CD11c+ dendritic cells mediate antigen-specific suppression in extracorporeal photopheresis.

Extracorporeal photopheresis (ECP) represents one of the most widespread and effective cell therapies for graft-versus-host disease and other T cell-mediated disorders. However, the key factors affecting the therapeutic efficacy of ECP remain unclear. We hypothesized that therapeutic effects are mediated by ECP-treated antigen-presenting dendritic cells (DC). To test this hypothesis, we used the experimental model of contact hypersensitivity (CHS). The ECP's therapeutic activity improved when the total cell dose of the ECP-treated cells was increased. We used different haptens during sensitization to demonstrate that the anti-inflammatory activity of ECP is antigen-specific. This confirmed the hypothesis that professional antigen-presenting cells are involved in the mode of action. Also, the ECP's therapeutic activity was abrogated by the depletion of CD11c+ DC, which represents fewer than 1% of all the ECP-exposed cells. Finally, we confirm the critical importance of CD11c+ DC for ECP activity by showing that only a few purified CD11c+ DC are sufficient to mediate its therapeutic effect. The finding that ECP-treated, physiological antigen-presenting DC alone mediate antigen-specific modulation of a pathological immune response may result in better-targeted interventions when treating patients.

Expression patterns of Notch receptors and their ligands Jagged and Delta in human placenta.

The establishment of an appropriate fetomaternal vessel system is a prerequisite for prevention of pregnancy associated pathologies. Notch receptors and ligands are manifoldly involved in vascular development and angiogenesis. To further characterize the process of human placental vasculo- and angiogenesis we investigated the expression pattern of Notch receptors and their ligands during pregnancy. Real time RT-PCR, immunohistochemistry and flow cytometry analysis were performed in early (6-12) weeks of gestation (w.o.g.) and late placenta (37-41 w.o.g.). To specify the exact cellular localization immunofluorescent labelling of epithelial and endothelial cells (EC), respectively, with cytokeratin-7 and vonWillebrand factor (vWF) was done. One placenta from a patient with Alagille syndrome (AGS) was examined with real time RT-PCR and immunohistochemistry. The receptors Notch2, -3, -4 and their ligands Jagged1, -2 and Delta1, -4 were detected at both the mRNA and protein level in early and late placenta. Notch1 was only detected at protein level. The expression was found mainly in the stromal compartment: placental EC expressed Notch1, Delta4, Jagged1 and Delta1. A strong Jagged1 expression was found in the endothelium of arteries and veins supporting a role in differentiation of capillaries. Hofbauer cells (HC) primarily displayed the receptors Notch2, -3 and -4. Placental stromal cells (SC) were positive for Jagged2. The syncytiotrophoblast (ST) and cytotrophoblast (CT) cells revealed a weak but detectable co-localization with cytokeratin-7 and Notch1, -3 and Delta1. These results were verified by flow cytometry of freshly isolated placental cells of placental tissue. Interestingly Jagged1 expression was absent in endothelial cells from an AGS placenta. The Notch receptors and their ligands are expressed in human placental ST, CT, EC, SC and HC. The distribution pattern of Notch receptors and their ligands suggests their involvement in the process of placental vasculo- and angiogenesis via cell-cell communication between trophoblast, -stroma and endothelial cells.

How to study placental vascular development?

Both exogenous and endogenous factors during pregnancy may impact placental vascular development and cause different malformations of placental vessels. In humans, consequences of abnormal vascular development have been associated with different pregnancy-related pathologies ranging from miscarriage to intrauterine growth restriction or preeclampsia. Pregnancy-associated exposure to bacterial or viral infections or pharmacologic or toxic agents may also influence vascular development of the placenta and lead to preterm labor and delivery. Several steps of vascular adaptation on both the fetal and maternal side are necessary and include such events as uterine vasodilation, remodeling by extravillous trophoblast, as well as vasculogenesis and angiogenesis within the placenta. Ubiquitous as well as pregnancy-specific angiogenic factors are involved. Morphologic and stereologic approaches, as well as experiments in established laboratory animals, cannot be applied to large domestic animals or humans without hesitation. Thus, further studies into the different aspects of this process will require an appropriate in vitro model of placental vascular development. Reflecting the core of placental vascular development, the in vitro model should facilitate the interactions between trophoblast and stromal cells with endothelial progenitor cells. The effects of viral or bacterial infection as well as pharmacologic or toxic agents may be studied more closely in the process. This report reviews major aspects of vascular development in the placenta and describes the establishment of a three-dimensional in vitro model of human placental vascular development.

