancer patients via increasing peritoneal permeability by downregulation of Claudin 5

OBJECTIVE To evaluate the role of VEGF-dependent Claudin 5 production for the development of ascites via influencing endothelial permeability in peritoneal tissue of ovarian cancer patients. METHODS This study investigates the mechanisms of formation of ascites in ovarian cancer patients, performing RT-PCR, VEGF-ELISA and immunohistochemical dual staining for CD31 and Claudin 5. In addition, in order to analyze the connectivity of VEGF, Claudin 5, and endothelial permeability, an endothelial cell/ovarian cancer cell-co-culture-system was established and evaluated performing Western blot analysis and a permeability assay. RESULTS Firstly, VEGF-gene expression was demonstrated for all ovarian cancer and peritoneal biopsies. In addition, quantification of VEGF in the serum and ascites of ovarian cancer patients revealed significantly increased values. We subsequently demonstrated Claudin 5 production in the peritoneal vessels, which was weaker than in the vessels of the controls. Evaluation of endothelial permeability finally showed a VEGF-dependent regulation via Claudin 5 suggesting a mechanism for the development of ascites in ovarian cancer patients. CONCLUSION VEGF induces ascites in ovarian cancer patients. This instance happens due to increased peritoneal permeability, caused by downregulation of the tight junction protein Claudin 5 in the peritoneal endothelium.

Local Renin-Angiotensin System in the Reproductive System

The renin-angiotensin system (RAS) is well known as regulator of electrolytes and blood pressure. Besides this function, there are numerous studies supporting the idea of a local tissue RAS. This system controls the local activity of the different RAS family members, especially of the functional proteins Angiotensin II and Angiotensin (1–7). Those antagonistically acting proteins have been described to be expressed in different organ systems including the human reproductive tract. Therefore, this local RAS has been suspected to be involved in the control and regulation of physiological and pathological conditions in the female reproduction tract. This review of the available literature summarizes the physiological influence of the RAS on the follicular development, ovarian angiogenesis, and placental- and uterine function. In addition, in the second part the role of the RAS concerning ovarian- and endometrial cancer becomes elucidated. This section includes possible novel therapeutic strategies via inhibition of RAS-mediated tumor growth and angiogenesis. Looking at a very complex system of agonistic and antagonistic tissue factors, it may be supposed that the RAS in the female reproduction tract will be of rising scientific interest in the upcoming years.

Does VEGF facilitate local tumor growth and spread into the abdominal cavity by suppressing endothelial cell adhesion, thus increasing vascular peritoneal permeability followed by ascites production in ovarian cancer?

BackgroundOvarian cancer is mostly associated with pathologically regulated permeability of peritoneal vessels, leading to ascites. Here, we investigated the molecular regulation of endothelial permeability by the vascular endothelial growth factor (VEGF) and both tight and adherens junction proteins (VE-cadherin and claudin 5) with regards to the tumor biology of different ovarian cancer types.MethodsSerum and ascites samples before and after surgery, as well as peritoneal biopsies of 68 ovarian cancer patients and 20 healthy controls were collected. In serum and ascites VEGF protein was measured by ELISA. In peritoneal biopsies co-localization of VE-cadherin and claudin 5 was investigated using immunohistochemical dual staining. In addition, the gene expression of VE-cadherin and claudin 5 was quantified by Real-time PCR. Differences in VEGF levels, VE-cadherin and claudin 5 gene expression were analyzed in relation to various tumor characteristics (tumor stage, grading, histological subtypes, resection status after surgery) and then compared to controls. Furthermore, human primary ovarian cancer cells were co-cultured with human umbilical vein endothelial cells (HUVEC) and changes in VE-cadherin and claudin 5 were investigated after VEGF inhibition.ResultsVEGF was significantly increased in tumor patients in comparison to controls and accumulates in ascites. The highest VEGF levels were found in patients diagnosed with advanced tumor stages, with tumors of poor differentiation, or in the group of solid / cystic-solid tumors. Patients with residual tumor after operation showed significantly higher levels of VEGF both before and after surgery as compared to tumor-free resected patients. Results of an immunohistochemical double-staining experiment indicated co-localization of VE-cadherin and claudin 5 in the peritoneal vasculature. Compared to controls, expression of VE-cadherin and claudin 5 was significantly suppressed in peritoneal vessels of tumor patients, but there were no significant differences regarding VE-cadherin and claudin 5 expression in relation to different tumor characteristics. A significant positive correlation was found between VE-cadherin and claudin 5 expression. VEGF inhibition in vitro was associated with significant increase in VE-cadherin and claudin 5.ConclusionsOur results indicate that increased peritoneal permeability in ovarian cancer is due to down-regulation of adhesion proteins via tumor derived VEGF. Advanced ovarian cancer with aggressive tumor biology may be associated with early dysregulation of vascular permeability leading to ascites. These patients may benefit from therapeutic VEGF inhibition.

