Marginal zone B cells emerge as a critical component of pregnancy well-being

The success of eutherian mammal evolution was certainly supported by the ability of the already existing immune system to adapt to the presence of the semi-allogeneic fetus without losing the capability to defend the mother against infections. This required the acquisition of highly regulated and coordinated immunological mechanisms. Failures in the development of these strategies not only lead to the interruption of pregnancy but also compromise maternal health. Alongside changes on the cytokine profile - expansion of tolerogenic dendritic and regulatory T cells - a profound adaptation of the B cell compartment during pregnancy was recently described. Among others, the suppression of B cell lymphopoiesis and B cell lymphopenia were proposed to be protective mechanisms tending to reduce the occurrence of autoreactive B cells that might recognize fetal structures and put pregnancy on risk. On the other hand, expansion of the pre-activated marginal zone (MZ) B cell phenotype was described as a compensatory strategy launched to overcome B cell lymphopenia thus ensuring a proper defense. In this work, using an animal model of pregnancy disturbances, we demonstrated that the suppression of B cell lymphopoiesis as well as splenic B cell lymphopenia occur independently of pregnancy outcome. However, only animals undergoing normal pregnancies, but not those suffering from pregnancy disturbances, could induce an expansion and activation of the MZ B cells. Hence, our results clearly show that MZ B cells, probably due to the production of natural protective antibodies, participate in the fine balance of immune activation required for pregnancy well-being.Fil: Muzzio, Damián Oscar. University of Greifswald; AlemaniaFil: Ziegler, Katharina B.. University of Greifswald; AlemaniaFil: Ehrhardt, Jens. University of Greifswald; AlemaniaFil: Zygmunt, Marek. University of Greifswald; AlemaniaFil: Jensen, Cristian Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; Argentina. University of Greifswald; Alemania. Universidad Nacional Arturo Jauretche; Argentin

Expression of IL-33 Receptor Is Significantly Up-Regulated in B Cells During Pregnancy and in the Acute Phase of Preterm Birth in Mice

Interleukin-33 (IL-33) is a mucosal alarmin belonging to the IL-1 cytokine family and is now recognized to have a key role in innate and adaptive immunity, contributing to tissue homeostasis and response to environmental stresses. In addition, IL-33 has also been shown to work as a positive regulator that initiates and maintains a Th2 immune response. In the context of pregnancy, it has been recently demonstrated that upon certain stress conditions, such as an infection induced inflammation, IL-33 is released from the uterine mucosa and triggers decidual B cells to produce anti-inflammatory molecules, which in turn restore immune homeostasis and prevents the development of preterm birth. In this study we therefore performed a detailed characterization of IL-33 receptor (Il1rl1 or ST2) expression in B cells during normal pregnancy, as well as in a mouse model of preterm birth. We observed that splenic B cells significantly up-regulate the expression of Il1rl1 during pregnancy and identified the B1 B cell population as the main ST2-expressing B cell subset. A further kinetic analysis showed that percentages of ST2-expressing B1 B cells are significantly augmented on days 12 and 14 of pregnancy, both in the spleen and peritoneal cavity of pregnant mice, and then drop toward the end of pregnancy to the levels observed in non-pregnant animals. Furthermore, using a mouse model of LPS-induced preterm birth, we demonstrated that not only are the percentages of ST2-expressing B1 B cells significantly enlarged in the spleen during the acute phase of preterm birth, but decidual B cells also significantly up-regulate ST2 expression as compared to term-pregnant mice. Overall, our results suggest a functional role of ST2 expression in B cells during pregnancy and reinforce the importance of the IL-33/ST2 axis in B cells as a critical mechanism to control inflammation-induced preterm birth.Fil: Valeff, Natalin Jimena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaFil: Juriol, Lorena Vanesa. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaFil: Quadrana, Florencia. Laboratory For Immunology Of Pregnancy; ArgentinaFil: Muzzio, Damián Oscar. Universität Greifswald; AlemaniaFil: Zygmunt, Marek. Universität Greifswald; AlemaniaFil: Quiroga, María Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Ventimiglia, María Silvia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaFil: Jensen, Cristian Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; Argentina. Universidad Nacional Arturo Jauretche; Argentin

