Bettina Toth

Recurrent miscarriage: current concepts in diagnosis and treatment

Although recurrent miscarriage (RM) affects only 1-3% of couples, it has a major influence on the wellbeing and psychosocial status of patients. Therefore, research into improved diagnosis and development of new treatment strategies is essential. In this review, we summarize current concepts on diagnosis and treatment in RM, drawing upon research reports and international guidelines to provide insights into the pathophysiology of pregnancy disrupted by repeated miscarriage. Anatomical malformations, infectious diseases, endocrine disorders, autoimmune defects as well as acquired and inherited thrombophilia are established risk factors in RM. In addition, our recent findings indicate an impact on miscarriage incidence of glycoproteins such as glycodelin, and nuclear hormone receptors such as the peroxisome proliferator-activated receptors (PPARs). Significantly reduced glycodelin expression is associated with miscarriage, whereas up-regulation of PPARs appears to compensate for either the activated immune response or the disturbed cytotrophoblast differentiation in RM patients. There is also evidence that circulating placental microparticles are increased in a subgroup of RM patients, indicating an acquired procoagulant state even outside pregnancy. Treatment strategies like aspirin and low molecular weight heparin (LMWH) are standard medications in RM, although only a few placebo-controlled trials have proven their benefit in respect to live birth rate. There is emerging evidence that new treatment options, including drugs like TNFalpha inhibitors and granulocyte colony-stimulating factor (G-CSF) might be beneficial in some cases of RM. However, larger clinical trials must be completed to further prove or disprove benefits of these drugs in the treatment of RM patients.

Platelet-derived microparticles and coagulation activation in breast cancer patients

In the mid 1800s Trousseau observed cancer-associated thrombosis, of which the underlying pathogenesis still remains unknown. We performed a prospective study on platelet-derived microparticles (PMP) and their procoagulant potential in breast cancer patients. Fifty-eight breast cancer patients and 13 women with benign breast tumors were included in the study. Microparticles (MP) were examined by electron microscopy and FACS analysis using labels for annexin V (total numbers), CD61 (PMP), CD62P and CD63 (activated platelets), CD62E (endothelial cells), CD45 (leukocytes) as well as CD142 (tissue factor). Prothrombin fragment 1+2 (F1+2) and thrombin generation were measured as blood coagulation markers. Numbers of annexin V+-MP were highest in breast cancer patients with larger tumor size (T2; median = 5,637 x 10(6)/l; range = 2,852-8,613) and patients with distant metastases (M1; median = 6,102 x 10(6)/l; range = 3,350-7,445), and differed significantly from patients with in-situ tumor (Tis; median = 3,220 x 10(6)/l; range = 2,277-4,124; p = 0.019), small tumor size (T1; median = 3,281 x 10(6)/l; range = 2,356-4,861; p = 0.043) and women with benign breast tumor (median = 4,108 x 10(6)/l; range = 2,530-4,874; p = 0.040). A total of 82.3% of MP were from platelets, 14.6 % from endothelial cells and 0.3% from leukocytes. Less than 10% of PMP showed degranulation markers. Larger tumor size (T2) and metastases correlated with high counts of PMP and with highest F1+2 levels. Since prothrombin levels and thrombin generation did not parallel MP levels, we speculate that MP act in the microenvironment of tumor tissue and may thus not be an exclusive parameter reflecting in-vivo procoagulant activity.

New approaches to embryo selection

Embryo selection has been an important topic since the introduction of assisted reproduction, with embryo morphology being the most obvious criterion. Although morphology serves as indicator for overall IVF laboratory quality, its statistical assessment limits the possibility to identify the most implantation-competent embryos. In order to reach a direct picture of the developing embryo, invasive procedures such as preimplantation genetic screening or transcriptome and proteome analysis of biopsied embryonic tissue were initially prioritized and are still under investigation. More recently, focus has shifted towards noninvasive techniques that maintain the integrity of the embryo. Metabolomic profiling of culture medium from growing embryos attracted much research. Although successful in a pilot study, that approach failed in a randomized controlled trial. Other metabolomics studies are on their way but not yet available for routine clinical use. The most promising strategy at present is the combined evaluation of morphology and developmental kinetics using time-lapse imaging. This has brought new insights into certain characteristics that enable deselection of embryos at an early stage of development and to identify others with high potential for successful implantation. However, there is still considerable room for improvement. Further strategies will most likely involve the combination of several different approaches.

Governing the invasive trophoblast: current aspects on intra- and extracellular regulation.