Studies of placental vasculogenesis: a way to understand pregnancy pathology?

Undisturbed development and growth of the fetus depends on an adequate vascular development in the fetomaternal unit. Several steps of vascular adaptation both on fetal and maternal side are necessary and involve uterine vasodilation and remodelling by extravillous trophoblast as well as vasculo- and angiogenesis within the placenta. Ubiquitous (e. g. VEGF, bFGF) as well as pregnancy specific (PlGF, hCG, IGF-II, AFP) angiogenic factors are involved. Consequences of abnormal vascular development have been associated with different pregnancy-related pathologies ranging from miscarriage to intrauterine growth restriction or preeclampsia. Pregnancy-associated exposure to bacterial and viral infections or toxic agents (e. g. alcohol, nicotine or drugs) may also influence vascular development of the placenta and often lead to preterm labour and delivery. Different methods of study placental vascular development exist. Morphological and stereological approaches as well as animal models reveal significant limitations and can not be applied to in vivo human situation without hesitation. There is a need to design in vitro models of human placental vascular development allowing studies into the different aspects of this process including: trophoblast and stromal cells, interaction with endothelial progenitor cells, influence of viral or bacterial infection of trophoblast as well as influence of toxic agents. Our manuscript reviews major aspects of vascular development in the placenta and describes author's efforts to establish a three-dimensional model of this process in vitro.

The gonadotropins: tissue-specific angiogenic factors?

The gonadotropins, whose members are human chorionic gonadotropin (hCG), lutenizing hormone (LH) and follicle-stimulating hormone (FSH) are a well characterized hormone family known to regulate reproductive functions in both females and males. Recent studies indicate that they can modulate the vascular system of reproductive organs. It was shown that gonadotropins not only influence the expression of vascular endothelial growth factor (VEGF) and both its receptors VEGFR-1 and -2, but also modulate other ubiquitously expressed angiogenic factors like the angiopoietins and their receptor Tie-2, basic fibroblast growth factor or placental-derived growth factor. Some recent data indicates a possible direct action of gonadotropins on endothelial cells. Thus, the gonadotropins act as tissue-specific angiogenic factors providing an optimal vascular supply during the menstrual cycle and early pregnancy in the female reproductive tract as well as in testis. In pathological conditions (e.g. preeclampsia, intrauterine growth restriction, ovarian hyperstimulation or endometriosis), these tightly regulated interactions between the gonadotropins and the ubiquitous angiogenic factors appear to be disturbed. The intent of this short manuscript is to review the current knowledge of the regulatory role of the gonadotropins in vasculo- and angiogenesis. We also review angiogenic actions of thyroid-stimulating hormone (TSH), a glycoprotein closely related to gonadotropins, which display strong gonodal actions.

HCG in the regulation of placental angiogenesis. Results of an in vitro study.

Placental vascular development is essential for fetal growth and development. Inadequate placental vascular development is associated with early pregnancy losses and other pregnancy related pathologies. In addition to the ubiquitous, well-characterized angiogenic factors like vascular endothelial growth factor (VEGF) or basic fibroblast growth factor (bFGF), some pregnancy-specific factors (e.g. human chorionic gonadotropin (hCG), insulin-like growth factor-II (IGF-II) or alpha fetoprotein (AFP) were recently described to play a possible regulatory role in this process. In the present study we described an improved separation method for human placental microvascular endothelial cells (HPMVEC) and their functional characterization. Using the combination of enzymatic digestion and multistep immunomagnetic sorting with CD31 antibodies a model for villous vascularization was established. Isolated cells took up ac-dil-LDL, spontaneously formed capillary-like structures, and expressed common endothelial markers such as vascular endothelial growth factor receptor-2 (VEGFR-2), angiopoetin-1 and -2, Tie-2, CD144, thrombomodulin, and von Willebrand factor (vWF) as shown by RT-PCR, flow cytometry and Western blot analysis. The expression of the hCG/LH receptor in the placental vascular tree was verified both in vitro and in vivo. hCG stimulated proliferation of HPMVEC in a dose specific manner. While hCG alone had no significant effect on endothelial cell apoptosis, the combination of VEGF-A and hCG protected HPMVEC from staurosporine-induced apoptosis. hCG significantly stimulated sprout formation when compared to controls in a spheroid angiogenesis assay. Our results demonstrate a modified and reproducible method allowing studies of placental vascular development and provide new insights into the possible role of trophoblastic factors in this process.