Association between β2-adrenoceptor (ADRB2) haplotypes and insulin resistance in PCOS

OBJECTIVE: The aim of this study was to explore β2-adrenoceptor (ADRB2) haplotype associations with phenotypes and quantitative traits related to insulin resistance (IR) and the metabolic syndrome (MS) in a polycystic ovary syndrome (PCOS) population. A secondary purpose was to assess the association between ADRB2 haplotype and PCOS. DESIGN: Genetic polymorphism analysis. Cross-sectional case-control association study. SETTING: Medical University Hospital and research laboratory. PATIENTS: One hundred and sixty-five unrelated women with PCOS and 116 unrelated women without PCOS (control sample). MEASUREMENTS: Clinical and biochemical measurements, and ADRB2 genotyping in PCOS patients and control subjects. METHODS: ADRB2 haplotypes (comprising rs1042711, rs1801704, rs1042713 and rs1042714 in that order), genotyping and statistical analysis to evaluate associations with continuous variables and traits related to IR and MS in a PCOS population. Associations between ADRB2 haplotypes and PCOS were also assessed. RESULTS: We observed an age-adjusted association between ADRB2 haplotype CCGG and lower insulin (P = 0·018) and HOMA (P = 0·008) in the PCOS sample. Interestingly, the expected differences in surrogate measures of IR between cases and controls were not significant in CCGG/CCGG carriers. In the case-control study, genotype CCGG/CCGG was associated with a 14% decrease in PCOS risk (P = 0·043), taking into account confounding variables. CONCLUSIONS: Haplotype I (CCGG) has a protective role for IR and MS in PCOS.Fil: Tellechea, Mariana Lorena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; Argentina. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Microbiología, Inmunología y Biotecnología. Cátedra de Genética y Biología Molecular; ArgentinaFil: Muzzio, Damián Oscar. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Microbiología, Inmunología y Biotecnología. Cátedra de Genética y Biología Molecular; ArgentinaFil: Iglesias Molli, Andrea Elena. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Microbiología, Inmunología y Biotecnología. Cátedra de Genética y Biología Molecular; ArgentinaFil: Belli, Susana H.. Hospital Carlos Durand; ArgentinaFil: Graffigna, Mabel N.. Hospital Carlos Durand; ArgentinaFil: Levalle, Oscar A.. Hospital Carlos Durand; ArgentinaFil: Frechtel, Gustavo Daniel. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Microbiología, Inmunología y Biotecnología. Cátedra de Genética y Biología Molecular; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Cerrone, Gloria Edith. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Microbiología, Inmunología y Biotecnología. Cátedra de Genética y Biología Molecular; Argentin

Cytochrome P450 17A1 inhibitor abiraterone attenuates cellular growth of prostate cancer cells independently from androgen receptor signaling by modulation of oncogenic and apoptotic pathways

Abiraterone provides significant survival advantages in prostate cancer (PC), however, the current understanding of the molecular mechanisms of abiraterone is still limited. Therefore, the abiraterone impact on androgen receptor (AR)-positive LNCaP and AR-negative PC-3 cells was assessed by cellular and molecular analyses. The present study demonstrated, that abiraterone treatment significantly decreased cell growth, AR expression, and AR activity of AR-positive LNCaP cells. Notably, AR-negative PC-3 cells exhibited comparable reductions in cellular proliferation, associated with DNA fragmentation and pro-apoptotic modulation of p21, caspase-3, survivin, and transforming growth factor β (TGFβ). Our observations suggest that the attenuation of AR signaling is not the only rationale to explain the abiraterone anticancer activity. Abiraterone efficacy may play a more global role in PC progression control than originally hypothesized. In this regard, abiraterone is not only a promising drug for treatment of AR-negative PC stages, even more, abiraterone may represent an alternative for treatment of other malignancies besides prostate cancer.Fil: Grossebrummel, Hannah. University Medicine Greifswald; AlemaniaFil: Peter, Tilmann. University Medicine Greifswald; AlemaniaFil: Mandelkow, Robert. University Medicine Greifswald; AlemaniaFil: Weiss, Martin. University Medicine Greifswald; AlemaniaFil: Muzzio, Damián Oscar. University Medicine Greifswald; ArgentinaFil: Zimmermann, Uwe. University Medicine Greifswald; AlemaniaFil: Walther, Reinhard. University Medicine Greifswald; AlemaniaFil: Jensen, Cristian Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; Argentina. University Medicine Greifswald; AlemaniaFil: Knabbe, Cornelius. Ruhr-universität Bochum; AlemaniaFil: Zygmunt, Marek. University Medicine Greifswald; AlemaniaFil: Burchardt, Martin. University Medicine Greifswald; AlemaniaFil: Stope, Matthias B.. University Medicine Greifswald; Alemani