Citation Fitzgerald JS, Germeyer A, Huppertz B, Jeschke U, Knöfler M, Moser G, Scholz C, Sonderegger S, Toth B, Markert UR. Governing the invasive trophoblast: current aspects on intra‐ and extracellular regulation. Am J Reprod Immunol 2010

Endothelial cell-derived microparticles in allogeneic hematopoietic stem cell recipients

Background. Alterations of microparticles derived from different cell types are described in a number of diseases associated with inflammation and hemostatic disorders. Methods. In this prospective study, we firstly analyzed endothelial cell derived microparticles (EMP) in 19 hematopoietic stem cell recipients. Cultured human umbilical vein endothelial cells (HUVEC) stimulated with tumor necrosis factor-alpha (TNF-alpha) served as positive controls. EMP were analyzed by fluorescent activated cell sorting (FACS), detecting the particels via expression of CD62 (E-selectin) and anionic phospholipids binding to annexin V. Results. EMP were not significantly influenced by conditioning regimens with non-myeloablative chemotherapy and 4 Gy total body irradiation (TBI) or by myeloablative regimens containing 12 Gy TBI. During acute graft versus host disease (aGVHD), significantly higher levels of EMP were detected than in patients without aGVHD (18.5/&mgr;l s=10.1 vs. 14.6/&mgr;l SD=11.5; P=0.004) while infectious complications did not alter EMP levels significantly. Immunosuppressive therapy with corticosteroids tendentially elevated EMP levels. HUVEC treated with TNF-alpha 1 ng/ml, 10 ng/ml and 100 ng/ml released significantly more EMP than unstimulated cultures (30.0/&mgr;l ss=13.6 vs. 126.8/&mgr;l SD=66.9, P=0.032 / vs. 683.3/&mgr;l SD=349.9; P=0.03 / vs. 489.3 s=184.4; P=0.013). Conclusions. Elevation of EMP during aGVHD might express severe endothelial cell injury within this complication after hematopoietic stem cell transplantation and might serve as a diagnostic test for early differentiation of aGVHD from other transplanted related complications.

REVIEW ARTICLE: Governing the Invasive Trophoblast: Current Aspects on Intra- and Extracellular Regulation

Citation Fitzgerald JS, Germeyer A, Huppertz B, Jeschke U, Knöfler M, Moser G, Scholz C, Sonderegger S, Toth B, Markert UR. Governing the invasive trophoblast: current aspects on intra‐ and extracellular regulation. Am J Reprod Immunol 2010

The Association between Vitamin D Receptor Expression and Prolonged Overall Survival in Breast Cancer

In this study, we analyzed vitamin D receptor (VDR) expression and survival in a breast cancer patient cohort of 82 breast cancer patients. Immunohistochemical analysis was possible in 91.5% of the patients (75/82). Staining was evaluated using the semi-quantitative assay according to Remmele and Stegner (immunoreactivity score [IRS]). IRS 0–1 was negative/very low, IRS 2–4 was moderate to high, and IRS 6–12 was high. Statistical analysis was performed by Spearman’s correlation test (p<0.05 significant). Overall survival was analyzed using Kaplan-Meier estimations. Only 6 patients had a negative IRS. Moderate IRS values were present in 20 patients. Most of the patients had a high IRS (49). For survival analysis, data were dichotomized (IRS 0–4: negative to moderate and IRS 6–12: high VDR expression). In univariate analysis, VDR expression showed significant differences in progression-free survival (PFS) and overall survival (OS). Patients with high IRS scores showed significantly better PFS and OS than patients with moderate/negative IRS scores for VDR expression. Tumor size was significantly correlated to PFS. When analyzed separately, the three different IRS groups showed significant differences in VDR expression. The present data suggest that VDR expression in breast cancer tissue may be of clinical significance, and the results provide evidence that VDR may be a factor with prognostic relevance.

Polar Body Biopsy

Polar body biopsy combined with array comparative genomic hybridization allows detection of maternal chromosomal aberrations. Although it has limitations, it can be seen as an alternative to blastomere and trophectoderm biopsy.

Recent advances in understanding immunology of reproductive failure.

Aspects of the immunological relationship between mother and conceptus still remain a mystery, although the recent advances in molecular biology have enlightened some of the parameters that participate in fetomaternal cross-talk during implantation. The atypical expression of major histocompatibility complex (MHC), the specific role of some hormones and cytokines, as well as the temporal and spatial distributions of uterine natural killer cells, represent substantive parameters of fetomaternal immunotolerance during implantation. Although human maternal and fetal immunology is difficult to investigate, aberrant immune responses and an imbalanced cytokine network may be related to infertility, implantation failures after IVF and recurrent pregnancy losses. In this review, immunological and interacting factors involved in human reproductive failure are summarized and critically evaluated.

Apheresis platelet concentrates contain platelet-derived and endothelial cell-derived microparticles.

Background and Objectives  Microparticles (MP) are membrane vesicles with thrombogenic and immunomodulatory properties. We determined MP subgroups from resting platelets, activated platelets and endothelial cells in donors and apheresis platelet concentrates (PC).