Regulation of endothelial permeability in the primate corpora lutea: implications for ovarian hyperstimulation syndrome

In a developing human corpus luteum, a closely regulated cellular communication system exists between the luteal steroidogenic cells and endothelial cells. This system guaranties the vascularization process during luteal formation. The process is combined with rapid release of large amounts of progesterone into the bloodstream. The regulation of endothelial proliferation and permeability by LH and human chorionic gonadotropin (hCG) is integral to this process. On the cellular level, endothelial permeability is regulated by intercellular junctions, such as adherens junctions (AJ) and tight junctions (TJ), which act as zipper-like structures between interacting endothelial cells. Several cell junctional proteins are localized to the corpus luteum, including Occludin, Nectin 2, Claudin 1, and Claudin 5, as well as, vascular endothelial (VE)-Cadherin. It has been assumed that regulation of AJ- and TJ-proteins is of particular importance for permeability, and accordingly, for the functionality of the corpus luteum in early pregnancy, because treatment with hCG induces downregulation of juntional proteins in the luteal vessels. The effect of hCG on the adhesive molecules is mediated by VE growth factor (VEGF). On a functional level, the hCG-dependent and VEGF-mediated decrease in junctional proteins causes a decrease in the density of cell-cell closure and, accordingly, an increase in endothelial permeability. In doing so, the different junctional proteins are not only directly influenced by VEGF but also interact among themselves and influence each other reciprocally. Disturbances in this strictly, regulated interactions may explain the development of pathologies with increased vascular permeability, such as the ovarian hyperstimulation syndrome.

Slit2/Robo4 Signaling: Potential Role of a VEGF-Antagonist Pathway to Regulate Luteal Permeability

Introduction The corpus luteum (CL) is dependent on luteal vascular permeability, which is controlled by human chorionic gonadotropin (hCG) via vascular endothelial growth factor (VEGF). In this study we investigated the role of a potential VEGF antagonist pathway - Slit2/Robo4 - and its influence on endothelial cell adhesion. Materials and Methods Luteinized granulosa cells (LGCs) were stimulated with hCG in the absence or presence of a VEGF inhibitor. The expression of VEGF and Slit2 were measured. Human umbilical vein endothelial cells (HUVECs) were stimulated with Slit2 or VEGF, and gene expressions of cadherin 5 (CDH5) and claudin 5 (CLDN5) were measured. Following Robo4 knockdown, CDH5, CLDN5 and endothelial permeability were measured. Results Stimulation of human LGCs with hCG significantly increased VEGF while Slit2 expression was significantly suppressed. Inhibition of VEGF action after hCG stimulation did not change Slit2 suppression. Slit2 knockdown did not affect VEGF expression. While VEGF stimulation of HUVECs significantly suppressed CDH5 and CLDN5 gene expression, stimulation of HUVECs with Slit2 resulted in a significant increase in CDH5 and CLDN5. Robo4 knockdown was done, leading to downregulation of CDH5 and CLDN5 which resulted in significantly increased permeability. Conclusions Our results indicate the existence of a VEGF-antagonist pathway in the CL that decreases vascular permeability. During the functional life of the CL the pathway is suppressed by hCG. It is possible that stimulation of this pathway could be used to treat ovarian hyperstimulation syndrome.

Controlling Vascular Permeability: How Does It Work and What Is the Impact on Normal and Pathological Angiogenesis