Funktionalität von B-Lymphozyten in der Schwangerschaft

Introduction: For a successful pregnancy, a set of physiological requirements has to be fulfilled. The mother has to provide enough nutrients and the proper anatomical environment for the developing fetus and protect him and herself against pathogens. The cells of the im-mune system constantly monitor the organism in search for pathogens and mount a response to eradicate the threat. The favourable outcome of an immune response re-lays on the capacity of those cells to recognize structures that shouldn’t be present in the organism and the speed or strength at which the cells react. During pregnancy, however, a fetus is able to establish a firm contact with the endometrium of the mother and then grow for an extended period of time. This “exception to the rule” hides behind a set of fine-tuned regulations of the immune responses which are not completely un-derstood. Though many cell types have been extensively investigated in the past dec-ades, B cells play yet enigmatic roles. The aim of this work is to uncover the events occurring within the B cell development during pregnancy and to study the role of certain subtypes in healthy pregnancy and pregnancy miscarriage. Methods: For all experiments, 8-weeks-old female mice either non-pregnant, having normal preg-nancies or miscarriage were used. Organs were removed and cells isolated using standard protocols. The analysis of the population distribution was performed by Flow Cytometry. For in vitro experiments, specific cell subsets were isolated using MACS Cell Separation. Bio-plex method was used for the assessment of Immunoglobulin isotypes in serum, while CBA Array was the method used to measure cytokine levels in the supernatant of cell cultures. Statistical analysis was done using GraphPad Prism software. Results: Pregnancy had a strong impact on the murine B cell development. The restructuration of the B cell compartment could be appreciated already from the bone marrow progeni-tors, reduced in pregnant mice. Peripheral subsets drastically adapted their develop-mental pathways, with a drift towards the generation of marginal zone B cells. B cells also showed functional adaptations to gravidity, as evidenced by the changes in the immunoglobulin production and immunomodulatory capacity. Conclusions: For the first time a deep investigation of the consequences of pregnancy on the B cell development was performed, covering several aspects of B cell functionality. This work shows that B lymphocyte compartment is remodelled during pregnancy. Aberration of this process may lead to pregnancy complications including miscarriage.Einleitung: Um eine erfolgreiche Schwangerschaft zu ermöglichen, müssen bestimmte physiologische Anforderungen erfüllt werden. Die Mutter muss sowohl genügend Nährstoffe sowie eine gute anatomische Umgebung für die Entwicklung des Fötus bereitstellen, als auch ihn und sich gegen Krankheitserreger schützen. Die Zellen des Immunsystems überwachen ständig den Organismus auf der Suche nach Krankheitserregern um diese bei einer Bedrohung zu beseitigen. Eine erfolgreiche Immunantwort benötigt Zellen, mit der Fähigkeit Organismusfremde Strukturen zu erkennen und mit der passenden Geschwindigkeit und Stärke dagegen zu reagieren. Während der Schwangerschaft ist ein Fötus trotzdem in der Lage, einen festen Kontakt mit dem Endometrium der Mutter aufzubauen und dann über einen längeren Zeitraum zu wachsen. Diese "Ausnahme von der Regel" versteckt sich hinter einer Reihe von fein abgestimmten Regulierungen der Immunreaktionen, die nicht vollständig verstanden sind. Obwohl viele Zelltypen in den letzten Jahrzehnten ausführlich untersucht wurden, spielen B-Zellen eine noch bisher unbekannte Rolle. Das Ziel dieser Arbeit ist es, die Ereignisse innerhalb der B-Zell-Entwicklung während der Schwangerschaft aufzudecken und die Rolle bestimmter Subtypen in gesunder Schwangerschaft und Schwangerschaft mit Fehlgeburt zu untersuchen. Methoden: Bei allen Experimenten wurden 8-Wochen-alte weibliche Mäuse entweder nicht schwanger, mit normalen Schwangerschaft oder mit Fehlgeburt verwendet. Organe wurden entfernt und die Zellen isoliert unter Nutzung von Standardprotokollen. Die Analyse der Verteilung der Grundgesamtheit wurde mittels Durchflusszytometrie durchgeführt. Für in-vitro-Experimente wurden spezifische Zell-Untergruppen mit MACS Zellseparation isoliert. Eine Bio-plex-Methode wurde für die Bestimmung der Immunoglobulin-Isotypen im Serum verwendet, ein CBA Array wurde zur Zytokinkonzentration im Überstand der Zellkulturen benutzt. Die statistische Analyse wurde nach Verwendung von GraphPad Prism Software durchgeführt. Ergebnisse: Die Schwangerschaft hatte einen starken Einfluss auf die Maus-B-Zell-Entwicklung. Die Umstrukturierung des B-Zell-Kompartiments konnte bereits aus der Reduktion der Knochenmarkvorläuferzellen in schwangeren Mäusen beobachtet werden. Periphere Untergruppen adaptierten drastisch ihrer Entwicklungswege mit einer Drift Richtung der Reifung zu Marginalzonen-B-Zellen. B-Zellen zeigten auch funktionale Anpassungen an Gravidität, Veränderungen in der Immunglobulinproduktion sowie immunmodulatorische Fähigkeiten. Schlussfolgerungen: Zum ersten Mal wurde eine tiefe Untersuchung zum Einfluss der Schwangerschaft auf die B-Zell-Entwicklung durchgeführt, die mehrere Aspekte der B-Zell-Funktionen umfasst. Diese Arbeit zeigt, dass B-Lymphozyten-Kompartiment wurde während der Schwangerschaft umgebaut. Abweichungen bei diesem Prozess können zu Schwangerschaftskomplikationen einschließlich Fehlgeburten führen

Oxygen regulates ILC3 antigen presentation potential and pregnancy-related hormone actions

Early pregnancy is marked by placentation and embryogenesis, which take place under physiological low oxygen concentrations. This oxygen condition is crucial for many aspects of placentation, trophoblast function, vascularization and immune function. Recently, a new family of innate lymphoid cells has been found to be expressed at the fetomaternal interface. Among these, type 3 innate lymphoid cells (ILC3) are important antigen presenting cells in the context of MHC-II. The expression of MHC-II on ILC3s during pregnancy is reduced. We tested the hypothesis that low oxygen concentrations reduce the potential of ILC3s to present antigens promoting fetal tolerance. Using an in vitro approach, NCR+ ILC3s generated from cord blood stem cell precursors were incubated under different O2 concentrations in the presence or absence of the pregnancy-related hormones hCG and TGF-β1. The expression of MHC-II, accessory molecules and an activation marker were assessed by flow cytometry. We observed that 1% O2 reduced the expression of the MHC-II molecule HLA-DR as compared to 21% O2 and modulated the relative effects of hCG and TGF-β1. Our data indicate that low oxygen concentrations reduce the antigen presentation potential of NCR+ ILC3s and suggest that it may promote fetal tolerance during the first trimester of pregnancy

Progesterone and estradiol exert an inhibitory effect on the production of anti-inflammatory cytokine IL-10 by activated MZ B cells