The permeability of the vasculature is a property of the capillary wall to obstruct movement of fluid or solutes driven by physiological force. The vasculature is essential for the health of normal tissues, hemostasis, lipid transport, and immune surveillance and is also an influential characteristic of many diseases in which it is greatly increased. The control mechanism of vascular permeability is a complex process that needs to be tightly regulated in order to preserve not only the vascular homeostasis but also its integrity. Here, transcellular and paracellular pathways play an important role as well as direct and indirect influence of the vascular permeability bymolecules or blood pressure. In pathologies the vasculature is often I. Bekes (*) Department of Obstetrics and Gynecology, University of Ulm, Ulm, Germany e-mail: inga.bekes@uniklinik-ulm.de C. Wulff Department of Obstetrics and Gynecology, University of Würzburg, Würzburg, Germany e-mail: wulff_c@ukw.de # Springer Nature Switzerland AG 2019 D. Marmé (ed.), Tumor Angiogenesis, https://doi.org/10.1007/978-3-319-31215-6_48-1 1 affected by the disease process. This may result in neoangiogenesis, where an excessive formation of new, unstable, and hyperpermeable vessels with poor blood flow takes place. In this scenario the vascular endothelial growth factor (VEGF) plays a key role.

Assoziationen zwischen Lymphknotenbefall und Tumoreigenschaften beim Endometriumkarzinom – eine Single-Center Erfahrung

Zielsetzung: Beim Endometriumkarzinom stellt der Lymphknotenbefall einen entscheidenden Prognosefaktor dar; nach aktueller Leitlinie sollte bei allen Stadien – auser T1a, G1 oder G2 – die komplette pelvine und paraaortale Lymphonodektomie Teil des operativen Stagings sein. Die Datenlage hinsichtlich operativer Morbiditat und therapeutischem Nutzen einer kompletten Lymphonodektomie bleibt jedoch letztlich kontrovers. Ziel unserer Untersuchung war es, Subgruppen mit niedrigem Risiko fur einen Lymphknotenbefall zu identifizieren. Materialien/Methoden: In einer retrospektiv beschreibenden Studie wurden die Daten von Patientinnen mit Endometriumkarzinom analysiert, welche an der Universitatsfrauenklinik Ulm zwischen 2000 und 2013 mittels totaler Hysterektomie, bilateraler Salpingo-Oophorektomie und pelviner sowie paraaortaler Lymphonodektomie operiert wurden. Ergebnisse: Es standen Datensatze von 134 Patientinnen fur die Auswertung zur Verfugung. Bei insgesamt 35 (26%) Patientinnen wurde ein Befall pelviner und/oder paraaortaler Lymphknoten festgestellt. Lymphknotenbefall war signifikant mit dem Tumorstadium assoziiert (Chi-Quadrat Test: p < 0,001), wobei pT1a G1/G2 Tumore und pT1a G3 Tumore mit 7% bzw. 5% am seltensten und pT3 Tumore mit 67% am haufigsten befallene Lymphknoten aufwiesen, wahrend pT1b und pT2 Tumore mittlere Haufigkeiten von Lymphknotenbefall zeigten (26% bzw. 19%). Bei 84 (63%) der Patientinnen lag eine Invasion ins Myometrium vor; eine Invasionstiefe von mehr als der Halfte der Myometriumsdicke war signifikant haufiger mit einem positiven Lymphknotenbefall assoziiert als darunter liegende Invasionstiefen (25% vs. 8%; Chi-Quadrat Test: p = 0,048). Zusammenfassung: 7% in der pT1a G1/G2 Gruppe (endometrioide Histologie) wiesen Lymphknotenbefall auf. Diese Patientinnen waren ohne Lymphonodektomie prognostisch unterschatzt worden. Eine Sentinelbiopsie konnte bei geringerer Morbiditat das Staging in diesen Fruhstadien deutlich verbessern.

Diagnostik und Resektion des Mammakarzinoms – State of the Art

Die Diagnostik des Mammakarzinoms basiert neben der klinischen Untersuchung auf bildgebenden Verfahren. Die wichtigste Untersuchungsmethode stellt hierbei die Mammografie dar, welche durch die Mammasonografie und ggf. die KM-MRT erganzt wird. Die Mammografie spielt dabei vor allem eine wichtige Rolle im Rahmen des Screenings asymptomatischer Frauen. Die histologische Diagnosesicherung erfolgt durch die sonografische Stanzbiopsie, stereotaktische Vakuumbiopsie oder selten auch durch die offene Biopsie. Im Falle eines Karzinoms bestehen dann operativ grundsatzlich 2 Moglichkeiten: eine Mastektomie oder ein brusterhaltendes Vorgehen, jeweils in Kombination mit der Sentinellymphknoten-Biopsie und ggf. Axilladissektion. Letztere hat grose diagnostische Bedeutung, da der Nodalstatus als wichtigster Prognoseparameter in hohem Mase Einfluss auf die adjuvante Systemtherapie hat.