The main message of this work is the fact that female sex hormones, progesterone and estradiol, whose levels significantly rise during pregnancy, inhibit the production of anti-inflammatory cytokine IL-10 with no apparent effect on pro-inflammatory cytokine TNF-α by activated MZ B cells. This is an important piece of information and helps to better understand how the maternal immune system controls the balance between immune tolerance and immune activation during pregnancy leading to the simultaneously acceptance of the semi-allogeneic fetus and the proper defense of the mother against pathogens during this critical period of time.Fil: Bommer, Imke. University of Greifswald; AlemaniaFil: Muzzio, Damián Oscar. University of Greifswald; AlemaniaFil: Zygmunt, Marek. University of Greifswald; AlemaniaFil: Jensen, Cristian Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; Argentin

3D in vitro models of early pregnancy: How to choose the right scaffolding material?

Following fertilization, the blastocyst has to complete two distinct steps to assure further development of pregnancy. After apposition it establishes a firm connection with the luminal epithelium of the endometrium (attachment) and subsequently enters the decidualizing stroma (invasion). If this step is not achieved successfully, fertility problems arise. Development of the placenta ensures an adequate supply of nutrients and gas between the mother and the fetus. Preeclampsia is a prevalent disorder arising from defects in the process of placentation. It is associated with an increase of maternal morbidity and mortality. Numerous attempts have been made in order to elucidate the etiology of the syndrome and identify women at risk. The lack of reliable animal models has turned the attention to the development of in vitro assays, which could provide a better insight into the individual processes that will later trigger preeclampsia symptoms. In particular, 3D in vitro models more closely resemble the complexity of the extracellular environment. The choice of the scaffolding material should be done carefully as cell-matrix interactions are very often as important as cell-cell interactions for the correct attachment, proliferation and differentiation of cells. The following review is aimed to provide a general overview of the scaffolds available for the in vitro modeling of these complicated systems as well as to discuss the importance surrounding the choice of the scaffolding material and its influence on the results obtained.Fil: Muzzio, Damián. Universitätsmedizin Greifswald; AlemaniaFil: Foglia, María Lucía. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Metabolismo del Fármaco. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Metabolismo del Fármaco; ArgentinaFil: Desimone, Martín Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Metabolismo del Fármaco. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Metabolismo del Fármaco; ArgentinaFil: Zygmunt, Marek. Universitätsmedizin Greifswald; Alemani

Expression analysis of cannabinoid receptors 1 and 2 in B cells during pregnancy and their role on cytokine production

The endocannabinoid system consists in a family of lipids that binds to and activates cannabinoid receptors. There are two receptors so far described, the cannabinoid receptor 1 (CB1) and 2 (CB2). In the context of pregnancy, the endocannabinoid system was shown participates in different key aspects of reproductive events. B-lymphocytes are pleiotropic cells belonging to the adaptive arm of the immune system. Besides immunoglobulin production, B-lymphocytes were recently shown to be actively involved in antigen presentation as well as cytokine production, thus playing a central role in immunity. In this study we first aimed to characterize the expression of CB1 and CB2 receptors in B cells during pregnancy and then analyze the impact of their activation in term of cytokine production by B cells from pregnant and non-pregnant mice. We observed that the expression of CB1 and CB2 receptors in B-lymphocytes is differentially regulated during pregnancy. While CB2 expression is down regulated CB1 is augmented in B-lymphocytes of pregnant mice. Additionally, the treatment of activated B-lymphocytes with specific CB1 and CB2 agonists, showed a different response in term of cytokine production. Particularly, CB1 against boosted the production of the anti-inflammatory cytokine IL-10 by activated B-lymphocytes from pregnant mice.Fil: Wolfson, Manuel Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaFil: Muzzio, Damián Oscar. University of Greifswald; AlemaniaFil: Ehrhardt, Jens. University of Greifswald; AlemaniaFil: Franchi, Ana Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaFil: Zygmunt, Marek. University of Greifswald; AlemaniaFil: Jensen, Cristian Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; Argentina. University of Greifswald; Alemania. Universidad Nacional Arturo Jauretche; Argentin